The uses of AFLP for detecting DNA polymorphism, genotype identification and genetic diversity between yeasts isolated from Mexican agave-distilled beverages and from grape musts.

PubMed

Flores Berrios, E P; Alba González, J F; Arrizon Gaviño, J P; Romano, P; Capece, A; Gschaedler Mathis, A

2005-01-01

The objectives were to determine the variability and to compare the genetic diversity obtained using amplified fragment length polymorphism (AFLP) markers in analyses of wine, tequila, mezcal, sotol and raicilla yeasts. A molecular characterization of yeasts isolated from Mexican agave musts, has been performed by AFLP marker analysis, using reference wine strains from Italian and South African regions. A direct co-relation between genetic profile, origin and fermentation process of strains was found especially in strains isolated from agave must. In addition, unique molecular markers were obtained for all the strains using six combination primers, confirming the discriminatory power of AFLP markers. This is the first report of molecular characterization between yeasts isolated from different Mexican traditional agave-distilled beverages, which shows high genetic differences with respect to wine strains.

Genetic characterization of three varieties of Astragalus lentiginosus (Fabaceae).

Treesearch

Brian J. Knaus; Rich C. Cronn; Aaron Liston

2005-01-01

Astragalus lentiginosus is a polymorphic species that occurs in geologically young habitats and whose varietal circumscription implies active morphological and genetic differentiation. In this preliminary study, we evaluate the potential of amplified fragment length polymorphism (AFLP) markers to resolve infraspecific taxa in three varieties of...

Characterization of North American Armillaria species: Genetic relationships determined by ribosomal DNA sequences and AFLP markers

Treesearch

M. -S. Kim; N. B. Klopfenstein; J. W. Hanna; G. I. McDonald

2006-01-01

Phylogenetic and genetic relationships among 10 North American Armillaria species were analysed using sequence data from ribosomal DNA (rDNA), including intergenic spacer (IGS-1), internal transcribed spacers with associated 5.8S (ITS + 5.8S), and nuclear large subunit rDNA (nLSU), and amplified fragment length polymorphism (AFLP) markers. Based on rDNA sequence data,...

Epigenetic variation predicts regional and local intraspecific functional diversity in a perennial herb.

PubMed

Medrano, Mónica; Herrera, Carlos M; Bazaga, Pilar

2014-10-01

The ecological significance of epigenetic variation has been generally inferred from studies on model plants under artificial conditions, but the importance of epigenetic differences between individuals as a source of intraspecific diversity in natural plant populations remains essentially unknown. This study investigates the relationship between epigenetic variation and functional plant diversity by conducting epigenetic (methylation-sensitive amplified fragment length polymorphisms, MSAP) and genetic (amplified fragment length polymorphisms, AFLP) marker-trait association analyses for 20 whole-plant, leaf and regenerative functional traits in a large sample of wild-growing plants of the perennial herb Helleborus foetidus from ten sampling sites in south-eastern Spain. Plants differed widely in functional characteristics, and exhibited greater epigenetic than genetic diversity, as shown by per cent polymorphism of MSAP fragments (92%) or markers (69%) greatly exceeding that for AFLP ones (41%). After controlling for genetic structuring and possible cryptic relatedness, every functional trait considered exhibited a significant association with at least one AFLP or MSAP marker. A total of 27 MSAP (13.0% of total) and 12 AFLP (4.4%) markers were involved in significant associations, which explained on average 8.2% and 8.0% of trait variance, respectively. Individual MSAP markers were more likely to be associated with functional traits than AFLP markers. Between-site differences in multivariate functional diversity were directly related to variation in multilocus epigenetic diversity after multilocus genetic diversity was statistically accounted for. Results suggest that epigenetic variation can be an important source of intraspecific functional diversity in H. foetidus, possibly endowing this species with the capacity to exploit a broad range of ecological conditions despite its modest genetic diversity. © 2014 John Wiley & Sons Ltd.

Genetic and epigenetic status of triple exotic consanguinity cotton introgression lines.

PubMed

He, S P; Sun, J L; Du, X M

2011-10-03

Introgression lines are some of the most important germplasm for breeding applications and other research conducted on cotton crops. The DNA methylation level among 10 introgression lines of cotton (Gossypium hirsutum) and three exotic parental species (G. arboreum, G. thurberi and G. barbadense) were assessed by methylation-sensitive amplified polymorphism (MSAP) technology. The methylation level in the introgression lines ranged from 33.3 to 51.5%. However, the lines PD0111 and PD0113 had the lowest methylation level (34.6 and 33.3%, respectively) due to demethylation of most non-coding sequences. Amplified fragment length polymorphism (AFLP) was used to evaluate the genetic polymorphism in the cotton introgression lines. A high degree of polymorphism was observed in all introgression lines (mean 47.2%) based on AFLP and MSAP analyses. This confirmed the effects of genetic improvement on cotton introgression lines. The low methylation varieties, PD0111 and PD0113 (introgression lines), clustered outside of the introgression lines based on MSAP data, which was incongruent with an AFLP-based dendrogram. This phenomenon could be caused by environmental changes or introgression of exotic DNA fragments.

DNA damage and genetic methylation changes caused by Cd in Arabidopsis thaliana seedlings.

PubMed

Li, Zhaoling; Liu, Zhihong; Chen, Ruijuan; Li, Xiaojun; Tai, Peidong; Gong, Zongqiang; Jia, Chunyun; Liu, Wan

2015-09-01

Amplified fragment length polymorphism (AFLP) and methylation-sensitive amplification polymorphism (MASP) techniques are sensitive to deoxyribonucleic acid (DNA) damage and genetic methylation, respectively. Using these 2 techniques, Arabidopsis thaliana cultured with 0 mg/L (control), 0.5 mg/L, 1.5 mg/L, and 5.0 mg/L Cd(2+) for 16 d was used to analyze the DNA damage and methylation changes as a result of cadmium (Cd). The DNA was amplified by 14 AFLP primer pairs and 13 MSAP primer combinations. In the AFLP experiment, 62 polymorphic sites were found in the patterns of 11 primer combinations and a total of 1116 fragments were obtained in these patterns. There were no polymorphic bands in the remaining 3 pairs. The proportions of polymorphic sites in the 0.5-mg/L Cd(2+) and 5.0-mg/L Cd(2+) treatments were significantly different. Seven polymorphic fragments were then separated and successfully sequenced, yielding 6 nucleobase substitutions and 1 nucleobase deletion. Similarly, in the MSAP experiment, the MSAP% and number of demethylated-type bands were unchanged after Cd treatment, but the number of methylated-type bands was increased significantly in the 5.0-mg/L Cd(2+) treatment group, a finding that may be associated with the AFLP results. The polymorphic bands were also sequenced and the functions of their homologous genes were determined. The DNA damage and methylation changes may be the primary cause of certain pathology changes as a result of Cd uptake in plants. © 2015 SETAC.

Photobacterium damselae ssp. piscicida: detection by direct amplification of 16S rRNA gene sequences and genotypic variation as determined by amplified fragment length polymorphism (AFLP).

PubMed

Kvitt, H; Ucko, M; Colorni, A; Batargias, C; Zlotkin, A; Knibb, W

2002-04-05

A PCR protocol for the rapid diagnosis of fish 'pasteurellosis' based on 16S rRNA gene sequences was developed. The procedure combines low annealing temperature that detects low titers of Photobacterium damselae but also related species, and high annealing temperature for the specific identification of P. damselae directly from infected fish. The PCR protocol was validated on 19 piscine isolates of P. damselae ssp. piscicida from different geographic regions (Japan, Italy, Spain, Greece and Israel), on spontaneously infected sea bream Sparus aurata and sea bass Dicentrarchus labrax, and on closely related American Type Culture Collection (ATCC) reference strains. PCR using high annealing temperature (64 degrees C) discriminated between P. damselae and closely related reference strains, including P. histaminum. Sixteen isolates of P. damselae ssp. piscicida, 2 P. damselae ssp. piscicida reference strains and 1 P. damselae ssp. damselae reference strain were subjected to Amplified Fragment Length Polymorphism (AFLP) analysis, and a similarity matrix was produced. Accordingly, the Japanese isolates of P. damselae ssp. piscicida were distinguished from the Mediterranean/European isolates at a cut-off value of 83% similarity. A further subclustering at a cut-off value of 97% allowed discrimination between the Israeli P. damselae ssp. piscicida isolates and the other Mediterranean/European isolates. The combination of PCR direct amplification and AFLP provides a 2-step procedure, where P. damselae is rapidly identified at genus level on the basis of its 16S rRNA gene sequence and then grouped into distinct clusters on the basis of AFLP polymorphisms. The first step of direct amplification is highly sensitive and has immediate practical consequences, offering fish farmers a rapid diagnosis, while the AFLP is more specific and detects intraspecific variation which, in our study, also reflected geographic correspondence. Because of its superior discriminative properties, AFLP can be an important tool for epidemiological and taxonomic studies of this highly homogeneous genus.

Genetic Diversity of Hibiscus tiliaceus (Malvaceae) in China Assessed using AFLP Markers

PubMed Central

TANG, TIAN; ZHONG, YANG; JIAN, SHUGUANG; SHI, SUHUA

2003-01-01

Amplified fragment length polymorphism (AFLP) markers were used to investigate the genetic variations within and among nine natural populations of Hibiscus tiliaceus in China. DNA from 145 individuals was amplified with eight primer pairs. No polymorphisms were found among the 20 samples of a marginal population of recent origin probably due to a founder effect. Across the other 125 individuals, 501 of 566 bands (88·5 %) were polymorphic, and 125 unique AFLP phenotypes were observed. Estimates of genetic diversity agreed with life history traits of H. tiliaceus and geographical distribution. AMOVA analysis revealed that most genetic diversity resided within populations (84·8 %), which corresponded to results reported for outcrossing plants. The indirect estimate of gene flow based on ϕST was moderate (Nm = 1·395). Long-distance dispersal of floating seeds and local environments may play an important role in shaping the genetic diversity of the population and the genetic structure of this species. PMID:12930729

Amplified fragment length polymorphism analysis to assess crossover interference and homozygosity in gynogenetic diploid Pacific abalone (Haliotis discus hannai).

PubMed

Nie, H-T; Li, Q; Kong, L-F

2014-06-01

Recombination analysis in gynogenetic diploids is a powerful tool for assessing the degree of inbreeding, investigating crossover events and understanding chiasma interference during meiosis. To estimate the marker-centromere recombination rate, the inheritance pattern of 654 amplified fragment length polymorphism (AFLP) markers was examined in the 72-h veliger larvae of two meiogynogenetic diploid families in the Pacific abalone (Haliotis discus hannai). The second-division segregation frequency (y) of the AFLP loci ranged from 0.00 to 0.96, with 23.9% of loci showing y-values higher than 0.67, evidencing the existence of interference. The average recombination frequency across the 654 AFLP loci was 0.45, allowing estimation of the fixation index of 0.55, indicating that meiotic gynogenesis could provide an effective means of rapid inbreeding in the Pacific abalone. The AFLP loci have a small proportion (4.4%) of y-values greater than 0.90, suggesting that a relatively low or intermediate degree of chiasma interference occurred in the abalone chromosomes. The information obtained in this study will enhance our understanding of the abalone genome and will be useful for genetic studies in the species. © 2014 Stichting International Foundation for Animal Genetics.

Using amplified fragment length polymorphism analysis to differentiate isolates of Pasteurella multocida serotype 1

USGS Publications Warehouse

Blehert, D.S.; Jefferson, K.L.; Heisey, D.M.; Samuel, M.D.; Berlowski, B.M.; Shadduck, D.J.

2008-01-01

Avian cholera, an infectious disease caused by the bacterium Pasteurella multocida, kills thousands of North American wild waterfowl annually. Pasteurella multocida serotype 1 isolates cultured during a laboratory challenge study of Mallards (Anas platyrhynchos) and collected from wild birds and environmental samples during avian cholera outbreaks were characterized using amplified fragment length polymorphism (AFLP) analysis, a whole-genome DNA fingerprinting technique. Comparison of the AFLP profiles of 53 isolates from the laboratory challenge demonstrated that P. multocida underwent genetic changes during a 3-mo period. Analysis of 120 P. multocida serotype 1 isolates collected from wild birds and environmental samples revealed that isolates were distinguishable from one another based on regional and temporal genetic characteristics. Thus, AFLP analysis had the ability to distinguish P. multocida isolates of the same serotype by detecting spatiotemporal genetic changes and provides a tool to advance the study of avian cholera epidemiology. Further application of AFLP technology to the examination of wild bird avian cholera outbreaks may facilitate more effective management of this disease by providing the potential to investigate correlations between virulence and P. multocida genotypes, to identify affiliations between bird species and bacterial genotypes, and to elucidate the role of specific bird species in disease transmission. ?? Wildlife Disease Association 2008.

Hybridization in the section Mentha (Lamiaceae) inferred from AFLP markers.

PubMed

Gobert, V; Moja, S; Colson, M; Taberlet, P

2002-12-01

The amplified fragment length polymorphism (AFLP) method was used to evaluate genetic diversity and to assess genetic relationships within the section Mentha in order to clarify the taxonomy of several interspecific mint hybrids with molecular markers. To this end, genetic diversity of 62 Mentha accessions from different geographic origins, representing five species and three hybrids, was assessed. Three EcoRI/MseI AFLP primer combinations generated an average of 40 AFLP markers per primer combination, ranging in size from 50 to 500 base pairs (bp). The percentage of markers polymorphic ranged from 50% to 60% across all accessions studied. According to phenetic and cladistic analysis, the 62 mint accessions were grouped into two major clusters. Principal coordinates analysis separated species into well-defined groups, and clear relationships between species and hybrids could be described. Our AFLP analysis supports taxonomic classification established among Mentha species by conventional (morphological, cytological, and chemical) methods. It allows the assessment of phenetic relationships between species and the hybrids M. spicata and M. × piperita, largely cultivated all over the world for their menthol source, and provides new insights into the subdivision of M. spicata, based for the first time on molecular markers.

An analysis on DNA fingerprints of thirty papaya cultivars (Carica papaya L.), grown in Thailand with the use of amplified fragment length polymorphisms technique.

PubMed

Ratchadaporn, Janthasri; Sureeporn, Katengam; Khumcha, U

2007-09-15

The experiment was carried out at the Department of Horticulture, Ubon Ratchathani University, Ubon Ratchathani province, Northeast Thailand during June 2002 to May 2003 aims to identify DNA fingerprints of thirty papaya cultivars with the use of Amplified Fragment Length Polymorphisms (AFLP) technique. Papaya cultivars were collected from six different research centers in Thailand. Papaya plants of each cultivar were grown under field conditions up to four months then leaf numbers 2 and 3 of each cultivar (counted from top) were chosen for DNA extraction and the samples were used for AFLP analysis. Out of 64 random primers being used, 55 pairs gave an increase in DNA bands but only 12 pairs of random primers were randomly chosen for the final analysis of the experiment. The results showed that AFLP markers gave Polymorphic Information Contents (PIC) of three ranges i.e., AFLP markers of 235 lied on a PIC range of 0.003-0.05, 47 for a PIC range of 0.15-0.20 and 12 for a PIC range of 0.35-0.40. The results on dendrogram cluster analysis revealed that the thirty papaya cultivars were classified into six groups i.e., (1) Kaeg Dum and Malador (2) Kaeg Nuan (3) Pakchong and Solo (4) Taiwan (5) Co Coa Hai Nan and (6) Sitong. Nevertheless, in spite of the six papaya groups all papaya cultivars were genetically related to each other where diversity among the cultivars was not significantly found.

Genetic diversity and relationship of Hedychium from Northeast India as dissected using PCA analysis and hierarchical clustering.

PubMed

Basak, Supriyo; Ramesh, Aadi Moolam; Kesari, Vigya; Parida, Ajay; Mitra, Sudip; Rangan, Latha

2014-12-01

Molecular genetic fingerprints of eleven Hedychium species from Northeast India were developed using PCR based markers. Fifteen inter-simple sequence repeats (ISSRs) and five amplified fragment length polymorphism (AFLP) primers produced 547 polymorphic fragments. Positive correlation (r = 0.46) was observed between the mean genetic similarity and genetic diversity parameters at the inter-species level. AFLP and ISSR markers were able to group the species according to its altitude and intensity of flower aroma. Cophenetic correlation coefficients between the dendrogram and the original similarity matrix were significant for ISSR (r = 0.89) compared to AFLP (r = 0.83) markers. This genetic characterization of Hedychium from Northeast India contributes to the knowledge of genetic structure of the species and can be used to define strategies for their conservation and management.

Isoschizomers and amplified fragment length polymorphism for the detection of specific cytosine methylation changes.

PubMed

Ruiz-García, Leonor; Cabezas, Jose Antonio; de María, Nuria; Cervera, María-Teresa

2010-01-01

Different molecular techniques have been developed to study either the global level of methylated cytosines or methylation at specific gene sequences. One of them is a modification of the Amplified Fragment Length Polymorphism (AFLP) technique that has been used to study methylation of anonymous CCGG sequences in different fungi, plant and animal species. The main variation of this technique is based on the use of isoschizomers with different methylation sensitivity (such as HpaII and MspI) as a frequent cutter restriction enzyme. For each sample, AFLP analysis is performed using both EcoRI/HpaII and EcoRI/MspI digested samples. Comparative analysis between EcoRI/HpaII and EcoRI/MspI fragment patterns allows the identification of two types of polymorphisms: (1) "Methylation-insensitive polymorphisms" that show common EcoRI/HpaII and EcoRI/MspI patterns but are detected as polymorphic amplified fragments among samples; and (2) "Methylation-sensitive polymorphisms" that are associated with amplified fragments differing in their presence or absence or in their intensity between EcoRI/HpaII and EcoRI/MspI patterns. This chapter describes a detailed protocol of this technique and discusses modifications that can be applied to adjust the technology to different species of interest.

AFLP analysis of a worldwide collection of Didymella bryoniae.

PubMed

Kothera, Ronald T; Keinath, Anthony P; Dean, Ralph A; Farnham, Mark W

2003-03-01

Didymella bryoniae (anamorph Phoma cucurbitacearum) is an ascomycete that causes gummy stem blight, a foliar disease that occurs on cucurbits in greenhouses and fields throughout the world. In a previous study using RAPD analysis, little genetic diversity was found among isolates of D. bryoniae from New York and South Carolina, USA. Here we report the use of amplified fragment length polymorphism (AFLP) analysis to assess the genetic variation within a worldwide collection of D. bryoniae, 102 field and greenhouse isolates from ten states in the USA (California, Delaware, Florida, Georgia, Indiana, Maryland, Michigan, Oklahoma, South Carolina, and Texas) and seven other countries (Australia, Canada, China, Greece, Israel, Sweden, and The Netherlands) were examined. Seven different AFLP primer-pair combinations generated 450 bands, of which 134 were polymorphic (30%). Using cluster analysis, two groups and a total of seven subgroups were delineated. Representative isolates varied in their virulence on muskmelon and watermelon seedlings, but the degree of virulence was not strongly associated with AFLP groupings. However, isolates from the northern USA grouped separately from isolates originating from the southern USA.

Amplified fragment length polymorphism of Streptococcus suis strains correlates with their profile of virulence-associated genes and clinical background.

PubMed

Rehm, Thomas; Baums, Christoph G; Strommenger, Birgit; Beyerbach, Martin; Valentin-Weigand, Peter; Goethe, Ralph

2007-01-01

Amplified fragment length polymorphism (AFLP) typing was applied to 116 Streptococcus suis isolates with different clinical backgrounds (invasive/pneumonia/carrier/human) and with known profiles of virulence-associated genes (cps1, -2, -7 and -9, as well as mrp, epf and sly). A dendrogram was generated that allowed identification of two clusters (A and C) with different subclusters (A1, A2, C1 and C2) and two heterogeneous groups of strains (B and D). For comparison, three strains from each AFLP subcluster and group were subjected to multilocus sequence typing (MLST) analysis. The closest relationship and lowest diversity were found for patterns clustering within AFLP subcluster A1, which corresponded with sequence type (ST) complex 1. Strains within subcluster A1 were mainly invasive cps1 and mrp+ epf+ (or epf*) sly+ cps2+ strains of porcine or human origin. A new finding of this study was the clustering of invasive mrp* cps9 isolates within subcluster A2. MLST analysis suggested that A2 correlates with a single ST complex (ST87). In contrast to A1 and A2, subclusters C1 and C2 contained mainly pneumonia isolates of genotype cps7 or cps2 and epf- sly-. In conclusion, this study demonstrates that AFLP allows identification of clusters of S. suis strains with clinical relevance.

Clonal structure in Ichthyobacterium seriolicida, the causative agent of bacterial haemolytic jaundice in yellowtail, Seriola quinqueradiata, inferred from molecular epidemiological analysis.

PubMed

Matsuyama, T; Fukuda, Y; Sakai, T; Tanimoto, N; Nakanishi, M; Nakamura, Y; Takano, T; Nakayasu, C

2017-08-01

Bacterial haemolytic jaundice caused by Ichthyobacterium seriolicida has been responsible for mortality in farmed yellowtail, Seriola quinqueradiata, in western Japan since the 1980s. In this study, polymorphic analysis of I. seriolicida was performed using three molecular methods: amplified fragment length polymorphism (AFLP) analysis, multilocus sequence typing (MLST) and multiple-locus variable-number tandem repeat analysis (MLVA). Twenty-eight isolates were analysed using AFLP, while 31 isolates were examined by MLST and MLVA. No polymorphisms were identified by AFLP analysis using EcoRI and MseI, or by MLST of internal fragments of eight housekeeping genes. However, MLVA revealed variation in repeat numbers of three elements, allowing separation of the isolates into 16 sequence types. The unweighted pair group method using arithmetic averages cluster analysis of the MLVA data identified four major clusters, and all isolates belonged to clonal complexes. It is likely that I. seriolicida populations share a common ancestor, which may be a recently introduced strain. © 2016 John Wiley & Sons Ltd.

Genetic Variation among Isolates of Sarcocystis neurona, the Agent of Protozoal Myeloencephalitis, as Revealed by Amplified Fragment Length Polymorphism Markers

PubMed Central

Elsheikha, H. M.; Schott, H. C.; Mansfield, L. S.

2006-01-01

Sarcocystis neurona causes serious neurological disease in horses and other vertebrates in the Americas. Based on epidemiological data, this parasite has recently emerged. Here, the genetic diversity of Sarcocystis neurona was evaluated using the amplified fragment length polymorphism (AFLP) method. Fifteen S. neurona taxa from different regions collected over the last 10 years were used; six isolates were from clinically diseased horses, eight isolates were from wild-caught opossums (Didelphis virginiana), and one isolate was from a cowbird (Molothrus ater). Additionally, four outgroup taxa were also fingerprinted. Nine primer pairs were used to generate AFLP patterns, with a total number of amplified fragments ranging from 30 to 60, depending on the isolate and primers tested. Based on the presence/absence of amplified AFLP fragments and pairwise similarity values, all the S. neurona isolates tested were clustered in one monophyletic group. No significant correlation could be found between genomic similarity and host origin of the S. neurona isolates. AFLP revealed significant intraspecific genetic variations, and S. neurona appeared as a highly variable species. Furthermore, linkage disequilibrium analysis suggested that S. neurona populations within Michigan have an intermediate type of population structure that includes characteristics of both clonal and panamictic population structures. AFLP is a reliable molecular technique that has provided one of the most informative approaches to ascertain phylogenetic relationships in S. neurona and its closest relatives, allowing them to be clustered by relative similarity using band matching and unweighted pair group method with arithmetic mean analysis, which may be applicable to other related protozoal species. PMID:16714575

Genetic variation among isolates of Sarcocystis neurona, the agent of protozoal myeloencephalitis, as revealed by amplified fragment length polymorphism markers.

PubMed

Elsheikha, H M; Schott, H C; Mansfield, L S

2006-06-01

Sarcocystis neurona causes serious neurological disease in horses and other vertebrates in the Americas. Based on epidemiological data, this parasite has recently emerged. Here, the genetic diversity of Sarcocystis neurona was evaluated using the amplified fragment length polymorphism (AFLP) method. Fifteen S. neurona taxa from different regions collected over the last 10 years were used; six isolates were from clinically diseased horses, eight isolates were from wild-caught opossums (Didelphis virginiana), and one isolate was from a cowbird (Molothrus ater). Additionally, four outgroup taxa were also fingerprinted. Nine primer pairs were used to generate AFLP patterns, with a total number of amplified fragments ranging from 30 to 60, depending on the isolate and primers tested. Based on the presence/absence of amplified AFLP fragments and pairwise similarity values, all the S. neurona isolates tested were clustered in one monophyletic group. No significant correlation could be found between genomic similarity and host origin of the S. neurona isolates. AFLP revealed significant intraspecific genetic variations, and S. neurona appeared as a highly variable species. Furthermore, linkage disequilibrium analysis suggested that S. neurona populations within Michigan have an intermediate type of population structure that includes characteristics of both clonal and panamictic population structures. AFLP is a reliable molecular technique that has provided one of the most informative approaches to ascertain phylogenetic relationships in S. neurona and its closest relatives, allowing them to be clustered by relative similarity using band matching and unweighted pair group method with arithmetic mean analysis, which may be applicable to other related protozoal species.

Genetic and epigenetic variation in Spartina alterniflora following the Deepwater Horizon oil spill.

PubMed

Robertson, Marta; Schrey, Aaron; Shayter, Ashley; Moss, Christina J; Richards, Christina

2017-09-01

Catastrophic events offer unique opportunities to study rapid population response to stress in natural settings. In concert with genetic variation, epigenetic mechanisms may allow populations to persist through severe environmental challenges. In 2010, the Deepwater Horizon oil spill devastated large portions of the coastline along the Gulf of Mexico. However, the foundational salt marsh grass, Spartina alterniflora , showed high resilience to this strong environmental disturbance. Following the spill, we simultaneously examined the genetic and epigenetic structure of recovering populations of S. alterniflora to oil exposure. We quantified genetic and DNA methylation variation using amplified fragment length polymorphism and methylation sensitive fragment length polymorphism (MS-AFLP) to test the hypothesis that response to oil exposure in S. alterniflora resulted in genetically and epigenetically based population differentiation. We found high genetic and epigenetic variation within and among sites and found significant genetic differentiation between contaminated and uncontaminated sites, which may reflect nonrandom mortality in response to oil exposure. Additionally, despite a lack of genomewide patterns in DNA methylation between contaminated and uncontaminated sites, we found five MS-AFLP loci (12% of polymorphic MS-AFLP loci) that were correlated with oil exposure. Overall, our findings support genetically based differentiation correlated with exposure to the oil spill in this system, but also suggest a potential role for epigenetic mechanisms in population differentiation.

AFLP and MS-AFLP Analysis of the Variation within Saffron Crocus (Crocus sativus L.) Germplasm

PubMed Central

Busconi, Matteo; Colli, Licia; Sánchez, Rosa Ana; Santaella, Marcela; De-Los-Mozos Pascual, Marcelino; Santana, Omar; Roldán, Marta; Fernández, José-Antonio

2015-01-01

The presence and extent of genetic variation in saffron crocus are still debated, as testified by several contradictory articles providing contrasting results about the monomorphism or less of the species. Remarkably, phenotypic variations have been frequently observed in the field, such variations are usually unstable and can change from one growing season to another. Considering that gene expression can be influenced both by genetic and epigenetic changes, epigenetics could be a plausible cause of the alternative phenotypes. In order to obtain new insights into this issue, we carried out a molecular marker analysis of 112 accessions from the World Saffron and Crocus Collection. The accessions were grown for at least three years in the same open field conditions. The same samples were analysed using Amplified Fragment Length Polymorphism (AFLP) and Methyl Sensitive AFLP in order to search for variation at the genetic (DNA sequence) and epigenetic (cytosine methylation) level. While the genetic variability was low (4.23% polymorphic peaks and twelve (12) effective different genotypes), the methyl sensitive analysis showed the presence of high epigenetic variability (33.57% polymorphic peaks and twenty eight (28) different effective epigenotypes). The pattern obtained by Factorial Correspondence Analysis of AFLP and, in particular, of MS-AFLP data was consistent with the geographical provenance of the accessions. Very interestingly, by focusing on Spanish accessions, it was observed that the distribution of the accessions in the Factorial Correspondence Analysis is not random but tends to reflect the geographical origin. Two clearly defined clusters grouping accessions from the West (Toledo and Ciudad Real) and accessions from the East (Cuenca and Teruel) were clearly recognised. PMID:25885113

A DNA microarray-based methylation-sensitive (MS)-AFLP hybridization method for genetic and epigenetic analyses.

PubMed

Yamamoto, F; Yamamoto, M

2004-07-01

We previously developed a PCR-based DNA fingerprinting technique named the Methylation Sensitive (MS)-AFLP method, which permits comparative genome-wide scanning of methylation status with a manageable number of fingerprinting experiments. The technique uses the methylation sensitive restriction enzyme NotI in the context of the existing Amplified Fragment Length Polymorphism (AFLP) method. Here we report the successful conversion of this gel electrophoresis-based DNA fingerprinting technique into a DNA microarray hybridization technique (DNA Microarray MS-AFLP). By performing a total of 30 (15 x 2 reciprocal labeling) DNA Microarray MS-AFLP hybridization experiments on genomic DNA from two breast and three prostate cancer cell lines in all pairwise combinations, and Southern hybridization experiments using more than 100 different probes, we have demonstrated that the DNA Microarray MS-AFLP is a reliable method for genetic and epigenetic analyses. No statistically significant differences were observed in the number of differences between the breast-prostate hybridization experiments and the breast-breast or prostate-prostate comparisons.

Molecular characterization of the U.S. Phaseolus acutifolius A. Gray collection using Amplified Fragment Length Polymorphism (AFLP) and Targeted Region Amplification Polymorphism (TRAP) markers.

USDA-ARS?s Scientific Manuscript database

Tepary bean (Phaseolus acutifolius A. Gray), a truly Native American crop, is a short life-cycle annual desert legume indigenous to northwestern Mexico and the southwestern USA and is considered drought and heat tolerant. The Western Regional Plant Introduction Station currently maintains 211 acce...

Genotypic Analysis of Escherichia coli Strains from Poultry Carcasses and Their Susceptibilities to Antimicrobial Agents

PubMed Central

Geornaras, Ifigenia; Hastings, John W.; von Holy, Alexander

2001-01-01

Plasmid profiling and amplified fragment length polymorphism (AFLP) analysis were used to genotype 50 Escherichia coli strains from poultry carcasses. Thirty different plasmid profiles were evident, and clustering of the AFLP data showed that they were a distinctly heterogeneous group of strains. Susceptibility testing against five antimicrobial agents used in the South African poultry industry showed all strains to be susceptible to danofloxacin and colistin, while the majority (96%) were resistant to two tetracyclines. PMID:11282652

Genetic diversity in populations of Isatis glauca Aucher ex Boiss. ssp. from Central Anatolia in Turkey, as revealed by AFLP analysis.

PubMed

Özbek, Özlem; Görgülü, Elçin; Yıldırımlı, Şinasi

2013-12-01

Isatidae L. is a complex and systematically difficult genus in Brassicaceae. The genus displays great morphological polymorphism, which makes the classification of species and subspecies difficult as it is observed in Isatis glauca Aucher ex Boiss. The aim of this study is characterization of the genetic diversity in subspecies of Isatis glauca Aucher ex Boiss. distributed widely in Central Anatolia, in Turkey by using Amplified Fragment Length Polymorphism (AFLP) technique. Eight different Eco RI-Mse I primer combinations produced 805 AFLP loci, 793 (98.5%) of which were polymorphic in 67 accessions representing nine different populations. The data obtained by AFLP was computed with using GDA (Genetic Data Analysis) and STRUCTURE (version 2.3.3) software programs for population genetics. The mean proportion of the polymorphic locus (P), the mean number of alleles (A), the number of unique alleles (U) and the mean value of gene diversity (He) were 0.59, 1.59, 20, and 0.23 respectively. The coancestry coefficient (ϴ) was 0.24. The optimal number of K was identified as seven. The principal component analysis (PCA) explained 85.61% of the total genetic variation. Isatis glauca ssp. populations showed a high level of genetic diversity, and the AFLP analysis revealed that high polymorphism and differentiated subspecies could be used conveniently for population genetic studies. The principal coordinate analysis (PCoA) based on the dissimilarity matrix, the dendrogram drawn with UPGMA method and STRUCTURE cluster analysis distinguished the accessions successfully. The accessions formed distinctive population structures for populations AA, AB, E, K, and S. Populations AG1 and AG2 seemed to have similar genetic content, in addition, in both populations several hybrid individuals were observed. The accessions did not formed distinctive population structures for both populations AI and ANP. Consequently, Ankara province might be the area, where species Isatis glauca Aucher ex Boiss. originated.

Identification of Amplified Fragment Length Polymorphism (AFLP) Markers Tightly Associated with Drought Stress Gene in Male Sterile and Fertile Salvia miltiorrhiza Bunge

PubMed Central

Zhang, Yuejin; Guo, Lijun; Shu, Zhiming; Sun, Yiyue; Chen, Yuanyuan; Liang, Zongsuo; Guo, Hongbo

2013-01-01

Consistent grain yield in drought environment has attracted wide attention due to global climate change. However, the important drought-related traits/genes in crops have been rarely reported. Many near-isogenic lines (NILs) of male sterile and fertile Salvia miltiorrhiza have been obtained in our previous work through testcross and backcross in continuous field experiments conducted in 2006–2009. Both segregating sterile and fertile populations were subjected to bulked segregant analysis (BSA) and amplified fragment length polymorphism (AFLP) with 384 and 170 primer combinations, respectively. One out of 14 AFLP markers (E9/M3246) was identified in treated fertile population as tightly linked to the drought stress gene with a recombination frequency of 6.98% and at a distance of 7.02 cM. One of 15 other markers (E2/M5357) was identified in a treated sterile population that is closely associated with the drought stress gene. It had a recombination frequency of 4.65% and at a distance of 4.66 cM. Interestingly, the E9/M3246 fragment was found to be identical to another AFLP fragment E11/M4208 that was tightly linked to the male sterile gene of S. miltiorrhiza with 95% identity and e-value 4 × 10−93. Blastn analysis suggested that the drought stress gene sequence showed higher identity with nucleotides in Arabidopsis chromosome 1–5. PMID:23525049

Genetic analysis of floating Enteromorpha prolifera in the Yellow Sea with AFLP marker

NASA Astrophysics Data System (ADS)

Liu, Cui; Zhang, Jing; Sun, Xiaoyu; Li, Jian; Zhang, Xi; Liu, Tao

2011-09-01

Extremely large accumulation of green algae Enteromorpha prolifera floated along China' coastal region of the Yellow Sea ever since the summer of 2008. Amplified Fragment Length Polymorphism (AFLP) analysis was applied to assess the genetic diversity and relationships among E. prolifera samples collected from 9 affected areas of the Yellow Sea. Two hundred reproducible fragments were generated with 8 AFLP primer combinations, of which 194 (97%) were polymorphic. The average Nei's genetic diversity, the coefficiency of genetic differentiation (Gst), and the average gene flow estimated from Gst in the 9 populations were 0.4018, 0.6404 and 0.2807 respectively. Cluster analysis based on the unweighed pair group method with arithmetic averages (UPGMA) showed that the genetic relationships within one population or among different populations were all related to their collecting locations and sampling time. Large genetic differentiation was detected among the populations. The E. prolifera originated from different areas and were undergoing a course of mixing.

Analysis of genetic diversity in banana cultivars (Musa cvs.) from the South of Oman using AFLP markers and classification by phylogenetic, hierarchical clustering and principal component analyses*

PubMed Central

Opara, Umezuruike Linus; Jacobson, Dan; Al-Saady, Nadiya Abubakar

2010-01-01

Banana is an important crop grown in Oman and there is a dearth of information on its genetic diversity to assist in crop breeding and improvement programs. This study employed amplified fragment length polymorphism (AFLP) to investigate the genetic variation in local banana cultivars from the southern region of Oman. Using 12 primer combinations, a total of 1094 bands were scored, of which 1012 were polymorphic. Eighty-two unique markers were identified, which revealed the distinct separation of the seven cultivars. The results obtained show that AFLP can be used to differentiate the banana cultivars. Further classification by phylogenetic, hierarchical clustering and principal component analyses showed significant differences between the clusters found with molecular markers and those clusters created by previous studies using morphological analysis. Based on the analytical results, a consensus dendrogram of the banana cultivars is presented. PMID:20443211

Amplified fragment length polymorphism mapping of quantitative trait loci for malaria parasite susceptibility in the yellow fever mosquito Aedes aegypti.

PubMed

Zhong, Daibin; Menge, David M; Temu, Emmanuel A; Chen, Hong; Yan, Guiyun

2006-07-01

The yellow fever mosquito Aedes aegypti has been the subject of extensive genetic research due to its medical importance and the ease with which it can be manipulated in the laboratory. A molecular genetic linkage map was constructed using 148 amplified fragment length polymorphism (AFLP) and six single-strand conformation polymorphism (SSCP) markers. Eighteen AFLP primer combinations were used to genotype two reciprocal F2 segregating populations. Each primer combination generated an average of 8.2 AFLP markers eligible for linkage mapping. The length of the integrated map was 180.9 cM, giving an average marker resolution of 1.2 cM. Composite interval mapping revealed a total of six QTL significantly affecting Plasmodium susceptibility in the two reciprocal crosses of Ae. aegypti. Two common QTL on linkage group 2 were identified in both crosses that had similar effects on the phenotype, and four QTL were unique to each cross. In one cross, the four main QTL accounted for 64% of the total phenotypic variance, and digenic epistasis explained 11.8% of the variance. In the second cross, the four main QTL explained 66% of the variance, and digenic epistasis accounted for 16% of the variance. The actions of these QTL were either dominance or underdominance. Our results indicated that at least three new QTL were mapped on chromosomes 1 and 3. The polygenic nature of susceptibility to P. gallinaceum and epistasis are important factors for significant variation within or among mosquito strains. The new map provides additional information useful for further genetic investigation, such as identification of new genes and positional cloning.

Invasion Dynamics and Genotypic Diversity of Cogongrass (Imperata cylindrica) at the Point of Introduction in the Southeastern United States

Treesearch

Ludovic J. A. Capo-chichi; Wilson H. Faircloth; A. G. Williamson; Michael G. Patterson; James H. Miller; Edzard van Santen

2008-01-01

Nine sites of cogongrass were included in a study of genotypic dimity and spread dynamics at the point of introduction and its adjacent areas in the southern United States. Clones evaluated with two primer pairs yielded a total of 137 amplified fragment length polymorphism (AFLP) hi of which 102 (74.4%) were polymorphic. Genetic diversity was measured as the percentage...

Optimization of cDNA-AFLP experiments using genomic sequence data.

PubMed

Kivioja, Teemu; Arvas, Mikko; Saloheimo, Markku; Penttilä, Merja; Ukkonen, Esko

2005-06-01

cDNA amplified fragment length polymorphism (cDNA-AFLP) is one of the few genome-wide level expression profiling methods capable of finding genes that have not yet been cloned or even predicted from sequence but have interesting expression patterns under the studied conditions. In cDNA-AFLP, a complex cDNA mixture is divided into small subsets using restriction enzymes and selective PCR. A large cDNA-AFLP experiment can require a substantial amount of resources, such as hundreds of PCR amplifications and gel electrophoresis runs, followed by manual cutting of a large number of bands from the gels. Our aim was to test whether this workload can be reduced by rational design of the experiment. We used the available genomic sequence information to optimize cDNA-AFLP experiments beforehand so that as many transcripts as possible could be profiled with a given amount of resources. Optimization of the selection of both restriction enzymes and selective primers for cDNA-AFLP experiments has not been performed previously. The in silico tests performed suggest that substantial amounts of resources can be saved by the optimization of cDNA-AFLP experiments.

Genetic variation and relationship among and within Withania species as revealed by AFLP markers.

PubMed

Negi, M S; Singh, A; Lakshmikumaran, M

2000-12-01

Withania somnifera is an important medicinal plant, and its anticancerous properties have been attributed to various classes of withanolide compounds. The objective of the present study was to investigate the inter- and intraspecific genetic variation present in 35 individuals of W. somnifera and 5 individuals of W. coagulans using AFLP (amplified fragment length polymorphism) marker technique. The information about genetic variation determined from AFLP data for 40 individuals was employed to estimate similarity matrix value based on Jaccard's coefficient. The similarity values were further used to construct a phenetic dendrogram revealing the genetic relationships. The dendrogram generated by UPGMA (unweighted pair group method of arithmetic averages) distinguished W. somnifera from W. coagulans and formed two major clusters. These two main clusters shared a similarity coefficient of 0.3, correlating with the high level of polymorphism detected. The dendrogram further separated W. somnifera into three subclasses corresponding to Kashmiri and Nagori groups and an intermediate type. The AFLP profile of Kashmiri individuals was distinct from that of the Nagori group of plants. The intermediate genotype was distinct as it shared bands with both the Kashmiri and Nagori individuals, even though it was identified as a Kashmiri morphotype. Furthermore, the intermediate type shared a similarity coefficient of 0.8 with the Kashmiri individuals. The present work revealed low levels of variation within a population though high levels of polymorphism were detected between Nagori and Kashmiri populations. The ability of AFLP markers for efficient and rapid detection of genetic variations at the species as well as intraspecific level qualifies it as an efficient tool for estimating genetic similarity in plant species and effective management of genetic resources.

Genetic differentiation of Octopus minor (Mollusca, Cephalopoda) off the northern coast of China as revealed by amplified fragment length polymorphisms.

PubMed

Yang, J M; Sun, G H; Zheng, X D; Ren, L H; Wang, W J; Li, G R; Sun, B C

2015-12-02

Octopus minor (Sasaki, 1920) is an economically important cephalopod that is found in the northern coastal waters of China. In this study, we investigated genetic differentiation in fishery populations using amplified fragment length polymorphisms (AFLPs). A total of 150 individuals were collected from five locations: Dalian (DL), Yan-tai (YT), Qingdao (QD), Lianyungang (LY), and Zhoushan (ZS), and 243 reproducible bands were amplified using five AFLP primer combinations. The percentage of polymorphic bands ranged from 53.33 to 76.08%. Nei's genetic identity ranged from 0.9139 to 0.9713, and the genetic distance ranged from 0.0291 to 0.0900. A phylogenetic tree was constructed using the unweighted pair group method with arithmetic mean, based on the genetic distance. The DL and YT populations originated from one clade, while the QD, LY, and ZS populations originated from another. The results indicate that the O. minor stock consisted of two genetic populations with an overall significantly analogous FST value (0.1088, P < 0.05). Most of the variance was within populations. These findings will be important for more sustainable octopus fisheries, so that this marine resource can be conserved for its long-term utilization.

Amplified Fragment Length Polymorphism Diversity in Cephalosporium maydis from Egypt.

PubMed

Saleh, Amgad A; Zeller, Kurt A; Ismael, Abou-Serie M; Fahmy, Zeinab M; El-Assiuty, Elhamy M; Leslie, John F

2003-07-01

ABSTRACT Cephalosporium maydis, the causal agent of late wilt of maize, was first described in Egypt in the 1960s, where it can cause yield losses of up to 40% in susceptible plantings. We characterized 866 isolates of C. maydis collected from 14 governates in Egypt, 7 in the Nile River Delta and 7 in southern (Middle and Upper) Egypt, with amplified fragment length polymorphism (AFLP) markers. The four AFLP primer-pair combinations generated 68 bands, 25 of which were polymorphic, resulting in 52 clonal haplotypes that clustered the 866 isolates into four phylogenetic lineages. Three lineages were found in both the Nile River Delta and southern Egypt. Lineage IV, the most diverse group (20 haplotypes), was recovered only from governates in the Nile River Delta. In some locations, one lineage dominated (up to 98% of the isolates recovered) and, from some fields, only a single haplotype was recovered. Under field conditions in Egypt, there is no evidence that C. maydis reproduces sexually. The nonuniform geographic distribution of the pathogen lineages within the country could be due to differences in climate or in the farming system, because host material differs in susceptibility and C. maydis lineages differ in pathogenicity.

Genetic characterization of hybridization and introgression between anadromous rainbow trout (oncorhynchus mykiss irideus) and coastal cutthroat trout (o. clarki clarki)

USGS Publications Warehouse

Young, W.P.; Ostberg, C.O.; Keim, P.; Thorgaard, G.H.

2001-01-01

Interspecific hybridization represents a dynamic evolutionary phenomenon and major conservation problem in salmonid fishes. In this study we used amplified fragment length polymorphisms (AFLP) and mitochondrial DNA (mtDNA) markers to describe the extent and characterize the pattern of hybridization and introgression between coastal rainbow trout (Oncorhynchus mykiss irideus) and coastal cutthroat trout (O. clarki clarki). Hybrid individuals were initially identified using principle coordinate analysis of 133 polymorphic AFLP markers. Subsequent analysis using 23 diagnostic AFLP markers revealed the presence of F1, rainbow trout backcross, cutthroat trout backcross and later-generation hybrids. mtDNA analysis demonstrated equal numbers of F1 hybrids with rainbow and cutthroat trout mtDNA indicating reciprocal mating of the parental types. In contrast, rainbow and cutthroat trout backcross hybrids always exhibited the mtDNA from the recurrent parent, indicating a male hybrid mating with a pure female. This study illustrates the usefulness of the AFLP technique for generating large numbers of species diagnostic markers. The pattern of hybridization raises many questions concerning the existence and action of reproductive isolating mechanisms between these two species. Our findings are consistent with the hypothesis that introgression between anadromous populations of coastal rainbow and coastal cutthroat trout is limited by an environment-dependent reduction in hybrid fitness.

Congruent climate-related genecological responses from molecular markers and quantitative traits for western white pine (Pinus monticola)

Treesearch

Bryce A. Richardson; Gerald E. Rehfeldt; Mee-Sook Kim

2009-01-01

Analyses of molecular and quantitative genetic data demonstrate the existence of congruent climate-related patterns in western white pine (Pinus monticola). Two independent studies allowed comparisons of amplified fragment length polymorphism (AFLP) markers with quantitative variation in adaptive traits. Principal component analyses...

Characterization of microsatellite loci and reliable genotyping in a polyploid plant, Mercurialis perennis (Euphorbiaceae).

PubMed

Pfeiffer, Tanja; Roschanski, Anna M; Pannell, John R; Korbecka, Grazyna; Schnittler, Martin

2011-01-01

For many applications in population genetics, codominant simple sequence repeats (SSRs) may have substantial advantages over dominant anonymous markers such as amplified fragment length polymorphisms (AFLPs). In high polyploids, however, allele dosage of SSRs cannot easily be determined and alleles are not easily attributable to potentially diploidized loci. Here, we argue that SSRs may nonetheless be better than AFLPs for polyploid taxa if they are analyzed as effectively dominant markers because they are more reliable and more precise. We describe the transfer of SSRs developed for diploid Mercurialis huetii to the clonal dioecious M. perennis. Primers were tested on a set of 54 male and female plants from natural decaploid populations. Eight of 65 tested loci produced polymorphic fragments. Binary profiles from 4 different scoring routines were used to define multilocus lineages (MLLs). Allowing for fragment differences within 1 MLL, all analyses revealed the same 14 MLLs without conflicting with merigenet, sex, or plot assignment. For semiautomatic scoring, a combination of as few as 2 of the 4 most polymorphic loci resulted in unambiguous discrimination of clones. Our study demonstrates that microsatellite fingerprinting of polyploid plants is a cost efficient and reliable alternative to AFLPs, not least because fewer loci are required than for diploids.

Detection of Variation in Long-Term Micropropagated Mature Pistachio via DNA-Based Molecular Markers.

PubMed

Akdemir, Hülya; Suzerer, Veysel; Tilkat, Engin; Onay, Ahmet; Çiftçi, Yelda Ozden

2016-12-01

Determination of genetic stability of in vitro-grown plantlets is needed for safe and large-scale production of mature trees. In this study, genetic variation of long-term micropropagated mature pistachio developed through direct shoot bud regeneration using apical buds (protocol A) and in vitro-derived leaves (protocol B) was assessed via DNA-based molecular markers. Randomly amplified polymorphic DNA (RAPD), inter-simple sequence repeat (ISSR), and amplified fragment length polymorphism (AFLP) were employed, and the obtained PIC values from RAPD (0.226), ISSR (0.220), and AFLP (0.241) showed that micropropagation of pistachio for different periods of time resulted in "reasonable polymorphism" among donor plant and its 18 clones. Mantel's test showed a consistence polymorphism level between marker systems based on similarity matrices. In conclusion, this is the first study on occurrence of genetic variability in long-term micropropagated mature pistachio plantlets. The obtained results clearly indicated that different marker approaches used in this study are reliable for assessing tissue culture-induced variations in long-term cultured pistachio plantlets.

Molecular Identification of Date Palm Cultivars Using Random Amplified Polymorphic DNA (RAPD) Markers.

PubMed

Al-Khalifah, Nasser S; Shanavaskhan, A E

2017-01-01

Ambiguity in the total number of date palm cultivars across the world is pointing toward the necessity for an enumerative study using standard morphological and molecular markers. Among molecular markers, DNA markers are more suitable and ubiquitous to most applications. They are highly polymorphic in nature, frequently occurring in genomes, easy to access, and highly reproducible. Various molecular markers such as restriction fragment length polymorphism (RFLP), amplified fragment length polymorphism (AFLP), simple sequence repeats (SSR), inter-simple sequence repeats (ISSR), and random amplified polymorphic DNA (RAPD) markers have been successfully used as efficient tools for analysis of genetic variation in date palm. This chapter explains a stepwise protocol for extracting total genomic DNA from date palm leaves. A user-friendly protocol for RAPD analysis and a table showing the primers used in different molecular techniques that produce polymorphisms in date palm are also provided.

Methylation-Sensitive Amplification Length Polymorphism (MS-AFLP) Microarrays for Epigenetic Analysis of Human Genomes.

PubMed

Alonso, Sergio; Suzuki, Koichi; Yamamoto, Fumiichiro; Perucho, Manuel

2018-01-01

Somatic, and in a minor scale also germ line, epigenetic aberrations are fundamental to carcinogenesis, cancer progression, and tumor phenotype. DNA methylation is the most extensively studied and arguably the best understood epigenetic mechanisms that become altered in cancer. Both somatic loss of methylation (hypomethylation) and gain of methylation (hypermethylation) are found in the genome of malignant cells. In general, the cancer cell epigenome is globally hypomethylated, while some regions-typically gene-associated CpG islands-become hypermethylated. Given the profound impact that DNA methylation exerts on the transcriptional profile and genomic stability of cancer cells, its characterization is essential to fully understand the complexity of cancer biology, improve tumor classification, and ultimately advance cancer patient management and treatment. A plethora of methods have been devised to analyze and quantify DNA methylation alterations. Several of the early-developed methods relied on the use of methylation-sensitive restriction enzymes, whose activity depends on the methylation status of their recognition sequences. Among these techniques, methylation-sensitive amplification length polymorphism (MS-AFLP) was developed in the early 2000s, and successfully adapted from its original gel electrophoresis fingerprinting format to a microarray format that notably increased its throughput and allowed the quantification of the methylation changes. This array-based platform interrogates over 9500 independent loci putatively amplified by the MS-AFLP technique, corresponding to the NotI sites mapped throughout the human genome.

An AFLP-based genetic linkage map of channel catfish (Ictalurus punctatus) constructed by using an interspecific hybrid resource family.

PubMed Central

Liu, Zhanjiang; Karsi, Attila; Li, Ping; Cao, Dongfeng; Dunham, R

2003-01-01

Catfish is the major aquaculture species in the United States. The hybrid catfish produced by crossing channel catfish females with blue catfish males exhibit a number of desirable production traits, but their mass production has been difficult. To introduce desirable genes from blue catfish into channel catfish through introgression, a genetic linkage map is helpful. In this project, a genetic linkage map was constructed using amplified fragment length polymorphism (AFLP). A total of 607 AFLP markers were analyzed using 65 primer combinations and an interspecific backcross resource family. A total of 418 AFLP markers were assigned to 44 linkage groups. Among the remaining 189 markers, 101 were not used because of significant segregation distortion, 29 were unlinked, and 59 were eliminated because they span very large distances. The 418 AFLP markers covered 1593 cM Kosambi. The AFLP markers showed a high level of clustering that appears to be related to certain primer combinations. This linkage map will serve as the basis for mapping a greater number of markers to provide a map with high enough resolution for it to be useful for selective breeding programs using introgression. PMID:14573480

Identification of QTLs Associated with Callogenesis and Embryogenesis in Oil Palm Using Genetic Linkage Maps Improved with SSR Markers

PubMed Central

Ting, Ngoot-Chin; Jansen, Johannes; Nagappan, Jayanthi; Ishak, Zamzuri; Chin, Cheuk-Weng; Tan, Soon-Guan; Cheah, Suan-Choo; Singh, Rajinder

2013-01-01

Clonal reproduction of oil palm by means of tissue culture is a very inefficient process. Tissue culturability is known to be genotype dependent with some genotypes being more amenable to tissue culture than others. In this study, genetic linkage maps enriched with simple sequence repeat (SSR) markers were developed for dura (ENL48) and pisifera (ML161), the two fruit forms of oil palm, Elaeis guineensis. The SSR markers were mapped onto earlier reported parental maps based on amplified fragment length polymorphism (AFLP) and restriction fragment length polymorphism (RFLP) markers. The new linkage map of ENL48 contains 148 markers (33 AFLPs, 38 RFLPs and 77 SSRs) in 23 linkage groups (LGs), covering a total map length of 798.0 cM. The ML161 map contains 240 markers (50 AFLPs, 71 RFLPs and 119 SSRs) in 24 LGs covering a total of 1,328.1 cM. Using the improved maps, two quantitative trait loci (QTLs) associated with tissue culturability were identified each for callusing rate and embryogenesis rate. A QTL for callogenesis was identified in LGD4b of ENL48 and explained 17.5% of the phenotypic variation. For embryogenesis rate, a QTL was detected on LGP16b in ML161 and explained 20.1% of the variation. This study is the first attempt to identify QTL associated with tissue culture amenity in oil palm which is an important step towards understanding the molecular processes underlying clonal regeneration of oil palm. PMID:23382832

Morphological and genetic analysis of four color morphs of bean leaf beetle, Cerotoma trifurcata (Coleoptera: Chrysomelidae)

USDA-ARS?s Scientific Manuscript database

Bean leaf beetle (BLB) exhibits a relatively large amount of morphological variation in terms of color but little is known about the underlying genetic structure and gene flow. Genetic variation among four color phenotypes of the BLB was analyzed using amplified fragment length polymorphisms (AFLP) ...

An integrated PCR colony hybridization approach to screen cDNA libraries for full-length coding sequences.

PubMed

Pollier, Jacob; González-Guzmán, Miguel; Ardiles-Diaz, Wilson; Geelen, Danny; Goossens, Alain

2011-01-01

cDNA-Amplified Fragment Length Polymorphism (cDNA-AFLP) is a commonly used technique for genome-wide expression analysis that does not require prior sequence knowledge. Typically, quantitative expression data and sequence information are obtained for a large number of differentially expressed gene tags. However, most of the gene tags do not correspond to full-length (FL) coding sequences, which is a prerequisite for subsequent functional analysis. A medium-throughput screening strategy, based on integration of polymerase chain reaction (PCR) and colony hybridization, was developed that allows in parallel screening of a cDNA library for FL clones corresponding to incomplete cDNAs. The method was applied to screen for the FL open reading frames of a selection of 163 cDNA-AFLP tags from three different medicinal plants, leading to the identification of 109 (67%) FL clones. Furthermore, the protocol allows for the use of multiple probes in a single hybridization event, thus significantly increasing the throughput when screening for rare transcripts. The presented strategy offers an efficient method for the conversion of incomplete expressed sequence tags (ESTs), such as cDNA-AFLP tags, to FL-coding sequences.

Preliminary genetic linkage map of the abalone Haliotis diversicolor Reeve

NASA Astrophysics Data System (ADS)

Shi, Yaohua; Guo, Ximing; Gu, Zhifeng; Wang, Aimin; Wang, Yan

2010-05-01

Haliotis diversicolor Reeve is one of the most important mollusks cultured in South China. Preliminary genetic linkage maps were constructed with amplified fragment length polymorphism (AFLP) markers. A total of 2 596 AFLP markers were obtained from 28 primer combinations in two parents and 78 offsprings. Among them, 412 markers (15.9%) were polymorphic and segregated in the mapping family. Chi-square tests showed that 151 (84.4%) markers segregated according to the expected 1:1 Mendelian ratio ( P<0.05) in the female parent, and 200 (85.8%) in the male parent. For the female map, 179 markers were used for linkage analysis and 90 markers were assigned to 17 linkage groups with an average interval length of 25.7 cm. For the male map, 233 markers were used and 94 were mapped into 18 linkage groups, with an average interval of 25.0 cm. The estimated genome length was 2 773.0 cm for the female and 2 817.1 cm for the male map. The observed length of the linkage map was 1 875.2 cm and 1 896.5 cm for the female and male maps, respectively. When doublets were considered, the map length increased to 2 152.8 cm for the female and 2 032.7 cm for the male map, corresponding to genome coverage of 77.6% and 72.2%, respectively.

Mapping of AFLP markers linked to seed coat colour loci in Brassica juncea (L.) Czern.

PubMed

Sabharwal, V; Negi, M S; Banga, S S; Lakshmikumaran, M

2004-06-01

Association mapping of the seed-coat colour with amplified fragment length polymorphism (AFLP) markers was carried out in 39 Brassica juncea lines. The lines had genetically diverse parentages and varied for seed-coat colour and other morphological characters. Eleven AFLP primer combinations were used to screen the 39 B. juncea lines, and a total of 335 polymorphic bands were detected. The bands were analysed for association with seed-coat colour using multiple regression analysis. This analysis revealed 15 markers associated with seed-coat colour, obtained with eight AFLP primer combinations. The marker E-ACA/M-CTG(350 )explained 69% of the variation in seed-coat colour. This marker along with markers E-AAC/M-CTC(235 )and E-AAC/M-CTA(250) explained 89% of the total variation. The 15 associated markers were validated for linkage with the seed-coat colour loci using a recombinant inbred line (RIL) mapping population. Bands were amplified with the eight AFLP primer combinations in 54 RIL progenies. Of the 15 associated markers, 11 mapped on two linkage groups. Eight markers were placed on linkage group 1 at a marker density of 6.0 cM, while the remaining three were mapped on linkage group 2 at a marker density of 3.6 cM. Marker E-ACA/M-CTG(350 )co-segregated with Gene1 controlling seed-coat colour; it was specific for yellow seed-coat colour and mapped to linkage group 1. Marker E-AAC/M-CTC(235) (AFLP8), which had been studied previously, was present on linkage group 2; it was specific for brown seed-coat colour. Since AFLP markers are not adapted for large-scale applications in plant breeding, it is important to convert these to sequence-characterised amplified region (SCAR) markers. Marker E-AAC/M-CTC(235) (AFLP8) had been previously converted into a SCAR. Work is in progress to convert the second of the linked markers, E-ACA/M-CTG(350), to a SCAR. The two linked AFLP markers converted to SCARs will be useful for developing yellow-seeded B. juncea lines by means of marker-assisted selection.

Space environment induced mutations prefer to occur at polymorphic sites of rice genomes

NASA Astrophysics Data System (ADS)

Li, Y.; Liu, M.; Cheng, Z.; Sun, Y.

To explore the genomic characteristics of rice mutants induced by space environment, space-induced mutants 971-5, 972-4, and R955, which acquired new traits after space flight such as increased yield, reduced resistance to rice blast, and semi-dwarfism compared with their on-ground controls, 971ck, 972ck, and Bing95-503, respectively, together with other 8 japonica and 3 indica rice varieties, 17 in total, were analyzed by amplified fragment length polymorphism (AFLP) method. We chose 16 AFLP primer-pairs which generated a total of 1251 sites, of which 745 (59.6%) were polymorphic over all the genotypes. With the 16 pairs of primer combinations, 54 space-induced mutation sites were observed in 971-5, 86 in 972-4, and 5 in R955 compared to their controls, and the mutation rates were 4.3%, 6.9% and 0.4%, respectively. Interestingly, 75.9%, 84.9% and 100% of the mutation sites identified in 971-5, 972-4, and R955 occurred in polymorphic sites. This result suggests that the space environment preferentially induced mutations at polymorphic sites in rice genomes and might share a common mechanism with other types of mutagens. It also implies that polymorphic sites in genomes are potential "hotspots" for mutations induced by the space environment.

Genetic variability in Brazilian Capsicum baccatum germplasm collection assessed by morphological fruit traits and AFLP markers

PubMed Central

Giacomin, Renata M.; Ruas, Paulo M.; Ruas, Eduardo A.; Barbieri, Rosa L.; Rodrigues, Rosana

2018-01-01

Capsicum baccatum is one of the main pepper species grown and consumed in South America. In Brazil, it is commonly cultivated by family farmers, using mostly the genotypes bishop's hat genotypes (locally cambuci) and red chili pepper (dedo-de-moça). This study had the objective of characterizing 116 C. baccatum accessions from different regions of Brazil, based on morphological fruit descriptors and AFLP (Amplified Fragment Length Polymorphisms) markers. Broad phenotypic variability among the C. baccatum accessions was detected when using morphological fruit descriptors. The Ward modified location model (Ward-MLM) discriminated five groups, based mainly on fruit shape. Six combinations of AFLP primers detected polymorphism in 97.93% of the 2466 identified bands, indicating the high genetic variability in the accessions. The UPGMA coincided with the Bayesian clustering analysis and three large groups were formed, separating the wild variety C. baccatum var. praetermissum from the other accessions. There was no relation between genetic distance and geographical origin of the accessions, probably due to the intense exchange of fruits and seeds between farmers. Morphological descriptors used together with AFLP markers proved efficient in detecting the levels of genetic variability among the accessions maintained in the germplasm collections. These results can be used as an additional source of helpful information to be exploited in C. baccatum breeding programs. PMID:29758023

Determining Phylogenetic Relationships Among Date Palm Cultivars Using Random Amplified Polymorphic DNA (RAPD) and Inter-Simple Sequence Repeat (ISSR) Markers.

PubMed

Haider, Nadia

2017-01-01

Investigation of genetic variation and phylogenetic relationships among date palm (Phoenix dactylifera L.) cultivars is useful for their conservation and genetic improvement. Various molecular markers such as restriction fragment length polymorphisms (RFLPs), simple sequence repeat (SSR), representational difference analysis (RDA), and amplified fragment length polymorphism (AFLP) have been developed to molecularly characterize date palm cultivars. PCR-based markers random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) are powerful tools to determine the relatedness of date palm cultivars that are difficult to distinguish morphologically. In this chapter, the principles, materials, and methods of RAPD and ISSR techniques are presented. Analysis of data generated from these two techniques and the use of these data to reveal phylogenetic relationships among date palm cultivars are also discussed.

Applicability of SCAR markers to food genomics: olive oil traceability.

PubMed

Pafundo, Simona; Agrimonti, Caterina; Maestri, Elena; Marmiroli, Nelson

2007-07-25

DNA analysis with molecular markers has opened a shortcut toward a genomic comprehension of complex organisms. The availability of micro-DNA extraction methods, coupled with selective amplification of the smallest extracted fragments with molecular markers, could equally bring a breakthrough in food genomics: the identification of original components in food. Amplified fragment length polymorphisms (AFLPs) have been instrumental in plant genomics because they may allow rapid and reliable analysis of multiple and potentially polymorphic sites. Nevertheless, their direct application to the analysis of DNA extracted from food matrixes is complicated by the low quality of DNA extracted: its high degradation and the presence of inhibitors of enzymatic reactions. The conversion of an AFLP fragment to a robust and specific single-locus PCR-based marker, therefore, could extend the use of molecular markers to large-scale analysis of complex agro-food matrixes. In the present study is reported the development of sequence characterized amplified regions (SCARs) starting from AFLP profiles of monovarietal olive oils analyzed on agarose gel; one of these was used to identify differences among 56 olive cultivars. All the developed markers were purposefully amplified in olive oils to apply them to olive oil traceability.

Paleoclimate effects and geographic barriers shape regional population genetic structure of blackbrush (Coleogyne ramosissima: Rosaceae)

Treesearch

Bryce A. Richardson; Susan E. Meyer

2012-01-01

Coleogyne ramosissima Torr. (blackbrush) is a dominant xerophytic shrub species in the ecotone between the warm and cold deserts of interior western North America. Amplified fragment length polymorphisms (AFLPs) were used to survey genetic diversity and population genetic structure at 14 collection sites across the species range. Analysis revealed significant...

Effect of white pine blister rust (Cronartium ribicola) and rust-resistance breeding on genetic variation in western white pine Pinus monticola)

Treesearch

M. -S. Kim; S. J. Brunsfeld; G. I. McDonald; N. B. Klopfenstein

2003-01-01

Western white pine (Pinus monticola) is an economically and ecologically important species from western North America that has declined over the past several decades mainly due to the introduction of blister rust (Cronartium ribicola) and reduced opportunities for regeneration. Amplified fragment length polymorphism (AFLP) was used...

Comparative ITS and AFLP Analysis of Diploid Cardamine (Brassicaceae) Taxa from Closely Related Polyploid Complexes

PubMed Central

MARHOLD, KAROL; LIHOVÁ, JUDITA; PERNÝ, MARIÁN; BLEEKER, WALTER

2004-01-01

• Background and Aims Diploid representatives from the related polyploid complexes of Cardamine amara, C. pratensis and C. raphanifolia (Brassicaceae), were studied to elucidate phylogenetic relationships among the complexes and among the individual taxa included. • Methods Two independent molecular data sets were used: nucleotide sequences from the internal transcribed spacers (ITS) of nrDNA, and amplified fragment length polymorphism (AFLP) markers. Seventeen diploid taxa from the studied groups were sampled. • Key Results Both ITS and AFLP analyses provided congruent results in inferred relationships, and revealed two main lineages. While the C. amara group, consisting of C. wiedemanniana and four subspecies of C. amara, was resolved as a well‐supported monophyletic group, taxa from the C. pratensis and C. tenera groups (the latter representing diploid taxa of the complex of C. raphanifolia) all appeared together in a single clade/cluster with no support for the recognition of either of the groups. Intra‐individual polymorphisms and patterns of nucleotide variation in the ITS region in C. uliginosa and C. tenera, together with the distribution of AFLP bands, indicate ancient hybridization and introgression among these Caucasian diploids. • Conclusions The lack of supported hierarchical structure suggests that extensive reticulate evolution between these groups, even at the diploid level, has occurred (although an alternative explanation, namely ancestral polymorphism in ITS data, cannot be completely excluded). Several implications for the investigation of the polyploid complexes of concern are drawn. When tracing origins of polyploid taxa, a much more complex scenario should be expected, taking into account all relatives as potential parents, irrespective of the group in which they are classified. PMID:15037449

Nuclear markers reveal that inter-lake cichlids' similar morphologies do not reflect similar genealogy.

PubMed

Kassam, Daud; Seki, Shingo; Horic, Michio; Yamaoka, Kosaku

2006-08-01

The apparent inter-lake morphological similarity among East African Great Lakes' cichlid species/genera has left evolutionary biologists asking whether such similarity is due to sharing of common ancestor or mere convergent evolution. In order to answer such question, we first used Geometric Morphometrics, GM, to quantify morphological similarity and then subsequently used Amplified Fragment Length Polymorphism, AFLP, to determine if similar morphologies imply shared ancestry or convergent evolution. GM revealed that not all presumed morphological similar pairs were indeed similar, and the dendrogram generated from AFLP data indicated distinct clusters corresponding to each lake and not inter-lake morphological similar pairs. Such results imply that the morphological similarity is due to convergent evolution and not shared ancestry. The congruency of GM and AFLP generated dendrograms imply that GM is capable of picking up phylogenetic signal, and thus GM can be potential tool in phylogenetic systematics.

Evaluation of a Modified Single-Enzyme Amplified-Fragment Length Polymorphism Technique for Fingerprinting and Differentiating of Mycobacterium kansasii Type I Isolates

PubMed Central

Gaafar, Ayman; Josebe Unzaga, M.; Cisterna, Ramón; Clavo, Felicitas Elena; Urra, Elena; Ayarza, Rafael; Martín, Gloria

2003-01-01

The usefulness of single-enzyme amplified-fragment length polymorphism (AFLP) analysis for the subtyping of Mycobacterium kansasii type I isolates was evaluated. This simplified technique classified 253 type I strains into 12 distinct clusters. The discriminating power of this technique was high, and the technique easily distinguished between the epidemiologically unrelated control strains and our clinical isolates. Overall, the technique was relatively rapid and technically simple, yet it gave reproducible and discriminatory results. This technique provides a powerful typing tool which may be helpful in solving many questions concerning the reservoirs, pathogenicities, and modes of transmission of these isolates. PMID:12904399

Levels of Intra-specific AFLP Diversity in Tuber-Bearing Potato Species with Different Breeding Systems and Ploidy Levels

PubMed Central

Bryan, Glenn J.; McLean, Karen; Waugh, Robbie; Spooner, David M.

2017-01-01

DNA-based marker analysis of plant genebank material has become a useful tool in the evaluation of levels of genetic diversity and for the informed use and maintenance of germplasm. In this study, we quantify levels of amplified fragment length polymorphism (AFLP) in representative accessions of wild and cultivated potato species of differing geographic origin, ploidy, and breeding system. We generated 449 polymorphic AFLP fragments in 619 plants, representing multiple plants (16–23) from 17 accessions of 14 potato taxa as well as single plants sampled from available accessions (from 3 to 56) of the same 14 taxa. Intra-accession diversities were compared to those of a synthetic ‘taxon-wide’ population comprising a single individual from a variable number of available accessions of each sampled taxon. Results confirm the expected considerably lower levels of polymorphism within accessions of self-compatible as compared to self-incompatible taxa. We observed broadly similar levels of ‘taxon-wide’ polymorphism among self-compatible and self-incompatible species, with self-compatible taxa showing only slightly lower rates of polymorphism. The most diverse accessions were the two cultivated potato accessions examined, the least diverse being the Mexican allohexaploids Solanum demissum and S. iopetalum. Generally allopolyploid self-compatible accessions exhibited lower levels of diversity. Some purported self-incompatible accessions showed relatively low levels of marker diversity, similar to the more diverse self-compatible material surveyed. Our data indicate that for self-compatible species a single plant is highly representative of a genebank accession. The situation for self-incompatible taxa is less clear, and sampling strategies used will depend on the type of investigation. These results have important implications for those seeking novel trait variation (e.g., disease resistance) in gene banks as well as for the selection of individuals for genomics studies. We also show that AFLPs, despite having been largely replaced by other marker types, is highly suitable for the evaluation of within and between accession diversity in genebanks. PMID:28983315

Does epigenetic polymorphism contribute to phenotypic variances in Jatropha curcas L.?

PubMed

Yi, Chengxin; Zhang, Shilu; Liu, Xiaokun; Bui, Ha T N; Hong, Yan

2010-11-23

There is a growing interest in Jatropha curcas L. (jatropha) as a biodiesel feedstock plant. Variations in its morphology and seed productivity have been well documented. However, there is the lack of systematic comparative evaluation of distinct collections under same climate and agronomic practices. With the several reports on low genetic diversity in jatropha collections, there is uncertainty on genetic contribution to jatropha morphology. In this study, five populations of jatropha plants collected from China (CN), Indonesia (MD), Suriname (SU), Tanzania (AF) and India (TN) were planted in one farm under the same agronomic practices. Their agronomic traits (branching pattern, height, diameter of canopy, time to first flowering, dormancy, accumulated seed yield and oil content) were observed and tracked for two years. Significant variations were found for all the agronomic traits studied. Genetic diversity and epigenetic diversity were evaluated using florescence Amplified Fragment Length Polymorphism (fAFLP) and methylation sensitive florescence AFLP (MfAFLP) methods. Very low level of genetic diversity was detected (polymorphic band <0.1%) within and among populations. In contrast, intermediate but significant epigenetic diversity was detected (25.3% of bands were polymorphic) within and among populations. More than half of CCGG sites surveyed by MfAFLP were methylated with significant difference in inner cytosine and double cytosine methylation among populations. Principal coordinates analysis (PCoA) based on Nei's epigenetic distance showed Tanzania/India group distinct from China/Indonesia/Suriname group. Inheritance of epigenetic markers was assessed in one F1 hybrid population between two morphologically distinct parent plants and one selfed population. 30 out of 39 polymorphic markers (77%) were found heritable and followed Mendelian segregation. One epiallele was further confirmed by bisulphite sequencing of its corresponding genomic region. Our study confirmed climate and practice independent differences in agronomic performance among jatropha collections. Such agronomic trait variations, however, were matched by very low genetic diversity and medium level but significant epigenetic diversity. Significant difference in inner cytosine and double cytosine methylation at CCGG sites was also found among populations. Most epigenetic differential markers can be inherited as epialleles following Mendelian segregation. These results suggest possible involvement of epigenetics in jatropha development.

Does epigenetic polymorphism contribute to phenotypic variances in Jatropha curcas L.?

PubMed Central

2010-01-01

Background There is a growing interest in Jatropha curcas L. (jatropha) as a biodiesel feedstock plant. Variations in its morphology and seed productivity have been well documented. However, there is the lack of systematic comparative evaluation of distinct collections under same climate and agronomic practices. With the several reports on low genetic diversity in jatropha collections, there is uncertainty on genetic contribution to jatropha morphology. Result In this study, five populations of jatropha plants collected from China (CN), Indonesia (MD), Suriname (SU), Tanzania (AF) and India (TN) were planted in one farm under the same agronomic practices. Their agronomic traits (branching pattern, height, diameter of canopy, time to first flowering, dormancy, accumulated seed yield and oil content) were observed and tracked for two years. Significant variations were found for all the agronomic traits studied. Genetic diversity and epigenetic diversity were evaluated using florescence Amplified Fragment Length Polymorphism (fAFLP) and methylation sensitive florescence AFLP (MfAFLP) methods. Very low level of genetic diversity was detected (polymorphic band <0.1%) within and among populations. In contrast, intermediate but significant epigenetic diversity was detected (25.3% of bands were polymorphic) within and among populations. More than half of CCGG sites surveyed by MfAFLP were methylated with significant difference in inner cytosine and double cytosine methylation among populations. Principal coordinates analysis (PCoA) based on Nei's epigenetic distance showed Tanzania/India group distinct from China/Indonesia/Suriname group. Inheritance of epigenetic markers was assessed in one F1 hybrid population between two morphologically distinct parent plants and one selfed population. 30 out of 39 polymorphic markers (77%) were found heritable and followed Mendelian segregation. One epiallele was further confirmed by bisulphite sequencing of its corresponding genomic region. Conclusion Our study confirmed climate and practice independent differences in agronomic performance among jatropha collections. Such agronomic trait variations, however, were matched by very low genetic diversity and medium level but significant epigenetic diversity. Significant difference in inner cytosine and double cytosine methylation at CCGG sites was also found among populations. Most epigenetic differential markers can be inherited as epialleles following Mendelian segregation. These results suggest possible involvement of epigenetics in jatropha development. PMID:21092236

Natural hybridization within seed sources of shortleaf pine (Pinus echinata Mill.) and loblolly pine (Pinus taeda L.)

Treesearch

Shiqin Xu; C.G. Tauer; C. Dana Nelson

2008-01-01

Shortleaf and loblolly pine trees (n=93 and 102, respectively) from 22 seed sources of the Southwide Southern Pine Seed Source Study plantings or equivalent origin were evaluated for amplified fragment length polymorphism (AFLP) variation. These sampled trees represent shortleaf pine and loblolly pine, as they existed across their native geographic ranges before...

Genetic variability of Brazilian isolates of Alternaria alternata detected by AFLP and RAPD techniques

PubMed Central

Dini-Andreote, Francisco; Pietrobon, Vivian Cristina; Andreote, Fernando Dini; Romão, Aline Silva; Spósito, Marcel Bellato; Araújo, Welington Luiz

2009-01-01

The Alternaria brown spot (ABS) is a disease caused in tangerine plants and its hybrids by the fungus Alternaria alternata f. sp. citri which has been found in Brazil since 2001. Due to the recent occurrence in Brazilian orchards, the epidemiology and genetic variability of this pathogen is still an issue to be addressed. Here it is presented a survey about the genetic variability of this fungus by the characterization of twenty four pathogenic isolates of A. alternata f. sp. citri from citrus plants and four endophytic isolates from mango (one Alternaria tenuissima and three Alternaria arborescens). The application of two molecular markers Random Amplified Polymorphic DNA (RAPD) and Amplified Fragment Length Polymorphism (AFLP) had revealed the isolates clustering in distinct groups when fingerprintings were analyzed by Principal Components Analysis (PCA). Despite the better assessment of the genetic variability through the AFLP, significant modifications in clusters components were not observed, and only slight shifts in the positioning of isolates LRS 39/3 and 25M were observed in PCA plots. Furthermore, in both analyses, only the isolates from lemon plants revealed to be clustered, differently from the absence of clustering for other hosts or plant tissues. Summarizing, both RAPD and AFLP analyses were both efficient to detect the genetic variability within the population of the pathogenic fungus Alternaria spp., supplying information on the genetic variability of this species as a basis for further studies aiming the disease control. PMID:24031413

A high degree of genetic diversity is revealed in Isatis spp. (dyer's woad) by amplified fragment length polymorphism (AFLP).

PubMed

Gilbert (nee Stoker), G.; Garton, S.; Karam, A.; Arnold, M.; Karp, A.; Edwards, J.; Cooke, T.; Barker, A.

2002-05-01

Genetic diversity in 38 genotypes, representing 28 individual genotypes from five landraces of Isatis tinctoria (three German: Tubingen, Potsdam and Erfurt, one Swiss and one English), five genotypes of Isatis indigotica (Chinese woad) and five genotypes of Isatis glauca, were investigated using AFLP analysis. Five primer combinations detected a total of 502 fragments of which 436 (86.9%) were polymorphic. The level of polymorphism recorded within each species was 29.8, 86.9 and 35.8% for I. indigotica, I. tinctoria and I. glauca, respectively. Clearly, genetic diversity within I. tinctoria was greater than that observed in I. indigotica or I. glauca. Cluster analyses of the AFLP data using UPGMA and PCO revealed the complete separation of the genotypes of each species into distinct groups. I. indigotica separated as an entirely independent group, whereas I. glauca formed a separate cluster within the I. tinctoria group. Indeed, I. tinctoria and I. glauca are more closely related to each other than either is to I. indigotica. In addition, the genotypes of each landrace, apart from one from the English group, were clearly discriminated. However, the anomalous genotype did associate with the rest of its group when it was linked with the Erfurt group. These results provide new and useful information about the make-up of the Isatis genome, which has not previously been evaluated. They will be useful in the selection of plant material for variety development and conservation of the gene-pool.

Methylated site display (MSD)-AFLP, a sensitive and affordable method for analysis of CpG methylation profiles.

PubMed

Aiba, Toshiki; Saito, Toshiyuki; Hayashi, Akiko; Sato, Shinji; Yunokawa, Harunobu; Maruyama, Toru; Fujibuchi, Wataru; Kurita, Hisaka; Tohyama, Chiharu; Ohsako, Seiichiroh

2017-03-09

It has been pointed out that environmental factors or chemicals can cause diseases that are developmental in origin. To detect abnormal epigenetic alterations in DNA methylation, convenient and cost-effective methods are required for such research, in which multiple samples are processed simultaneously. We here present methylated site display (MSD), a unique technique for the preparation of DNA libraries. By combining it with amplified fragment length polymorphism (AFLP) analysis, we developed a new method, MSD-AFLP. Methylated site display libraries consist of only DNAs derived from DNA fragments that are CpG methylated at the 5' end in the original genomic DNA sample. To test the effectiveness of this method, CpG methylation levels in liver, kidney, and hippocampal tissues of mice were compared to examine if MSD-AFLP can detect subtle differences in the levels of tissue-specific differentially methylated CpGs. As a result, many CpG sites suspected to be tissue-specific differentially methylated were detected. Nucleotide sequences adjacent to these methyl-CpG sites were identified and we determined the methylation level by methylation-sensitive restriction endonuclease (MSRE)-PCR analysis to confirm the accuracy of AFLP analysis. The differences of the methylation level among tissues were almost identical among these methods. By MSD-AFLP analysis, we detected many CpGs showing less than 5% statistically significant tissue-specific difference and less than 10% degree of variability. Additionally, MSD-AFLP analysis could be used to identify CpG methylation sites in other organisms including humans. MSD-AFLP analysis can potentially be used to measure slight changes in CpG methylation level. Regarding the remarkable precision, sensitivity, and throughput of MSD-AFLP analysis studies, this method will be advantageous in a variety of epigenetics-based research.

Amplified Fragment Length Polymorphism Fingerprinting of Pseudomonas Strains from a Poultry Processing Plant

PubMed Central

Geornaras, Ifigenia; Kunene, Nokuthula F.; von Holy, Alexander; Hastings, John W.

1999-01-01

Molecular typing has been used previously to identify and trace dissemination of pathogenic and spoilage bacteria associated with food processing. Amplified fragment length polymorphism (AFLP) is a novel DNA fingerprinting technique which is considered highly reproducible and has high discriminatory power. This technique was used to fingerprint 88 Pseudomonas fluorescens and Pseudomonas putida strains that were previously isolated from plate counts of carcasses at six processing stages and various equipment surfaces and environmental sources of a poultry abattoir. Clustering of the AFLP patterns revealed a high level of diversity among the strains. Six clusters (clusters I through VI) were delineated at an arbitrary Dice coefficient level of 0.65; clusters III (31 strains) and IV (28 strains) were the largest clusters. More than one-half (52.3%) of the strains obtained from carcass samples, which may have represented the resident carcass population, grouped together in cluster III. By contrast, 43.2% of the strains from most of the equipment surfaces and environmental sources grouped together in cluster IV. In most cases, the clusters in which carcass strains from processing stages grouped corresponded to the clusters in which strains from the associated equipment surfaces and/or environmental sources were found. This provided evidence that there was cross-contamination between carcasses and the abattoir environment at the DNA level. The AFLP data also showed that strains were being disseminated from the beginning to the end of the poultry processing operation, since many strains associated with carcasses at the packaging stage were members of the same clusters as strains obtained from carcasses after the defeathering stage. PMID:10473382

Amplified fragment length polymorphism fingerprinting of Pseudomonas strains from a poultry processing plant.

PubMed

Geornaras, I; Kunene, N F; von Holy, A; Hastings, J W

1999-09-01

Molecular typing has been used previously to identify and trace dissemination of pathogenic and spoilage bacteria associated with food processing. Amplified fragment length polymorphism (AFLP) is a novel DNA fingerprinting technique which is considered highly reproducible and has high discriminatory power. This technique was used to fingerprint 88 Pseudomonas fluorescens and Pseudomonas putida strains that were previously isolated from plate counts of carcasses at six processing stages and various equipment surfaces and environmental sources of a poultry abattoir. Clustering of the AFLP patterns revealed a high level of diversity among the strains. Six clusters (clusters I through VI) were delineated at an arbitrary Dice coefficient level of 0.65; clusters III (31 strains) and IV (28 strains) were the largest clusters. More than one-half (52.3%) of the strains obtained from carcass samples, which may have represented the resident carcass population, grouped together in cluster III. By contrast, 43.2% of the strains from most of the equipment surfaces and environmental sources grouped together in cluster IV. In most cases, the clusters in which carcass strains from processing stages grouped corresponded to the clusters in which strains from the associated equipment surfaces and/or environmental sources were found. This provided evidence that there was cross-contamination between carcasses and the abattoir environment at the DNA level. The AFLP data also showed that strains were being disseminated from the beginning to the end of the poultry processing operation, since many strains associated with carcasses at the packaging stage were members of the same clusters as strains obtained from carcasses after the defeathering stage.

Epigenetic differentiation and relationship to adaptive genetic divergence in discrete populations of the violet Viola cazorlensis.

PubMed

Herrera, Carlos M; Bazaga, Pilar

2010-08-01

*In plants, epigenetic variations based on DNA methylation are often heritable and could influence the course of evolution. Before this hypothesis can be assessed, fundamental questions about epigenetic variation remain to be addressed in a real-world context, including its magnitude, structuring within and among natural populations, and autonomy in relation to the genetic context. *Extent and patterns of cytosine methylation, and the relationship to adaptive genetic divergence between populations, were investigated for wild populations of the southern Spanish violet Viola cazorlensis (Violaceae) using the methylation-sensitive amplified polymorphism (MSAP) technique, a modification of the amplified fragment length polymorphism method (AFLP) based on the differential sensitivity of isoschizomeric restriction enzymes to site-specific cytosine methylation. *The genome of V. cazorlensis plants exhibited extensive levels of methylation, and methylation-based epigenetic variation was structured into distinct between- and within- population components. Epigenetic differentiation of populations was correlated with adaptive genetic divergence revealed by a Bayesian population-genomic analysis of AFLP data. Significant associations existed at the individual genome level between adaptive AFLP loci and the methylation state of methylation-susceptible MSAP loci. *Population-specific, divergent patterns of correlated selection on epigenetic and genetic individual variation could account for the coordinated epigenetic-genetic adaptive population differentiation revealed by this study.

Identification of Genes Related to Paulownia Witches’ Broom by AFLP and MSAP

PubMed Central

Cao, Xibing; Fan, Guoqiang; Deng, Minjie; Zhao, Zhenli; Dong, Yanpeng

2014-01-01

DNA methylation is believed to play important roles in regulating gene expression in plant growth and development. Paulownia witches’ broom (PaWB) infection has been reported to be related to gene expression changes in paulownia plantlets. To determine whether DNA methylation is associated with gene expression changes in response to phytoplasma, we investigated variations in genomic DNA sequence and methylation in PaWB plantlets treated with methyl methane sulfonate (MMS) using amplified fragment length polymorphism (AFLP) and methylation-sensitive amplification polymorphism (MSAP) techniques, respectively. The results indicated that PaWB seedings recovered a normal morphology after treatment with more than 15 mg·L−1 MMS. PaWB infection did not cause changes of the paulownia DNA sequence at the AFLP level; However, DNA methylation levels and patterns were altered. Quantitative real-time PCR (qRT-PCR) showed that three of the methylated genes were up-regulated and three were down-regulated in the MMS-treated PaWB plantlets that had regained healthy morphology. These six genes might be involved in transcriptional regulation, plant defense, signal transduction and energy. The possible roles of these genes in PaWB are discussed. The results showed that changes of DNA methylation altered gene expression levels, and that MSAP might help identify genes related to PaWB. PMID:25196603

Identification of genes related to Paulownia witches' broom by AFLP and MSAP.

PubMed

Cao, Xibing; Fan, Guoqiang; Deng, Minjie; Zhao, Zhenli; Dong, Yanpeng

2014-08-21

DNA methylation is believed to play important roles in regulating gene expression in plant growth and development. Paulownia witches' broom (PaWB) infection has been reported to be related to gene expression changes in paulownia plantlets. To determine whether DNA methylation is associated with gene expression changes in response to phytoplasma, we investigated variations in genomic DNA sequence and methylation in PaWB plantlets treated with methyl methane sulfonate (MMS) using amplified fragment length polymorphism (AFLP) and methylation-sensitive amplification polymorphism (MSAP) techniques, respectively. The results indicated that PaWB seedings recovered a normal morphology after treatment with more than 15 mg·L(-1) MMS. PaWB infection did not cause changes of the paulownia DNA sequence at the AFLP level; However, DNA methylation levels and patterns were altered. Quantitative real-time PCR (qRT-PCR) showed that three of the methylated genes were up-regulated and three were down-regulated in the MMS-treated PaWB plantlets that had regained healthy morphology. These six genes might be involved in transcriptional regulation, plant defense, signal transduction and energy. The possible roles of these genes in PaWB are discussed. The results showed that changes of DNA methylation altered gene expression levels, and that MSAP might help identify genes related to PaWB.

Evaluation of genetic diversity of Panicum turgidum Forssk from Saudi Arabia.

PubMed

Assaeed, Abdulaziz M; Al-Faifi, Sulieman A; Migdadi, Hussein M; El-Bana, Magdy I; Al Qarawi, Abdulaziz A; Khan, Mohammad Altaf

2018-01-01

The genetic diversity of 177 accessions of Panicum turgidum Forssk, representing ten populations collected from four geographical regions in Saudi Arabia, was analyzed using amplified fragment length polymorphism (AFLP) markers. A set of four primer-pairs with two/three selective nucleotides scored 836 AFLP amplified fragments (putative loci/genome landmarks), all of which were polymorphic. Populations collected from the southern region of the country showed the highest genetic diversity parameters, whereas those collected from the central regions showed the lowest values. Analysis of molecular variance (AMOVA) revealed that 78% of the genetic variability was attributable to differences within populations. Pairwise values for population differentiation and genetic structure were statistically significant for all variances. The UPGMA dendrogram, validated by principal coordinate analysis-grouped accessions, corresponded to the geographical origin of the accessions. Mantel's test showed that there was a significant correlation between the genetic and geographical distances ( r  = 0.35, P  < 0.04). In summary, the AFLP assay demonstrated the existence of substantial genetic variation in P. turgidum . The relationship between the genetic diversity and geographical source of P. turgidum populations of Saudi Arabia, as revealed through this comprehensive study, will enable effective resource management and restoration of new areas without compromising adaptation and genetic diversity.

A genetic linkage map of the Durum x Triticum dicoccoides backcross population based on SSRs and AFLP markers, and QTL analysis for milling traits.

PubMed

Elouafi, I; Nachit, M M

2004-02-01

Durum wheat ( Triticum turgidum L. var durum) is mainly produced and consumed in the Mediterranean region; it is used to produce several specific end-products; such as local pasta, couscous and burghul. To study the genetics of grain-milling quality traits, chromosomal locations, and interaction with the environment, a genetic linkage map of durum was constructed and the quantitative trait loci QTLs for the milling-related traits, test weight (TW) and thousand-kernel weight (TKW), were identified. The population constituted 114 recombinant inbred lines derived from the cross: Omrabi 5 /Triticum dicoccoides 600545// Omrabi 5. TW and TKW were analyzed over 18 environments (sites x years). Single-sequence-repeat markers (SSRs), Amplified-fragment-length-polymorphism markers (AFLPs), and seed storage proteins (SSPs) showed a high level of polymorphism (>60%). The map was constructed with 124 SSRs, 149 AFLPs and 6 SSPs; its length covered 2,288.8 cM (8.2 cM/marker). The map showed high synteny with previous wheat maps, and both SSRs and AFLPs mapped evenly across the genome, with more markers in the B genome. However, some rearrangements were observed. For TW, a high genotypic effect was detected and two QTLs with epistasic effect were identified on 7AS and 6BS, explaining 30% of the total variation. The TKW showed a significant transgressive inheritance and five QTLs were identified, explaining 32% of the total variation, out of which 25% was of a genetic nature, and showing QTLxE interaction. The major TKW-QTLs were around the centromere region of 6B. For both traits, Omrabi 5 alleles had a significant positive effect. This population will be used to determine other QTLs of interest, as its parents are likely to harbor different genes for diseases and drought tolerance.

Molecular Linkage Mapping and Marker-Trait Associations with NlRPT, a Downy Mildew Resistance Gene in Nicotiana langsdorffii

PubMed Central

Zhang, Shouan; Gao, Muqiang; Zaitlin, David

2012-01-01

Nicotiana langsdorffii is one of two species of Nicotiana known to express an incompatible interaction with the oomycete Peronospora tabacina, the causal agent of tobacco blue mold disease. We previously showed that incompatibility is due to the hypersensitive response (HR), and plants expressing the HR are resistant to P. tabacina at all stages of growth. Resistance is due to a single dominant gene in N. langsdorffii accession S-4-4 that we have named NlRPT. In further characterizing this unique host-pathogen interaction, NlRPT has been placed on a preliminary genetic map of the N. langsdorffii genome. Allelic scores for five classes of DNA markers were determined for 90 progeny of a “modified backcross” involving two N. langsdorffii inbred lines and the related species N. forgetiana. All markers had an expected segregation ratio of 1:1, and were scored in a common format. The map was constructed with JoinMap 3.0, and loci showing excessive transmission distortion were removed. The linkage map consists of 266 molecular marker loci defined by 217 amplified fragment length polymorphisms (AFLPs), 26 simple-sequence repeats (SSRs), 10 conserved orthologous sequence markers, nine inter-simple sequence repeat markers, and four target region amplification polymorphism markers arranged in 12 linkage groups with a combined length of 1062 cM. NlRPT is located on linkage group three, flanked by four AFLP markers and one SSR. Regions of skewed segregation were detected on LGs 1, 5, and 9. Markers developed for N. langsdorffii are potentially useful genetic tools for other species in Nicotiana section Alatae, as well as in N. benthamiana. We also investigated whether AFLPs could be used to infer genetic relationships within N. langsdorffii and related species from section Alatae. A phenetic analysis of the AFLP data showed that there are two main lineages within N. langsdorffii, and that both contain populations expressing dominant resistance to P. tabacina. PMID:22936937

Epigenetic Variance, Performing Cooperative Structure with Genetics, Is Associated with Leaf Shape Traits in Widely Distributed Populations of Ornamental Tree Prunus mume

PubMed Central

Ma, Kaifeng; Sun, Lidan; Cheng, Tangren; Pan, Huitang; Wang, Jia; Zhang, Qixiang

2018-01-01

Increasing evidence shows that epigenetics plays an important role in phenotypic variance. However, little is known about epigenetic variation in the important ornamental tree Prunus mume. We used amplified fragment length polymorphism (AFLP) and methylation-sensitive amplified polymorphism (MSAP) techniques, and association analysis and sequencing to investigate epigenetic variation and its relationships with genetic variance, environment factors, and traits. By performing leaf sampling, the relative total methylation level (29.80%) was detected in 96 accessions of P. mume. And the relative hemi-methylation level (15.77%) was higher than the relative full methylation level (14.03%). The epigenetic diversity (I∗ = 0.575, h∗ = 0.393) was higher than the genetic diversity (I = 0.484, h = 0.319). The cultivated population displayed greater epigenetic diversity than the wild populations in both southwest and southeast China. We found that epigenetic variance and genetic variance, and environmental factors performed cooperative structures, respectively. In particular, leaf length, width and area were positively correlated with relative full methylation level and total methylation level, indicating that the DNA methylation level played a role in trait variation. In total, 203 AFLP and 423 MSAP associated markers were detected and 68 of them were sequenced. Homologous analysis and functional prediction suggested that the candidate marker-linked genes were essential for leaf morphology development and metabolism, implying that these markers play critical roles in the establishment of leaf length, width, area, and ratio of length to width. PMID:29441078

Epigenetic Variance, Performing Cooperative Structure with Genetics, Is Associated with Leaf Shape Traits in Widely Distributed Populations of Ornamental Tree Prunus mume.

PubMed

Ma, Kaifeng; Sun, Lidan; Cheng, Tangren; Pan, Huitang; Wang, Jia; Zhang, Qixiang

2018-01-01

Increasing evidence shows that epigenetics plays an important role in phenotypic variance. However, little is known about epigenetic variation in the important ornamental tree Prunus mume . We used amplified fragment length polymorphism (AFLP) and methylation-sensitive amplified polymorphism (MSAP) techniques, and association analysis and sequencing to investigate epigenetic variation and its relationships with genetic variance, environment factors, and traits. By performing leaf sampling, the relative total methylation level (29.80%) was detected in 96 accessions of P . mume . And the relative hemi-methylation level (15.77%) was higher than the relative full methylation level (14.03%). The epigenetic diversity ( I ∗ = 0.575, h ∗ = 0.393) was higher than the genetic diversity ( I = 0.484, h = 0.319). The cultivated population displayed greater epigenetic diversity than the wild populations in both southwest and southeast China. We found that epigenetic variance and genetic variance, and environmental factors performed cooperative structures, respectively. In particular, leaf length, width and area were positively correlated with relative full methylation level and total methylation level, indicating that the DNA methylation level played a role in trait variation. In total, 203 AFLP and 423 MSAP associated markers were detected and 68 of them were sequenced. Homologous analysis and functional prediction suggested that the candidate marker-linked genes were essential for leaf morphology development and metabolism, implying that these markers play critical roles in the establishment of leaf length, width, area, and ratio of length to width.

Genetic Comparison of B. Anthracis and its Close Relatives Using AFLP and PCR Analysis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jackson, P.J.; Hill, K.K.; Laker, M.T.

1999-02-01

Amplified Fragment length Polymorphism (AFLP) analysis allows a rapid, relatively simple analysis of a large portion of a microbial genome, providing information about the species and its phylogenetic relationship to other microbes (Vos, et al., 1995). The method simply surveys the genome for length and sequence polymorphisms. The pattern identified can be used for comparison to the genomes of other species. Unlike other methods, it does not rely on analysis of a single genetic locus that may bias the interpretation of results and it does not require any prior knowledge of the targeted organism. Moreover, a standard set of reagentsmore » can be applied to any species without using species-specific information or molecular probes. The authors are using AFLP's to rapidly identify different bacterial species. A comparison of AFLP profiles generated from a large battery of B. anthracis strains shows very little variability among different isolates (Keim, et al., 1997). By contrast, there is a significant difference between AFLP profiles generated for any B. anthracis strain and even the most closely related Bacillus species. Sufficient variability is apparent among all known microbial species to allow phylogenetic analysis based on large numbers of genetically unlinked loci. These striking differences among AFLP profiles allow unambiguous identification of previously identified species and phylogenetic placement of newly characterized isolates relative to known species based on a large number of independent genetic loci. Data generated thus far show that the method provides phylogenetic analyses that are consistent with other widely accepted phylogenetic methods. However, AFLP analysis provides a more detailed analysis of the targets and samples a much larger portion of the genome. Consequently, it provides an inexpensive, rapid means of characterizing microbial isolates to further differentiate among strains and closely related microbial species. Such information cannot be rapidly generated by other means. AFLP sample analysis quickly generates a very large amount of molecular information about microbial genomes. However, this information cannot be analyzed rapidly using manual methods. The authors are developing a large archive of electronic AFLP signatures that is being used to identify isolates collected from medical, veterinary, forensic and environmental samples. They are also developing the computational packages necessary to rapidly and unambiguously analyze the AFLP profiles and conduct a phylogenetic comparison of these data relative to information already in the database. They will use this archive and the associated algorithms to determine the species identity of previously uncharacterized isolates and place them phylogenetically relative to other microbes based on their AFLP signatures. This study provides significant new information about microbes with environmental, veterinary and medical significance. This information can be used in further studies to understand the relationships among these species and the factors that distinguish them from one another. It should also allow identification of unique factors that contribute to important microbial traits including pathogenicity and virulence. They are also using AFLP data to identify, isolate and sequence DNA fragments that are unique to particular microbial species and strains. The fragment patterns and sequence information provide insights into the complexity and organization of bacterial genomes relative to one another. They also provide the information necessary for development of species-specific PCR primers that can be used to interrogate complex samples for the presence of B. anthracis, other microbial pathogens or their remnants.« less

Species phylogeny and diversification process of Northeast Asian Pungitius revealed by AFLP and mtDNA markers.

PubMed

Takahashi, Hiroshi; Møller, Peter R; Shedko, Sergei V; Ramatulla, Temirbekov; Joen, Sang-Rin; Zhang, Chun-Guang; Sideleva, Valentina G; Takata, Keisuke; Sakai, Harumi; Goto, Akira; Nishida, Mutsumi

2016-06-01

Pungitius is a highly diversified genus of sticklebacks (Gasterosteidae) occurring widely in northern parts of the Northern Hemisphere. Several ecologically and genetically divergent types that are largely isolated reproductively but occasionally hybridize in sympatry have been discovered in Northeast Asia, although the taxonomy and evolutionary relationships among them remain unclear. We used amplified fragment length polymorphism (AFLP) and mitochondrial DNA (mtDNA) markers to infer phylogenies among individuals collected from sympatric and allopatric populations, including the type localities of the described species. Phylogenetic analyses based on 2683 polymorphic AFLP loci confirmed seven species, each of which (except for one entirely allopatric species P. platygaster) was clearly differentiated from one or two other sympatric species and constituted a highly supported monophyletic clade with conspecific allopatric populations. The phylogeny showed that two lineages arose early; one gave rise to two species (circumpolar species P. pungitius and Paratethys species P. platygaster) and the other to five species endemic to Northeast Asia (P. sinensis, P. tymensis, P. polyakovi, P. kaibarae, and P. bussei). The brackish-water, freshwater, and Omono types previously discovered in Japan were reidentified as P. pungitius, P. sinensis, and P. kaibarae, respectively. A marked incongruence was noted between the phylogenies of AFLP and mtDNA markers, suggesting the occasional occurrence of hybridization and mtDNA introgression among distinct species. Our results highlight that the marginal seas of Northeast Asia played a key role as barriers to or facilitators of gene flow in the evolution of species diversity of Pungitius concentrated in this region. Copyright © 2016 Elsevier Inc. All rights reserved.

Acute and chronic gregarisation are associated with distinct DNA methylation fingerprints in desert locusts.

PubMed

Mallon, Eamonn B; Amarasinghe, Harindra E; Ott, Swidbert R

2016-10-18

Desert locusts (Schistocerca gregaria) show a dramatic form of socially induced phenotypic plasticity known as phase polyphenism. In the absence of conspecifics, locusts occur in a shy and cryptic solitarious phase. Crowding with conspecifics drives a behavioural transformation towards gregariousness that occurs within hours and is followed by changes in physiology, colouration and morphology, resulting in the full gregarious phase syndrome. We analysed methylation-sensitive amplified fragment length polymorphisms (MS-AFLP) to compare the effect of acute and chronic crowding on DNA methylation in the central nervous system. We find that crowd-reared and solitary-reared locusts show markedly different neural MS-AFLP fingerprints. However, crowding for a day resulted in neural MS-AFLP fingerprints that were clearly distinct from both crowd-reared and uncrowded solitary-reared locusts. Our results indicate that changes in DNA methylation associated with behavioural gregarisation proceed through intermediate states that are not simply partial realisations of the endpoint states.

Molecular types of Cryptococcus gattii/Cryptococcus neoformans species complex from clinical and environmental sources in Nairobi, Kenya.

PubMed

Kangogo, Mourine; Bader, Oliver; Boga, Hamadi; Wanyoike, Wanjiru; Folba, Claudia; Worasilchai, Navaporn; Weig, Michael; Groß, Uwe; Bii, Christine C

2015-11-01

Cryptococcal meningitis infections cause high mortality rates among HIV-infected patients in Sub-Saharan Africa. The high incidences of cryptococcal infections may be attributed to common environmental sources which, if identified, could lead to institution of appropriate control strategies. To determine the genotypes of Cryptococcus gattii/C. neoformans- species complex from Nairobi, Kenya, 123 clinical and environmental isolates were characterised. Typing was done using orotidine monophosphate pyrophosphorylase (URA5) gene restriction fragment length polymorphism (URA5-RFLP). The majority of the isolates [105/123; 85.4%] were C. neoformans genotype (AFLPI/VNI) and 1.6% AFLP1A/VNB/VNII, whereas (13%) were C. gattii (AFLP4/VGI). This is the first report on the genotypes of C. gattii/C. neoformans species complex from clinical and environmental sources in Nairobi, Kenya and the isolation of C. gattii genotype AFLP4/VGI from the environment in Kenya. © 2015 Blackwell Verlag GmbH.

Analysis of ploidy and the patterns of amplified fragment length polymorphism and methylation sensitive amplified polymorphism in strawberry plants recovered from cryopreservation.

PubMed

Hao, Yu-Jin; You, Chun-Xiang; Deng, Xui-Xin

2002-01-01

Shoot-tips of 10 strawberry genotypes were successfully cryopreserved using a modified encapsulation-dehydration method. All genotypes survived cryopreservation with high survival and regeneration rates. Eight Joho single-bud sibling lines were established as a model system for genetic analysis. Although cytological examination found chromosomal variation in both non-cryopreserved and cryopreserved samples, the ploidy constitution remained relatively stable after cryopreservation. DNA samples digested with MseI and PstI were used for amplified fragmentation length polymorphism (AFLP) assay. In 16 primer combinations, only one, namely, PCCA-MCAG, detected one site where band pattern changed after cryopreservation, which might be contributed to the change in DNA methylation status at PstI recognition site. Methylation sensitive amplified polymorphism (MSAP) assay was carried out for further investigation on the influence of cryopreservation on DNA methylation status. It was found that cryopreservation induced a significant change in DNA methylation status.

Genetic diversity and structure of western white pine (Pinus monticola) in North America: A baseline study for conservation, restoration, and addressing impacts of climate change

Treesearch

Mee-Sook Kim; Bryce A. Richardson; Geral I. McDonald; Ned B. Klopfenstein

2010-01-01

Western white pine (Pinus monticola) is an economically and ecologically important species in western North America that has declined in prominence over the past several decades, mainly due to the introduction of Cronartium ribicola (cause of white pine blister rust) and reduced opportunities for regeneration. Amplified fragment length polymorphism (AFLP) markers were...

Use of inter-simple sequence repeats and amplified fragment length polymorphisms to analyze genetic relationships among small grain-infecting species of ustilago.

PubMed

Menzies, J G; Bakkeren, G; Matheson, F; Procunier, J D; Woods, S

2003-02-01

ABSTRACT In the smut fungi, few features are available for use as taxonomic criteria (spore size, shape, morphology, germination type, and host range). DNA-based molecular techniques are useful in expanding the traits considered in determining relationships among these fungi. We examined the phylogenetic relationships among seven species of Ustilago (U. avenae, U. bullata, U. hordei, U. kolleri, U. nigra, U. nuda, and U. tritici) using inter-simple sequence repeats (ISSRs) and amplified fragment length polymorphisms (AFLPs) to compare their DNA profiles. Fifty-four isolates of different Ustilago spp. were analyzed using ISSR primers, and 16 isolates of Ustilago were studied using AFLP primers. The variability among isolates within species was low for all species except U. bullata. The isolates of U. bullata, U. nuda, and U. tritici were well separated and our data supports their speciation. U. avenae and U. kolleri isolates did not separate from each other and there was little variability between these species. U. hordei and U. nigra isolates also showed little variability between species, but the isolates from each species grouped together. Our data suggest that U. avenae and U. kolleri are monophyletic and should be considered one species, as should U. hordei and U. nigra.

Genetic diversity analysis of isolates belonging to the Photobacterium phosphoreum species group collected from salmon products using AFLP fingerprinting.

PubMed

Jérôme, Marc; Macé, Sabrina; Dousset, Xavier; Pot, Bruno; Joffraud, Jean-Jacques

2016-01-18

An accurate amplified fragment length polymorphism (AFLP) method, including three primer sets for the selective amplification step, was developed to display the phylogenetic position of Photobacterium isolates collected from salmon products. This method was efficient for discriminating the three species Photobacterium phosphoreum, Photobacterium iliopiscarium and Photobacterium kishitanii, until now indistinctly gathered in the P. phosphoreum species group known to be strongly responsible for seafood spoilage. The AFLP fingerprints enabled the isolates to be separated into two main clusters that, according to the type strains, were assigned to the two species P. phosphoreum and P. iliopiscarium. P. kishitanii was not found in the collection. The accuracy of the method was validated by using gyrB-gene sequencing and luxA-gene PCR amplification, which confirmed the species delineation. Most of the isolates of each species were clonally distinct and even those that were isolated from the same source showed some diversity. Moreover, this AFLP method may be an excellent tool for genotyping isolates in bacterial communities and for clarifying our knowledge of the role of the different members of the Photobacterium species group in seafood spoilage. Copyright © 2015 Elsevier B.V. All rights reserved.

Assessment of genetic and epigenetic variation during long-term Taxus cell culture.

PubMed

Fu, Chunhua; Li, Liqin; Wu, Wenjuan; Li, Maoteng; Yu, Xiaoqing; Yu, Longjiang

2012-07-01

Gradual loss of secondary metabolite production is a common obstacle in the development of a large-scale plant cell production system. In this study, cell morphology, paclitaxel (Taxol®) biosynthetic ability, and genetic and epigenetic variations in the long-term culture of Taxus media cv Hicksii cells were assessed over a 5-year period to evaluate the mechanisms of the loss of secondary metabolites biosynthesis capacity in Taxus cell. The results revealed that morphological variations, gradual loss of paclitaxel yield and decreased transcriptional level of paclitaxel biosynthesis key genes occurred during long-term subculture. Genetic and epigenetic variations in these cultures were also studied at different times during culture using amplified fragment-length polymorphism (AFLP), methylation-sensitive amplified polymorphism (MSAP), and high-performance liquid chromatography (HPLC) analyses. A total of 32 primer combinations were used in AFLP amplification, and none of the AFLP loci were found to be polymorphic, thus no major genetic rearrangements were detected in any of the tested samples. However, results from both MSAP and HPLC indicated that there was a higher level of DNA methylation in the low-paclitaxel yielding cell line after long-term culture. Based on these results, we proposed that accumulation of paclitaxel in Taxus cell cultures might be regulated by DNA methylation. To our knowledge, this is the first report of increased methylation with the prolongation of culture time in Taxus cell culture. It provides substantial clues for exploring the gradual loss of the taxol biosynthesis capacity of Taxus cell lines during long-term subculture. DNA methylation maybe involved in the regulation of paclitaxel biosynthesis in Taxus cell culture.

Genetic diversity analysis of common beans based on molecular markers

PubMed Central

Gill-Langarica, Homar R.; Muruaga-Martínez, José S.; Vargas-Vázquez, M.L. Patricia; Rosales-Serna, Rigoberto; Mayek-Pérez, Netzahualcoyotl

2011-01-01

A core collection of the common bean (Phaseolus vulgaris L.), representing genetic diversity in the entire Mexican holding, is kept at the INIFAP (Instituto Nacional de Investigaciones Forestales, Agricolas y Pecuarias, Mexico) Germplasm Bank. After evaluation, the genetic structure of this collection (200 accessions) was compared with that of landraces from the states of Oaxaca, Chiapas and Veracruz (10 genotypes from each), as well as a further 10 cultivars, by means of four amplified fragment length polymorphisms (AFLP) +3/+3 primer combinations and seven simple sequence repeats (SSR) loci, in order to define genetic diversity, variability and mutual relationships. Data underwent cluster (UPGMA) and molecular variance (AMOVA) analyses. AFLP analysis produced 530 bands (88.5% polymorphic) while SSR primers amplified 174 alleles, all polymorphic (8.2 alleles per locus). AFLP indicated that the highest genetic diversity was to be found in ten commercial-seed classes from two major groups of accessions from Central Mexico and Chiapas, which seems to be an important center of diversity in the south. A third group included genotypes from Nueva Granada, Mesoamerica, Jalisco and Durango races. Here, SSR analysis indicated a reduced number of shared haplotypes among accessions, whereas the highest genetic components of AMOVA variation were found within accessions. Genetic diversity observed in the common-bean core collection represents an important sample of the total Phaseolus genetic variability at the main Germplasm Bank of INIFAP. Molecular marker strategies could contribute to a better understanding of the genetic structure of the core collection as well as to its improvement and validation. PMID:22215964

Genetic diversity analysis of common beans based on molecular markers.

PubMed

Gill-Langarica, Homar R; Muruaga-Martínez, José S; Vargas-Vázquez, M L Patricia; Rosales-Serna, Rigoberto; Mayek-Pérez, Netzahualcoyotl

2011-10-01

A core collection of the common bean (Phaseolus vulgaris L.), representing genetic diversity in the entire Mexican holding, is kept at the INIFAP (Instituto Nacional de Investigaciones Forestales, Agricolas y Pecuarias, Mexico) Germplasm Bank. After evaluation, the genetic structure of this collection (200 accessions) was compared with that of landraces from the states of Oaxaca, Chiapas and Veracruz (10 genotypes from each), as well as a further 10 cultivars, by means of four amplified fragment length polymorphisms (AFLP) +3/+3 primer combinations and seven simple sequence repeats (SSR) loci, in order to define genetic diversity, variability and mutual relationships. Data underwent cluster (UPGMA) and molecular variance (AMOVA) analyses. AFLP analysis produced 530 bands (88.5% polymorphic) while SSR primers amplified 174 alleles, all polymorphic (8.2 alleles per locus). AFLP indicated that the highest genetic diversity was to be found in ten commercial-seed classes from two major groups of accessions from Central Mexico and Chiapas, which seems to be an important center of diversity in the south. A third group included genotypes from Nueva Granada, Mesoamerica, Jalisco and Durango races. Here, SSR analysis indicated a reduced number of shared haplotypes among accessions, whereas the highest genetic components of AMOVA variation were found within accessions. Genetic diversity observed in the common-bean core collection represents an important sample of the total Phaseolus genetic variability at the main Germplasm Bank of INIFAP. Molecular marker strategies could contribute to a better understanding of the genetic structure of the core collection as well as to its improvement and validation.

Chromosomal assignment of ALFP markers in upland cotton (Gossypium hirsutum L.)

USDA-ARS?s Scientific Manuscript database

In this research, we used two sets of cotton aneuploid (G. hirsutum × G. tomentosum and G. hirsutum × G. barbadense) plants to locate AFLP markers to chromosome using deletion analysis method. Thirty-eight primer combinations were used to generate 608 polymorphic AFLP markers. Ninety-eight AFLP mark...

Evidence of genotypic diversity among Candida auris isolates by multilocus sequence typing, matrix-assisted laser desorption ionization time-of-flight mass spectrometry and amplified fragment length polymorphism.

PubMed

Prakash, A; Sharma, C; Singh, A; Kumar Singh, P; Kumar, A; Hagen, F; Govender, N P; Colombo, A L; Meis, J F; Chowdhary, A

2016-03-01

Candida auris is a multidrug-resistant nosocomial bloodstream pathogen that has been reported from Asian countries and South Africa. Herein, we studied the population structure and genetic relatedness among 104 global C. auris isolates from India, South Africa and Brazil using multilocus sequence typing (MLST), amplified fragment length polymorphism (AFLP) fingerprinting and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS). RPB1, RPB2 and internal transcribed spacer (ITS) and D1/D2 regions of the ribosomal DNA were sequenced for MLST. Further, genetic variation and proteomic assessment was carried out using AFLP and MALDI-TOF MS, respectively. Both MLST and AFLP typing clearly demarcated two major clusters comprising Indian and Brazilian isolates. However, the South African isolates were randomly distributed, suggesting different genotypes. MALDI-TOF MS spectral profiling also revealed evidence of geographical clustering but did not correlate fully with the genotyping methods. Notably, overall the population structure of C. auris showed evidence of geographical clustering by all the three techniques analysed. Antifungal susceptibility testing by the CLSI microbroth dilution method revealed that fluconazole had limited activity against 87% of isolates (MIC90, 64 mg/L). Also, MIC90 of AMB was 4 mg/L. Candida auris is emerging as an important yeast pathogen globally and requires reproducible laboratory methods for identification and typing. Evaluation of MALDI-TOF MS as a typing method for this yeast is warranted. Copyright © 2015 European Society of Clinical Microbiology and Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Genetic analyses of captive Alala (Corvus hawaiiensis) using AFLP analyses

USGS Publications Warehouse

Jarvi, Susan I.; Bianchi, Kiara R.

2006-01-01

Population level studies of genetic diversity can provide information about population structure, individual genetic distinctiveness and former population size. They are especially important for rare and threatened species like the Alala, where they can be used to assess extinction risks and evolutionary potential. In an ideal situation multiple methods should be used to detect variation, and these methods should be comparable across studies. In this report, we discuss AFLP (Amplified Fragment Length Polymorphism) as a genetic approach for detecting variation in the Alala , describe our findings, and discuss these in relation to mtDNA and microsatellite data reported elsewhere in this same population. AFLP is a technique for DNA fingerprinting that has wide applications. Because little or no prior knowledge of the particular species is required to carry out this method of analysis, AFLP can be used universally across varied taxonomic groups. Within individuals, estimates of diversity or heterozygosity across genomes may be complex because levels of diversity differ between and among genes. One of the more traditional methods of estimating diversity employs the use of codominant markers such as microsatellites. Codominant markers detect each allele at a locus independently. Hence, one can readily distinguish heterozygotes from homozygotes, directly assess allele frequencies and calculate other population level statistics. Dominant markers (for example, AFLP) are scored as either present or absent (null) so heterozygotes cannot be directly distinguished from homozygotes. However, the presence or absence data can be converted to expected heterozygosity estimates which are comparable to those determined by codominant markers. High allelic diversity and heterozygosity inherent in microsatellites make them excellent tools for studies of wild populations and they have been used extensively. One limitation to the use of microsatellites is that heterozygosity estimates are affected by the mutation rate at microsatellite loci, thus introducing a bias. Also, the number of loci that can be studied is frequently limited to fewer than 10. This theoretically represents a maximum of one marker for each of 10 chromosomes. Dominant markers like AFLP allow a larger fraction of the genome to be screened. Large numbers of loci can be screened by AFLP to resolve very small individual differences that can be used for identification of individuals, estimates of pairwise relatedness and, in some cases, for parentage analyses. Since AFLP is a dominant marker (can not distinguish between +/+ homozygote versus +/- heterozygote), it has limitations for parentage analyses. Only when both parents are homozygous for the absence of alleles (-/-) and offspring show a presence (+/+ or +/-) can the parents be excluded. In this case, microsatellites become preferable as they have the potential to exclude individual parents when the other parent is unknown. Another limitation of AFLP is that the loci are generally less polymorphic (only two alleles/locus) than microsatellite loci (often >10 alleles/locus). While generally fewer than 10 highly polymorphic microsatellite loci are enough to exclude and assign parentage, it might require up to 100 or more AFLP loci. While there are pros and cons to different methodologies, the total number of loci evaluated by AFLP generally offsets the limitations imposed due to the dominant nature of this approach and end results between methods are generally comparable. Overall objectives of this study were to evaluate the level of genetic diversity in the captive population of Alala, to compare genetic data with currently available pedigree information, and to determine the extent of relatedness of mating pairs and among founding individuals.

Construction of integrated linkage map of a recombinant inbred line population of white lupin (Lupinus albus L.)

PubMed Central

Vipin, Cina Ann; Luckett, David J.; Harper, John D.I.; Ash, Gavin J.; Kilian, Andrzej; Ellwood, Simon R.; Phan, Huyen T.T.; Raman, Harsh

2013-01-01

We report the development of a Diversity Arrays Technology (DArT) marker panel and its utilisation in the development of an integrated genetic linkage map of white lupin (Lupinus albus L.) using an F8 recombinant inbred line population derived from Kiev Mutant/P27174. One hundred and thirty-six DArT markers were merged into the first genetic linkage map composed of 220 amplified fragment length polymorphisms (AFLPs) and 105 genic markers. The integrated map consists of 38 linkage groups of 441 markers and spans a total length of 2,169 cM, with an average interval size of 4.6 cM. The DArT markers exhibited good genome coverage and were associated with previously identified genic and AFLP markers linked with quantitative trait loci for anthracnose resistance, flowering time and alkaloid content. The improved genetic linkage map of white lupin will aid in the identification of markers for traits of interest and future syntenic studies. PMID:24273424

Phylogenetic analysis of Bacillus subtilis strains applicable to natto (fermented soybean) production.

PubMed

Kubo, Yuji; Rooney, Alejandro P; Tsukakoshi, Yoshiki; Nakagawa, Rikio; Hasegawa, Hiromasa; Kimura, Keitarou

2011-09-01

Spore-forming Bacillus strains that produce extracellular poly-γ-glutamic acid were screened for their application to natto (fermented soybean food) fermentation. Among the 424 strains, including Bacillus subtilis and B. amyloliquefaciens, which we isolated from rice straw, 59 were capable of fermenting natto. Biotin auxotrophism was tightly linked to natto fermentation. A multilocus nucleotide sequence of six genes (rpoB, purH, gyrA, groEL, polC, and 16S rRNA) was used for phylogenetic analysis, and amplified fragment length polymorphism (AFLP) analysis was also conducted on the natto-fermenting strains. The ability to ferment natto was inferred from the two principal components of the AFLP banding pattern, and natto-fermenting strains formed a tight cluster within the B. subtilis subsp. subtilis group.

Phylogenetic Analysis of Bacillus subtilis Strains Applicable to Natto (Fermented Soybean) Production ▿

PubMed Central

Kubo, Yuji; Rooney, Alejandro P.; Tsukakoshi, Yoshiki; Nakagawa, Rikio; Hasegawa, Hiromasa; Kimura, Keitarou

2011-01-01

Spore-forming Bacillus strains that produce extracellular poly-γ-glutamic acid were screened for their application to natto (fermented soybean food) fermentation. Among the 424 strains, including Bacillus subtilis and B. amyloliquefaciens, which we isolated from rice straw, 59 were capable of fermenting natto. Biotin auxotrophism was tightly linked to natto fermentation. A multilocus nucleotide sequence of six genes (rpoB, purH, gyrA, groEL, polC, and 16S rRNA) was used for phylogenetic analysis, and amplified fragment length polymorphism (AFLP) analysis was also conducted on the natto-fermenting strains. The ability to ferment natto was inferred from the two principal components of the AFLP banding pattern, and natto-fermenting strains formed a tight cluster within the B. subtilis subsp. subtilis group. PMID:21764950

Sequence-related amplified polymorphism (SRAP) markers: A potential resource for studies in plant molecular biology(1.).

PubMed

Robarts, Daniel W H; Wolfe, Andrea D

2014-07-01

In the past few decades, many investigations in the field of plant biology have employed selectively neutral, multilocus, dominant markers such as inter-simple sequence repeat (ISSR), random-amplified polymorphic DNA (RAPD), and amplified fragment length polymorphism (AFLP) to address hypotheses at lower taxonomic levels. More recently, sequence-related amplified polymorphism (SRAP) markers have been developed, which are used to amplify coding regions of DNA with primers targeting open reading frames. These markers have proven to be robust and highly variable, on par with AFLP, and are attained through a significantly less technically demanding process. SRAP markers have been used primarily for agronomic and horticultural purposes, developing quantitative trait loci in advanced hybrids and assessing genetic diversity of large germplasm collections. Here, we suggest that SRAP markers should be employed for research addressing hypotheses in plant systematics, biogeography, conservation, ecology, and beyond. We provide an overview of the SRAP literature to date, review descriptive statistics of SRAP markers in a subset of 171 publications, and present relevant case studies to demonstrate the applicability of SRAP markers to the diverse field of plant biology. Results of these selected works indicate that SRAP markers have the potential to enhance the current suite of molecular tools in a diversity of fields by providing an easy-to-use, highly variable marker with inherent biological significance.

Sequence-related amplified polymorphism (SRAP) markers: A potential resource for studies in plant molecular biology1

PubMed Central

Robarts, Daniel W. H.; Wolfe, Andrea D.

2014-01-01

In the past few decades, many investigations in the field of plant biology have employed selectively neutral, multilocus, dominant markers such as inter-simple sequence repeat (ISSR), random-amplified polymorphic DNA (RAPD), and amplified fragment length polymorphism (AFLP) to address hypotheses at lower taxonomic levels. More recently, sequence-related amplified polymorphism (SRAP) markers have been developed, which are used to amplify coding regions of DNA with primers targeting open reading frames. These markers have proven to be robust and highly variable, on par with AFLP, and are attained through a significantly less technically demanding process. SRAP markers have been used primarily for agronomic and horticultural purposes, developing quantitative trait loci in advanced hybrids and assessing genetic diversity of large germplasm collections. Here, we suggest that SRAP markers should be employed for research addressing hypotheses in plant systematics, biogeography, conservation, ecology, and beyond. We provide an overview of the SRAP literature to date, review descriptive statistics of SRAP markers in a subset of 171 publications, and present relevant case studies to demonstrate the applicability of SRAP markers to the diverse field of plant biology. Results of these selected works indicate that SRAP markers have the potential to enhance the current suite of molecular tools in a diversity of fields by providing an easy-to-use, highly variable marker with inherent biological significance. PMID:25202637

Molecular characterization and identification of markers for toxic and non-toxic varieties of Jatropha curcas L. using RAPD, AFLP and SSR markers.

PubMed

Sudheer Pamidimarri, D V N; Singh, Sweta; Mastan, Shaik G; Patel, Jalpa; Reddy, Muppala P

2009-07-01

Jatropha curcas L., a multipurpose shrub has acquired significant economic importance for its seed oil which can be converted to biodiesel, is emerging as an alternative to petro-diesel. The deoiled seed cake remains after oil extraction is toxic and cannot be used as a feed despite having best nutritional contents. No quantitative and qualitative differences were observed between toxic and non-toxic varieties of J. curcas except for phorbol esters content. Development of molecular marker will enable to differentiate non-toxic from toxic variety in a mixed population and also help in improvement of the species through marker assisted breeding programs. The present investigation was undertaken to characterize the toxic and non-toxic varieties at molecular level and to develop PCR based molecular markers for distinguishing non-toxic from toxic or vice versa. The polymorphic markers were successfully identified specific to non-toxic and toxic variety using RAPD and AFLP techniques. Totally 371 RAPD, 1,442 AFLP markers were analyzed and 56 (15.09%) RAPD, 238 (16.49%) AFLP markers were found specific to either of the varieties. Genetic similarity between non-toxic and toxic verity was found to be 0.92 by RAPD and 0.90 by AFLP fingerprinting. In the present study out of 12 microsatellite markers analyzed, seven markers were found polymorphic. Among these seven, jcms21 showed homozygous allele in the toxic variety. The study demonstrated that both RAPD and AFLP techniques were equally competitive in identifying polymorphic markers and differentiating both the varieties of J. curcas. Polymorphism of SSR markers prevailed between the varieties of J. curcas. These RAPD and AFLP identified markers will help in selective cultivation of specific variety and along with SSRs these markers can be exploited for further improvement of the species through breeding and Marker Assisted Selection (MAS).

Isolation and genetic diversity of endangered grey nurse shark (Carcharias taurus) populations.

PubMed

Stow, Adam; Zenger, Kyall; Briscoe, David; Gillings, Michael; Peddemors, Victor; Otway, Nicholas; Harcourt, Robert

2006-06-22

Anthropogenic impacts are believed to be the primary threats to the eastern Australian population of grey nurse sharks (Carcharias taurus), which is listed as critically endangered, and the most threatened population globally. Analyses of 235 polymorphic amplified fragment length polymorphisms (AFLP) loci and 700 base pairs of mitochondrial DNA control region provide the first account of genetic variation and geographical partitioning (east and west coasts of Australia, South Africa) in C. taurus. Assignment tests, analysis of relatedness and Fst values all indicate that the Australian populations are isolated from South Africa, with negligible migration between the east and west Australian coasts. There are significant differences in levels of genetic variation among regions. Australian C. taurus, particularly the eastern population, has significantly less AFLP variation than the other sampling localities. Further, the eastern Australian sharks possess only a single mitochondrial haplotype, also suggesting a small number of founding individuals. Therefore, historical, rather than anthropogenic processes most likely account for their depauperate genetic variation. These findings have implications for the viability of the eastern Australian population of grey nurse sharks.

Beyond an AFLP genome scan towards the identification of immune genes involved in plague resistance in Rattus rattus from Madagascar.

PubMed

Tollenaere, C; Jacquet, S; Ivanova, S; Loiseau, A; Duplantier, J-M; Streiff, R; Brouat, C

2013-01-01

Genome scans using amplified fragment length polymorphism (AFLP) markers became popular in nonmodel species within the last 10 years, but few studies have tried to characterize the anonymous outliers identified. This study follows on from an AFLP genome scan in the black rat (Rattus rattus), the reservoir of plague (Yersinia pestis infection) in Madagascar. We successfully sequenced 17 of the 22 markers previously shown to be potentially affected by plague-mediated selection and associated with a plague resistance phenotype. Searching these sequences in the genome of the closely related species Rattus norvegicus assigned them to 14 genomic regions, revealing a random distribution of outliers in the genome (no clustering). We compared these results with those of an in silico AFLP study of the R. norvegicus genome, which showed that outlier sequences could not have been inferred by this method in R. rattus (only four of the 15 sequences were predicted). However, in silico analysis allowed the prediction of AFLP markers distribution and the estimation of homoplasy rates, confirming its potential utility for designing AFLP studies in nonmodel species. The 14 genomic regions surrounding AFLP outliers (less than 300 kb from the marker) contained 75 genes encoding proteins of known function, including nine involved in immune function and pathogen defence. We identified the two interleukin 1 genes (Il1a and Il1b) that share homology with an antigen of Y. pestis, as the best candidates for genes subject to plague-mediated natural selection. At least six other genes known to be involved in proinflammatory pathways may also be affected by plague-mediated selection. © 2012 Blackwell Publishing Ltd.

Development of AFLP and RAPD markers linked to a locus associated with twisted growth in corkscrew willow (Salix matsudana 'Tortuosa').

PubMed

Lin, Juan; Gunter, Lee E; Harding, Scott A; Kopp, Richard F; McCord, Rachel P; Tsai, Chung-Jui; Tuskan, Gerald A; Smart, Lawrence B

2007-11-01

Salix matsudana Koidz. cultivar 'Tortuosa' (corkscrew willow) is characterized by extensive stem bending and curling of leaves. To investigate the genetic basis of this trait, controlled crosses were made between a corkscrew female (S. matsudana 'Tortuosa') and a straight-stemmed, wild-type male (Salix alba L. Clone 99010). Seventy-seven seedlings from this family (ID 99270) were grown in the field for phenotypic observation. Among the progeny, 39 had straight stems and leaves and 38 had bent stems and curled leaves, suggesting that a dominant allele at a single locus controls this phenotype. As a first step in characterizing the locus, we searched for amplified fragment length polymorphism (AFLP) and randomly amplified polymorphic DNA (RAPD) markers linked to the tortuosa allele using bulked segregant analysis. Samples of DNA from 10 corkscrew individuals were combined to produce a corkscrew pool, and DNA from 10 straight progeny was combined to make a wild-type pool. Sixty-four AFLP primer combinations and 640 RAPD primers were screened to identify marker bands amplified from the corkscrew parent and progeny pool, but not from the wild-type parent or progeny pool. An AFLP marker and a RAPD marker linked to and flanking the tortuosa locus were placed on a preliminary linkage map constructed based on segregation among the 77 progeny. Sectioning and analysis of shoot tips revealed that the corkscrew phenotype is associated with vascular cell collapse, smaller cell size in regions near the cambium and less developed phloem fibers than in wild-type progeny. Identification of a gene associated with this trait could lead to greater understanding of the control of normal stem development in woody plants.

AFLP analysis of genetic diversity and phylogenetic relationships of Brassica oleracea in Ireland.

PubMed

El-Esawi, Mohamed A; Germaine, Kieran; Bourke, Paula; Malone, Renee

2016-01-01

Brassica oleracea L. is one of the most economically important vegetable crop species of the genus Brassica L. This species is threatened in Ireland, without any prior reported genetic studies. The use of this species is being very limited due to its imprecise phylogeny and uncompleted genetic characterisation. The main objective of this study was to assess the genetic diversity and phylogenetic relationships of a set of 25 Irish B. oleracea accessions using the powerful amplified fragment length polymorphism (AFLP) technique. A total of 471 fragments were scored across all the 11 AFLP primer sets used, out of which 423 (89.8%) were polymorphic and could differentiate the accessions analysed. The dendrogram showed that cauliflowers were more closely related to cabbages than kales were, and accessions of some cabbage types were distributed among different clusters within cabbage subgroups. Approximately 33.7% of the total genetic variation was found among accessions, and 66.3% of the variation resided within accessions. The total genetic diversity (HT) and the intra-accessional genetic diversity (HS) were 0.251 and 0.156, respectively. This high level of variation demonstrates that the Irish B. oleracea accessions studied should be managed and conserved for future utilisation and exploitation in food and agriculture. In conclusion, this study addressed important phylogenetic questions within this species, and provided a new insight into the inclusion of four accessions of cabbages and kales in future breeding programs for improving varieties. AFLP markers were efficient for assessing genetic diversity and phylogenetic relationships in Irish B. oleracea species. Copyright © 2016 Académie des sciences. Published by Elsevier SAS. All rights reserved.

Analysis of genetic variation in Ganoderma Lucidum after space flight

NASA Astrophysics Data System (ADS)

Qi, Jian-Jun; Ma, Rong-Cai; Chen, Xiang-Dong; Lan, Jin

A modified CTAB method was used in the extraction of total cellular DNA of Ganoderma lucidum. Four strains Cx, Ch, C3 and C4, and their counterparts, four space flown strains Sx, Xh, S3 and S4, were analysed by amplified fragment length polymorphism (AFLP) with several primer combinations. Polymorphic bands were detected between Sx and Cx, S3 and C3, respectively. Somatic incompatibility tests further confirmed their heterogeneity. However, no disparity between Sh and Ch, S4 and C4 was detectable. The results suggest that spaceflight may be used to accelerate breeding of Ganoderma lucidum strains for commercial cultivation.

Assessing Date Palm Genetic Diversity Using Different Molecular Markers.

PubMed

Atia, Mohamed A M; Sakr, Mahmoud M; Adawy, Sami S

2017-01-01

Molecular marker technologies which rely on DNA analysis provide powerful tools to assess biodiversity at different levels, i.e., among and within species. A range of different molecular marker techniques have been developed and extensively applied for detecting variability in date palm at the DNA level. Recently, the employment of gene-targeting molecular marker approaches to study biodiversity and genetic variations in many plant species has increased the attention of researchers interested in date palm to carry out phylogenetic studies using these novel marker systems. Molecular markers are good indicators of genetic distances among accessions, because DNA-based markers are neutral in the face of selection. Here we describe the employment of multidisciplinary molecular marker approaches: amplified fragment length polymorphism (AFLP), start codon targeted (SCoT) polymorphism, conserved DNA-derived polymorphism (CDDP), intron-targeted amplified polymorphism (ITAP), simple sequence repeats (SSR), and random amplified polymorphic DNA (RAPD) to assess genetic diversity in date palm.

Genetic diversity and population structure of a protected species: Polygala tenuifolia Willd.

PubMed

Peng, Yan Qun; Fan, Ling Ling; Mao, Fu Ying; Zhao, Yun Sheng; Xu, Rui; Yin, Yu Jie; Chen, Xin; Wan, De Guang; Zhang, Xin Hui

2018-03-01

Polygala tenuifolia Willd. is an important protected species used in traditional Chinese medicine. In the present study, amplified fragment length polymorphism (AFLP) markers were employed to characterize the genetic diversity in wild and cultivated P. tenuifolia populations. Twelve primer combinations of AFLP produced 310 unambiguous and repetitious bands. Among these bands, 261 (84.2%) were polymorphic. The genetic diversity was high at the species level: percentage of polymorphic loci (PPL)=84.2%, Nei's gene diversity (h)=0.3296 and Shannon's information index (I)=0.4822. Between the two populations, the genetic differentiation of 0.1250 was low and the gene flow was relatively high, at 3.4989. The wild population (PPL=81.9%, h=0.3154, I=0.4635) showed a higher genetic diversity level than the cultivated population (PPL=63.9%, h=0.2507, I=0.3688). The results suggest that the major factors threatening the persistence of P. tenuifolia resources are ecological and human factors rather than genetic. These results will assist with the design of conservation and management programs, such as in natural habitat conservation, setting the excavation time interval for resource regeneration and the substitution of cultivated for wild plants. Copyright © 2018 Académie des sciences. Published by Elsevier Masson SAS. All rights reserved.

Genomic change, retrotransposon mobilization and extensive cytosine methylation alteration in Brassica napus introgressions from two intertribal hybridizations.

PubMed

Zhang, Xueli; Ge, Xianhong; Shao, Yujiao; Sun, Genlou; Li, Zaiyun

2013-01-01

Hybridization and introgression represent important means for the transfer and/or de novo origination of traits and play an important role in facilitating speciation and plant breeding. Two sets of introgression lines in Brassica napus L. were previously established by its intertribal hybridizations with two wild species and long-term selection. In this study, the methods of amplified fragment length polymorphisms (AFLP), sequence-specific amplification polymorphism (SSAP) and methylation-sensitive amplified polymorphism (MSAP) were used to determine their genomic change, retrotransposon mobilization and cytosine methylation alteration in these lines. The genomic change revealed by the loss or gain of AFLP bands occurred for ∼10% of the total bands amplified in the two sets of introgressions, while no bands specific for wild species were detected. The new and absent SSAP bands appeared for 9 out of 11 retrotransposons analyzed, with low frequency of new bands and their total percentage of about 5% in both sets. MSAP analysis indicated that methylation changes were common in these lines (33.4-39.8%) and the hypermethylation was more frequent than hypomethylation. Our results suggested that certain extents of genetic and epigenetic alterations were induced by hybridization and alien DNA introgression. The cryptic mechanism of these changes and potential application of these lines in breeding were also discussed.

Epigenetic Variability in the Genetically Uniform Forest Tree Species Pinus pinea L

PubMed Central

Sáez-Laguna, Enrique; Guevara, María-Ángeles; Díaz, Luis-Manuel; Sánchez-Gómez, David; Collada, Carmen; Aranda, Ismael; Cervera, María-Teresa

2014-01-01

There is an increasing interest in understanding the role of epigenetic variability in forest species and how it may contribute to their rapid adaptation to changing environments. In this study we have conducted a genome-wide analysis of cytosine methylation pattern in Pinus pinea, a species characterized by very low levels of genetic variation and a remarkable degree of phenotypic plasticity. DNA methylation profiles of different vegetatively propagated trees from representative natural Spanish populations of P. pinea were analyzed with the Methylation Sensitive Amplified Polymorphism (MSAP) technique. A high degree of cytosine methylation was detected (64.36% of all scored DNA fragments). Furthermore, high levels of epigenetic variation were observed among the studied individuals. This high epigenetic variation found in P. pinea contrasted with the lack of genetic variation based on Amplified Fragment Length Polymorphism (AFLP) data. In this manner, variable epigenetic markers clearly discriminate individuals and differentiates two well represented populations while the lack of genetic variation revealed with the AFLP markers fail to differentiate at both, individual or population levels. In addition, the use of different replicated trees allowed identifying common polymorphic methylation sensitive MSAP markers among replicates of a given propagated tree. This set of MSAPs allowed discrimination of the 70% of the analyzed trees. PMID:25084460

Epigenetic variability in the genetically uniform forest tree species Pinus pinea L.

PubMed

Sáez-Laguna, Enrique; Guevara, María-Ángeles; Díaz, Luis-Manuel; Sánchez-Gómez, David; Collada, Carmen; Aranda, Ismael; Cervera, María-Teresa

2014-01-01

There is an increasing interest in understanding the role of epigenetic variability in forest species and how it may contribute to their rapid adaptation to changing environments. In this study we have conducted a genome-wide analysis of cytosine methylation pattern in Pinus pinea, a species characterized by very low levels of genetic variation and a remarkable degree of phenotypic plasticity. DNA methylation profiles of different vegetatively propagated trees from representative natural Spanish populations of P. pinea were analyzed with the Methylation Sensitive Amplified Polymorphism (MSAP) technique. A high degree of cytosine methylation was detected (64.36% of all scored DNA fragments). Furthermore, high levels of epigenetic variation were observed among the studied individuals. This high epigenetic variation found in P. pinea contrasted with the lack of genetic variation based on Amplified Fragment Length Polymorphism (AFLP) data. In this manner, variable epigenetic markers clearly discriminate individuals and differentiates two well represented populations while the lack of genetic variation revealed with the AFLP markers fail to differentiate at both, individual or population levels. In addition, the use of different replicated trees allowed identifying common polymorphic methylation sensitive MSAP markers among replicates of a given propagated tree. This set of MSAPs allowed discrimination of the 70% of the analyzed trees.

Molecular epidemiology and in vitro antifungal susceptibility testing of 108 clinical Cryptococcus neoformans sensu lato and Cryptococcus gattii sensu lato isolates from Denmark.

PubMed

Hagen, Ferry; Hare Jensen, Rasmus; Meis, Jacques F; Arendrup, Maiken Cavling

2016-09-01

Cryptococcosis is mainly caused by members of the Cryptococcus gattii/Cryptococcus neoformans species complexes. Here, we report the molecular characterisation and in vitro antifungal susceptibility of Danish clinical cryptococcal isolates. Species, genotype, serotype and mating type were determined by amplified fragment length polymorphism (AFLP) fingerprinting and qPCR. EUCAST E.Def 7.2 MICs were determined for amphotericin B, flucytosine, fluconazole, voriconazole and isavuconazole. Most isolates were C. neoformans (serotype A; n = 66) and belonged to genotype AFLP1/VNI (n = 61) or AFLP1B/VNII (n = 5) followed by Cryptococcus deneoformans (serotype D; genotype AFLP2, n = 20), C. neoformans × C. deneoformans hybrids (serotype AD; genotype AFLP3, n = 13) and Cryptococcus curvatus (n = 2). Six isolates were C. gattii sensu lato, and one isolate was a C. deneoformans × C. gattii hybrid (genotype AFLP8). All isolates were amphotericin B susceptible. Flucytosine susceptibility was uniform MIC50 of 4-8 mg l(-1) except for C. curvatus (MICs >32 mg l(-1) ). Cryptococcus gattii sensu lato isolates were somewhat less susceptible to the azoles. MICs of fluconazole (>32 mg l(-1) ), voriconazole (≥0.5 mg l(-1) ) and isavuconazole (0.06 and 0.25 mg l(-1) respectively) were elevated compared to the wild-type population for 1/19 C. deneoformans and 1/2 C. curvatus isolates. Flucytosine MIC was elevated for 1/61 C. neoformans (>32 mg l(-1) ). Antifungal susceptibility revealed species-specific differential susceptibility, but suggested acquired resistance was an infrequent phenomenon. © 2016 Blackwell Verlag GmbH.

AFLP-Based Analysis of Genetic Diversity, Population Structure, and Relationships with Agronomic Traits in Rice Germplasm from North Region of Iran and World Core Germplasm Set.

PubMed

Sorkheh, Karim; Masaeli, Mohammad; Chaleshtori, Maryam Hosseini; Adugna, Asfaw; Ercisli, Sezai

2016-04-01

Analysis of the genetic diversity and population structure of crops is very important for use in breeding programs and for genetic resources conservation. We analyzed the genetic diversity and population structure of 47 rice genotypes from diverse origins using amplified fragment length polymorphism (AFLP) markers and morphological characters. The 47 genotypes, which were composed of four populations: Iranian native varieties, Iranian improved varieties, International Rice Research Institute (IRRI) rice varieties, and world rice collections, were analyzed using ten primer combinations. A total of 221 scorable bands were produced with an average of 22.1 alleles per pair of primers, of which 120 (54.30%) were polymorphic. The polymorphism information content (PIC) values varied from 0.32 to 0.41 with an average of 0.35. The high percentage of polymorphic bands (%PB) was found to be 64.71 and the resolving power (R p) collections were 63.36. UPGMA clustering based on numerical data from AFLP patterns clustered all 47 genotypes into three large groups. The genetic similarity between individuals ranged from 0.54 to 0.94 with an average of 0.74. Population genetic tree showed that Iranian native cultivars formed far distant cluster from the other populations, which may indicate that these varieties had minimal genetic change over time. Analysis of molecular variance (AMOVA) revealed that the largest proportion of the variation (84%) to be within populations showing the inbreeding nature of rice. Therefore, Iranian native varieties (landraces) may have unique genes, which can be used for future breeding programs and there is a need to conserve this unique diversity. Furthermore, crossing of Iranian genotypes with the genetically distant genotypes in the other three populations may result in useful combinations, which can be used as varieties and/or lines for future rice breeding programs.

Morphological and Genetic Analysis of Four Color Morphs of Bean Leaf Beetle.

PubMed

Tiroesele, Bamphitlhi; Skoda, Steven R; Hunt, Thomas E; Lee, Donald J; Ullah, Muhammad Irfan; Molina-Ochoa, Jaime; Foster, John E

2018-03-01

Bean leaf beetle (BLB), Cerotoma trifurcata (Forster; Coleoptera: Chrysomelidae), exhibits considerable color variation but little is known about the underlying genetic structure and gene flow among color phenotypes. Genetic and morphological variation among four color phenotypes-green with spots (G+S), green without spots (G-S), red with spots (R+S) and red without spots (R-S)-were analyzed using amplified fragment length polymorphisms (AFLP) and morphometrics, respectively. AFLP generated 175 markers that showed ≥80% polymorphism. Analysis of molecular variance (AMOVA) indicated that genetic variation was greatest within phenotypes (82.6-84.0%); gene flow among the four phenotypes was relatively high (Nm = 3.82). The dendrogram and STRUCTURE analysis indicated some population divergence of G-S from the other phenotypes. Morphological parameters were similar among phenotypes except that R+S showed significant differences in weight and body-length. Canonical variables 1 and 2, based on average morphometric characters, accounted for 98% of the total variation; some divergence was indicated between G+S and R+S from each other and from the G-S/R-S BLB color morphs. The pattern of genetic variation indicated potential divergence of G-S and G+S from each other and from R-S and R+S. Although these results indicate that the four different color morphs are not genetically or reproductively isolated, there is some genetic differentiation/structure and morphological dissimilarity suggesting weak/incomplete isolation.

Hierarchical Population Genetic Structure in a Direct Developing Antarctic Marine Invertebrate

PubMed Central

Hoffman, Joseph I.; Clarke, Andrew; Clark, Melody S.; Peck, Lloyd S.

2013-01-01

Understanding the relationship between life-history variation and population structure in marine invertebrates is not straightforward. This is particularly true of polar species due to the difficulty of obtaining samples and a paucity of genomic resources from which to develop nuclear genetic markers. Such knowledge, however, is essential for understanding how different taxa may respond to climate change in the most rapidly warming regions of the planet. We therefore used over two hundred polymorphic Amplified Fragment Length Polymorphisms (AFLPs) to explore population connectivity at three hierachical spatial scales in the direct developing Antarctic topshell Margarella antarctica. To previously published data from five populations spanning a 1500 km transect along the length of the Western Antarctic Peninsula, we added new AFLP data for four populations separated by up to 6 km within Ryder Bay, Adelaide Island. Overall, we found a nonlinear isolation-by-distance pattern, suggestive of weaker population structure within Ryder Bay than is present over larger spatial scales. Nevertheless, significantly positive F st values were obtained in all but two of ten pairwise population comparisons within the bay following Bonferroni correction for multiple tests. This is in contrast to a previous study of the broadcast spawner Nacella concinna that found no significant genetic differences among several of the same sites. By implication, the topshell's direct-developing lifestyle may constrain its ability to disperse even over relatively small geographic scales. PMID:23691125

Development of a SCAR marker for male gametophyte of Gracilariopsis lemaneiformis based on AFLP technique

NASA Astrophysics Data System (ADS)

Zhou, Wei; Ding, Hongye; Sui, Zhenghong; Wang, Zhongxia; Wang, Jinguo

2014-05-01

The red alga Gracilariopsis lemaneiformis (Bory) is an economically valuable macroalgae. As a means to identify the sex of immature Gracilariopsis lemaneiformis, the amplified fragment length polymorphism (AFLP) technique was used to search for possible sex- or phase-related markers in male gametophytes, female gametophytes, and tetrasporophytes, respectively. Seven AFLP selective amplification primers were used in this study. The primer combination E-TG/M-CCA detected a specific band linked to male gametophytes. The DNA fragment was recovered and a 402-bp fragment was sequenced. However, no DNA sequence match was found in public databases. Sequence characterized amplified region (SCAR) primers were designed from the sequence to test the repeatability of the relationship to the sex, using 69 male gametophytes, 139 female gametophytes, and 47 tetrasporophytes. The test results demonstrate a good linkage and repeatability of the SCAR marker to sex. The SCAR primers developed in this study could reduce the time required for sex identification of Gracilariopsis lemaneiformis by four to six months. This can reduce both the time investment and number of specimens required in breeding experiments.

Tissue culture-induced genetic and epigenetic variation in triticale (× Triticosecale spp. Wittmack ex A. Camus 1927) regenerants.

PubMed

Machczyńska, Joanna; Zimny, Janusz; Bednarek, Piotr Tomasz

2015-10-01

Plant regeneration via in vitro culture can induce genetic and epigenetic variation; however, the extent of such changes in triticale is not yet understood. In the present study, metAFLP, a variation of methylation-sensitive amplified fragment length polymorphism analysis, was used to investigate tissue culture-induced variation in triticale regenerants derived from four distinct genotypes using androgenesis and somatic embryogenesis. The metAFLP technique enabled identification of both sequence and DNA methylation pattern changes in a single experiment. Moreover, it was possible to quantify subtle effects such as sequence variation, demethylation, and de novo methylation, which affected 19, 5.5, 4.5% of sites, respectively. Comparison of variation in different genotypes and with different in vitro regeneration approaches demonstrated that both the culture technique and genetic background of donor plants affected tissue culture-induced variation. The results showed that the metAFLP approach could be used for quantification of tissue culture-induced variation and provided direct evidence that in vitro plant regeneration could cause genetic and epigenetic variation.

AFLP analysis of Cynodon dactylon (L.) Pers. var. dactylon genetic variation.

PubMed

Wu, Y Q; Taliaferro, C M; Bai, G H; Anderson, M P

2004-08-01

Cynodon dactylon (L.) Pers. var. dactylon (common bermudagrass) is geographically widely distributed between about lat 45 degrees N and lat 45 degrees S, penetrating to about lat 53 degrees N in Europe. The extensive variation of morphological and adaptive characteristics of the taxon is substantially documented, but information is lacking on DNA molecular variation in geographically disparate forms. Accordingly, this study was conducted to assess molecular genetic variation and genetic relatedness among 28 C. dactylon var. dactylon accessions originating from 11 countries on 4 continents (Africa, Asia, Australia, and Europe). A fluorescence-labeled amplified fragment length polymorphism (AFLP) DNA profiling method was used to detect the genetic diversity and relatedness. On the basis of 443 polymorphic AFLP fragments from 8 primer combinations, the accessions were grouped into clusters and subclusters associating with their geographic origins. Genetic similarity coefficients (SC) for the 28 accessions ranged from 0.53 to 0.98. Accessions originating from Africa, Australia, Asia, and Europe formed major groupings as indicated by cluster and principal coordinate analysis. Accessions from Australia and Asia, though separately clustered, were relatively closely related and most distantly related to accessions of European origin. African accessions formed two distant clusters and had the greatest variation in genetic relatedness relative to accessions from other geographic regions. Sampling the full extent of genetic variation in C. dactylon var. dactylon would require extensive germplasm collection in the major geographic regions of its distributional range.

Genetic and epigenetic stability of cryopreserved and cold-stored hops (Humulus lupulus L.).

PubMed

Peredo, Elena L; Arroyo-García, Rosa; Reed, Barbara M; Revilla, M Angeles

2008-12-01

Conventional cold storage and cryopreservation methods for hops (Humulus lupulus L.) are available but, to our knowledge, the genetic and epigenetic stability of the recovered plants have not been tested. This study analyzed 51 accessions of hop using the molecular techniques, Random Amplified DNA Polymorphism (RAPD) and Amplified Fragment Length Polymorphism (AFLP), revealing no genetic variation among greenhouse-grown controls and cold stored or cryopreserved plants. Epigenetic stability was evaluated using Methylation Sensitive Amplified Polymorphism (MSAP). Over 36% of the loci were polymorphic when the cold and cryo-treated plants were compared to greenhouse plants. The main changes were demethylation events and they were common to the cryopreserved and cold stored plants indicating the possible effect of the in vitro establishment process, an essential step in both protocols. Protocol-specific methylation patterns were also detected indicating that both methods produced epigenetic changes in plants following cold storage and cryopreservation.

Isolation of Cryptococcus gattii from a Castanopsis argyrophylla tree hollow (Mai-Kaw), Chiang Mai, Thailand.

PubMed

Khayhan, Kantarawee; Hagen, Ferry; Norkaew, Treepradab; Puengchan, Tanpalang; Boekhout, Teun; Sriburee, Pojana

2017-04-01

The pathogenic yeast Cryptococcus gattii was isolated from a tree hollow of a Castanopsis argyrophylla King ex Hook.f. (Fagaceae) in Chiang Mai, Thailand. Molecular characterization with amplified fragment length polymorphism analysis and multi-locus sequence typing showed that this isolate belonged to genotype AFLP4/VGI representing C. gattii sensu stricto. Subsequent comparison of the environmental isolate with those from clinical samples from Thailand showed that they grouped closely together in a single cluster.

Molecular mapping of powdery mildew resistance gene Eg-3 in cultivated oat (Avena sativa L. cv. Rollo).

PubMed

Mohler, Volker; Zeller, Friedrich J; Hsam, Sai L K

2012-05-01

Powdery mildew is a prevalent fungal disease affecting oat (Avena sativa L.) production in Europe. Common oat cultivar Rollo was previously shown to carry the powdery mildew resistance gene Eg-3 in common with cultivar Mostyn. The resistance gene was mapped with restriction fragment length polymorphism (RFLP) markers from Triticeae group-1 chromosomes using a population of F(3) lines from a cross between A. byzantina cv. Kanota and A. sativa cv. Rollo. This comparative mapping approach positioned Eg-3 between cDNA-RFLP marker loci cmwg706 and cmwg733. Since both marker loci were derived from the long arm of barley chromosome 1H, the subchromosomal location of Eg-3 was assumed to be on the long arm of oat chromosome 17. Amplified fragment length polymorphism (AFLP) marker technology featured as an efficient means for obtaining markers closely linked to Eg-3.

Genomic Change, Retrotransposon Mobilization and Extensive Cytosine Methylation Alteration in Brassica napus Introgressions from Two Intertribal Hybridizations

PubMed Central

Zhang, Xueli; Ge, Xianhong; Shao, Yujiao; Sun, Genlou; Li, Zaiyun

2013-01-01

Hybridization and introgression represent important means for the transfer and/or de novo origination of traits and play an important role in facilitating speciation and plant breeding. Two sets of introgression lines in Brassica napus L. were previously established by its intertribal hybridizations with two wild species and long-term selection. In this study, the methods of amplified fragment length polymorphisms (AFLP), sequence-specific amplification polymorphism (SSAP) and methylation-sensitive amplified polymorphism (MSAP) were used to determine their genomic change, retrotransposon mobilization and cytosine methylation alteration in these lines. The genomic change revealed by the loss or gain of AFLP bands occurred for ∼10% of the total bands amplified in the two sets of introgressions, while no bands specific for wild species were detected. The new and absent SSAP bands appeared for 9 out of 11 retrotransposons analyzed, with low frequency of new bands and their total percentage of about 5% in both sets. MSAP analysis indicated that methylation changes were common in these lines (33.4–39.8%) and the hypermethylation was more frequent than hypomethylation. Our results suggested that certain extents of genetic and epigenetic alterations were induced by hybridization and alien DNA introgression. The cryptic mechanism of these changes and potential application of these lines in breeding were also discussed. PMID:23468861

Identification of exotic genetic components and DNA methylation pattern analysis of three cotton introgression lines from Gossypium bickii.

PubMed

He, Shou-Pu; Sun, Jun-Ling; Zhang, Chao; Du, Xiong-Ming

2011-01-01

The impact of alien DNA fragments on plant genome has been studied in many species. However, little is known about the introgression lines of Gossypium. To study the consequences of introgression in Gossypium, we investigated 2000 genomic and 800 epigenetic sites in three typical cotton introgression lines, as well as their cultivar (Gossypium hirsutum) and wild parents (Gossypium bickii), by amplified fragment length polymorphism (AFLP) and methylation-sensitive amplified polymorphism (MSAP). The results demonstrate that an average of 0.5% of exotic DNA segments from wild cotton is transmitted into the genome of each introgression line, with the addition of other forms of genetic variation. In total, an average of 0.7% of genetic variation sites is identified in introgression lines. Simultaneously, the overall cytosine methylation level in each introgression line is very close to that of the upland cotton parent (an average of 22.6%). Further dividing patterns reveal that both hypomethylation and hypermethylation occurred in introgression lines in comparison with the upland cotton parent. Sequencing of nine methylation polymorphism fragments showed that most (7 of 9) of the methylation alternations occurred in the noncoding sequences. The molecular evidence of introgression from wild cotton into introgression lines in our study is identified by AFLP. Moreover, the causes of petal variation in introgression lines are discussed.

A Predominant and Virulent Legionella pneumophila Serogroup 1 Strain Detected in Isolates from Patients and Water in Queensland, Australia, by an Amplified Fragment Length Polymorphism Protocol and Virulence Gene-Based PCR Assays

PubMed Central

Huang, Bixing; Heron, Brett A.; Gray, Bruce R.; Eglezos, Sofroni; Bates, John R.; Savill, John

2004-01-01

In epidemiological investigations of community legionellosis outbreaks, knowledge of the prevalence, distribution, and clinical significance (virulence) of environmental Legionella isolates is crucial for interpretation of the molecular subtyping results. To obtain such information for Legionella pneumophila serogroup 1 isolates, we used the standardized amplified fragment length polymorphism (AFLP) protocol of the European Working Group on Legionella Infection to subtype L. pneumophila SG1 isolates obtained from patients and water sources in Queensland, Australia. An AFLP genotype, termed AF1, was predominant in isolates from both patients (40.5%) and water (49.0%). The second most common AFLP genotype found in water isolates was AF16 (36.5%), but this genotype was not identified in the patient isolates. When virulence gene-based PCR assays for lvh and rtxA genes were applied to the isolates from patients and water, nearly all (65 of 66) AF1 strains had both virulence genes, lvh and rtxA. In contrast, neither the lvh nor the rtxA gene was found in the AF16 strains, except for one isolate with the rtxA gene. It appears that this may explain the failure to find this genotype in the isolates from patients even though it may be common in the environment. In view of the evidence that the AF1 genotype is the most common genotype among strains found in patients and water sources in this region, any suggested epidemiological link derived from comparing the AF1 genotype from patient isolates with the AF1 genotype from environmental isolates must be interpreted and acted on with caution. The use of virulence gene-based PCR assays applied to environmental samples may be helpful in determining the infection potential of the isolates involved. PMID:15365006

Comparison of seven techniques for typing international epidemic strains of Clostridium difficile: restriction endonuclease analysis, pulsed-field gel electrophoresis, PCR-ribotyping, multilocus sequence typing, multilocus variable-number tandem-repeat analysis, amplified fragment length polymorphism, and surface layer protein A gene sequence typing.

PubMed

Killgore, George; Thompson, Angela; Johnson, Stuart; Brazier, Jon; Kuijper, Ed; Pepin, Jacques; Frost, Eric H; Savelkoul, Paul; Nicholson, Brad; van den Berg, Renate J; Kato, Haru; Sambol, Susan P; Zukowski, Walter; Woods, Christopher; Limbago, Brandi; Gerding, Dale N; McDonald, L Clifford

2008-02-01

Using 42 isolates contributed by laboratories in Canada, The Netherlands, the United Kingdom, and the United States, we compared the results of analyses done with seven Clostridium difficile typing techniques: multilocus variable-number tandem-repeat analysis (MLVA), amplified fragment length polymorphism (AFLP), surface layer protein A gene sequence typing (slpAST), PCR-ribotyping, restriction endonuclease analysis (REA), multilocus sequence typing (MLST), and pulsed-field gel electrophoresis (PFGE). We assessed the discriminating ability and typeability of each technique as well as the agreement among techniques in grouping isolates by allele profile A (AP-A) through AP-F, which are defined by toxinotype, the presence of the binary toxin gene, and deletion in the tcdC gene. We found that all isolates were typeable by all techniques and that discrimination index scores for the techniques tested ranged from 0.964 to 0.631 in the following order: MLVA, REA, PFGE, slpAST, PCR-ribotyping, MLST, and AFLP. All the techniques were able to distinguish the current epidemic strain of C. difficile (BI/027/NAP1) from other strains. All of the techniques showed multiple types for AP-A (toxinotype 0, binary toxin negative, and no tcdC gene deletion). REA, slpAST, MLST, and PCR-ribotyping all included AP-B (toxinotype III, binary toxin positive, and an 18-bp deletion in tcdC) in a single group that excluded other APs. PFGE, AFLP, and MLVA grouped two, one, and two different non-AP-B isolates, respectively, with their AP-B isolates. All techniques appear to be capable of detecting outbreak strains, but only REA and MLVA showed sufficient discrimination to distinguish strains from different outbreaks.

Gene expression analysis by cDNA-AFLP highlights a set of new signaling networks and translational control during seed dormancy breaking in Nicotiana plumbaginifolia.

PubMed

Bove, Jérôme; Lucas, Philippe; Godin, Béatrice; Ogé, Laurent; Jullien, Marc; Grappin, Philippe

2005-03-01

Seed dormancy in Nicotiana plumbaginifolia is characterized by an abscisic acid accumulation linked to a pronounced germination delay. Dormancy can be released by 1 year after-ripening treatment. Using a cDNA-amplified fragment length polymorphism (cDNA-AFLP) approach we compared the gene expression patterns of dormant and after-ripened seeds, air-dry or during one day imbibition and analyzed 15,000 cDNA fragments. Among them 1020 were found to be differentially regulated by dormancy. Of 412 sequenced cDNA fragments, 83 were assigned to a known function by search similarities to public databases. The functional categories of the identified dormancy maintenance and breaking responsive genes, give evidence that after-ripening turns in the air-dry seed to a new developmental program that modulates, at the RNA level, components of translational control, signaling networks, transcriptional control and regulated proteolysis.

Genetic diversity among brazilian isolates of beauveria bassiana: comparisons with non-brazilian isolates and other beauveria species

USGS Publications Warehouse

Fernandes, E.K.K.; Moraes, A.M.L.; Pacheco, R.S.; Rangel, D.E.N.; Miller, M.P.; Bittencourt, V.R.E.P.; Roberts, D.W.

2009-01-01

Aims: The genetic diversity of Beauveria bassiana was investigated by comparing isolates of this species to each other (49 from different geographical regions of Brazil and 4 from USA) and to other Beauveria spp. Methods and Results: The isolates were examined by multilocus enzyme electrophoresis (MLEE), amplified fragment length polymorphism (AFLP), and rDNA sequencing. MLEE and AFLP revealed considerable genetic variability among B. bassiana isolates. Several isolates from South and Southeast Brazil had high similarity coefficients, providing evidence of at least one population with clonal structure. There were clear genomic differences between most Brazilian and USA B. bassiana isolates. A Mantel test using data generated by AFLP provided evidence that greater geographical distances were associated with higher genetic distances. AFLP and rDNA sequencing demonstrated notable genotypic variation between B. bassiana and other Beauveria spp. Conclusion: Geographical distance between populations apparently is an important factor influencing genotypic variability among B. bassiana populations in Brazil. Significance and Impact of the Study: This study characterized many B. bassiana isolates. The results indicate that certain Brazilian isolates are considerably different from others and possibly should be regarded as separate species from B. bassiana sensu latu. The information on genetic variation among the Brazilian isolates, therefore, will be important to comprehending the population structure of B. bassiana in Brazil. ?? 2009 The Society for Applied Microbiology.

A preliminary assessment of genetic relationships among agronomically important cultivars of black pepper

PubMed Central

Joy, Nisha; Abraham, Z; Soniya, EV

2007-01-01

Background The impact of diseases such as Phytophthora foot rot and the replacement of unproductive cultivars by high yielding ones has brought about the disappearance of varieties in Piper species, like any other crop. Black pepper (King of spices), is a major spice crop consumed throughout the world. It is widely cultivated across various parts of the world apart from India. The different cultivars may be genetically related and could be a source of valuable genes for disease resistance and an increase in quantity and quality. Even though Western Ghats in India is believed to be the site of origin of this crop, numerous accessions from the NBPGR have not yet been evaluated. Our study aims to investigate the genetic relatedness in major cultivars of black pepper using Amplified Fragment Length Polymorphism. Results Amplified Fragment Length Polymorphic (AFLP) DNA analysis was performed in thirty popular cultivars of black pepper from National Bureau of Plant Genetic Resources (NBPGR), India. Fingerprint profiles were generated initially with, five different primer combinations, from which three primer pair combinations (EAGC/MCAA, EAGG/MCTA and EAGC/MCTG) gave consistent and scorable banding patterns. From 173 scorable markers, 158(> 90%) were polymorphic which shows there is considerable variation in the available germplasm. The dendrogram derived by unweighted pair group method analysis (UPGMA) grouped the accessions into three major clusters and four diverse cultivars with only 30% similarity. Karimunda, a widely grown and popular cultivar was unique in the fingerprint profiles obtained. Conclusion There are currently few fingerprinting studies using the valuable spice crop black pepper. We found considerable genetic variability among cultivars of black pepper. Fingerprinting analysis with AFLP proved to be an ideal tool for cultivar identification and phylogenetic studies. It shows the high level of polymorphism and the unique characterization of the major cultivars. An extensive range of similarity value between the cultivars was noted (6.01 to 98.13). Further screening of more cultivars will provide valuable information for current breeding programmes. PMID:17603884

Dense genetic linkage maps of three Populus species (Populus deltoides, P. nigra and P. trichocarpa) based on AFLP and microsatellite markers.

PubMed

Cervera, M T; Storme, V; Ivens, B; Gusmão, J; Liu, B H; Hostyn, V; Van Slycken, J; Van Montagu, M; Boerjan, W

2001-06-01

Populus deltoides, P. nigra, and P. trichocarpa are the most important species for poplar breeding programs worldwide. In addition, Populus has become a model for fundamental research on trees. Linkage maps were constructed for these three species by analyzing progeny of two controlled crosses sharing the same female parent, Populus deltoides cv. S9-2 x P. nigra cv. Ghoy and P. deltoides cv. S9-2 x P. trichocarpa cv. V24. The two-way pseudotestcross mapping strategy was used to construct the maps. Amplified fragment length polymorphism (AFLP) markers that segregated 1:1 were used to form the four parental maps. Microsatellites and sequence-tagged sites were used to align homoeologous groups between the maps and to merge linkage groups within the individual maps. Linkage analysis and alignment of the homoeologous groups resulted in 566 markers distributed over 19 groups for P. deltoides covering 86% of the genome, 339 markers distributed over 19 groups for P. trichocarpa covering 73%, and 369 markers distributed over 28 groups for P. nigra covering 61%. Several tests for randomness showed that the AFLP markers were randomly distributed over the genome.

Dense genetic linkage maps of three Populus species (Populus deltoides, P. nigra and P. trichocarpa) based on AFLP and microsatellite markers.

PubMed Central

Cervera, M T; Storme, V; Ivens, B; Gusmão, J; Liu, B H; Hostyn, V; Van Slycken, J; Van Montagu, M; Boerjan, W

2001-01-01

Populus deltoides, P. nigra, and P. trichocarpa are the most important species for poplar breeding programs worldwide. In addition, Populus has become a model for fundamental research on trees. Linkage maps were constructed for these three species by analyzing progeny of two controlled crosses sharing the same female parent, Populus deltoides cv. S9-2 x P. nigra cv. Ghoy and P. deltoides cv. S9-2 x P. trichocarpa cv. V24. The two-way pseudotestcross mapping strategy was used to construct the maps. Amplified fragment length polymorphism (AFLP) markers that segregated 1:1 were used to form the four parental maps. Microsatellites and sequence-tagged sites were used to align homoeologous groups between the maps and to merge linkage groups within the individual maps. Linkage analysis and alignment of the homoeologous groups resulted in 566 markers distributed over 19 groups for P. deltoides covering 86% of the genome, 339 markers distributed over 19 groups for P. trichocarpa covering 73%, and 369 markers distributed over 28 groups for P. nigra covering 61%. Several tests for randomness showed that the AFLP markers were randomly distributed over the genome. PMID:11404342

Genetic stability of ectomycorrhizal fungi is not affected by cryopreservation at -130 °C or cold storage with repeated sub-cultivations over a period of 2 years.

PubMed

Crahay, Charlotte; Munaut, Françoise; Colpaert, Jan V; Huret, Stéphanie; Declerck, Stéphane

2017-08-01

Cryopreservation is considered the most reliable method for storage of filamentous fungi including ectomycorrhizal (ECM) fungi. A number of studies, however, have reported genetic changes in fungus cultures following cryopreservation. In the present study, the genetic stability of six ECM fungus isolates was analyzed using amplified fragment length polymorphism (AFLP). The isolates were preserved for 2 years either by cryopreservation (at -130 °C) or by storage at 4 °C with regular sub-cultivation. A third preservation treatment consisting of isolates maintained on Petri dishes at 22-23 °C for 2 years (i.e., without any sub-cultivation) was included and used as a control. The differences observed in AFLP patterns between the three preservation methods remained within the range of the total error generated by the AFLP procedure (6.85%). Therefore, cryopreservation at -130 °C and cold storage with regular sub-cultivation did not affect the genetic stability of the ECM fungus isolates, and both methods can be used for the routine storage of ECM fungus isolates over a period of 2 years.

Characterization of Betula platyphylla gene transcripts associated with early development of male inflorescence.

PubMed

Xing, Lei; Liu, Xue-Mei

2012-02-01

Birch (Betula platyphylla), an eminent tree species in Northeast and Inner Mongolia of China, has been widely used in architecture, furniture, and paper making in recent years. In order to retrieve genes involved in early development of B. platyphylla male inflorescence, RNA populations extracted from early and late developmental stage were analyzed by cDNA-Amplified Fragment Length Polymorphism (cDNA-AFLP) technique. Following amplification of 256 pairs of primer combinations, ~7000 fragments were generated, of which 350 transcripts expressing more in early stage than late. Of 350 specific transcripts, 198 clear and reproducible electrophoresis bands were retrieved and sequenced successfully, 74 of them (37%) showing significant homologies to known genes after GO annotation. Majority of the predicted gene products were involved in metabolism (24.56%), cellular process (27.19%), response to stimulus (11.4%) and cell growth (8.7%). Transcripts ME56, ME108, ME206 and ME310, representing metabolism, cellular process, response to stimulus and cell growth, respectively, were selected for further study to validate cDNA-AFLP expression patterns via RT-PCR and qRT-PCR analysis. RT-PCR and qRT-PCR expression pattern results were consistent with cDNA-AFLP analysis results.

Strain variation and geographic endemism in Streptococcus iniae.

PubMed

Kvitt, H; Colorni, A

2004-10-21

Twenty-six Israeli isolates of Streptococcus iniae from both marine and fresh/brackish water sources were compared with each other and with 9 foreign isolates. All the isolates were tentatively identified according to their biochemical profile. Direct sequencing of approximately 600 bp PCR products of the 16S rDNA confirmed their identification as S. iniae at the molecular level and revealed a new (one-nucleotide) variant among Israeli isolates, in addition to 2 variants that had been previously reported. Strain variation was further examined by subjecting the isolates to randomly amplified polymorphic DNA (RAPD) and amplified fragment length polymorphism (AFLP) analyses. The RAPD method allowed separation of the isolates into only 2 groups, one including 5 Israeli fresh/brackish water isolates and one including all the other isolates. The AFLP method grouped the Israeli marine isolates into one homogeneous cluster, although they had been obtained in different years (1995 to 2001) from different species of fish, and from wild (Red Sea) as well as cultured (both Mediterranean and Red Sea) sources. The Israeli fresh/brackish water isolates and foreign isolates separated into distinct entities that clustered at generally high degrees of similarity. The distance between the clusters of the Israeli marine and fresh/brackish water isolates indicates that the S. iniae streptococcosis that has been afflicting the aquaculture industries in the 2 environments in recent years was caused by distinct strains. AFLP showed superior discriminative properties over RAPD in detecting intraspecific variation and proved to be an important tool for the characterization of S. iniae. A correlation between strain variation and geographic endemism was established.

An AFLP genetic linkage map of pacific abalone ( Haliotis discus hannai)

NASA Astrophysics Data System (ADS)

Qi, Li; Yanhong, Xu; Ruihai, Yu; Akihiro, Kijima

2007-07-01

A genetic linkage map of Pacific abalone ( Haliotis discus hannai) was constructed using AFLP markers based on a two-way pseudo-testeross strategy in a full-sib family. With 33 primer combinations, a total of 455 markers (225 from the female parent and 230 from the male parent) segregated in a 1:1 ratio, corresponding to DNA polymorphism: heterozygous in one parent and null in the other. The female framework map consisted of 174 markers distributed in 18 linkage groups, equivalent to the H. discus hannai haploid chromosome number, and spanning a total length of 2031.4 cM, with an average interval of 13.0 cM between adjacent markers. The male framework map consisted of 195 markers mapped on 19 linkage groups, spanning a total length of 2273.4 cM, with an average spacing of 12.9 cM between adjacent markers. The estimated coverage for the framework linkage maps was 81.2% for the female and 82.1% for the male, on the basis of two estimates of genome length. Fifty-two markers (11.4%) remained unlinked. The level of segregation distortion observed in this cross was 20.4%. These linkage maps will serve as a starting point for linkage studies in the Pacific abalone with potential application for marker-assisted selection in breeding programs.

Host-Specific Phenotypic Plasticity of the Turtle Barnacle Chelonibia testudinaria: A Widespread Generalist Rather than a Specialist

PubMed Central

Chu, Ka Hou; Cheng, I-Jiunn; Chan, Benny K. K.

2013-01-01

Turtle barnacles are common epibionts on marine organisms. Chelonibia testudinaria is specific on marine turtles whereas C. patula is a host generalist, but rarely found on turtles. It has been questioned why C. patula, being abundant on a variety of live substrata, is almost absent from turtles. We evaluated the genetic (mitochondrial COI, 16S and 12S rRNA, and amplified fragment length polymorphism (AFLP)) and morphological differentiation of C. testudinaia and C. patula from different hosts, to determine the mode of adaptation exhibited by Chelonibia species on different hosts. The two taxa demonstrate clear differences in shell morphology and length of 4–6th cirri, but very similar in arthropodal characters. Moreover, we detected no genetic differentiation in mitochondrial DNA and AFLP analyses. Outlier detection infers insignificant selection across loci investigated. Based on combined morphological and molecular evidence, we proposed that C. testudinaria and C. patula are conspecific, and the two morphs with contrasting shell morphologies and cirral length found on different host are predominantly shaped by developmental plasticity in response to environmental setting on different hosts. Chelonibia testudinaria is, thus, a successful general epibiotic fouler and the phenotypic responses postulated can increase the fitness of the animals when they attach on hosts with contrasting life-styles. PMID:23469208

High-density genetic maps for loci involved in nuclear male sterility (NMS1) and sporophytic self-incompatibility (S-locus) in chicory (Cichorium intybus L., Asteraceae).

PubMed

Gonthier, Lucy; Blassiau, Christelle; Mörchen, Monika; Cadalen, Thierry; Poiret, Matthieu; Hendriks, Theo; Quillet, Marie-Christine

2013-08-01

High-density genetic maps were constructed for loci involved in nuclear male sterility (NMS1-locus) and sporophytic self-incompatibility (S-locus) in chicory (Cichorium intybus L.). The mapping population consisted of 389 F1' individuals derived from a cross between two plants, K28 (male-sterile) and K59 (pollen-fertile), both heterozygous at the S-locus. This F1' mapping population segregated for both male sterility (MS) and strong self-incompatibility (SI) phenotypes. Phenotyping F1' individuals for MS allowed us to map the NMS1-locus to linkage group (LG) 5, while controlled diallel and factorial crosses to identify compatible/incompatible phenotypes mapped the S-locus to LG2. To increase the density of markers around these loci, bulked segregant analysis was used. Bulks and parental plants K28 and K59 were screened using amplified fragment length polymorphism (AFLP) analysis, with a complete set of 256 primer combinations of EcoRI-ANN and MseI-CNN. A total of 31,000 fragments were generated, of which 2,350 showed polymorphism between K59 and K28. Thirteen AFLP markers were identified close to the NMS1-locus and six in the vicinity of the S-locus. From these AFLP markers, eight were transformed into sequence-characterized amplified region (SCAR) markers and of these five showed co-dominant polymorphism. The chromosomal regions containing the NMS1-locus and the S-locus were each confined to a region of 0.8 cM. In addition, we mapped genes encoding proteins similar to S-receptor kinase, the female determinant of sporophytic SI in the Brasicaceae, and also markers in the vicinity of the putative S-locus of sunflower, but none of these genes or markers mapped close to the chicory S-locus.

Genetic Map Construction and Quantitative Trait Locus (QTL) Detection of Growth-Related Traits in Litopenaeus vannamei for Selective Breeding Applications

PubMed Central

Andriantahina, Farafidy; Liu, Xiaolin; Huang, Hao

2013-01-01

Growth is a priority trait from the point of view of genetic improvement. Molecular markers linked to quantitative trait loci (QTL) have been regarded as useful for marker-assisted selection (MAS) in complex traits as growth. Using an intermediate F2 cross of slow and fast growth parents, a genetic linkage map of Pacific whiteleg shrimp, Litopenaeusvannamei , based on amplified fragment length polymorphisms (AFLP) and simple sequence repeats (SSR) markers was constructed. Meanwhile, QTL analysis was performed for growth-related traits. The linkage map consisted of 451 marker loci (429 AFLPs and 22 SSRs) which formed 49 linkage groups with an average marker space of 7.6 cM; they spanned a total length of 3627.6 cM, covering 79.50% of estimated genome size. 14 QTLs were identified for growth-related traits, including three QTLs for body weight (BW), total length (TL) and partial carapace length (PCL), two QTLs for body length (BL), one QTL for first abdominal segment depth (FASD), third abdominal segment depth (TASD) and first abdominal segment width (FASW), which explained 2.62 to 61.42% of phenotypic variation. Moreover, comparison of linkage maps between L . vannamei and Penaeus japonicus was applied, providing a new insight into the genetic base of QTL affecting the growth-related traits. The new results will be useful for conducting MAS breeding schemes in L . vannamei . PMID:24086466

Assessment of genetic and epigenetic variation in hop plants regenerated from sequential subcultures of organogenic calli.

PubMed

Peredo, Elena L; Revilla, M Angeles; Arroyo-García, Rosa

2006-10-01

Organogenic calli induced from internodal segments were subcultured three times. Regenerated plants obtained from each subculture were analysed by molecular methods. No major genetic rearrangements were detected in the callus-derived plants since none of the amplified fragment-length polymorphism (AFLP) loci were found to be polymorphic. However, epigenetic changes due to a demethylation process were detected by methylation-sensitive amplified polymorphism (MSAP) technique. The results allowed inference of the possible relationship among the plants derived from different calli subcultures and the in vitro control. The plants recovered from the first and second callus subcultures clustered with the in vitro control pools in the phenogram while the regenerants from the third callus subculture showed the highest genetic distance with the controls. This is the first study reporting data about the genetic stability of callus-derived Humulus lupulus L. plants.

Next-generation sequencing for typing and detection of resistance genes: performance of a new commercial method during an outbreak of extended-spectrum-beta-lactamase-producing Escherichia coli.

PubMed

Veenemans, J; Overdevest, I T; Snelders, E; Willemsen, I; Hendriks, Y; Adesokan, A; Doran, G; Bruso, S; Rolfe, A; Pettersson, A; Kluytmans, J A J W

2014-07-01

Next-generation sequencing (NGS) has the potential to provide typing results and detect resistance genes in a single assay, thus guiding timely treatment decisions and allowing rapid tracking of transmission of resistant clones. We evaluated the performance of a new NGS assay (Hospital Acquired Infection BioDetection System; Pathogenica) during an outbreak of sequence type 131 (ST131) Escherichia coli infections in a nursing home in The Netherlands. The assay was performed on 56 extended-spectrum-beta-lactamase (ESBL) E. coli isolates collected during 2 prevalence surveys (March and May 2013). Typing results were compared to those of amplified fragment length polymorphism (AFLP), whereby we visually assessed the agreement of the BioDetection phylogenetic tree with clusters defined by AFLP. A microarray was considered the gold standard for detection of resistance genes. AFLP identified a large cluster of 31 indistinguishable isolates on adjacent departments, indicating clonal spread. The BioDetection phylogenetic tree showed that all isolates of this outbreak cluster were strongly related, while the further arrangement of the tree also largely agreed with other clusters defined by AFLP. The BioDetection assay detected ESBL genes in all but 1 isolate (sensitivity, 98%) but was unable to discriminate between ESBL and non-ESBL TEM and SHV beta-lactamases or to specify CTX-M genes by group. The performance of the hospital-acquired infection (HAI) BioDetection System for typing of E. coli isolates compared well with the results of AFLP. Its performance with larger collections from different locations, and for typing of other species, was not evaluated and needs further study. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

Adaptation to a steep environmental gradient and an associated barrier to gene exchange in Littorina saxatilis.

PubMed

Grahame, John W; Wilding, Craig S; Butlin, Roger K

2006-02-01

Steep environmental gradients offer important opportunities to study the interaction between natural selection and gene flow. Allele frequency clines are expected to form at loci under selection, but unlinked neutral alleles may pass easily across these clines unless a generalized barrier evolves. Here we consider the distribution of forms of the intertidal gastropod Littorina saxatilis, analyzing shell shape and amplified fragment length polymorphism (AFLP) loci on two rocky shores in Britain. On the basis of previous work, the AFLP loci were divided into differentiated and undifferentiated groups. On both shores, we have shown a sharp cline in allele frequencies between the two morphs for differentiated AFLP loci. This is coincident with a habitat transition on the shore where the two habitats (cliff and boulder field) are immediately contiguous. The allele frequency clines coincide with a cline in shell morphology. In the middle of the cline, linkage disequilibrium for the differentiated loci rises in accordance with expectation. The clines are extremely narrow relative to dispersal, probably as a result of both strong selection and habitat choice. An increase in F(ST) for undifferentiated AFLPs between morphs, relative to within-morph comparisons, is consistent with there being a general barrier to gene flow across the contact zone. These features are consistent either with an episode of allopatric divergence followed by secondary contact or with primary, nonallopatric divergence. Further data will be needed to distinguish between these alternatives.

Species-specific AFLP markers for identification of Zingiber officinale, Z. montanum and Z. zerumbet (Zingiberaceae).

PubMed

Ghosh, S; Majumder, P B; Sen Mandi, S

2011-02-08

The Zingiber genus, which includes the herbs known as gingers, commonly used in cooking, is well known for its medicinal properties, as described in the Indian pharmacopoeia. Different members of this genus, although somewhat similar in morphology, differ widely in their pharmacological and therapeutic properties. The most important species of this genus, with maximal therapeutic properties, is Zingiber officinale (garden ginger), which is often adulterated with other less-potent Zingiber sp. There is an existing demand in the herbal drug industry for an authentication system for the Zingiber sp in order to facilitate their commercial use as genuine phytoceuticals. To this end, we used amplified fragment length polymorphism (AFLP) to produce DNA fingerprints for three Zingiber species. Sixteen collections (six of Z. officinale, five of Z. montanum, and five of Z. zerumbet) were used in the study. Seven selective primer pairs were found to be useful for all the accessions. A total of 837 fragments were produced by these primer pairs. Species-specific markers were identified for all three Zingiber species (91 for Z. officinale, 82 for Z. montanum, and 55 for Z. zerumbet). The dendogram analysis generated from AFLP patterns showed that Z. montanum and Z. zerumbet are phylogenetically closer to each other than to Z. officinale. The AFLP fingerprints of the Zingiber species could be used to authenticate Zingiber sp-derived drugs and to resolve adulteration-related problems faced by the commercial users of these herbs.

DNA Methylation and Methylation Polymorphism in Genetically Stable In vitro Regenerates of Jatropha curcas L. Using Methylation-Sensitive AFLP Markers.

PubMed

Rathore, Mangal S; Jha, Bhavanath

2016-03-01

The present investigation aimed to evaluate the degree and pattern of DNA methylation using methylation-sensitive AFLP (MS-AFLP) markers in genetically stable in vitro regenerates of Jatropha curcas L.. The genetically stable in vitro regenerates were raised through direct organogenesis via enhanced axillary shoot bud proliferation (Protocol-1) and in vitro-derived leaf regeneration (Protocol-2). Ten selective combinations of MS-AFLP primers produced 462 and 477 MS-AFLP bands in Protocol-1 (P-1) and Protocol-2 (P-2) regenerates, respectively. In P-1 regenerates, 15.8-31.17 % DNA was found methylated with an average of 25.24 %. In P-2 regenerates, 15.93-32.7 % DNA was found methylated with an average of 24.11 %. Using MS-AFLP in P-1 and P-2 regenerates, 11.52-25.53 % and 13.33-25.47 % polymorphism in methylated DNA was reported, respectively. Compared to the mother plant, P-1 regenerates showed hyper-methylation while P-2 showed hypo-methylation. The results clearly indicated alternation in degree and pattern of DNA methylation; hence, epigenetic instability in the genetically stable in vitro regenerates of J. curcas, developed so far using two different regeneration systems and explants of two different origins. The homologous nucleotide fragments in genomes of P-1 and P-2 regenerates showing methylation re-patterning might be involved in immediate adaptive responses and developmental processes through differential regulation of transcriptome under in vitro conditions.

Clonality, genetic diversity and support for the diversifying selection hypothesis in natural populations of a flower-living yeast.

PubMed

Herrera, C M; Pozo, M I; Bazaga, P

2011-11-01

Vast amounts of effort have been devoted to investigate patterns of genetic diversity and structuring in plants and animals, but similar information is scarce for organisms of other kingdoms. The study of the genetic structure of natural populations of wild yeasts can provide insights into the ecological and genetic correlates of clonality, and into the generality of recent hypotheses postulating that microbial populations lack the potential for genetic divergence and allopatric speciation. Ninety-one isolates of the flower-living yeast Metschnikowia gruessii from southeastern Spain were DNA fingerprinted using amplified fragment length polymorphism (AFLP) markers. Genetic diversity and structuring was investigated with band-based methods and model- and nonmodel-based clustering. Linkage disequilibrium tests were used to assess reproduction mode. Microsite-dependent, diversifying selection was tested by comparing genetic characteristics of isolates from bumble bee vectors and different floral microsites. AFLP polymorphism (91%) and genotypic diversity were very high. Genetic diversity was spatially structured, as shown by amova (Φ(st)  = 0.155) and clustering. The null hypothesis of random mating was rejected, clonality seeming the prevailing reproductive mode in the populations studied. Genetic diversity of isolates declined from bumble bee mouthparts to floral microsites, and frequency of five AFLP markers varied significantly across floral microsites, thus supporting the hypothesis of diversifying selection on clonal lineages. Wild populations of clonal fungal microbes can exhibit levels of genetic diversity and spatial structuring that are not singularly different from those shown by sexually reproducing plants or animals. Microsite-dependent, divergent selection can maintain high local and regional genetic diversity in microbial populations despite extensive clonality. © 2011 Blackwell Publishing Ltd.

Untangling individual variation in natural populations: ecological, genetic and epigenetic correlates of long-term inequality in herbivory.

PubMed

Herrera, C M; Bazaga, P

2011-04-01

Individual variation in ecologically important features of organisms is a crucial element in ecology and evolution, yet disentangling its underlying causes is difficult in natural populations. We applied a genomic scan approach using amplified fragment length polymorphism (AFLP) markers to quantify the genetic basis of long-term individual differences in herbivory by mammals at a wild population of the violet Viola cazorlensis monitored for two decades. In addition, methylation-sensitive amplified polymorphism (MSAP) analyses were used to investigate the association between browsing damage and epigenetic characteristics of individuals, an aspect that has been not previously explored for any wild plant. Structural equation modelling was used to identify likely causal structures linking genotypes, epigenotypes and herbivory. Individuals of V. cazorlensis differed widely in the incidence of browsing mammals over the 20-year study period. Six AFLP markers (1.6% of total) were significantly related to herbivory, accounting altogether for 44% of population-wide variance in herbivory levels. MSAP analyses revealed considerable epigenetic variation among individuals, and differential browsing damage was significantly related to variation in multilocus epigenotypes. In addition, variation across plants in epigenetic characteristics was related to variation in several herbivory-related AFLP markers. Statistical comparison of alternative causal models suggested that individual differences in herbivory are the outcome of a complex causal structure where genotypes and epigenotypes are interconnected and have direct and indirect effects on herbivory. Insofar as methylation states of MSAP markers influential on herbivory are transgenerationally heritable, herbivore-driven evolutionary changes at the study population will involve correlated changes in genotypic and epigenotypic distributions. © 2011 Blackwell Publishing Ltd.

Genetic diversity of Kenyan native oyster mushroom (Pleurotus).

PubMed

Otieno, Ojwang D; Onyango, Calvin; Onguso, Justus Mungare; Matasyoh, Lexa G; Wanjala, Bramwel W; Wamalwa, Mark; Harvey, Jagger J W

2015-01-01

Members of the genus Pleurotus, also commonly known as oyster mushroom, are well known for their socioeconomic and biotechnological potentials. Despite being one of the most important edible fungi, the scarce information about the genetic diversity of the species in natural populations has limited their sustainable utilization. A total of 71 isolates of Pleurotus species were collected from three natural populations: 25 isolates were obtained from Kakamega forest, 34 isolates from Arabuko Sokoke forest and 12 isolates from Mount Kenya forest. Amplified fragment length polymorphism (AFLP) was applied to thirteen isolates of locally grown Pleurotus species obtained from laboratory samples using five primer pair combinations. AFLP markers and internal transcribed spacer (ITS) sequences of the ribosomal DNA were used to estimate the genetic diversity and evaluate phylogenetic relationships, respectively, among and within populations. The five primer pair combinations generated 293 polymorphic loci across the 84 isolates. The mean genetic diversity among the populations was 0.25 with the population from Arabuko Sokoke having higher (0.27) diversity estimates compared to Mount Kenya population (0.24). Diversity between the isolates from the natural population (0.25) and commercial cultivars (0.24) did not differ significantly. However, diversity was greater within (89%; P > 0.001) populations than among populations. Homology search analysis against the GenBank database using 16 rDNA ITS sequences randomly selected from the two clades of AFLP dendrogram revealed three mushroom species: P. djamor, P. floridanus and P. sapidus; the three mushrooms form part of the diversity of Pleurotus species in Kenya. The broad diversity within the Kenyan Pleurotus species suggests the possibility of obtaining native strains suitable for commercial cultivation. © 2015 by The Mycological Society of America.

Multilocus analyses indicate a mosaic distribution of hybrid populations in ground squirrels (genus Ictidomys)

PubMed Central

Thompson, Cody W; Anwarali Khan, Faisal Ali; Stangl, Frederick B; Baker, Robert J; Bradley, Robert D

2013-01-01

DNA sequence data from mitochondrial cytochrome-b (Cytb) and Y-linked structural maintenance of chromosomes (SmcY) genes were combined with 478 nuclear loci obtained from amplified fragment length polymorphisms (AFLP) to assess the extent of hybridization and genetic spatial structure of populations in two hybridizing species of ground squirrel (Ictidomys parvidens and Ictidomys tridecemlineatus). Based on AFLP analyses of 134 individuals from 28 populations, 10 populations were identified that possessed hybrid individuals. Overall estimates of FST values revealed strong support for population structure in the Cytb data set; however, analyses of the SmcY gene and the AFLP data indicated ongoing gene flow between species. Pairwise FST comparisons of populations were not significant for the SmcY gene; although they were significant for the Cytb gene, indicating that these populations were structured and that gene flow was minimal. Therefore, gene flow between I. parvidens and I. tridecemlineatus appeared to be restricted to populations that exhibited hybridization. In addition, the fragmented nature of the geographic landscape suggested limited gene flow between populations. As a result, the distributional pattern of interspersed parental and hybrid populations were compatible with a mosaic hybrid zone model. Because ground squirrels display female philopatry and male-biased dispersal, the ecology of these species is compatible with this hypothesis. PMID:24340186

GISH and AFLP analyses of novel Brassica napus lines derived from one hybrid between B. napus and Orychophragmus violaceus.

PubMed

Ma, Ni; Li, Zai-Yun; Cartagena, J A; Fukui, K

2006-10-01

New Brassica napus inbred lines with different petal colors and with canola quality and increased levels of oleic (approximately 70%, 10% higher than that of B. napus parent) and linoleic (28%) acids have been developed in the progenies of one B. napus cv. Oro x Orychophragmus violaceus F5 hybrid plant (2n = 31). Their genetic constituents were analyzed by using the methods of genomic in situ hybridization (GISH) and amplified fragments length polymorphism (AFLP). No intact chromosomes of O. violaceus origin were detected by GISH in their somatic cells of ovaries and root tips (2n = 38) and pollen mother cells (PMCs) with normal chromosome pairing (19 bivalents) and segregation (19:19), though signals of variable sizes and intensities were located mainly at terminal and centromeric parts of some mitotic chromosomes and meiotic bivalents at diakinesis or chromosomes in anaphase I groups and one large patch of chromatin was intensively labeled and separated spatially in some telophase I nuclei and metaphase II PMCs. AFLP analysis revealed that substantial genomic changes have occurred in these lines and O. violaceus-specific bands, deleted bands in 'Oro' and novel bands for two parents were detected. The possible mechanisms for these results were discussed.

Genetic Variability and Population Structure of Disanthus cercidifolius subsp. longipes (Hamamelidaceae) Based on AFLP Analysis

PubMed Central

Yu, Yi; Fan, Qiang; Shen, Rujiang; Guo, Wei; Jin, Jianhua; Cui, Dafang; Liao, Wenbo

2014-01-01

Disanthus cercidifolius subsp. longipes is an endangered species in China. Genetic diversity and structure analysis of this species was investigated using amplified fragments length polymorphism (AFLP) fingerprinting. Nei's gene diversity ranged from 0.1290 to 0.1394. The AMOVA indicated that 75.06% of variation was distributed within populations, while the between-group component 5.04% was smaller than the between populations-within-group component 19.90%. Significant genetic differentiation was detected between populations. Genetic and geographical distances were not correlated. PCA and genetic structure analysis showed that populations from East China were together with those of the Nanling Range. These patterns of genetic diversity and levels of genetic variation may be the result of D. c. subsp. longipes restricted to several isolated habitats and “excess flowers production, but little fruit set”. It is necessary to protect all existing populations of D. c. subsp. longipes in order to preserve as much genetic variation as possible. PMID:25250583

A High-throughput AFLP-based Method for Constructing Integrated Genetic and Physical Maps: Progress Toward a Sorghum Genome Map

PubMed Central

Klein, Patricia E.; Klein, Robert R.; Cartinhour, Samuel W.; Ulanch, Paul E.; Dong, Jianmin; Obert, Jacque A.; Morishige, Daryl T.; Schlueter, Shannon D.; Childs, Kevin L.; Ale, Melissa; Mullet, John E.

2000-01-01

Sorghum is an important target for plant genomic mapping because of its adaptation to harsh environments, diverse germplasm collection, and value for comparing the genomes of grass species such as corn and rice. The construction of an integrated genetic and physical map of the sorghum genome (750 Mbp) is a primary goal of our sorghum genome project. To help accomplish this task, we have developed a new high-throughput PCR-based method for building BAC contigs and locating BAC clones on the sorghum genetic map. This task involved pooling 24,576 sorghum BAC clones (∼4× genome equivalents) in six different matrices to create 184 pools of BAC DNA. DNA fragments from each pool were amplified using amplified fragment length polymorphism (AFLP) technology, resolved on a LI-COR dual-dye DNA sequencing system, and analyzed using Bionumerics software. On average, each set of AFLP primers amplified 28 single-copy DNA markers that were useful for identifying overlapping BAC clones. Data from 32 different AFLP primer combinations identified ∼2400 BACs and ordered ∼700 BAC contigs. Analysis of a sorghum RIL mapping population using the same primer pairs located ∼200 of the BAC contigs on the sorghum genetic map. Restriction endonuclease fingerprinting of the entire collection of sorghum BAC clones was applied to test and extend the contigs constructed using this PCR-based methodology. Analysis of the fingerprint data allowed for the identification of 3366 contigs each containing an average of 5 BACs. BACs in ∼65% of the contigs aligned by AFLP analysis had sufficient overlap to be confirmed by DNA fingerprint analysis. In addition, 30% of the overlapping BACs aligned by AFLP analysis provided information for merging contigs and singletons that could not be joined using fingerprint data alone. Thus, the combination of fingerprinting and AFLP-based contig assembly and mapping provides a reliable, high-throughput method for building an integrated genetic and physical map of the sorghum genome. [The sequence data described in this paper have been submitted to the GenBank data library under accession no. AF218263.] PMID:10854411

SNP Discovery by Illumina-Based Transcriptome Sequencing of the Olive and the Genetic Characterization of Turkish Olive Genotypes Revealed by AFLP, SSR and SNP Markers

PubMed Central

Kaya, Hilal Betul; Cetin, Oznur; Kaya, Hulya; Sahin, Mustafa; Sefer, Filiz; Kahraman, Abdullah; Tanyolac, Bahattin

2013-01-01

Background The olive tree (Olea europaea L.) is a diploid (2n = 2x = 46) outcrossing species mainly grown in the Mediterranean area, where it is the most important oil-producing crop. Because of its economic, cultural and ecological importance, various DNA markers have been used in the olive to characterize and elucidate homonyms, synonyms and unknown accessions. However, a comprehensive characterization and a full sequence of its transcriptome are unavailable, leading to the importance of an efficient large-scale single nucleotide polymorphism (SNP) discovery in olive. The objectives of this study were (1) to discover olive SNPs using next-generation sequencing and to identify SNP primers for cultivar identification and (2) to characterize 96 olive genotypes originating from different regions of Turkey. Methodology/Principal Findings Next-generation sequencing technology was used with five distinct olive genotypes and generated cDNA, producing 126,542,413 reads using an Illumina Genome Analyzer IIx. Following quality and size trimming, the high-quality reads were assembled into 22,052 contigs with an average length of 1,321 bases and 45 singletons. The SNPs were filtered and 2,987 high-quality putative SNP primers were identified. The assembled sequences and singletons were subjected to BLAST similarity searches and annotated with a Gene Ontology identifier. To identify the 96 olive genotypes, these SNP primers were applied to the genotypes in combination with amplified fragment length polymorphism (AFLP) and simple sequence repeats (SSR) markers. Conclusions/Significance This study marks the highest number of SNP markers discovered to date from olive genotypes using transcriptome sequencing. The developed SNP markers will provide a useful source for molecular genetic studies, such as genetic diversity and characterization, high density quantitative trait locus (QTL) analysis, association mapping and map-based gene cloning in the olive. High levels of genetic variation among Turkish olive genotypes revealed by SNPs, AFLPs and SSRs allowed us to characterize the Turkish olive genotype. PMID:24058483

Genetic structure of Eurasian and North American Leymus (Triticeae) wildryes assessed by chloroplast DNA sequences and AFLP profiles

Treesearch

C. Mae Culumber; Steve R. Larson; Kevin B. Jensen; Thomas A. Jones

2011-01-01

Leymus is a genomically defined allopolyploid of genus Triticeae with two distinct subgenomes. Chloroplast DNA sequences of Eurasian and North American species are distinct and polyphyletic. However, phylogenies derived from chloroplast and nuclear DNA sequences are confounded by polyploidy and lack of polymorphism among many taxa. The AFLP technique can resolve...

Characterization of ten date palm (Phoenix dactylifera L.) cultivars from Saudi Arabia using AFLP and ISSR markers.

PubMed

Sabir, Jamal S M; Abo-Aba, Salah; Bafeel, Sameera; Zari, Talal A; Edris, Sherif; Shokry, Ahmed M; Atef, Ahmed; Gadalla, Nour O; Ramadan, Ahmed M; Al-Kordy, Magdy A; El-Domyati, Fotouh M; Jansen, Robert K; Bahieldin, Ahmed

2014-01-01

Date palm is the most economically important plant in the Middle East due to its nutritionally valuable fruit. The development of accurate DNA fingerprints to characterize cultivars and the detection of genetic diversity are of great value for breeding programs. The present study explores the usefulness of ISSR and AFLP molecular markers to detect relationships among 10 date palm (Phoenix dactylifera L.) cultivars from Saudi Arabia. Thirteen ISSR primers and six AFLP primer combinations were examined. The level of polymorphism among cultivars for ISSRs ranged from 20% to 100% with an average of 85%. Polymorphism levels for AFLPs ranged from 63% to 84% with an average of 76%. The total number of cultivar-specific markers was 241, 208 of which were generated from AFLP analysis. AJWA cultivar had the highest number of cultivar-specific ISSR markers, whereas DEK, PER, SUK-Q, SHA and MOS-H cultivars had the lowest. RAB and SHA cultivars had the most and least AFLP cultivar-specific markers, respectively. The highest pairwise similarity indices for ISSRs, AFLPs and combined markers were 84% between DEK (female) and PER (female), 81% between SUK-Q (male) and RAB (male), and 80% between SUK-Q (male) and RAB (male), respectively. The lowest similarity indices were 65% between TAB (female) and SUK-Q (male), 67% between SUK-A (female) and SUK-Q (male), and 67% between SUK-A (female) and SUK-Q (male). Cultivars of the same sex had higher pairwise similarities than those between cultivars of different sex. The Neighbor-Joining (NJ) tree generated from the ISSR dataset was not well resolved and bootstrap support for resolved nodes in the tree was low. AFLP and combined data generated completely resolved trees with high levels of bootstrap support. In conclusion, AFLP and ISSR approaches enabled discrimination among 10 date palm cultivars of from Saudi Arabia, which will provide valuable information for future improvement of this important crop. Copyright © 2013 Académie des sciences. All rights reserved.

Molecular evidence for a founder effect in invasive house finch (Carpodacus mexicanus) populations experiencing an emergent disease epidemic.

PubMed

Hawley, Dana M; Hanley, Daniel; Dhondt, André A; Lovette, Irby J

2006-01-01

The impact of founder events on levels of genetic variation in natural populations remains a topic of significant interest. Well-documented introductions provide a valuable opportunity to examine how founder events influence genetic diversity in invasive species. House finches (Carpodacus mexicanus) are passerine birds native to western North America, with the large eastern North American population derived from a small number of captive individuals released in the 1940s. Previous comparisons using amplified fragment length polymorphism (AFLP) markers found equivalent levels of diversity in eastern and western populations, suggesting that any genetic effects of the founder event were ameliorated by the rapid growth of the newly established population. We used an alternative marker system, 10 highly polymorphic microsatellites, to compare levels of genetic diversity between four native and five introduced house finch populations. In contrast to the AFLP comparisons, we found significantly lower allelic richness and heterozygosity in introduced populations across all loci. Three out of five introduced populations showed significant reductions in the ratio of the number of alleles to the allele size range, a within-population characteristic of recent bottlenecks. Finally, native and introduced populations showed significant pairwise differences in allele frequencies in every case, with stronger isolation by distance within the introduced than native range. Overall, our results provide compelling molecular evidence for a founder effect during the introduction of eastern house finches that reduced diversity levels at polymorphic microsatellite loci and may have contributed to the emergence of the Mycoplasma epidemic which recently swept the eastern range of this species.

Genomics and Ethics: The Case of Cloned and/or Transgenic Animals

PubMed Central

2003-01-01

The point of the present study is to illustrate and, if possible, promote the existing link between genomics and ethics, taking the example of cloned and transgenic animals. These ‘new animals’ raise theoretical and practical problems that concern applied ethics. We will explore more particularly an original strategy showing that it is possible, starting from philosophical questioning about the nature of identity, to use a genomic approach, based on amplification fragment length polymorphism (AFLP) and methylation-sensitive amplification polymorphism (MSAP) detection, to provide useful tools to define more rigorously what cloned animals are, by testing their genetic and epigenetic identity. We expect from the future results of this combined approach to stimulate the creativity of the philosophical and ethical reflection about the impact of biotechnology on animals, and to increase scientific involvement in such issues. PMID:18629111

Analyses of amplified fragment length polymorphisms (AFLP) indicate rapid radiation of Diospyros species (Ebenaceae) endemic to New Caledonia

PubMed Central

2013-01-01

Background Radiation in some plant groups has occurred on islands and due to the characteristic rapid pace of phenotypic evolution, standard molecular markers often provide insufficient variation for phylogenetic reconstruction. To resolve relationships within a clade of 21 closely related New Caledonian Diospyros species and evaluate species boundaries we analysed genome-wide DNA variation via amplified fragment length polymorphisms (AFLP). Results A neighbour-joining (NJ) dendrogram based on Dice distances shows all species except D. minimifolia, D. parviflora and D. vieillardii to form unique clusters of genetically similar accessions. However, there was little variation between these species clusters, resulting in unresolved species relationships and a star-like general NJ topology. Correspondingly, analyses of molecular variance showed more variation within species than between them. A Bayesian analysis with BEAST produced a similar result. Another Bayesian method, this time a clustering method, Structure, demonstrated the presence of two groups, highly congruent with those observed in a principal coordinate analysis (PCO). Molecular divergence between the two groups is low and does not correspond to any hypothesised taxonomic, ecological or geographical patterns. Conclusions We hypothesise that such a pattern could have been produced by rapid and complex evolution involving a widespread progenitor for which an initial split into two groups was followed by subsequent fragmentation into many diverging populations, which was followed by range expansion of then divergent entities. Overall, this process resulted in an opportunistic pattern of phenotypic diversification. The time since divergence was probably insufficient for some species to become genetically well-differentiated, resulting in progenitor/derivative relationships being exhibited in a few cases. In other cases, our analyses may have revealed evidence for the existence of cryptic species, for which more study of morphology and ecology are now required. PMID:24330478

Free-Ranging Synanthropic Birds (Ardea alba and Columba livia domestica) as Carriers of Salmonella spp. and Diarrheagenic Escherichia coli in the Vicinity of an Urban Zoo.

PubMed

de Oliveira, Mirela C V; Camargo, Beatriz Q; Cunha, Marcos P V; Saidenberg, Andre Becker; Teixeira, Rodrigo H F; Matajira, Carlos E C; Moreno, Luisa Z; Gomes, Vasco T M; Christ, Ana P G; Barbosa, Mikaela R F; Sato, Maria I Z; Moreno, Andrea M; Knöbl, Terezinha

2018-01-01

The presence of free-ranging urban birds is a risk factor for transmitting pathogens to captive animals and humans alike, including Salmonella spp. and diarrheagenic Escherichia coli. Cloacal swabs from 156 synanthropic Great egrets (Ardea alba) and feral pigeons (Columba livia domestica) that inhabit the surroundings of an urban zoo were processed for the identification of Salmonella spp. and diarrheagenic E. coli pathotypes. Bacterial species identification and genotypic characterization employed the matrix-assisted laser desorption ionization-time of flight mass spectrometry and PCR techniques, respectively, comparing their phylogenetic profiles through amplified fragment length polymorphism (AFLP) analysis. A total of 11 birds were positive for Salmonella Typhimurium (7%) and 9 individuals (5.8%) for diarrheagenic E. coli (enteropathogenic E. coli/Shiga-toxin producing E. coli [EPEC/STEC]) strains. S. Typhimurium strains presented highly similar AFLP profiles (85-100%), whereas EPEC/STEC strains showed more polymorphism. The results show free-ranging birds as carriers for both microorganisms in a zoo environment in Brazil for the first time and suggest these species as possible sources of infection to other animals as well as exposing personnel and visitors to potential zoonotic microorganisms. The presence of carriers highlights the importance of a surveillance system and the need for preventive measures to avoid attracting growing number of synanthropic avian species.

Production and genetic analysis of resynthesized Brassica napus from a B. rapa landrace from the Qinghai-Tibet Plateau and B. alboglabra.

PubMed

Liu, H D; Zhao, Z G; Du, D Z; Deng, C R; Fu, G

2016-01-08

This study aimed to reveal the genetic and epigenetic variations involved in a resynthesized Brassica napus (AACC) generated from a hybridization between a B. rapa (AA) landrace and B. alboglabra (CC). Amplified fragment length polymorphism (AFLP), methylation-sensitive amplified polymorphism, and the cDNA-AFLP technique were performed to detect changes between different generations at the genome, methylation, and transcription levels. We obtained 30 lines of resynthesized B. napus with a mean 1000-seed weight of over 7.50 g. All of the lines were self-compatible, probably because both parents were self-compatible. At the genome level, the S0 generation had the lowest frequency of variations (0.18%) and the S3 generation had the highest (6.07%). The main variation pattern was the elimination of amplified restriction fragments on the CC genome from the S0 to the S4 generations. At the methylation level, we found three loci that exhibited altered methylation patterns on the parental A genome; the variance rate was 1.35%. At the transcription level, we detected 43.77% reverse mutations and 37.56% deletion mutations that mainly occurred on the A and C genomes, respectively, in the S3 generation. Our results highlight the genetic variations that occur during the diploidization of resynthesized B. napus.

Genetic diversity in wild populations of Paulownia fortune.

PubMed

Li, H Y; Ru, G X; Zhang, J; Lu, Y Y

2014-11-01

The genetic diversities of 16 Paulownia fortunei populations involving 143 individuals collected from 6 provinces in China were analyzed using amplified fragment length polymorphism (AFLP). A total of 9 primer pairs with 1169 polymorphic loci were screened out, and each pair possessed 132 bands on average. The percentage of polymorphic bands (98.57%), the effective number of alleles (1.2138-1.2726), Nei's genetic diversity (0.1566-0.1887), and Shannon's information index (0.2692-0.3117) indicated a plentiful genetic diversity and different among Paulownia fortunei populations. The genetic differentiation coefficient between populations was 0.2386, while the gene flow was 1.0954, and the low gene exchange promoted genetic differentiation. Analysis of variance indicated that genetic variation mainly occurred within populations (81.62% of total variation) rather than among populations (18.38%). The 16 populations were divided by unweighted pair-group method with arithmetic means (UPGMA) into 4 groups with obvious regionalism, in which the populations with close geographical locations (latitude) were clustered together.

AFLP fragment isolation technique as a method to produce random sequences for single nucleotide polymorphism discovery in the green turtle, Chelonia mydas.

PubMed

Roden, Suzanne E; Dutton, Peter H; Morin, Phillip A

2009-01-01

The green sea turtle, Chelonia mydas, was used as a case study for single nucleotide polymorphism (SNP) discovery in a species that has little genetic sequence information available. As green turtles have a complex population structure, additional nuclear markers other than microsatellites could add to our understanding of their complex life history. Amplified fragment length polymorphism technique was used to generate sets of random fragments of genomic DNA, which were then electrophoretically separated with precast gels, stained with SYBR green, excised, and directly sequenced. It was possible to perform this method without the use of polyacrylamide gels, radioactive or fluorescent labeled primers, or hybridization methods, reducing the time, expense, and safety hazards of SNP discovery. Within 13 loci, 2547 base pairs were screened, resulting in the discovery of 35 SNPs. Using this method, it was possible to yield a sufficient number of loci to screen for SNP markers without the availability of prior sequence information.

Analysis of DNA Cytosine Methylation Patterns Using Methylation-Sensitive Amplification Polymorphism (MSAP).

PubMed

Guevara, María Ángeles; de María, Nuria; Sáez-Laguna, Enrique; Vélez, María Dolores; Cervera, María Teresa; Cabezas, José Antonio

2017-01-01

Different molecular techniques have been developed to study either the global level of methylated cytosines or methylation at specific gene sequences. One of them is the methylation-sensitive amplified polymorphism technique (MSAP) which is a modification of amplified fragment length polymorphism (AFLP). It has been used to study methylation of anonymous CCGG sequences in different fungi, plants, and animal species. The main variation of this technique resides on the use of isoschizomers with different methylation sensitivity (such as HpaII and MspI) as a frequent-cutter restriction enzyme. For each sample, MSAP analysis is performed using both EcoRI/HpaII- and EcoRI/MspI-digested samples. A comparative analysis between EcoRI/HpaII and EcoRI/MspI fragment patterns allows the identification of two types of polymorphisms: (1) methylation-insensitive polymorphisms that show common EcoRI/HpaII and EcoRI/MspI patterns but are detected as polymorphic amplified fragments among samples and (2) methylation-sensitive polymorphisms which are associated with the amplified fragments that differ in their presence or absence or in their intensity between EcoRI/HpaII and EcoRI/MspI patterns. This chapter describes a detailed protocol of this technique and discusses the modifications that can be applied to adjust the technology to different species of interest.

Radiative evolution of polyploid races of the Iberian carnation Dianthus broteri (Caryophyllaceae).

PubMed

Balao, Francisco; Valente, Luis M; Vargas, Pablo; Herrera, Javier; Talavera, Salvador

2010-07-01

*The micro-evolutionary mechanisms that drive large-scale radiations are not completely understood, partly because of a shortage of population-level studies aimed at identifying putative causes of rapid evolutionary change. The Dianthus broteri complex, representing the largest polyploid series known to date for any species in the genus (2x, 4x, 6x and 12x cytotypes), belongs to a lineage that was recently found to have diversified at unusually rapid rates. *We used a combination of genome sequencing (internal transcribed spacer (ITS), plus chloroplast DNA (cpDNA) regions trnH-psbA, psbA-trnK and trnK-matK) and amplified fragment length polymorphism (AFLP) fingerprinting in 25 populations to infer the evolutionary history of extant polyploid races. *The haplotype, ribotype and AFLP reconstructions showed a star-shaped arrangement suggesting a pattern of radiative evolution. The major, widespread haplotype occurred at all ploidy levels, whereas 20 minor haplotypes were restricted to single populations and cytotypes. In addition, AFLP analyses retrieved well-supported cytogeographic groups: six clades were clearly differentiated in terms of ploidy level and geography. Molecular data indicate that gene flow among different cytotypes is rare or nonexistent. *Our study supports a scenario of rapid diversification in carnations in which autopolyploidy and allopolyploidy, in interaction with geography and/or isolation, have played prominent roles.

Detecting the influence of ornamental Berberis thunbergii var. atropurpurea in invasive populations of Berberis thunbergii (Berberidaceae) using AFLP1.

PubMed

Lubell, Jessica D; Brand, Mark H; Lehrer, Jonathan M; Holsinger, Kent E

2008-06-01

Japanese barberry (Berberis thunbergii DC.) is a widespread invasive plant that remains an important landscape shrub represented by ornamental, purple-leaved forms of the botanical variety atropurpurea. These forms differ greatly in appearance from feral plants, bringing into question whether they contribute to invasive populations or whether the invasions represent self-sustaining populations derived from the initial introduction of the species in the late 19th century. In this study we used amplified fragment length polymorphism (AFLP) markers to determine whether genetic contributions from B. t. var. atropurpurea are found within naturalized Japanese barberry populations in southern New England. Bayesian clustering of AFLP genotypes and principal coordinate analysis distinguished B. t. var. atropurpurea genotypes from 85 plants representing five invasive populations. While a single feral plant resembled B. t. var. atropurpurea phenotypically and fell within the same genetic cluster, all other naturalized plants sampled were genetically distinct from the purple-leaved genotypes. Seven plants from two different sites possessed morphology consistent with Berberis vulgaris (common barberry) or B. ×ottawensis (B. thunbergii × B. vulgaris). Genetic analysis placed these plants in two clusters separate from B. thunbergii. Although the Bayesian analysis indicated some introgression of B. t. var. atropurpurea and B. vulgaris, these genotypes have had limited influence on extant feral populations of B. thunbergii.

Identification of genes associated with low furanocoumarin content in grapefruit.

PubMed

Chen, Chunxian; Yu, Qibin; Wei, Xu; Cancalon, Paul F; Gmitter, Fred G

2014-10-01

Some furanocoumarins in grapefruit (Citrus paradisi) are associated with the so-called grapefruit juice effect. Previous phytochemical quantification and genetic analysis suggested that the synthesis of these furanocoumarins may be controlled by a single gene in the pathway. In this study, cDNA-amplified fragment length polymorphism (cDNA-AFLP) analysis of fruit tissues was performed to identify the candidate gene(s) likely associated with low furanocoumarin content in grapefruit. Fifteen tentative differentially expressed fragments were cloned through the cDNA-AFLP analysis of the grapefruit variety Foster and its spontaneous low-furanocoumarin mutant Low Acid Foster. Sequence analysis revealed a cDNA-AFLP fragment, Contig 6, was homologous to a substrate-proved psoralen synthase gene, CYP71A22, and was part of citrus unigenes Cit.3003 and Csi.1332, and predicted genes Ciclev10004717m in mandarin and orange1.1g041507m in sweet orange. The two predicted genes contained the highly conserved motifs at one of the substrate recognition sites of CYP71A22. Digital gene expression profile showed the unigenes were expressed only in fruit and seed. Quantitative real-time PCR also proved Contig 6 was down-regulated in Low Acid Foster. These results showed the differentially expressed Contig 6 was related to the reduced furanocoumarin levels in the mutant. The identified fragment, homologs, unigenes, and genes may facilitate further furanocoumarin genetic study and grapefruit variety improvement.

DNA methylation and methylation polymorphism in ecotypes of Jatropha curcas L. using methylation-sensitive AFLP markers.

PubMed

Mastan, Shaik G; Rathore, Mangal S; Bhatt, Vacha D; Chikara, J; Ghosh, A

2014-12-01

We investigated DNA methylation and polymorphism in the methylated DNA using AFLP based methylation-sensitive amplification polymorphism (MS-AFLP) markers in ecotypes of Jatropha curcas L. growing in similar and different geo-ecological conditions. Three ecotypes growing in different geo-ecological conditions with environmental heterogeneity (Group-1) and five ecotypes growing in similar environmental conditions (Group-2) were assessed. In ecotypes growing in group-1, 44.32 % DNA was methylated and of which 93.59 % DNA was polymorphic. While in group-2, 32.27 % DNA was methylated, of which 51.64 % DNA was polymorphic. In site 1 and site 2 of group-1, overall methylation was 18.94 and 22.44 % respectively with difference of 3.5 %, while overall polymorphism was 41.14 and 39.23 % with a difference of 1.91 %. In site 1 and site 2 of group-2, overall methylation was 24.68 and 24.18 % respectively with difference of 0.5 %, while overall polymorphism was 12.19 and 12.65 % with a difference of 0.46 %. The difference of methylation percentage and percentage of methylation polymorphism throughout the genome of J. curcas at site 1 and 2 of group-1 is higher than that of J. curcas at site 1 and 2 of group-2. These results correlated the physico-chemical properties of soil at these sites. The variations of physico-chemical properties of soil at Chorwadla (site 1 in group-1 and site 2 in group-2) compared to the soil at Brahmapur (site 2 in group-1) is higher than that of soil at Neswad (site 1 in group-2). The study suggests that these homologous nucleotide sequences probably play important role in ecotype adaptation to environmental heterogeneity by creating epiallelic variations hence in evolution of ecotypes/clines or forms of species showing phenotypic/genotypic differences in different geographical areas.

Intraspecific diversity in Sinningia speciosa (Gesneriaceae: Sinningieae), and possible origins of the cultivated florist's gloxinia

PubMed Central

Zaitlin, David

2012-01-01

Background and aims The florist's gloxinia is a familiar houseplant in the Gesneriaceae, the botanical family that includes the African violet (Saintpaulia) and other ornamental species. The gloxinia's wild progenitor is Sinningia speciosa (Lodd.) Hiern, a Brazilian endemic. Although it has been cultivated for almost 200 years, little is known about the genetic diversity in S. speciosa, how the wild populations relate to one another or even where the cultivated forms originated. Using available wild collections, preliminary phenetic and phylogenetic investigations were conducted to elucidate the interspecific relationships within S. speciosa and to infer the origins of the cultivars. Methodology Amplified fragment length polymorphism (AFLP) analysis was applied to 24 accessions of S. speciosa (17 wild collections, seven cultivars) and one accession each of Sinningia guttata and Sinningia macrophylla. A maximum likelihood (ML) tree was also calculated from an alignment of the nuclear ribosomal internal transcribed spacer sequence from the same 26 accessions. Principal results Dice/UPGMA and principal coordinates analysis of the AFLP data partitioned S. speciosa into several distinct clusters, one of which included S. macrophylla. All cultivated ‘gloxinias’ grouped together in a major cluster with plants from Rio de Janeiro. The AFLP results were compared with a phylogenetic analysis of the ribosomal spacer region, which was informative in S. speciosa. The ML tree generally supported the AFLP results, although several clades lacked strong statistical support. Conclusions Independent analyses of two different data sets show that S. speciosa is a diverse species comprised of several lineages. Genetic distance estimates calculated from the AFLP data were positively correlated with geographic distances between populations, indicating that reproductive isolation could be driving speciation in this taxon. Molecular markers are under development for population genetic studies in S. speciosa, which will make it possible to define evolutionarily significant units for purposes of conservation. PMID:23755350

Glacial survival east and west of the 'Mekong-Salween Divide' in the Himalaya-Hengduan Mountains region as revealed by AFLPs and cpDNA sequence variation in Sinopodophyllum hexandrum (Berberidaceae).

PubMed

Li, Yong; Zhai, Sheng-Nan; Qiu, Ying-Xiong; Guo, Yan-Ping; Ge, Xue-Jun; Comes, Hans Peter

2011-05-01

Molecular phylogeographic studies have recently begun to elucidate how plant species from the Qinghai-Tibetan Plateau (QTP) and adjacent regions responded to the Quaternary climatic oscillations. In this regard, however, far less attention has been paid to the southern and south-eastern declivities of the QTP, i.e. the Himalaya-Hengduan Mountains (HHM) region. Here, we report a survey of amplified fragment length polymorphisms (AFLPs) and chloroplast DNA (cpDNA) sequence variation in the HHM endemic Sinopodophyllum hexandrum, a highly selfing alpine perennial herb with mainly gravity-dispersed berries (105 individuals, 19 localities). We specifically aimed to test a vicariant evolutionary hypothesis across the 'Mekong-Salween Divide', a known biogeographic and phytogeographic boundary of north-to-south trending river valleys separating the East Himalayas and Hengduan Mts. Both cpDNA and AFLPs identified two divergent phylogroups largely congruent with these mountain ranges. There was no genetic depauperation in the more strongly glaciated East Himalayas (AFLPs: H(E)=0.031; cpDNA: h(S)=0.133) compared to the mainly ice-free Hengduan Mts. (AFLPs: H(E)=0.037; cpDNA: h(S)=0.082), while population differentiation was consistently higher in the former region (AFLPs: Φ(ST)=0.522 vs. 0.312; cpDNA: Φ(ST)=0.785 vs. 0.417). Our results suggest that East Himalayan and Hengduan populations of S. hexandrum were once fragmented, persisted in situ during glacials in both areas, and have not merged again, except for a major instance of inter-lineage chloroplast capture identified at the MSD boundary. Our coalescent time estimate for all cpDNA haplotypes (c. 0.37-0.48 mya), together with paleogeological evidence, strongly rejects paleo-drainage formation as a mechanism underlying allopatric fragmentation, whereas mountain glaciers following the ridges of the MSD during glacials (and possible interglacials) could have been responsible. This study thus indicates an important role for mountain glaciers in driving (incipient) allopatric speciation across the MSD in the HHM region by causing vicariant lineage divergence and acting as barriers to post-divergence gene flow. Copyright © 2011 Elsevier Inc. All rights reserved.

Genetic mapping and QTL analysis of agronomic traits in Indian Mucuna pruriens using an intraspecific F₂population.

PubMed

Mahesh, S; Leelambika, M; Jaheer, Md; Anithakumari, A M; Sathyanarayana, N

2016-03-01

Mucuna pruriens is a well-recognized agricultural and horticultural crop with important medicinal use. However, antinutritional factors in seed and adverse morphological characters have negatively affected its cultivation. To elucidate the genetic control of agronomic traits, an intraspecific genetic linkage map of Indian M. pruriens has been developed based on amplified fragment length polymorphism (AFLP) markers using 200 F₂ progenies derived from a cross between wild and cultivated genotypes. The resulting linkage map comprised 129 AFLP markers dispersed over 13 linkage groups spanning a total distance of 618.88 cM with an average marker interval of 4.79 cM. For the first time, three QTLs explaining about 6.05-14.77% of the corresponding total phenotypic variation for three quantitative (seed) traits and, eight QTLs explaining about 25.96% of the corresponding total phenotypic variation for three qualitative traits have been detected on four linkage groups. The map presented here will pave a way for mapping of genes/QTLs for the important agronomic and horticultural traits contrasting between the parents used in this study.

Genetic diversity and population structure of an extremely endangered species: the world's largest Rhododendron.

PubMed

Wu, Fu Qin; Shen, Shi Kang; Zhang, Xin Jun; Wang, Yue Hua; Sun, Wei Bang

2014-12-04

Comprehensive studies on the genetic diversity and structure of endangered species are urgently needed to promote effective conservation and management activities. The big tree rhododendron, Rhododendron protistum var. giganteum, is a highly endangered species with only two known endemic populations in a small area in the southern part of Yunnan Province in China. Unfortunately, limited information is available regarding the population genetics of this species. Therefore, we conducted amplified fragment length polymorphism (AFLP) analysis to characterize the genetic diversity and variation of this species within and between remaining populations. Twelve primer combinations of AFLP produced 447 unambiguous and repetitious bands. Among these bands, 298 (66.67 %) were polymorphic. We found high genetic diversity at the species level (percentage of polymorphic loci = 66.67 %, h = 0.240, I = 0.358) and low genetic differentiation (Gst = 0.110) between the two populations. Gene flow between populations (Nm) was relatively high at 4.065. Analysis of molecular variance results revealed that 22 % of the genetic variation was partitioned between populations and 78 % of the genetic variation was within populations. The presence of moderate to high genetic diversity and low genetic differentiation in the two populations can be explained by life history traits, pollen dispersal and high gene flow (Nm = 4.065). Bayesian structure and principal coordinate analysis revealed that 56 sampled trees were clustered into two groups. Our results suggest that some rare and endangered species are able to maintain high levels of genetic diversity even at small population sizes. These results will assist with the design of conservation and management programmes, such as in situ and ex situ conservation, seed collection for germplasm conservation and reintroduction. Published by Oxford University Press on behalf of the Annals of Botany Company.

Spatial genetic structure of the sea sandwort (Honckenya peploides) on Surtsey: an immigrant's journey

NASA Astrophysics Data System (ADS)

Árnason, S. H.; Thórsson, Ae. Th.; Magnússon, B.; Philipp, M.; Adsersen, H.; Anamthawat-Jónsson, K.

2014-11-01

Sea sandwort (Honckenya peploides) was one of the first plants to successfully colonize and reproduce on the volcanic island Surtsey, formed in 1963 off the southern coast of Iceland. Using amplified fragment length polymorphic (AFLP) markers, we examined levels of genetic variation and differentiation among populations of H. peploides on Surtsey in relation to populations on the nearby island Heimaey and from the southern coast of Iceland. Selected populations from Denmark and Greenland were used for comparison. In addition, we tested whether the effects of isolation by distance could be seen in the Surtsey populations. Using two primer combinations, we obtained 173 AFLP markers from a total of 347 plant samples. The resulting binary matrix was then analysed statistically. The main results include the following: (i) Surtsey had the highest proportion of polymorphic markers as well as a comparatively high genetic diversity (55.5% proportion of polymorphic loci, PLP; 0.1974 HE) and Denmark the lowest (31.8% PLP; 0.132 HE), indicating rapid expansion during an early stage of population establishment on Surtsey and/or multiple origins of immigrants; (ii) the total genetic differentiation (FST) among Surtsey (0.0714) and Heimaey (0.055) populations was less than half of that found among the mainland populations in Iceland (0.1747), indicating substantial gene flow on the islands; (iii) most of the genetic variation (79%, p < 0.001) was found within localities, possibly due to the outcrossing and subdioecious nature of the species; (iv) a significant genetic distance was found within Surtsey, among sites, and this appeared to correlate with the age of plant colonization; and (v) the genetic structure analysis indicated multiple colonization episodes on Surtsey, whereby H. peploides most likely immigrated from the nearby island of Heimaey and directly from the southern coast of Iceland.

Revisiting AFLP fingerprinting for an unbiased assessment of genetic structure and differentiation of taurine and zebu cattle

PubMed Central

2014-01-01

Background Descendants from the extinct aurochs (Bos primigenius), taurine (Bos taurus) and zebu cattle (Bos indicus) were domesticated 10,000 years ago in Southwestern and Southern Asia, respectively, and colonized the world undergoing complex events of admixture and selection. Molecular data, in particular genome-wide single nucleotide polymorphism (SNP) markers, can complement historic and archaeological records to elucidate these past events. However, SNP ascertainment in cattle has been optimized for taurine breeds, imposing limitations to the study of diversity in zebu cattle. As amplified fragment length polymorphism (AFLP) markers are discovered and genotyped as the samples are assayed, this type of marker is free of ascertainment bias. In order to obtain unbiased assessments of genetic differentiation and structure in taurine and zebu cattle, we analyzed a dataset of 135 AFLP markers in 1,593 samples from 13 zebu and 58 taurine breeds, representing nine continental areas. Results We found a geographical pattern of expected heterozygosity in European taurine breeds decreasing with the distance from the domestication centre, arguing against a large-scale introgression from European or African aurochs. Zebu cattle were found to be at least as diverse as taurine cattle. Western African zebu cattle were found to have diverged more from Indian zebu than South American zebu. Model-based clustering and ancestry informative markers analyses suggested that this is due to taurine introgression. Although a large part of South American zebu cattle also descend from taurine cows, we did not detect significant levels of taurine ancestry in these breeds, probably because of systematic backcrossing with zebu bulls. Furthermore, limited zebu introgression was found in Podolian taurine breeds in Italy. Conclusions The assessment of cattle diversity reported here contributes an unbiased global view to genetic differentiation and structure of taurine and zebu cattle populations, which is essential for an effective conservation of the bovine genetic resources. PMID:24739206

Continental-scale footprint of balancing and positive selection in a small rodent (Microtus arvalis).

PubMed

Fischer, Martin C; Foll, Matthieu; Heckel, Gerald; Excoffier, Laurent

2014-01-01

Genetic adaptation to different environmental conditions is expected to lead to large differences between populations at selected loci, thus providing a signature of positive selection. Whereas balancing selection can maintain polymorphisms over long evolutionary periods and even geographic scale, thus leads to low levels of divergence between populations at selected loci. However, little is known about the relative importance of these two selective forces in shaping genomic diversity, partly due to difficulties in recognizing balancing selection in species showing low levels of differentiation. Here we address this problem by studying genomic diversity in the European common vole (Microtus arvalis) presenting high levels of differentiation between populations (average F ST = 0.31). We studied 3,839 Amplified Fragment Length Polymorphism (AFLP) markers genotyped in 444 individuals from 21 populations distributed across the European continent and hence over different environmental conditions. Our statistical approach to detect markers under selection is based on a Bayesian method specifically developed for AFLP markers, which treats AFLPs as a nearly codominant marker system, and therefore has increased power to detect selection. The high number of screened populations allowed us to detect the signature of balancing selection across a large geographic area. We detected 33 markers potentially under balancing selection, hence strong evidence of stabilizing selection in 21 populations across Europe. However, our analyses identified four-times more markers (138) being under positive selection, and geographical patterns suggest that some of these markers are probably associated with alpine regions, which seem to have environmental conditions that favour adaptation. We conclude that despite favourable conditions in this study for the detection of balancing selection, this evolutionary force seems to play a relatively minor role in shaping the genomic diversity of the common vole, which is more influenced by positive selection and neutral processes like drift and demographic history.

AFLP-based genetic diversity and its comparison with diversity based on SSR, SAMPL, and phenotypic traits in bread wheat.

PubMed

Roy, J K; Lakshmikumaran, M S; Balyan, H S; Gupta, P K

2004-02-01

Data on AFLP (eight primer pairs) and 14 phenotypic traits, collected on 55 elite and exotic bread wheat genotypes, were utilized for estimations of genetic diversity. We earlier used these 55 genotypes for a similar study using SSRs and SAMPL. As many as 615 scorable AFLP bands visualized included 287 (46.6%) polymorphic bands. The phenotypic traits included yield and its component traits, as well as physiomorphological traits like flag leaf area. Dendrograms were prepared using cluster analysis based on Jaccard's similarity coefficients in case of AFLP and on squared Euclidean distances in case of phenotypic traits. PCA was conducted using AFLP data and a PCA plot was prepared, which was compared with clustering patterns in two dendrograms, one each for AFLP and phenotypic traits. The results were also compared with published results that included studies conducted elsewhere using entirely different wheat germplasm and our own SSR and SAMPL studies based on the same 55 genotypes used in the present study. It was shown that molecular markers are superior to phenotypic traits and that AFLP and SAMPL are superior to other molecular markers for estimation of genetic diversity. On the basis of AFLP analysis and keeping in view the yield performance and stability, a pair of genotypes (E3876 and E677) was recommended for hybridization in order to develop superior cultivars.

Population genetic structure of rare and endangered plants using molecular markers

USGS Publications Warehouse

Raji, Jennifer; Atkinson, Carter T.

2013-01-01

This study was initiated to assess the levels of genetic diversity and differentiation in the remaining populations of Phyllostegia stachyoides and Melicope zahlbruckneri in Hawai`i Volcanoes National Park and determine the extent of gene flow to identify genetically distinct individuals or groups for conservation purposes. Thirty-six Amplified Fragment Length Polymorphic (AFLP) primer combinations generated a total of 3,242 polymorphic deoxyribonucleic acid (DNA) fragments in the P. stachyoides population with a percentage of polymorphic bands (PPB) ranging from 39.3 to 65.7% and 2,780 for the M. zahlbruckneri population with a PPB of 18.8 to 64.6%. Population differentiation (Fst) of AFLP loci between subpopulations of P. stachyoides was low (0.043) across populations. Analysis of molecular variance of P. stachyoides showed that 4% of the observed genetic differentiation occurred between populations in different kīpuka and 96% when individuals were pooled from all kīpuka. Moderate genetic diversity was detected within the M. zahlbruckneri population. Bayesian and multivariate analyses both classified the P. stachyoides and M. zahlbruckneri populations into genetic groups with considerable sub-structuring detected in the P. stachyoides population. The proportion of genetic differentiation among populations explained by geographical distance was estimated by Mantel tests. No spatial correlation was found between genetic and geographic distances in both populations. Finally, a moderate but significant gene flow that could be attributed to insect or bird-mediated dispersal of pollen across the different kīpuka was observed. The results of this study highlight the utility of a multi-allelic DNA-based marker in screening a large number of polymorphic loci in small and closely related endangered populations and revealed the presence of genetically unique groups of individuals in both M. zahlbruckneri and P. stachyoides populations. Based on these findings, approaches that can assist conservation efforts of these species are proposed. 

Pollinator-mediated gene flow fosters genetic variability in a narrow alpine endemic, Abronia alpina (Nyctaginaceae).

PubMed

Jabis, Meredith D; Ayers, Tina J; Allan, Gerard J

2011-10-01

For rare and endemic plants that exist in small, isolated habitats, natural selection is expected to favor self-compatibility, which can result in low genetic diversity due to inbreeding and genetic drift. Using Abronia alpina, a rare alpine endemic of the California Floristic Province, we demonstrate that there are exceptions to these predictions. We present the results of both a pollination experiment and a genetic study using AFLPs (amplified fragment length polymorphisms). Using controlled hand-pollination and pollinator observations, we examined the breeding system, pollination ecology, and mechanism for self-incompatibility in A. alpina. Abronia alpina exhibits an allogamous mating system with probable self-incompatibility resulting from limited growth of pollen tubes originating from self-pollination. Only xenogamous crosses and open-pollinated controls produced seed, and only xenogamous crosses produced pollen tubes that reached the ovary. The molecular study shows that A. alpina has substantial genetic diversity for a rare, endemic species, evidenced by the high percentage of polymorphic loci and average expected heterozygosity. Gene flow among subpopulations, as inferred from AFLP markers, appears to be substantial, although the Kern River is an important physical barrier. Our results indicate that A. alpina is dependent on insects for both seed production and the maintenance of genetic diversity. This finding suggests that pollinators may be crucial to the long-term adaptive potential of rare, endemic plants and that conservation of rare endemics is, in part, dependent on community-level interactions such as plant-pollinator mutualisms.

Genetic and Epigenetic Variations Induced by Wheat-Rye 2R and 5R Monosomic Addition Lines

PubMed Central

Fu, Shulan; Sun, Chuanfei; Yang, Manyu; Fei, Yunyan; Tan, Feiqun; Yan, Benju; Ren, Zhenglong; Tang, Zongxiang

2013-01-01

Background Monosomic alien addition lines (MAALs) can easily induce structural variation of chromosomes and have been used in crop breeding; however, it is unclear whether MAALs will induce drastic genetic and epigenetic alterations. Methodology/Principal Findings In the present study, wheat-rye 2R and 5R MAALs together with their selfed progeny and parental common wheat were investigated through amplified fragment length polymorphism (AFLP) and methylation-sensitive amplification polymorphism (MSAP) analyses. The MAALs in different generations displayed different genetic variations. Some progeny that only contained 42 wheat chromosomes showed great genetic/epigenetic alterations. Cryptic rye chromatin has introgressed into the wheat genome. However, one of the progeny that contained cryptic rye chromatin did not display outstanding genetic/epigenetic variation. 78 and 49 sequences were cloned from changed AFLP and MSAP bands, respectively. Blastn search indicated that almost half of them showed no significant similarity to known sequences. Retrotransposons were mainly involved in genetic and epigenetic variations. Genetic variations basically affected Gypsy-like retrotransposons, whereas epigenetic alterations affected Copia-like and Gypsy-like retrotransposons equally. Genetic and epigenetic variations seldom affected low-copy coding DNA sequences. Conclusions/Significance The results in the present study provided direct evidence to illustrate that monosomic wheat-rye addition lines could induce different and drastic genetic/epigenetic variations and these variations might not be caused by introgression of rye chromatins into wheat. Therefore, MAALs may be directly used as an effective means to broaden the genetic diversity of common wheat. PMID:23342073

Genetic and epigenetic variations induced by wheat-rye 2R and 5R monosomic addition lines.

PubMed

Fu, Shulan; Sun, Chuanfei; Yang, Manyu; Fei, Yunyan; Tan, Feiqun; Yan, Benju; Ren, Zhenglong; Tang, Zongxiang

2013-01-01

Monosomic alien addition lines (MAALs) can easily induce structural variation of chromosomes and have been used in crop breeding; however, it is unclear whether MAALs will induce drastic genetic and epigenetic alterations. In the present study, wheat-rye 2R and 5R MAALs together with their selfed progeny and parental common wheat were investigated through amplified fragment length polymorphism (AFLP) and methylation-sensitive amplification polymorphism (MSAP) analyses. The MAALs in different generations displayed different genetic variations. Some progeny that only contained 42 wheat chromosomes showed great genetic/epigenetic alterations. Cryptic rye chromatin has introgressed into the wheat genome. However, one of the progeny that contained cryptic rye chromatin did not display outstanding genetic/epigenetic variation. 78 and 49 sequences were cloned from changed AFLP and MSAP bands, respectively. Blastn search indicated that almost half of them showed no significant similarity to known sequences. Retrotransposons were mainly involved in genetic and epigenetic variations. Genetic variations basically affected Gypsy-like retrotransposons, whereas epigenetic alterations affected Copia-like and Gypsy-like retrotransposons equally. Genetic and epigenetic variations seldom affected low-copy coding DNA sequences. The results in the present study provided direct evidence to illustrate that monosomic wheat-rye addition lines could induce different and drastic genetic/epigenetic variations and these variations might not be caused by introgression of rye chromatins into wheat. Therefore, MAALs may be directly used as an effective means to broaden the genetic diversity of common wheat.

Genetic diversity of populations and clones of Rhopilema esculentum in China based on AFLP analysis

NASA Astrophysics Data System (ADS)

Qiao, Hongjin; Liu, Xiangquan; Zhang, Xijia; Jiang, Haibin; Wang, Jiying; Zhang, Limin

2013-03-01

Amplified fragment length polymorphisms (AFLP) markers were developed to assess the genetic variation of populations and clones of Rhopilema esculentum Kishinouye (Scyphozoa, Rhizostomatidae). One hundred and seventy-nine loci from 56 individuals of two hatchery populations and two wild populations were genotyped with five primer combinations. The polymorphic ratio, Shannon's diversity index and average heterozygosity were 70.3%, 0.346 and 0.228 for the white hatchery population, 74.3%, 0.313, and 0.201 for the red hatchery population, 79.3%, 0.349, and 0.224 for the Jiangsu wild population, and 74.9%, 0.328 and 0.210 for the Penglai wild population, respectively. Thus, all populations had a relatively high level of genetic diversity. A specific band was identified that could separate the white from the red hatchery population. There was 84.85% genetic differentiation within populations. Individual cluster analysis using unweighted pair-group method with arithmetic mean (UPGMA) suggested that hatchery populations and wild populations could be divided. For the hatchery populations, the white and red populations clustered separately; however, for the wild populations, Penglai and Jiangsu populations clustered together. The genetic diversity at the clone level was also determined. Our data suggest that there are relatively high genetic diversities within populations but low genetic differentiation between populations, which may be related to the long-term use of germplasm resources from Jiangsu Province for artificial seeding and releasing. These findings will benefit the artificial seeding and conservation of the germplasm resources.

Genetic diversity and structure of Brazilian ginger germplasm (Zingiber officinale) revealed by AFLP markers.

PubMed

Blanco, Eleonora Zambrano; Bajay, Miklos Maximiliano; Siqueira, Marcos Vinícius Bohrer Monteiro; Zucchi, Maria Imaculada; Pinheiro, José Baldin

2016-12-01

Ginger is a vegetable with medicinal and culinary properties widely cultivated in the Southern and Southeastern Brazil. The knowledge of ginger species' genetic variability is essential to direct correctly future studies of conservation and genetic improvement, but in Brazil, little is known about this species' genetic variability. In this study, we analyzed the genetic diversity and structure of 55 Brazilian accessions and 6 Colombian accessions of ginger, using AFLP (Amplified Fragment Length Polymorphism) molecular markers. The molecular characterization was based on 13 primers combinations, which generated an average of 113.5 polymorphic loci. The genetic diversity estimates of Nei (Hj), Shannon-Weiner index (I) and an effective number of alleles (n e ) were greater in the Colombian accessions in relation to the Brazilian accessions. The analysis of molecular variance showed that most of the genetic variation occurred between the two countries while in the Brazilian populations there is no genetic structure and probably each region harbors 100 % of genetic variation found in the samples. The bayesian model-based clustering and the dendrogram using the dissimilarity's coefficient of Jaccard were congruent with each other and showed that the Brazilian accessions are highly similar between themselves, regardless of the geographic region of origin. We suggested that the exploration of the interspecific variability and the introduction of new varieties of Z.officinale are viable alternatives for generating diversity in breeding programs in Brazil. The introduction of new genetic materials will certainly contribute to a higher genetic basis of such crop.

Geographical parthenogenesis and population genetic structure in the alpine species Ranunculus kuepferi (Ranunculaceae).

PubMed

Cosendai, A-C; Wagner, J; Ladinig, U; Rosche, C; Hörandl, E

2013-06-01

Geographical parthenogenesis describes the enigmatic phenomenon that asexual organisms have larger distribution areas than their sexual relatives, especially in previously glaciated areas. Classical models suggest temporary advantages to asexuality in colonization scenarios because of uniparental reproduction and clonality. We analyzed population genetic structure and self-fertility of the plant species Ranunculus kuepferi on 59 populations from the whole distribution area (European Alps, Apennines and Corsica). Amplified fragment length polymorphisms (AFLPs) and five microsatellite loci revealed individual genotypes for all populations and mostly insignificant differences between diploid sexuals and tetraploid apomicts in all measures of genetic diversity. Low frequencies of private AFLP fragments/simple sequence repeat alleles, and character incompatibility analyses suggest that facultative recombination explains best the unexpectedly high genotypic diversity of apomicts. STRUCTURE analyses using AFLPs revealed a higher number of partitions and a stronger geographical subdivision for diploids than for tetraploids, which contradicts expectations of standard gene flow models, but indicates a reduction of genetic structure in asexuals. Apomictic populations exhibited high admixture near the sexual area, but appeared rather uniform in remote areas. Bagging experiments and analyses of pollen tube growth confirmed self-fertility for pollen-dependent apomicts, but self-sterility for diploid sexuals. Facultative apomixis combines advantages of both modes of reproduction: uniparental reproduction allows for rapid colonization of remote areas, whereas facultative sexuality and polyploidy maintains genetic diversity within apomictic populations. The density dependence of outcrossing limits range expansions of sexual populations.

Geographical parthenogenesis and population genetic structure in the alpine species Ranunculus kuepferi (Ranunculaceae)

PubMed Central

Cosendai, A-C; Wagner, J; Ladinig, U; Rosche, C; Hörandl, E

2013-01-01

Geographical parthenogenesis describes the enigmatic phenomenon that asexual organisms have larger distribution areas than their sexual relatives, especially in previously glaciated areas. Classical models suggest temporary advantages to asexuality in colonization scenarios because of uniparental reproduction and clonality. We analyzed population genetic structure and self-fertility of the plant species Ranunculus kuepferi on 59 populations from the whole distribution area (European Alps, Apennines and Corsica). Amplified fragment length polymorphisms (AFLPs) and five microsatellite loci revealed individual genotypes for all populations and mostly insignificant differences between diploid sexuals and tetraploid apomicts in all measures of genetic diversity. Low frequencies of private AFLP fragments/simple sequence repeat alleles, and character incompatibility analyses suggest that facultative recombination explains best the unexpectedly high genotypic diversity of apomicts. STRUCTURE analyses using AFLPs revealed a higher number of partitions and a stronger geographical subdivision for diploids than for tetraploids, which contradicts expectations of standard gene flow models, but indicates a reduction of genetic structure in asexuals. Apomictic populations exhibited high admixture near the sexual area, but appeared rather uniform in remote areas. Bagging experiments and analyses of pollen tube growth confirmed self-fertility for pollen-dependent apomicts, but self-sterility for diploid sexuals. Facultative apomixis combines advantages of both modes of reproduction: uniparental reproduction allows for rapid colonization of remote areas, whereas facultative sexuality and polyploidy maintains genetic diversity within apomictic populations. The density dependence of outcrossing limits range expansions of sexual populations. PMID:23403961

Propagule pressure, genetic structure, and geographic origins of Chondrilla juncea (Asteraceae): an apomictic invader on three continents.

PubMed

Gaskin, John F; Schwarzländer, Mark; Kinter, C Lynn; Smith, James F; Novak, Stephen J

2013-09-01

Assessing propagule pressure and geographic origins of invasive species provides insight into the invasion process. Rush skeletonweed (Chondrilla juncea; Asteraceae) is an apomictic, perennial plant that is invasive in Australia, South America (Argentina), and North America (Canada and the United States). This study comprehensively compares propagule pressure and geographic structure of genotypes to improve our understanding of a clonal invasion and enhance management strategies. • We analyzed 1056 native range plants from Eurasia and 1156 plants from three invaded continents using amplified fragment length polymorphism (AFLP) techniques. We used measures of diversity (Simpson's D) and evenness (E), analysis of molecular variance, and Mantel tests to compare invasions, and genotype similarity to determine origins of invasive genotypes. • We found 682 unique genotypes in the native range, but only 13 in the invaded regions. Each invaded region contained distinct AFLP genotypes, suggesting independent introduction events, probably with different geographic origins. Relatively low propagule pressure was associated with each introduction around the globe, but levels of among-population variation differed. We found exact AFLP genotype matches between the native and invaded ranges for five of the 13 invasive genotypes. • Invasion dynamics can vary across invaded ranges within a species. Intensive sampling for molecular analyses can provide insight for understanding intraspecific invasion dynamics, which can hold significance for the management of plant species, especially by finding origins and distributions of invasive genotypes for classical biological control efforts.

Molecular epidemiology of infant botulism in California and elsewhere, 1976-2010.

PubMed

Dabritz, Haydee A; Hill, Karen K; Barash, Jason R; Ticknor, Lawrence O; Helma, Charles H; Dover, Nir; Payne, Jessica R; Arnon, Stephen S

2014-12-01

Infant botulism (IB), first identified in California in 1976, results from Clostridium botulinum spores that germinate, multiply, and produce botulinum neurotoxin (BoNT) in the immature intestine. From 1976 to 2010 we created an archive of 1090 BoNT-producing isolates consisting of 1012 IB patient (10 outpatient, 985 hospitalized, 17 sudden death), 25 food, 18 dust/soils, and 35 other strains. The mouse neutralization assay determined isolate toxin type (56% BoNT/A, 32% BoNT/B). Amplified fragment-length polymorphism (AFLP) analysis of the isolates was combined with epidemiologic information. The AFLP dendrogram, the largest to date, contained 154 clades; 52% of isolates clustered in just 2 clades, 1 BoNT/A (n=418) and 1 BoNT/B (n=145). These clades constituted an endemic C. botulinum population that produced the entire clinical spectrum of IB. Isolates from the patient's home environment (dust/soil, honey) usually located to the same AFLP clade as the patient's isolate, thereby identifying the likely source of infective spores. C. botulinum A(B) strains were identified in California for the first time. Combining molecular methods and epidemiological data created an effective tool that yielded novel insights into the genetic diversity of C. botulinum and the clinical spectrum, occurrence, and distribution of IB in California. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Molecular typing of Sarcocystis neurona: current status and future trends.

PubMed

Elsheikha, Hany M; Mansfield, Linda S

2007-10-21

Sarcocystis neurona is an important protozoal pathogen because it causes the serious neurological disease equine protozoal myeloencephalitis (EPM). The capacity of this organism to cause a wide spectrum of neurological signs in horses and the broad geographic distribution of observed cases in the Americas drive the need for sensitive, reliable and rapid typing methods to characterize strains. Various molecular methods have been developed and used to diagnose EPM due to S. neurona, to identify S. neurona isolates and to determine the heterogeneity and evolutionary relatedness within this species and related Sarcocystis spp. These methods included sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), immuno-fluorescent assay (IFA), slide agglutination test (SAT), SnSAG-specific ELISA, random amplified polymorphic DNA (RAPD), PCR-based restriction fragment length polymorphism (RFLP), amplified fragment length polymorphism (AFLP) fingerprinting, and sequence analysis of surface protein genes, ribosomal genes, microsatellite alleles and other molecular markers. Here, the utility of these molecular methods is reviewed and evaluated with respect to the need for molecular approaches that utilize well-characterized polymorphic, simple, independent, and stable genetic markers. These tools have the potential to add to knowledge of the genetic population structure of S. neurona and to provide new insights into the pathogenesis of EPM and S. neurona epidemiology. In particular, these methods provide new tools to address the hypothesis that particular genetic variants are associated with adverse clinical outcomes (severe pathotypes). The ultimate goal is to utilize them in future studies to improve treatment and prevention strategies.

Transcriptome and proteome analysis of ovaries of arrhenotokous and thelytokous Venturia canescens.

PubMed

Mateo Leach, I; Hesseling, A; Huibers, W H C; Witsenboer, H; Beukeboom, L W; van de Zande, L

2009-08-01

Under arrhenotoky, unfertilized haploid eggs develop as males but under thelytoky they develop into diploid females after they have undergone diploidy restoration. In the parasitoid wasp Venturia canescens both reproductive modes occur. Thelytoky is genetically determined but the underlying genetics of diploidy restoration remain unknown. In this study we aim to identify the genes and/or proteins that control thelytoky. cDNA-amplified fragment length polymorphism (cDNA-AFLP) analysis of total ovarian RNA and two-dimensional protein electrophoresis in combination with mass spectrometry revealed putative transcripts and proteins involved in arrhenotokous and thelytokous development. The detected tubulin and actin protein differences are most likely functionally related to the two types of reproduction.

Campylobacter subantarcticus sp. nov., isolated from birds in the sub-Antarctic region.

PubMed

Debruyne, Lies; Broman, Tina; Bergström, Sven; Olsen, Björn; On, Stephen L W; Vandamme, Peter

2010-04-01

Six Gram-stain-negative, spiral-shaped, microaerobic isolates were obtained during a sampling from wild birds in the sub-Antarctic region. Based on initial observations, these isolates were classified as Campylobacter lari-like. The isolates were further characterized by whole-cell protein and amplified fragment length polymorphism (AFLP) analysis, which revealed that they were distinct from C. lari and all other known species of the genus Campylobacter. Here, we present comprehensive phylogenetic, genomic and phenotypic evidence that these isolates represent a novel species within the genus Campylobacter, for which the name Campylobacter subantarcticus sp. nov. is proposed. The type strain is R-3023(T) (=LMG 24377(T) =CCUG 38513(T)).

Improving AFLP analysis of large-scale patterns of genetic variation--a case study with the Central African lianas Haumania spp (Marantaceae) showing interspecific gene flow.

PubMed

Ley, A C; Hardy, O J

2013-04-01

AFLP markers are often used to study patterns of population genetic variation and gene flow because they offer a good coverage of the nuclear genome, but the reliability of AFLP scoring is critical. To assess interspecific gene flow in two African rainforest liana species (Haumania danckelmaniana, H. liebrechtsiana) where previous evidence of chloroplast captures questioned the importance of hybridization and species boundaries, we developed new AFLP markers and a novel approach to select reliable bands from their degree of reproducibility. The latter is based on the estimation of the broad-sense heritability of AFLP phenotypes, an improvement over classical scoring error rates, which showed that the polymorphism of most AFLP bands was affected by a substantial nongenetic component. Therefore, using a quantitative genetics framework, we also modified an existing estimator of pairwise kinship coefficient between individuals correcting for the limited heritability of markers. Bayesian clustering confirms the recognition of the two Haumania species. Nevertheless, the decay of the relatedness between individuals of distinct species with geographic distance demonstrates that hybridization affects the nuclear genome. In conclusion, although we showed that AFLP markers might be substantially affected by nongenetic factors, their analysis using the new methods developed considerably advanced our understanding of the pattern of gene flow in our model species. © 2013 Blackwell Publishing Ltd.

Genetic variation and genetic structure of the endangered species Sinowilsonia henryi Hemsi. (Hamamelidaceae) revealed by amplified fragment length polymorphism (AFLP) markers.

PubMed

Zhang, H; Ji, W L; Li, M; Zhou, L Y

2015-10-14

Comprehensive research of genetic variation is crucial in designing conservation strategies for endangered and threatened species. Sinowilsonia henryi Hemsi. is a tertiary relic with a limited geographical distribution in the central and western areas of China. It is endangered because of climate change and habitat fragmentation over the last thousands of years. In this study, amplified fragment length polymorphism markers were utilized to estimate genetic diversity and genetic structure in and among S. henryi. In this study, Nei's genetic diversity and Shannon's information index were found to be 0.192 and 0.325 respectively, indicating a moderate-to-high genetic diversity in species. According to analysis of molecular variation results, 32% of the genetic variation was shown to be partitioned among populations, demonstrating a relatively high genetic divergence; this was supported by principal coordinate analysis and unweighted pair-group method with arithmetic average analysis. Moreover, the Mantel test showed that there was no significant correlation between genetic and geographical distances. The above results can be explained by the effects of habitat fragmentation, history traits, and gene drift. Based on the results, several implications were indicated and suggestions proposed for preservation strategies for this species.

Extensive genetic and DNA methylation variation contribute to heterosis in triploid loquat hybrids.

PubMed

Liu, Chao; Wang, Mingbo; Wang, Lingli; Guo, Qigao; Liang, Guolu

2018-04-24

We aim to overcome the unclear origin of the loquat and elucidate the heterosis mechanism of the triploid loquat. Here we investigated the genetic and epigenetic variations between the triploid plant and its parental lines using amplified fragment length polymorphism (AFLP) and methylation-sensitive amplified fragment length polymorphism (MSAP) analyses. We show that in addition to genetic variations, extensive DNA methylation variation occurred during the formation process of triploid loquat, with the triploid hybrid having increased DNA methylation compared to the parents. Furthermore, a correlation existed between genetic variation and DNA methylation remodeling, suggesting that genome instability may lead to DNA methylation variation or vice versa. Sequence analysis of the MSAP bands revealed that over 53% of them overlap with protein-coding genes, which may indicate a functional role of the differential DNA methylation in gene regulation and hence heterosis phenotypes. Consistent with this, the genetic and epigenetic alterations were associated closely to the heterosis phenotypes of triploid loquat, and this association varied for different traits. Our results suggested that the formation of triploid is accompanied by extensive genetic and DNA methylation variation, and these changes contribute to the heterosis phenotypes of the triploid loquats from the two cross lines.

Molecular Diagnostics of Arthroconidial Yeasts, Frequent Pulmonary Opportunists.

PubMed

Kaplan, Engin; Al-Hatmi, Abdullah M S; Ilkit, Macit; Gerrits van den Ende, A H G; Hagen, Ferry; Meis, Jacques F; de Hoog, G Sybren

2018-01-01

Magnusiomyces capitatus and Saprochaete clavata are members of the clade of arthroconidial yeasts that represent emerging opportunistic pulmonary pathogens in immunocompromised patients. Given that standard ribosomal DNA (rDNA) identification often provides confusing results, in this study, we analyzed 34 isolates with the goal of finding new genetic markers for classification using multilocus sequencing and amplified fragment length polymorphism (AFLP). The interspecific similarity obtained using rDNA markers (the internal transcribed spacer [ITS] and large subunit regions) was in the range of 96 to 99%, whereas that obtained using protein-coding loci ( Rbp2 , Act , and Tef1α ) was lower at 89.4 to 95.2%. Ultimately, Rbp2 was selected as the best marker for species distinction. On the basis of cloned ITS data, some strains proved to be misidentified in comparison with the identities obtained with phenotypic characters, protein sequences, and AFLP profiles, indicating that different copies of the ribosomal operon were present in a single species. Antifungal susceptibility testing revealed that voriconazole had the lowest MIC against M. capitatus , while amphotericin B had the lowest MIC against S. clavata Both species exhibited in vitro resistance to fluconazole and micafungin. Copyright © 2017 American Society for Microbiology.

Genetic diversity reflects geographical origin of Ralstonia solanacearum strains isolated from plant and water sources in Spain.

PubMed

Caruso, Paola; Biosca, Elena G; Bertolini, Edson; Marco-Noales, Ester; Gorris, María Teresa; Licciardello, Concetta; López, María M

2017-12-01

The characterization and intraspecific diversity of a collection of 45 Ralstonia solanacearum strains isolated in Spain from different sources and geographical origins is reported. To test the influence of the site and the host on strain diversity, phenotypic and genotypic analysis were performed by a polyphasic approach. Biochemical and metabolic profiles were compared. Serological relationship was evaluated by Indirect-ELISA using polyclonal and monoclonal antibodies. For genotypic analysis, hrpB and egl DNA sequence analysis, repetitive sequences (rep-PCR), amplified fragment length polymorphism (AFLP) profiles and macrorestriction with XbaI followed by pulsed field gel electrophoresis (PFGE) were performed. The biochemical and metabolic characterization, serological tests, rep-PCR typing and phylogenetic analysis showed that all analysed strains belonged to phylotype II sequevar 1 and shared homogeneous profiles. However, interesting differences among strains were found by AFLP and macrorestriction with XbaI followed by PFGE techniques, some profiles being related to the geographical origin of the strains. Diversity results obtained offer new insights into the biogeography of this quarantine organism and its possible sources and reservoirs in Spain and Mediterranean countries. Copyright© by the Spanish Society for Microbiology and Institute for Catalan Studies.

Differential gene expression for Curvularia eragrostidis pathogenic incidence in crabgrass (Digitaria sanguinalis) revealed by cDNA-AFLP analysis.

PubMed

Wang, Jianshu; Wang, Xuemin; Yuan, Bohua; Qiang, Sheng

2013-01-01

Gene expression profiles of Digitaria sanguinalis infected by Curvularia eragrostidis strain QZ-2000 at two concentrations of conidia and two dew durations were analyzed by cDNA amplified fragment length polymorphisms (cDNA-AFLP). Inoculum strength was more determinant of gene expression than dew duration. A total of 256 primer combinations were used for selective amplification and 1214 transcript-derived fragments (TDFs) were selected for their differential expression. Of these, 518 up-regulated differentially expressed TDFs were identified. Forty-six differential cDNA fragments were chosen to be cloned and 35 of them were successfully cloned and sequenced, of which 25 were homologous to genes of known function according to the GenBank database. Only 6 genes were up-regulated in Curvularia eragrostidis-inoculated D. sanguinalis, with functions involved in signal transduction, energy metabolism, cell growth and development, stress responses, abscisic acid biosynthesis and response. It appears that a few pathways may be important parts of the pathogenic strategy of C. eragrostidis strain QZ-2000 on D. sanguinalis. Our study provides the fundamentals to further study the pathogenic mechanism, screen for optimal C. eragrostidis strains as potential mycoherbicide and apply this product to control D. sanguinalis.

Improved coverage of cDNA-AFLP by sequential digestion of immobilized cDNA.

PubMed

Weiberg, Arne; Pöhler, Dirk; Morgenstern, Burkhard; Karlovsky, Petr

2008-10-13

cDNA-AFLP is a transcriptomics technique which does not require prior sequence information and can therefore be used as a gene discovery tool. The method is based on selective amplification of cDNA fragments generated by restriction endonucleases, electrophoretic separation of the products and comparison of the band patterns between treated samples and controls. Unequal distribution of restriction sites used to generate cDNA fragments negatively affects the performance of cDNA-AFLP. Some transcripts are represented by more than one fragment while other escape detection, causing redundancy and reducing the coverage of the analysis, respectively. With the goal of improving the coverage of cDNA-AFLP without increasing its redundancy, we designed a modified cDNA-AFLP protocol. Immobilized cDNA is sequentially digested with several restriction endonucleases and the released DNA fragments are collected in mutually exclusive pools. To investigate the performance of the protocol, software tool MECS (Multiple Enzyme cDNA-AFLP Simulation) was written in Perl. cDNA-AFLP protocols described in the literature and the new sequential digestion protocol were simulated on sets of cDNA sequences from mouse, human and Arabidopsis thaliana. The redundancy and coverage, the total number of PCR reactions, and the average fragment length were calculated for each protocol and cDNA set. Simulation revealed that sequential digestion of immobilized cDNA followed by the partitioning of released fragments into mutually exclusive pools outperformed other cDNA-AFLP protocols in terms of coverage, redundancy, fragment length, and the total number of PCRs. Primers generating 30 to 70 amplicons per PCR provided the highest fraction of electrophoretically distinguishable fragments suitable for normalization. For A. thaliana, human and mice transcriptome, the use of two marking enzymes and three sequentially applied releasing enzymes for each of the marking enzymes is recommended.

The possible physical barrier and coastal dispersal strategy for Japanese grenadier anchovy, Coilia nasus in the East China Sea and Yellow Sea: evidence from AFLP markers.

PubMed

Han, Zhi-Qiang; Han, Gang; Wang, Zhi-Yong; Gao, Tian-Xiang

2015-02-03

In order to ascertain the taxonomic status of the Ariake Sea population of Japanese grenadier anchovy, Coilia nasus, and assess the contemporary possible genetic barrier between the west and east coastal waters of the East China Sea, we used amplified fragment length polymorphism (AFLP) markers to detect the genetic structure of C. nasus, in the East China Sea and Yellow Sea. Eighty-one individuals of C. nasus were collected from five locations and 12 individuals of Coilia mystus were sampled from the Yangtze River Estuary. A total of 371 loci were detected by five primer combinations, 310 of which were polymorphic (83.56%). Analysis of molecular variation (AMOVA) and pairwise fixation index (FST) revealed significant genetic differentiation among five samples, indicating limited gene flow among populations. The dendrogram for populations by neighbor-joining (NJ) cluster analysis provided evidence of a clear relationship between genetic and geographic patterns, supporting significant genetic differentiation between China coastal populations and Ariake Sea populations. Compared to the genetic divergence between C. nasus and C. mystus, the level of genetic differentiation between China and the Ariake Sea populations of C. nasus is obvious below the species level, indicating isolated populations of C. nasus in the Ariake Sea. Isolation by distance analysis revealed that direct ocean distance with deep-water at the continental slope and high salinity between west and east coastal waters of the East China Sea served as major physical barrier to C. nasus, supporting the coastal dispersal pattern in this estuarine species, and rejecting offshore dispersal strategy.

Phylogeography and seed dispersal in islands: the case of Rumex bucephalophorus subsp. canariensis (Polygonaceae).

PubMed

Talavera, María; Navarro-Sampedro, Laura; Ortiz, Pedro L; Arista, Montserrat

2013-02-01

Rumex bucephalophorus subsp. canariensis is an endemic taxon to Macaronesia with diaspore polymorphism. The origin and colonizing route of this taxon in Macaronesia was studied using molecular data and information on diaspore types. Amplified fragment length polymorphism (AFLP) was used in 260 plants from 22 populations of R. bucephalophorus subsp. canariensis, four from the Madeiran archipelago and 18 from the Canary archipelago. Diaspore production was analysed in 9-50 plants from each population used for AFLP analysis. One hundred and one plants from the Madeiran archipelago and 375 plants from the Canary Islands were studied. For each plant the type of diaspore produced was recorded. Overall populations had low genetic diversity but they showed a geographical pattern of genetic diversity that was higher in the older eastern islands than in the younger western ones. Two types of dispersible diaspores were found: in the eastern Canary islands (Lanzarote, Fuerteventura and Gran Canaria), plants produced exclusively long-dispersible diaspores, whereas in the western Canary islands (Tenerife, La Gomera, El Hierro) and the Madeiran archipelago plants produced exclusively short-dispersible diaspores. Genetically, the studied populations fell into four main island groups: Lanzarote-Fuerteventura, Gran Canaria, Tenerife-El Hierro and La Gomera-Madeira archipelago. A Moroccan origin of R. bucephalophorus subsp. canariensis is hypothesized with a colonization route from the eastern to the western islands. In addition, at least one gene flow event from La Gomera to the Madeiran archipelago has taken place. During the colonization process the type of dispersible diaspore changed so that dispersability decreased in populations of the westernmost islands.

Pleistocene refugia and polytopic replacement of diploids by tetraploids in the Patagonian and Subantarctic plant Hypochaeris incana (Asteraceae, Cichorieae).

PubMed

Tremetsberger, Karin; Urtubey, Estrella; Terrab, Anass; Baeza, Carlos M; Ortiz, María Angeles; Talavera, María; König, Christiane; Temsch, Eva M; Kohl, Gudrun; Talavera, Salvador; Stuessy, Tod F

2009-09-01

We report the phylogeographic pattern of the Patagonian and Subantarctic plant Hypochaeris incana endemic to southeastern South America. We applied amplified fragment length polymorphism (AFLP) and chloroplast DNA (cpDNA) analysis to 28 and 32 populations, respectively, throughout its distributional range and assessed ploidy levels using flow cytometry. While cpDNA data suggest repeated or simultaneous parallel colonization of Patagonia and Tierra del Fuego by several haplotypes and/or hybridization, AFLPs reveal three clusters corresponding to geographic regions. The central and northern Patagonian clusters (approximately 38-51 degrees S), which are closer to the outgroup, contain mainly tetraploid, isolated and highly differentiated populations with low genetic diversity. To the contrary, the southern Patagonian and Fuegian cluster (approximately 51-55 degrees S) contains mainly diploid populations with high genetic diversity and connected by high levels of gene flow. The data suggest that H. incana originated at the diploid level in central or northern Patagonia, from where it migrated south. All three areas, northern, central and southern, have similar levels of rare and private AFLP bands, suggesting that all three served as refugia for H. incana during glacial times. In southern Patagonia and Tierra del Fuego, the species seems to have expanded its populational system in postglacial times, when the climate became warmer and more humid. In central and northern Patagonia, the populations seem to have become restricted to favourable sites with increasing temperature and decreasing moisture and there was a parallel replacement of diploids by tetraploids in local populations.

Inter-island movements and population differentiation in a pelagic seabird.

PubMed

Dearborn, Donald C; Anders, Angela D; Schreiber, E A; Adams, Rachelle M M; Mueller, Ulrich G

2003-10-01

We used mark-resight data and amplified fragment length polymorphism (AFLP) markers to assess movements and gene flow between Central Pacific breeding colonies of the great frigatebird, Fregata minor. Of 715 adult frigatebirds marked on Tern Island and Johnston Atoll, 21.3% were resighted at other frigatebird colonies at least 582 km away. Mark-resight data indicated regular movement of males and females between Tern Island and Johnston Atoll (873 km apart), and less frequent movements to other islands; no birds marked on Tern or Johnston were seen on Christmas Island, but one was seen in the Philippines, 7627 km from where it was marked. Despite the regular occurrence of interisland movements, Bayesian analyses of AFLP data showed significant genetic differentiation between Tern Island and Johnston Atoll, and more pronounced differentiation between these two islands and the more distant Christmas Island. The AFLP profiles of three birds breeding on Tern Island fell within the profile-cluster typical for Christmas Island birds, both in a nonmetric multidimensional scaling analysis and in a population assignment test, suggesting dispersal events from Christmas Island to Tern Island. Several factors could explain the persistence of genetic structure despite frequent movements between colonies: many movements occurred during the nonbreeding season, many breeding-season movements did not involve mate-acquisition behaviours and individuals that do disperse may be selected against, as suggested by morphometric differences between colonies. The persistence of genetic structure among breeding colonies despite significant interisland movements suggests limits to the effectiveness of migration as a homogenizing force in this broadly distributed, extremely mobile species.

Vicariance, long-distance dispersal, and regional extinction-recolonization dynamics explain the disjunct circumpolar distribution of the arctic-alpine plant Silene acaulis.

PubMed

Gussarova, Galina; Allen, Geraldine A; Mikhaylova, Yulia; McCormick, Laurie J; Mirré, Virginia; Marr, Kendrick L; Hebda, Richard J; Brochmann, Christian

2015-10-01

Many arctic-alpine species have vast geographic ranges, but these may encompass substantial gaps whose origins are poorly understood. Here we address the phylogeographic history of Silene acaulis, a perennial cushion plant with a circumpolar distribution except for a large gap in Siberia. We assessed genetic variation in a range-wide sample of 103 populations using plastid DNA (pDNA) sequences and AFLPs (amplified fragment length polymorphisms). We constructed a haplotype network and performed Bayesian phylogenetic analyses based on plastid sequences. We visualized AFLP patterns using principal coordinate analysis, identified genetic groups using the program structure, and estimated genetic diversity and rarity indices by geographic region. The history of the main pDNA lineages was estimated to span several glaciations. AFLP data revealed a distinct division between Beringia/North America and Europe/East Greenland. These two regions shared only one of 17 pDNA haplotypes. Populations on opposite sides of the Siberian range gap (Ural Mountains and Chukotka) were genetically distinct and appear to have resulted from postglacial leading-edge colonizations. We inferred two refugia in North America (Beringia and the southern Rocky Mountains) and two in Europe (central-southern Europe and northern Europe/East Greenland). Patterns in the East Atlantic region suggested transoceanic long-distance dispersal events. Silene acaulis has a highly dynamic history characterized by vicariance, regional extinction, and recolonization, with persistence in at least four refugia. Long-distance dispersal explains patterns across the Atlantic Ocean, but we found no evidence of dispersal across the Siberian range gap. © 2015 Botanical Society of America.

Genetic and epigenetic diversity and structure of Phragmites australis from local habitats of the Songnen Prairie using amplified fragment length polymorphism markers.

PubMed

Qiu, T; Jiang, L L; Yang, Y F

2016-08-19

The genetic and epigenetic diversity and structure of naturally occurring Phragmites australis populations occupying two different habitats on a small spatial scale in the Songnen Prairie in northeastern China were investigated by assessing amplified fragment length polymorphisms (AFLPs) and methylation-sensitive amplified polymorphisms (MSAPs) through fluorescent capillary detection. The two groups of P. australis were located in a seasonal waterlogged low-lying and alkalized meadow with a pH of 8-8.5 and in an alkaline patch without accumulated rainwater and with a pH greater than 10. These groups showed high levels of genetic diversity at the habitat level based on the percentage of polymorphic bands (90.32, 82.56%), Nei's gene diversity index (0.262, 0.248), and the Shannon diversity index (0.407, 0.383). Although little is known about the between-habitat genetic differentiation of P. australis on a small spatial scale, our results implied significant genetic differentiation between habitats. Extensive epigenetic diversity within habitats, along with clear differentiation, was found. Specifically, the former habitat (Habitat 1, designated H1) harbored higher levels of genetic and epigenetic diversity than the latter (Habitat 2, designated H2), and population-level diversity was also high. This study represents one of few attempts to predict habitat-based genetic differentiation of reeds on a small scale. These assessments of genetic and epigenetic variation are integral aspects of molecular ecological studies on P. australis. Possible causes for within- and between-habitat genetic and epigenetic variations are discussed.

Construction of the first genetic linkage map of Japanese gentian (Gentianaceae)

PubMed Central

2012-01-01

Background Japanese gentians (Gentiana triflora and Gentiana scabra) are amongst the most popular floricultural plants in Japan. However, genomic resources for Japanese gentians have not yet been developed, mainly because of the heterozygous genome structure conserved by outcrossing, the long juvenile period, and limited knowledge about the inheritance of important traits. In this study, we developed a genetic linkage map to improve breeding programs of Japanese gentians. Results Enriched simple sequence repeat (SSR) libraries from a G. triflora double haploid line yielded almost 20,000 clones using 454 pyrosequencing technology, 6.7% of which could be used to design SSR markers. To increase the number of molecular markers, we identified three putative long terminal repeat (LTR) sequences using the recently developed inter-primer binding site (iPBS) method. We also developed retrotransposon microsatellite amplified polymorphism (REMAP) markers combining retrotransposon and inter-simple sequence repeat (ISSR) markers. In addition to SSR and REMAP markers, modified amplified fragment length polymorphism (AFLP) and random amplification polymorphic DNA (RAPD) markers were developed. Using 93 BC1 progeny from G. scabra backcrossed with a G. triflora double haploid line, 19 linkage groups were constructed with a total of 263 markers (97 SSR, 97 AFLP, 39 RAPD, and 30 REMAP markers). One phenotypic trait (stem color) and 10 functional markers related to genes controlling flower color, flowering time and cold tolerance were assigned to the linkage map, confirming its utility. Conclusions This is the first reported genetic linkage map for Japanese gentians and for any species belonging to the family Gentianaceae. As demonstrated by mapping of functional markers and the stem color trait, our results will help to explain the genetic basis of agronomic important traits, and will be useful for marker-assisted selection in gentian breeding programs. Our map will also be an important resource for further genetic analyses such as mapping of quantitative trait loci and map-based cloning of genes in this species. PMID:23186361

Analysis of Genetic Diversity and Structure Pattern of Indigofera Pseudotinctoria in Karst Habitats of the Wushan Mountains Using AFLP Markers.

PubMed

Fan, Yan; Zhang, Chenglin; Wu, Wendan; He, Wei; Zhang, Li; Ma, Xiao

2017-10-16

Indigofera pseudotinctoria Mats is an agronomically and economically important perennial legume shrub with a high forage yield, protein content and strong adaptability, which is subject to natural habitat fragmentation and serious human disturbance. Until now, our knowledge of the genetic relationships and intraspecific genetic diversity for its wild collections is still poor, especially at small spatial scales. Here amplified fragment length polymorphism (AFLP) technology was employed for analysis of genetic diversity, differentiation, and structure of 364 genotypes of I. pseudotinctoria from 15 natural locations in Wushan Montain, a highly structured mountain with typical karst landforms in Southwest China. We also tested whether eco-climate factors has affected genetic structure by correlating genetic diversity with habitat features. A total of 515 distinctly scoreable bands were generated, and 324 of them were polymorphic. The polymorphic information content (PIC) ranged from 0.694 to 0.890 with an average of 0.789 per primer pair. On species level, Nei's gene diversity ( H j ), the Bayesian genetic diversity index ( H B ) and the Shannon information index ( I ) were 0.2465, 0.2363 and 0.3772, respectively. The high differentiation among all sampling sites was detected ( F ST = 0.2217, G ST = 0.1746, G' ST = 0.2060, θ B = 0.1844), and instead, gene flow among accessions ( N m = 1.1819) was restricted. The population genetic structure resolved by the UPGMA tree, principal coordinate analysis, and Bayesian-based cluster analyses irrefutably grouped all accessions into two distinct clusters, i.e., lowland and highland groups. The population genetic structure resolved by the UPGMA tree, principal coordinate analysis, and Bayesian-based cluster analyses irrefutably grouped all accessions into two distinct clusters, i.e., lowland and highland groups. This structure pattern may indicate joint effects by the neutral evolution and natural selection. Restricted N m was observed across all accessions, and genetic barriers were detected between adjacent accessions due to specifically geographical landform.

Analysis of DNA methylation in Arabidopsis thaliana based on methylation-sensitive AFLP markers.

PubMed

Cervera, M T; Ruiz-García, L; Martínez-Zapater, J M

2002-12-01

AFLP analysis using restriction enzyme isoschizomers that differ in their sensitivity to methylation of their recognition sites has been used to analyse the methylation state of anonymous CCGG sequences in Arabidopsis thaliana. The technique was modified to improve the quality of fingerprints and to visualise larger numbers of scorable fragments. Sequencing of amplified fragments indicated that detection was generally associated with non-methylation of the cytosine to which the isoschizomer is sensitive. Comparison of EcoRI/ HpaII and EcoRI/ MspI patterns in different ecotypes revealed that 35-43% of CCGG sites were differentially digested by the isoschizomers. Interestingly, the pattern of digestion among different plants belonging to the same ecotype is highly conserved, with the rate of intra-ecotype methylation-sensitive polymorphisms being less than 1%. However, pairwise comparisons of methylation patterns between samples belonging to different ecotypes revealed differences in up to 34% of the methylation-sensitive polymorphisms. The lack of correlation between inter-ecotype similarity matrices based on methylation-insensitive or methylation-sensitive polymorphisms suggests that whatever the mechanisms regulating methylation may be, they are not related to nucleotide sequence variation.

The genetic map of finger millet, Eleusine coracana.

PubMed

Dida, Mathews M; Srinivasachary; Ramakrishnan, Sujatha; Bennetzen, Jeffrey L; Gale, Mike D; Devos, Katrien M

2007-01-01

Restriction fragment length polymorphism (RFLP), amplified fragment length polymorphism (AFLP), expressed-sequenced tag (EST), and simple sequence repeat (SSR) markers were used to generate a genetic map of the tetraploid finger millet (Eleusine coracana subsp. coracana) genome (2n = 4x = 36). Because levels of variation in finger millet are low, the map was generated in an inter-subspecific F(2) population from a cross between E. coracana subsp. coracana cv. Okhale-1 and its wild progenitor E. coracana subsp. africana acc. MD-20. Duplicated loci were used to identify homoeologous groups. Assignment of linkage groups to the A and B genome was done by comparing the hybridization patterns of probes in Okhale-1, MD-20, and Eleusine indica acc. MD-36. E. indica is the A genome donor to E. coracana. The maps span 721 cM on the A genome and 787 cM on the B genome and cover all 18 finger millet chromosomes, at least partially. To facilitate the use of marker-assisted selection in finger millet, a first set of 82 SSR markers was developed. The SSRs were identified in small-insert genomic libraries generated using methylation-sensitive restriction enzymes. Thirty-one of the SSRs were mapped. Application of the maps and markers in hybridization-based breeding programs will expedite the improvement of finger millet.

Comparison of biotyping methods as alternative identification tools to molecular typing of pathogenic Cryptococcus species in sub-Saharan Africa

PubMed Central

Nyazika, Tinashe K.; Robertson, Valerie J.; Nherera, Brenda; Mapondera, Prichard T.; Meis, Jacques F.; Hagen, Ferry

2015-01-01

Summary Cryptococcal meningitis is the leading fungal infection and AIDS defining opportunistic illness in patients with late stage HIV infection, particularly in South-East Asia and sub-Saharan Africa. Given the high mortality, clinical differences and the extensive ecological niche of Cryptococcus neoformans and Cryptococcus gattii species complexes, there is need for laboratories in sub-Sahara African countries to adopt new and alternative reliable diagnostic algorithms that rapidly identify and distinguish these species. We biotyped 74 and then amplified fragment length polymorphism (AFLP) genotyped 66 Cryptococcus isolates from a cohort of patients with HIV-associated cryptococcal meningitis. Cryptococcus gattii sensu lato was isolated at a prevalence of 16.7% (n = 11/66) and C. neoformans sensu stricto was responsible for 83.3% (n = 55/66) of the infections. l-Canavanine glycine bromothymol blue, yeast-carbon-base-d-proline-d-tryptophan and creatinine dextrose bromothymol blue thymine were able to distinguish pathogenic C. gattii sensu lato from C. neoformans sensu stricto species when compared with amplified fragment length polymorphism genotyping. This study demonstrates high C. gattii sensu lato prevalence in Zimbabwe. In addition, biotyping methods can be used as alternative diagnostic tools to molecular typing in resource-limited areas for differentiating pathogenic Cryptococcus species. PMID:26661484

Genetic structure and breeding system of a rare understory herb, Dysosma versipellis (Berberidaceae), from temperate deciduous forests in China.

PubMed

Guan, Bi-Cai; Fu, Cheng-Xing; Qiu, Ying-Xiong; Zhou, Shi-Liang; Comes, Hans Peter

2010-01-01

To evaluate the role of Quaternary refugial isolation in allopatric (incipient) speciation of East Asian temperate forest biotas, we analyzed amplified fragment length polymorphisms (AFLPs) and the breeding system in Dysosma versipellis. The study revealed that D. versipellis is mostly self-incompatible, genetically highly subdivided and depauperate at the population level (e.g., Φ(ST) = 0.572/H(E) = 0.083), and characterized by a low pollen-to-seed migration ratio (r ≈ 4.0). The latter outcome likely reflects limited pollen flow in a low-seed disperser whose hypothesized "sapromyophilous" flowers undergo scarce, inefficient, and likely specialized cross-pollination by small Anoplodera beetles, rather than carrion flies as assumed previously. In consequence, fruit set in D. versipellis was strongly pollen-limited. Our AFLP data support the hypothesis of a long-standing cessation of gene flow between western and central eastern populations, consistent with previous chloroplast DNA data. This phylogeographic pattern supports the role of the Sichuan Basin as a floristic boundary separating the Sino-Himalayan vs. Sino-Japanese Forest subkingdoms. Our genetic data of D. versipellis also imply that temperate deciduous forest elements to the west and the east of this basin responded differently to Quaternary climate change, which may have triggered or is leading to allopatric (incipient) speciation.

Identification on commercialized products of AFLP markers able to discriminate slow- from fast-growing chicken strains.

PubMed

Fumière, Olivier; Dubois, Marc; Grégoire, Dimitrie; Théwis, André; Berben, Gilbert

2003-02-26

The European chicken meat market is characterized by numerous quality marks: "Label de Qualité Wallon" in Belgium, "Label Rouge" in France, denominations of geographical origin, organic agriculture, etc. Most of those certified productions have specifications requiring the use of slow-growing chicken strains. The amplified fragment length polymorphism (AFLP) technique has been used to search molecular markers able to discriminate slow-growing chicken strains from fast-growing ones and to authenticate certified products. Two pairs of restriction enzymes (EcoRI/MseI and EcoRI/TaqI) and 121 selective primer combinations were tested on individual DNA samples from chicken products essentially in carcass form that were ascribed as belonging to either slow- or fast-growing strains. Within the resulting fingerprints, two fragments were identified as type-strains specific markers. One primer combination gives a band (333 bp) that is specific for slow-growing chickens, and another primer pair generates a band (372 bp) that was found to be characteristic of fast-growing chickens. The two markers were isolated, cloned, and sequenced. The effectiveness and the specificity of the two interesting determinants were assessed on individuals of two well-known strains (ISA 657 and Cobb 500) and on commercialized products coming from various origins.

Bearded dragons (Pogona vitticeps) asymptomatically infected with Devriesea agamarum are a source of persistent clinical infection in captive colonies of dab lizards (Uromastyx sp.).

PubMed

Devloo, R; Martel, A; Hellebuyck, T; Vranckx, K; Haesebrouck, F; Pasmans, F

2011-06-02

Devriesea agamarum causes dermatitis and septicaemia in a variety of lizards, notably those belonging to the genus Uromastyx, whereas other species such as bearded dragons (Pogona vitticeps) seem to be asymptomatic carriers. Using amplified fragment length polymorphism (AFLP), the relatedness between 69 D. agamarum isolates was examined. The isolates derived from 44 diseased lizards, of which 31 belonged to the genus Uromastyx, and from 25 healthy lizards, of which 21 were bearded dragons. Eight AFLP genotypes were obtained, four of which comprised 93% of the isolates. These four genotypes were each present in 2, 2, 8 and 13 different captive colonies. Up to three genotypes were isolated from a single infected colony simultaneously. On two occasions, the same genotype was found in healthy bearded dragons and diseased Uromastyx lizards from the same colony, confirming the role of the former as an asymptomatic source of infection for the latter. Two genotypes, comprising 12 isolates, were exclusively associated with diseased Uromastyx lizards, suggesting strain dependent host adaptation. Finally, D. agamarum was shown to be able to persist for at least seven years in a lizard colony, persistently causing severe disease in several lizard species. Copyright © 2011 Elsevier B.V. All rights reserved.

Broad-scale adaptive genetic variation in alpine plants is driven by temperature and precipitation

PubMed Central

MANEL, STÉPHANIE; GUGERLI, FELIX; THUILLER, WILFRIED; ALVAREZ, NADIR; LEGENDRE, PIERRE; HOLDEREGGER, ROLF; GIELLY, LUDOVIC; TABERLET, PIERRE

2014-01-01

Identifying adaptive genetic variation is a challenging task, in particular in non-model species for which genomic information is still limited or absent. Here, we studied distribution patterns of amplified fragment length polymorphisms (AFLPs) in response to environmental variation, in 13 alpine plant species consistently sampled across the entire European Alps. Multiple linear regressions were performed between AFLP allele frequencies per site as dependent variables and two categories of independent variables, namely Moran’s eigenvector map MEM variables (to account for spatial and unaccounted environmental variation, and historical demographic processes) and environmental variables. These associations allowed the identification of 153 loci of ecological relevance. Univariate regressions between allele frequency and each environmental factor further showed that loci of ecological relevance were mainly correlated with MEM variables. We found that precipitation and temperature were the best environmental predictors, whereas topographic factors were rarely involved in environmental associations. Climatic factors, subject to rapid variation as a result of the current global warming, are known to strongly influence the fate of alpine plants. Our study shows, for the first time for a large number of species, that the same environmental variables are drivers of plant adaptation at the scale of a whole biome, here the European Alps. PMID:22680783

Carry-over of thermophilic Campylobacter spp. between sequential and adjacent poultry flocks.

PubMed

Alter, Thomas; Weber, Rita Margarete; Hamedy, Ahmad; Glünder, Gerhard

2011-01-10

Nineteen flocks of four poultry species were monitored at a veterinary field station to investigate the distribution and spread of Campylobacter genotypes between sequential and adjacent flocks. Caecal and liver samples were obtained at frequent intervals from birds of all flocks and examined for Campylobacter. Amplified fragment length polymorphism (AFLP) analysis was performed to genotype Campylobacter isolates. Of the 1643 caecal and liver samples investigated, 452 (27.5%) caecal samples and 11 (0.7%) liver samples contained Campylobacter. Of the caecal isolates 76.3% were identified as Campylobacter jejuni and 23.7% were identified as Campylobacter coli. Poultry flocks were largely colonized by more than one AFLP type and an intense exchange of Campylobacter genotypes between different poultry flocks occurred. These findings indicate that multiple genotypes can constitute the Campylobacter population within single poultry flocks, hinting to different sources of exposure and/or genetic drifts within the Campylobacter population. Nevertheless, in most flocks single Campylobacter genotypes predominated. Some strains superseded others resulting in colonization by successive Campylobacter genotypes during the observation period. In conclusion, the data demonstrate that the large genetic diversity of Campylobacter must be considered in epidemiological evaluations and microbial risk assessments of Campylobacter in poultry. Copyright © 2010 Elsevier B.V. All rights reserved.

Phylogeography above the species level for perennial species in a composite genus

PubMed Central

Tremetsberger, Karin; Ortiz, María Ángeles; Terrab, Anass; Balao, Francisco; Casimiro-Soriguer, Ramón; Talavera, María; Talavera, Salvador

2016-01-01

In phylogeography, DNA sequence and fingerprint data at the population level are used to infer evolutionary histories of species. Phylogeography above the species level is concerned with the genealogical aspects of divergent lineages. Here, we present a phylogeographic study to examine the evolutionary history of a western Mediterranean composite, focusing on the perennial species of Helminthotheca (Asteraceae, Cichorieae). We used molecular markers (amplified fragment length polymorphism (AFLP), internal transcribed spacer and plastid DNA sequences) to infer relationships among populations throughout the distributional range of the group. Interpretation is aided by biogeographic and molecular clock analyses. Four coherent entities are revealed by Bayesian mixture clustering of AFLP data, which correspond to taxa previously recognized at the rank of subspecies. The origin of the group was in western North Africa, from where it expanded across the Strait of Gibraltar to the Iberian Peninsula and across the Strait of Sicily to Sicily. Pleistocene lineage divergence is inferred within western North Africa as well as within the western Iberian region. The existence of the four entities as discrete evolutionary lineages suggests that they should be elevated to the rank of species, yielding H. aculeata, H. comosa, H. maroccana and H. spinosa, whereby the latter two necessitate new combinations. PMID:26644340

Diversification of Bromelioideae (Bromeliaceae) in the Brazilian Atlantic rainforest: A case study in Aechmea subgenus Ortgiesia.

PubMed

Goetze, Márcia; Schulte, Katharina; Palma-Silva, Clarisse; Zanella, Camila M; Büttow, Miriam V; Capra, Fernanda; Bered, Fernanda

2016-05-01

Aechmea subgenus Ortgiesia comprises ca. 20 species distributed in Brazil, Argentina, Paraguay, and Uruguay, with a center of diversity in the Brazilian Atlantic rainforest. We examined interspecific relationships of Ortgiesia based on Amplified Fragment Length Polymorphisms (AFLP). Ninety-six accessions belonging to 14 species of Ortgiesia were sampled, and genotyped with 11 AFLP primer combinations. The neighbor joining (NJ) tree depicted two main genetic groups within Aechmea subgenus Ortgiesia, and four subgroups. The NJ tree showed short internal branches, indicating an overall shallow genetic divergence among Ortgiesia species as expected for the recently radiated subfamily Bromelioideae. Our results suggest that hybridization and/or incomplete lineage sorting may have hampered the reconstruction of interspecific relationships in Aechmea subgenus Ortgiesia. The mapping of petal color (yellow, blue, pink, or white), inflorescence type (simple or compound), and inflorescence shape (ellipsoid, subcylindric, cylindric, or pyramidal) against the NJ tree indicated that these characters are of limited taxonomic use in Aechmea subgenus Ortgiesia due to homoplasy. An analysis of the current distribution of Ortgiesia identified the southern region of the Brazilian Atlantic rainforest, between latitudes of 26° and 27°S, as the center of diversity for the subgenus. Copyright © 2016 Elsevier Inc. All rights reserved.

Identification of aluminum-regulated genes by cDNA-AFLP analysis of roots in two contrasting genotypes of highbush blueberry (Vaccinium corymbosum L.).

PubMed

Inostroza-Blancheteau, Claudio; Aquea, Felipe; Reyes-Díaz, Marjorie; Alberdi, Miren; Arce-Johnson, Patricio

2011-09-01

To investigate the molecular mechanisms of Al(3+)-stress in blueberry, a cDNA-amplified fragment length polymorphism (cDNA-AFLP) analysis was employed to identify Al-regulated genes in roots of contrasting genotypes of highbush blueberry (Brigitta, Al(3+)-resistant and Bluegold, Al(3+)-sensitive). Plants grown in hydroponic culture were treated with 0 and 100 μM Al(3+) and collected at different times over 48 h. Seventy transcript-derived fragments (TDFs) were identified as being Al(3+) responsive, 31 of which showed significant homology to genes with known or putative functions. Twelve TDFs were homologous to uncharacterized genes and 27 did not have significant matches. The expression pattern of several of the genes with known functions in other species was confirmed by quantitative relative real-time RT-PCR. Twelve genes of known or putative function were related to cellular metabolism, nine associated to stress responses and other transcription and transport facilitation processes. Genes involved in signal transduction, photosynthetic and energy processes were also identified, suggesting that a multitude of processes are implicated in the Al(3+)-stress response as reported previously for other species. The Al(3+)-stress response genes identified in this study could be involved in Al(3+)-resistance in woody plants.

Comparison of biotyping methods as alternative identification tools to molecular typing of pathogenic Cryptococcus species in sub-Saharan Africa.

PubMed

Nyazika, Tinashe K; Robertson, Valerie J; Nherera, Brenda; Mapondera, Prichard T; Meis, Jacques F; Hagen, Ferry

2016-03-01

Cryptococcal meningitis is the leading fungal infection and AIDS defining opportunistic illness in patients with late stage HIV infection, particularly in South-East Asia and sub-Saharan Africa. Given the high mortality, clinical differences and the extensive ecological niche of Cryptococcus neoformans and Cryptococcus gattii species complexes, there is need for laboratories in sub-Sahara African countries to adopt new and alternative reliable diagnostic algorithms that rapidly identify and distinguish these species. We biotyped 74 and then amplified fragment length polymorphism (AFLP) genotyped 66 Cryptococcus isolates from a cohort of patients with HIV-associated cryptococcal meningitis. C. gattii sensu lato was isolated at a prevalence of 16.7% (n = 11/66) and C. neoformans sensu stricto was responsible for 83.3% (n = 55/66) of the infections. l-Canavanine glycine bromothymol blue, yeast-carbon-base-d-proline-d-tryptophan and creatinine dextrose bromothymol blue thymine were able to distinguish pathogenic C. gattii sensu lato from C. neoformans sensu stricto species when compared with AFLP genotyping. This study demonstrates high C. gattii sensu lato prevalence in Zimbabwe. In addition, biotyping methods can be used as alternative diagnostic tools to molecular typing in resource-limited areas for differentiating pathogenic Cryptococcus species. © 2015 Blackwell Verlag GmbH.

Molecular evidence of hybrid zones of Cedrela (Meliaceae) in the Yungas of Northwestern Argentina.

PubMed

Zelener, Noga; Tosto, Daniela; de Oliveira, Luiz Orlando; Soldati, María Cristina; Inza, María Virginia; Fornes, Luis Fernando

2016-09-01

In the Yungas of Northwestern Argentina, three endangered species of Cedrela (C. angustifolia, C. saltensis, and C. balansae) follow altitudinal gradients of distribution with contact zones between them. We sampled 210 individuals from 20 populations that spanned most of Cedrela's geographical range in the Yungas, and used Amplified Fragment Length Polymorphism (AFLP) markers and DNA sequences of the nuclear Internal Transcribed Spacer (ITS) to investigate hybrid zones. Data analyses employed an array of complementary methods, including principal coordinate analyses, Bayesian clustering analyses, maximum likelihood tree-building, and network techniques. Both nuclear molecular systems - AFLP and ITS - provided insights into the evolutionary history of Cedrela in the Yungas in a congruent manner. We uncovered strong support for the occurrence of natural hybridization between C. balansae and C. saltensis. Additionally, we identified hybrid zones in areas of sympatry (at both the Calilegua National Park and the San Andrés farm) and in transition zones from 820 to 1100meters above sea level (localities of Pintascayo and Acambuco). There was no evidence for hybridization of either C. balansae or C. saltensis with C. angustifolia. The role of hybrid populations in conservation and use of genetic resources in the Yungas were discussed. Copyright © 2016 Elsevier Inc. All rights reserved.

Spatial genetic structure of the Sea Sandwort on Surtsey: an immigrant's journey

NASA Astrophysics Data System (ADS)

Árnason, S. H.; Thórsson, Th.; Magnússon, B.; Philipp, M.; Adsersen, H. E.; Anamthawat-Jónsson, K.

2014-06-01

Sea Sandwort (Honckenya peploides) is one of the first plants to successfully colonize and reproduce on the volcanic island Surtsey, formed in 1963 off the southern coast of Iceland. Using amplified fragment length polymorphic (AFLP) markers we examined levels of genetic variation and differentiation among populations of H. peploides on Surtsey in relation to populations on the nearby island Heimaey and from the southern coast of Iceland. Selected populations from Denmark and Greenland were used for comparison. In addition, we tested whether the effects of isolation by distance can be seen in the Surtsey populations. Using two primer combinations, we obtained 173 AFLP markers from a total of 347 plant samples. The resulting binary matrix was then analyzed statistically. Main results include the followings: (i) Surtsey has the highest proportion of polymorphic markers as well as a comparatively high genetic diversity (55.5% PLP; 0.1974 HE) and Denmark the lowest (31.8% PLP; 0.132 HE), indicating rapid expansion during an early stage of population establishment on Surtsey and/or multiple origins of immigrants; (ii) the total genetic differentiation (FST) among Surtsey (0.0714) and Heimaey (0.055) populations was less than half of that found among the mainland populations in Iceland (0.1747), indicating substantial gene flow on the islands; (iii) most of the genetic variation (79%, p<0.001) was found within localities, possibly due to the outcrossing and subdioecious nature of the species; (iv) a significant genetic distance was found within Surtsey, among sites, and this appears to correlate with the age of plant colonization; and (v) the genetic structure analysis indicated multiple colonization episodes on Surtsey, whereby H. peploides most likely immigrated from the nearby island of Heimaey and directly from the southern coast of Iceland.

Phylogeography and seed dispersal in islands: the case of Rumex bucephalophorus subsp. canariensis (Polygonaceae)

PubMed Central

Talavera, María; Navarro-Sampedro, Laura; Ortiz, Pedro L.; Arista, Montserrat

2013-01-01

Background and Aims Rumex bucephalophorus subsp. canariensis is an endemic taxon to Macaronesia with diaspore polymorphism. The origin and colonizing route of this taxon in Macaronesia was studied using molecular data and information on diaspore types. Methods Amplified fragment length polymorphism (AFLP) was used in 260 plants from 22 populations of R. bucephalophorus subsp. canariensis, four from the Madeiran archipelago and 18 from the Canary archipelago. Diaspore production was analysed in 9–50 plants from each population used for AFLP analysis. One hundred and one plants from the Madeiran archipelago and 375 plants from the Canary Islands were studied. For each plant the type of diaspore produced was recorded. Key Results Overall populations had low genetic diversity but they showed a geographical pattern of genetic diversity that was higher in the older eastern islands than in the younger western ones. Two types of dispersible diaspores were found: in the eastern Canary islands (Lanzarote, Fuerteventura and Gran Canaria), plants produced exclusively long-dispersible diaspores, whereas in the western Canary islands (Tenerife, La Gomera, El Hierro) and the Madeiran archipelago plants produced exclusively short-dispersible diaspores. Genetically, the studied populations fell into four main island groups: Lanzarote–Fuerteventura, Gran Canaria, Tenerife–El Hierro and La Gomera–Madeira archipelago. Conclusions A Moroccan origin of R. bucephalophorus subsp. canariensis is hypothesized with a colonization route from the eastern to the western islands. In addition, at least one gene flow event from La Gomera to the Madeiran archipelago has taken place. During the colonization process the type of dispersible diaspore changed so that dispersability decreased in populations of the westernmost islands. PMID:23267005

A first linkage map of globe artichoke (Cynara cardunculus var. scolymus L.) based on AFLP, S-SAP, M-AFLP and microsatellite markers.

PubMed

Lanteri, S; Acquadro, A; Comino, C; Mauro, R; Mauromicale, G; Portis, E

2006-05-01

We present the first genetic maps of globe artichoke (Cynara cardunculus var. scolymus L. 2n=2x=34), constructed with a two-way pseudo-testcross strategy. A F1 mapping population of 94 individuals was generated between a late-maturing, non-spiny type and an early-maturing spiny type. The 30 AFLP, 13 M-AFLP and 9 S-SAP primer combinations chosen identified, respectively, 352, 38 and 41 polymorphic markers. Of 32 microsatellite primer pairs tested, 12 identified heterozygous loci in one or other parent, and 7 were fully informative as they segregated in both parents. The female parent map comprised 204 loci, spread over 18 linkage groups and spanned 1330.5 cM with a mean marker density of 6.5 cM. The equivalent figures for the male parent map were 180 loci, 17 linkage groups, 1239.4 and 6.9 cM. About 3% of the AFLP and AFLP-derived markers displayed segregation distortion with a P value below 0.01, and were not used for map construction. All the SSR loci were included in the linkage analysis, although one locus did show some segregation distortion. The presence of 78 markers in common to both maps allowed the alignment of 16 linkage groups. The maps generated provide a firm basis for the mapping of agriculturally relevant traits, which will then open the way for the application of a marker-assisted selection breeding strategy in this species.

Genetic identification of spotted owls, barred owls, and their hybrids: Legal implications of hybrid identity

USGS Publications Warehouse

Haig, Susan M.; Wennerberg, Liv; Mullins, Thomas D.; Forsman, E.D.; Trail, P.

2004-01-01

Recent population expansion of Barred Owls ( Strix varia) into western North America has led to concern that they may compete with and further harm the Northern Spotted Owl ( S. occidentalis caurina), which is already listed as threatened under the U.S. Endangered Species Act (ESA). Because they hybridize, there is a legal need under the ESA for forensic identification of both species and their hybrids. We used mitochondrial control-region DNA and amplified fragment-length polymorphism (AFLP) analyses to assess maternal and biparental gene flow in this hybridization process. Mitochondrial DNA sequences (524 base pairs) indicated large divergence between Barred and Spotted Owls (13.9%). Further, the species formed two distinct clades with no signs of previous introgression. Fourteen diagnostic AFLP bands also indicated extensive divergence between the species, including markers differentiating them. Principal coordinate analyses and assignment tests clearly supported this differentiation. We found that hybrids had unique genetic combinations, including AFLP markers from both parental species, and identified known hybrids as well as potential hybrids with unclear taxonomic status. Our analyses corroborated the findings of extensive field studies that most hybrids genetically sampled resulted from crosses between female Barred Owls and male Spotted Owls. These genetic markers make it possible to clearly identify these species as well as hybrids and can now be used for research, conservation, and law enforcement. Several legal avenues may facilitate future conservation of Spotted Owls and other ESA-listed species that hybridize, including the ESA similarity-of-appearance clause (section 4[e]) and the Migratory Bird Treaty Act. The Migratory Bird Treaty Act appears to be the most useful route at this time.

Genetic characterization and barcoding of taxa in the genus Wolffia Horkel ex Schleid. (Lemnaceae) as revealed by two plastidic markers and amplified fragment length polymorphism (AFLP).

PubMed

Bog, Manuela; Schneider, Philipp; Hellwig, Frank; Sachse, Svea; Kochieva, Elena Z; Martyrosian, Elena; Landolt, Elias; Appenroth, Klaus-J

2013-01-01

The genus Wolffia of the duckweed family (Lemnaceae) contains the smallest flowering plants. Presently, 11 species are recognized and categorized mainly on the basis of morphology. Because of extreme reduction of structure of all species, molecular methods are especially required for barcoding and identification of species and clones of this genus. We applied AFLP combined with Bayesian analysis of population structure to 66 clones covering all 11 species. Nine clusters were identified: (1) W. angusta and W. microscopica (only one clone), (2) W. arrhiza, (3) W. cylindracea (except one clone that might be a transition form), (4) W. australiana, (5) W. globosa, (6) W. globosa, W. neglecta, and W. borealis, (7) W. brasiliensis, and W. columbiana, (8) W. columbiana, (9) W. elongata. Furthermore, we investigated the sequences of plastidic regions rps16 (54 clones) and rpl16 (55 clones), and identified the following species: W. angusta, W. australiana, W. brasiliensis, W. cylindracea, W. elongata, W. microscopica, and W. neglecta. Wolffia globosa has been separated into two groups by both methods. One group which consists only of clones from North America and East Asia was labelled here "typical W. globosa". The other group of W. globosa, termed operationally "W. neglecta", contains also clones of W. neglecta and shows high similarity to W. borealis. None of the methods recognized W. borealis as a distinct species. Although each clone could be characterized individually by AFLP and plastidic sequences, and most species could be bar-coded, the presently available data are not sufficient to identify all taxa of Wolffia.

Molecular typing of Legionella pneumophila serogroup 1 clinical strains isolated in Italy.

PubMed

Fontana, Stefano; Scaturro, Maria; Rota, Maria Cristina; Caporali, Maria Grazia; Ricci, Maria Luisa

2014-07-01

Molecular typing methods for discriminating different bacterial isolates are essential epidemiological tools in prevention and control of Legionella infections and outbreaks. A selection of 56 out of 184 Legionella pneumophila serogroup 1 (Lp1) clinical isolates, collected from different Italian regions between 1987 and 2012, and stored at the National Reference Laboratory for Legionella, were typed by monoclonal antibody (MAb) subgrouping, amplified fragment length polymorphism (AFLP) and sequence based typing (SBT). These strains were isolated from 39 community (69.6%), 14 nosocomial (25%) and 3 travel associated (5.4%) Legionnaires'disease cases. MAb typing results showed a prevalence of MAb 3/1 positive isolates (75%) with the Philadelphia subgroup representing 35.7%, followed by Knoxville (23.2%), Benidorm (12.5%), Allentown/France (1.8%), Allentown/France-Philadelphia (1.8%). The remaining 25% were MAb 3/1 negative, namely 11 Olda (19.6%), 2 Oxford (3.6%) and 1 Bellingham (1.8%) subgroups. AFLP analysis detected 20 different genomic profiles. SBT analysis revealed 32 different sequence types (STs) with high diversity of STs (IODSTs=0.952) 12 of which were never described before. ST1 and ST23 were most frequently isolated as observed worldwide. A helpful analysis of data from SBT, MAb subgrouping and AFLP is provided, as well as a comparison to the Lp1 types investigated from other countries. This study describes the first Italian Lp1 strains database, providing molecular epidemiology data useful for future epidemiological investigations, especially of travel associated Legionnaires' diseases (TALD) cases, Italy being the country associated with the highest number of clusters. Copyright © 2014 Elsevier GmbH. All rights reserved.

Whole genome sequencing as a tool to investigate a cluster of seven cases of listeriosis in Austria and Germany, 2011-2013.

PubMed

Schmid, D; Allerberger, F; Huhulescu, S; Pietzka, A; Amar, C; Kleta, S; Prager, R; Preußel, K; Aichinger, E; Mellmann, A

2014-05-01

A cluster of seven human cases of listeriosis occurred in Austria and in Germany between April 2011 and July 2013. The Listeria monocytogenes serovar (SV) 1/2b isolates shared pulsed-field gel electrophoresis (PFGE) and fluorescent amplified fragment length polymorphism (fAFLP) patterns indistinguishable from those from five food producers. The seven human isolates, a control strain with a different PFGE/fAFLP profile and ten food isolates were subjected to whole genome sequencing (WGS) in a blinded fashion. A gene-by-gene comparison (multilocus sequence typing (MLST)+) was performed, and the resulting whole genome allelic profiles were compared using SeqSphere(+) software version 1.0. On analysis of 2298 genes, the four human outbreak isolates from 2012 to 2013 had different alleles at ≤6 genes, i.e. differed by ≤6 genes from each other; the dendrogram placed these isolates in between five Austrian unaged soft cheese isolates from producer A (≤19-gene difference from the human cluster) and two Austrian ready-to-eat meat isolates from producer B (≤8-gene difference from the human cluster). Both food products appeared on grocery bills prospectively collected by these outbreak cases after hospital discharge. Epidemiological results on food consumption and MLST+ clearly separated the three cases in 2011 from the four 2012-2013 outbreak cases (≥48 different genes). We showed that WGS is capable of discriminating L. monocytogenes SV1/2b clones not distinguishable by PFGE and fAFLP. The listeriosis outbreak described clearly underlines the potential of sequence-based typing methods to offer enhanced resolution and comparability of typing systems for public health applications. © 2014 The Authors Clinical Microbiology and Infection © 2014 European Society of Clinical Microbiology and Infectious Diseases.

Genet-specific DNA methylation probabilities detected in a spatial epigenetic analysis of a clonal plant population.

PubMed

Araki, Kiwako S; Kubo, Takuya; Kudoh, Hiroshi

2017-01-01

In sessile organisms such as plants, spatial genetic structures of populations show long-lasting patterns. These structures have been analyzed across diverse taxa to understand the processes that determine the genetic makeup of organismal populations. For many sessile organisms that mainly propagate via clonal spread, epigenetic status can vary between clonal individuals in the absence of genetic changes. However, fewer previous studies have explored the epigenetic properties in comparison to the genetic properties of natural plant populations. Here, we report the simultaneous evaluation of the spatial structure of genetic and epigenetic variation in a natural population of the clonal plant Cardamine leucantha. We applied a hierarchical Bayesian model to evaluate the effects of membership of a genet (a group of individuals clonally derived from a single seed) and vegetation cover on the epigenetic variation between ramets (clonal plants that are physiologically independent individuals). We sampled 332 ramets in a 20 m × 20 m study plot that contained 137 genets (identified using eight SSR markers). We detected epigenetic variation in DNA methylation at 24 methylation-sensitive amplified fragment length polymorphism (MS-AFLP) loci. There were significant genet effects at all 24 MS-AFLP loci in the distribution of subepiloci. Vegetation cover had no statistically significant effect on variation in the majority of MS-AFLP loci. The spatial aggregation of epigenetic variation is therefore largely explained by the aggregation of ramets that belong to the same genets. By applying hierarchical Bayesian analyses, we successfully identified a number of genet-specific changes in epigenetic status within a natural plant population in a complex context, where genotypes and environmental factors are unevenly distributed. This finding suggests that it requires further studies on the spatial epigenetic structure of natural populations of diverse organisms, particularly for sessile clonal species.

Prevalence and Population Structure of Vibrio vulnificus on Fishes from the Northern Gulf of Mexico

PubMed Central

Tao, Zhen; Larsen, Andrea M.; Bullard, Stephen A.; Wright, Anita C.

2012-01-01

The prevalence of Vibrio vulnificus on the external surfaces of fish from the northern Gulf of Mexico was determined in this study. A collection of 242 fish comprising 28 species was analyzed during the course of 12 sampling trips over a 16-month period. The prevalence of V. vulnificus was 37% but increased up to 69% in summer. A positive correlation was found between the percentages of V. vulnificus-positive fish and water temperatures, while salinity and V. vulnificus-positive fish prevalence were inversely correlated. A general lineal model (percent V. vulnificus-positive fish = 0.5930 − 0.02818 × salinity + 0.01406 × water temperature) was applied to best fit the data. Analysis of the population structure was carried out using 244 isolates recovered from fish. Ascription to 16S rRNA gene types indicated that 157 isolates were type A (62%), 72 (29%) were type B, and 22 (9%) were type AB. The percentage of type B isolates, considered to have greater virulence potential, was higher than that previously reported in oyster samples from the northern Gulf of Mexico. Amplified fragment length polymorphism (AFLP) was used to resolve the genetic diversity within the species. One hundred twenty-one unique AFLP profiles were found among all analyzed isolates, resulting in a calculated Simpson's index of diversity of 0.991. AFLP profiles were not grouped on the basis of collection date, fish species, temperature, or salinity, but isolates were clustered into two main groups that correlated precisely with 16S rRNA gene type. The population of V. vulnificus associated with fishes from the northern Gulf of Mexico is heterogeneous and includes strains of great virulence potential. PMID:22923394

cDNA-AFLP analysis of transcripts induced in chickpea plants by TiO2 nanoparticles during cold stress.

PubMed

Amini, Saeed; Maali-Amiri, Reza; Mohammadi, Rahmat; Kazemi-Shahandashti, Seyyedeh-Sanam

2017-02-01

We evaluated the effect of TiO 2 nanoparticles (NPs) on cold tolerance (CT) development in two chickpea (Cicer arietinum L.) genotypes (Sel96Th11439, cold tolerant, and ILC533, cold susceptible) by using cDNA-amplified fragment length polymorphism (cDNA-AFLP) technique during the first and sixth days of cold stress (CS) at 4 °C. Selective amplification by primer combinations generated 4200 transcript-derived fragments (TDFs) while 100 of them (2.62%) were differentially expressed. During CS, 60 differentially expressed TDFs of TiO 2 NPs-treated plants were cloned and 10 of them produced successfully readable sequences. These data represented different groups of genes involved in metabolism pathways, cellular defense, cell connections and signaling, transcriptional regulation and chromatin architecture. Two out of 10 TDFs were unknown genes with uncharacterized functions or sequences without homology to known ones. The network-based analysis showed a gene-gene relationship in response to CS. Quantitative reverse-transcriptase polymerase chain reaction (qPCR) confirmed differential expression of identified genes (six out of 10 TDFs) with potential functions in CT and showed similar patterns with cDNA-AFLP results. An increase in transcription level of these TDFs, particularly on the first day of CS, was crucial for developing CT through decreasing electrolyte leakage index (ELI) content in tolerant plants compared to susceptible ones, as well as in TiO 2 NPs-treated plants compared to control ones. It could also indicate probable role of TiO 2 NPs against CS-induced oxidative stress. Therefore, a new application of TiO 2 NPs in CT development is suggested for preventing or controlling the damages in field conditions and increasing crop productivity. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

The effect of antimicrobial photodynamic therapy on the expression of novel methicillin resistance markers determined using cDNA-AFLP approach in Staphylococcus aureus.

PubMed

Hoorijani, Mohammad Neshvan; Rostami, Hosein; Pourhajibagher, Maryam; Chiniforush, Nasim; Heidari, Mansour; Pourakbari, Babak; Kazemian, Hossein; Davari, Kambiz; Amini, Vahid; Raoofian, Reza; Bahador, Abbas

2017-09-01

Widespread methicillin resistant Staphylococcus aureus (MRSA) and absence of effective antimicrobial agents has led to limited therapeutic options for treating MRSA infection. We aimed to evaluate the effect of antimicrobial photodynamic therapy (aPDT) on the expression of novel identified methicillin resistance markers (NIMRMs) in S. aureus using complementary DNA-Amplified Fragment Length Polymorphism (cDNA-AFLP) approaches to address the therapeutic alternatives for MRSA infections. We used cDNA-AFLP to compare MRSA and methicillin susceptible S. aureus (MSSA) for identification of target genes implicated in methicillin resistance. To determine the sub-lethal aPDT (sPDT), MRSA and MSSA clinical isolates photosensitized with toluidine blue O (TBO), and then were irradiated with diode laser. After sPDT, the colony forming units/mL was quantified. Antimicrobial susceptibility against methicillin was assessed for cell-surviving aPDT. Effects of sPDT on the expression of NIMRMs were evaluated by real-time quantitative reverse transcription PCR. According to our results, serine hydrolase family protein (Shfp) encoding gene and a gene encoding a conserved hypothetical protein (Chp) were implicated in methicillin resistance in MRSA. sPDT reduced the minimum inhibitory concentrations of methicillin by 3-fold in MRSA. sPDT could lead to about 10- and 6.2- fold suppression of expression of the Chp and Shfp encoding genes, respectively. sPDT would lead to reduction in resistance to methicillin of MRSA in surviving cells by suppressing the expression of the Shfp and Chp encoding genes associated with methicillin resistance. This may have potential implications of aPDT for the treatment of MRSA infections. Copyright © 2017 Elsevier B.V. All rights reserved.

Spatial patterns of AFLP diversity in Bulbophyllum occultum (Orchidaceae) indicate long-term refugial isolation in Madagascar and long-distance colonization effects in La Réunion.

PubMed

Jaros, U; Fischer, G A; Pailler, T; Comes, H P

2016-05-01

Bulbophyllum occultum, an epiphytic orchid mainly distributed in the rainforests of (north)eastern Madagascar and La Réunion, represents an interesting model case for testing the effects of anthropogenic vs historical (e.g., climate induced) habitat isolation and long-distance colonization on the genetic structure of plant species with disjunct distributions in the Madagascan region. To this aim, we surveyed amplified fragment length polymorphisms (AFLPs) across 13 populations in Madagascar and nine in La Réunion (206 individuals in total). We found overall high levels of population subdivision (Φ(PT)=0.387) and low within-population diversity (H(E), range: 0.026-0.124), indicating non-equilibrium conditions in a mainly selfing species. There was no impact of recent deforestation (Madagascar) or habitat disturbance (La Réunion) detectable on AFLP diversity. K-means clustering and BARRIER analyses identified multiple gene pools and several genetic breaks, both within and among islands. Inter-island levels of population genetic diversity and subdivision were similar, whereby inter-individual divergence in flower colour explained a significant part of gene pool divergence in La Réunion. Our results suggest that (i) B. occultum persisted across multiple isolated ('refugial') regions along the eastern rainforest corridor of Madagascar over recent climatic cycles and (ii) populations in La Réunion arose from either single or few independent introductions from Madagascar. High selfing rates and sufficient time for genetic drift likely promoted unexpectedly high population genetic and phenotypic (flower colour) differentiation in La Réunion. Overall, this study highlights a strong imprint of history on the genetic structure of a low-gene-dispersing epiphytic orchid from the Madagascan region.

Profiling and Quantifying Differential Gene Transcription Provide Insights into Ganoderic Acid Biosynthesis in Ganoderma lucidum in Response to Methyl Jasmonate

PubMed Central

Shi, Liang; Mu, Da-Shuai; Jiang, Ai-Liang; Han, Qin; Zhao, Ming-Wen

2013-01-01

Ganoderma lucidum is a mushroom with traditional medicinal properties that has been widely used in China and other countries in Eastern Asia. Ganoderic acids (GA) produced by G. lucidum exhibit important pharmacological activities. Previous studies have demonstrated that methyl jasmonate (MeJA) is a potent inducer of GA biosynthesis and the expression of genes involved in the GA biosynthesis pathway in G. lucidum. To further explore the mechanism of GA biosynthesis, cDNA-Amplified Fragment Length Polymorphism (cDNA-AFLP) was used to identify genes that are differentially expressed in response to MeJA. Using 64 primer combinations, over 3910 transcriptionally derived fragments (TDFs) were obtained. Reliable sequence data were obtained for 390 of 458 selected TDFs. Ninety of these TDFs were annotated with known functions through BLASTX searching the GenBank database, and 12 annotated TDFs were assigned into secondary metabolic pathways by searching the KEGGPATHWAY database. Twenty-five TDFs were selected for qRT-PCR analysis to confirm the expression patterns observed with cDNA-AFLP. The qRT-PCR results were consistent with the altered patterns of gene expression revealed by the cDNA-AFLP technique. Additionally, the transcript levels of 10 genes were measured at the mycelium, primordia, and fruiting body developmental stages of G. lucidum. The greatest expression levels were reached during primordia for all of the genes except cytochrome b2 reached its highest expression level in the mycelium stage. This study not only identifies new candidate genes involved in the regulation of GA biosynthesis but also provides further insight into MeJA-induced gene expression and secondary metabolic response in G. lucidum. PMID:23762280

Rapid genetic divergence in response to 15 years of simulated climate change.

PubMed

Ravenscroft, Catherine H; Whitlock, Raj; Fridley, Jason D

2015-11-01

Genetic diversity may play an important role in allowing individual species to resist climate change, by permitting evolutionary responses. Our understanding of the potential for such responses to climate change remains limited, and very few experimental tests have been carried out within intact ecosystems. Here, we use amplified fragment length polymorphism (AFLP) data to assess genetic divergence and test for signatures of evolutionary change driven by long-term simulated climate change applied to natural grassland at Buxton Climate Change Impacts Laboratory (BCCIL). Experimental climate treatments were applied to grassland plots for 15 years using a replicated and spatially blocked design and included warming, drought and precipitation treatments. We detected significant genetic differentiation between climate change treatments and control plots in two coexisting perennial plant study species (Festuca ovina and Plantago lanceolata). Outlier analyses revealed a consistent signature of selection associated with experimental climate treatments at individual AFLP loci in P. lanceolata, but not in F. ovina. Average background differentiation at putatively neutral AFLP loci was close to zero, and genomewide genetic structure was associated neither with species abundance changes (demography) nor with plant community-level responses to long-term climate treatments. Our results demonstrate genetic divergence in response to a suite of climatic environments in reproductively mature populations of two perennial plant species and are consistent with an evolutionary response to climatic selection in P. lanceolata. These genetic changes have occurred in parallel with impacts on plant community structure and may have contributed to the persistence of individual species through 15 years of simulated climate change at BCCIL. © 2015 The Authors. Global Change Biology Bioenergy Published by John Wiley & Sons Ltd.

Widespread utility of highly informative AFLP molecular markers across divergent shark species.

PubMed

Zenger, Kyall R; Stow, Adam J; Peddemors, Victor; Briscoe, David A; Harcourt, Robert G

2006-01-01

Population numbers of many shark species are declining rapidly around the world. Despite the commercial and conservation significance, little is known on even the most fundamental aspects of their population biology. Data collection that relies on direct observation can be logistically challenging with sharks. Consequently, molecular methods are becoming increasingly important to obtain knowledge that is critical for conservation and management. Here we describe an amplified fragment length polymorphism method that can be applied universally to sharks to identify highly informative genome-wide polymorphisms from 12 primer pairs. We demonstrate the value of our method on 15 divergent shark species within the superorder Galeomorphii, including endangered species which are notorious for low levels of genetic diversity. Both the endangered sand tiger shark (Carcharodon taurus, N = 18) and the great white shark (Carcharodon carcharias, N = 7) displayed relatively high levels of allelic diversity. A total of 59 polymorphic loci (H(e) = 0.373) and 78 polymorphic loci (H(e) = 0.316) were resolved in C. taurus and C. carcharias, respectively. Results from other sharks (e.g., Orectolobus ornatus, Orectolobus sp., and Galeocerdo cuvier) produced remarkably high numbers of polymorphic loci (106, 94, and 86, respectively) from a limited sample size of only 2. A major constraint to obtaining much needed genetic data from sharks is the time-consuming process of developing molecular markers. Here we demonstrate the general utility of a technique that provides large numbers of informative loci in sharks.

Molecular identification of Mango, Mangifera indica L.var. totupura

PubMed Central

Jagarlamudi, Sankar; G, Rosaiah; Kurapati, Ravi Kumar; Pinnamaneni, Rajasekhar

2011-01-01

Mango (>Mangifera indica) belonging to Anacardiaceae family is a fruit that grows in tropical regions. It is considered as the King of fruits. The present work was taken up to identify a tool in identifying the mango species at the molecular level. The chloroplast trnL-F region was amplified from extracted total genomic DNA using the polymerase chain reaction (PCR) and sequenced. Sequence of the dominant DGGE band revealed that Mangifera indica in tested leaves was Mangifera indica (100% similarity to the ITS sequences of Mangifera indica). This sequence was deposited in NCBI with the accession no. GQ927757. Abbreviations AFLP - Amplified fragment length polymorphism , cpDNA - Chloroplast DNA, DDGE - Denaturing gradient gel electrophoresis, DNA - Deoxyribo nucleic acid, EDTA - Ethylenediamine tetraacetic acid, HCl - Hydrochloric acid, ISSR - Inter simple sequence repeats, ITS - Internal transcribed spacer, MATAB - Methyl Ammonium Bromide, Na2SO3 - Sodium sulphite, NaCl - Sodium chloride, NCBI - National Centre for Biotechnology Information, PCR - Polymerase chain reaction, PEG - Polyethylene glycol, RAPD - Randomly amplified polymorphic DNA, trnL-F - Transfer RNA genes start codon- termination codon. PMID:21423885

Genetic similarity analysis within Pyropia yezoensis blades developed from both conchospores and blade archeospores using AFLP(1).

PubMed

He, Linwen; Zhu, Jianyi; Lu, Qinqin; Niu, Jianfeng; Zhang, Baoyu; Lin, Apeng; Wang, Guangce

2013-06-01

Pyropia yezoensis (Ueda) M. S. Hwang et H. G. Choi (previously called Porphyra yezoensis) is an economically important alga. The blades generated from conchospores are genetic chimeras, which are not suitable for genetic similarity analysis. In this study, two types of blades from a single filament of P. yezoensis sporophyte filament were obtained. One type, ConB, consisted of 40 blades that had germinated from conchospores. The other type, ArcB, consisted of 88 blades that had germinated from archeospores released from ConB. Both of them were analyzed by amplified fragment length polymorphism. The low genetic similarity levels for both conchospore-germinated and archeospore-germinated blades demonstrated that the conchcelis we used was cross-fertilized. Furthermore, a higher polymorphic loci ratio (98.6%) was detected in ArcB than in ConB (80.7%), and the average genetic similarity of ArcB (average 0.61) was lower than that of ConB (average 0.71). These differences indicated that genetic analysis using ArcB gives more accurate results. © 2013 Phycological Society of America.

Significant population genetic structure detected in the rock bream Oplegnathus fasciatus (Temminck & Schlegel, 1844) inferred from fluorescent-AFLP analysis

NASA Astrophysics Data System (ADS)

Xiao, Yongshuang; Ma, Daoyuan; Xu, Shihong; Liu, Qinghua; Wang, Yanfeng; Xiao, Zhizhong; Li, Jun

2016-05-01

Oplegnathus fasciatus (rock bream) is a commercial rocky reef fish species in East Asia that has been considered for aquaculture. We estimated the population genetic diversity and population structure of the species along the coastal waters of China using fluorescent-amplified fragment length polymorphisms technology. Using 53 individuals from three populations and four pairs of selective primers, we amplified 1 264 bands, 98.73% of which were polymorphic. The Zhoushan population showed the highest Nei's genetic diversity and Shannon genetic diversity. The results of analysis of molecular variance (AMOVA) showed that 59.55% of genetic variation existed among populations and 40.45% occurred within populations, which indicated that a significant population genetic structure existed in the species. The pairwise fixation index F st ranged from 0.20 to 0.63 and were significant after sequential Bonferroni correction. The topology of an unweighted pair group method with arithmetic mean tree showed two significant genealogical branches corresponding to the sampling locations of North and South China. The AMOVA and STRUCTURE analyses suggested that the O. fasciatus populations examined should comprise two stocks.

AFLP marking and polymorphism among progenies of Gymnema sylvestre: an important medicinal plant of India.

PubMed

Osman, Magda Abbaker; Dhawan, Sunita Singh; Bahl, Janak Raj; Darokar, Mahendra P; Khanuja, Suman P S

2011-11-01

The level of polymorphism among twelve selected progenies of Gymnema sylvestre was investigated through AFLP markers by multiplexing PCR reactions using 64 (8x8) primer combinations. Fourteen primer combinations were selected as the most suitable combination for G. sylvestre. Analysis of the 12 progenies with these 14 primer pairs produced 1689 fragments of which 972 (57.5%) were polymorphic and 485 (28.7%) were unique to a particular genotype. The number of fragments produced by individual primer pairs was in the range of 55 to 225. Out of these, polymorphic fragments were in the range of 34 (E-ACC/M-CAC) to 157 (E-AGG/M-CAG) and unique bands observed were 8 (E-ACC / M-CAC) to 69 (E-AGG/M-CAC). Different primer combinations detected different levels of polymorphism, ranging from 33% (E-AGG/ M-CAC) to 69.8% (E-AGG/ M-CAC). From the observations, it appears that the primer combinations E-AGG/M-CAC, E-AGG/CTG, E-AGG/CAG and E-ACA/CAT were the most informative for the detection of polymorphism among the progenies compared with others, since they produced a high number of unique fragments. The similarity coefficient ranged from 0.212 to 0.731. High similarity was observed between progeny S8 and S9 (73%) and high divergence between progenies S3 and S11. Among the selected progeny, S9 was found to be the most similar to the parent (63%), while genotype S11 was the most distant (36.9%).

Analysis of DNA Methylation of Gracilariopsis lemaneiformis Under Temperature Stress Using the Methylation Sensitive Amplification Polymorphism (MSAP) Technique

NASA Astrophysics Data System (ADS)

Peng, Chong; Sui, Zhenghong; Zhou, Wei; Hu, Yiyi; Mi, Ping; Jiang, Minjie; Li, Xiaodong; Ruan, Xudong

2018-06-01

Gracilariopsis lemaneiformis is an economically important agarophyte, which contains high quality gel and shows a high growth rate. Wild population of G. lemaneiformis displayed resident divergence, though with a low genetic diversity as was revealed by amplified fragment length polymorphism (AFLP) and simple sequence repeat (SSR) analyses. In addition, different strains of G. lemaneiformis are diverse in morphology. The highly inconsistence between genetic background and physiological characteristics recommends strongly to the regulation at epigenetic level. In this study, the DNA methylation change in G. lemaneiformis among different generation branches and under different temperature stresses was assessed using methylation sensitive amplified polymorphism (MSAP) technique. It was shown that DNA methylation level among different generation branches was diverse. The full and total methylated DNA level was the lowest in the second generation branch and the highest in the third generation. The total methylation level was 61.11%, 60.88% and 64.12% at 15°C, 22°C and 26°C, respectively. Compared with the control group (22°C), the fully methylated and totally methylated ratios were increased in both experiment groups (15°C and 26°C). All of the cytosine methylation/demethylation transform (CMDT) was further analyzed. High temperature treatment could induce more CMDT than low temperature treatment did.

Lactobacillus delbrueckii subsp. sunkii subsp. nov., isolated from sunki, a traditional Japanese pickle.

PubMed

Kudo, Yuko; Oki, Kaihei; Watanabe, Koichi

2012-11-01

Although four strains of bacteria isolated from sunki, a traditional Japanese, non-salted pickle, were initially identified as Lactobacillus delbrueckii, the molecular and phenotypic characteristics of the strains did not match those of any of the four recognized subspecies of L. delbrueckii. Together, the results of phenotypic characterization, DNA-DNA hybridizations (in which the relatedness values between the novel strains and type strains of the recognized subspecies of L. delbrueckii were all >88.7%) and 16S rRNA gene sequence, amplified fragment length polymorphism (AFLP) and whole-cell MALDI-TOF/MS spectral pattern analyses indicated that the four novel strains represented a single, novel subspecies, for which the name Lactobacillus delbrueckii subsp. sunkii subsp. nov. is proposed. The type strain is YIT 11221(T) (=JCM 17838(T) =DSM 24966(T)).

Global Spread of Human Chromoblastomycosis Is Driven by Recombinant Cladophialophora carrionii and Predominantly Clonal Fonsecaea Species.

PubMed

Deng, Shuwen; Tsui, Clement K M; Gerrits van den Ende, A H G; Yang, Liyue; Najafzadeh, Mohammad Javad; Badali, Hamid; Li, Ruoyu; Hagen, Ferry; Meis, Jacques F; Sun, Jiufeng; Dolatabadi, Somayeh; Papierok, Bernard; Pan, Weihua; de Hoog, G S; Liao, Wanqing

2015-01-01

Global distribution patterns of Cladophialophora carrionii, agent of human chromoblastomycosis in arid climates of Africa, Asia, Australia, Central-and South-America, were compared with similar data of the vicarious Fonsecaea spp., agents of the disease in tropical rain forests. Population diversities among 73 C. carrionii strains and 60 strains of three Fonsecaea species were analyzed for rDNA ITS, partial β-tubulin, and amplified fragment-length polymorphism (AFLP) fingerprints. Populations differed significantly between continents. Lowest haplotype diversity was found in South American populations, while African strains were the most diverse. Gene flow was noted between the African population and all other continents. The general pattern of Fonsecaea agents of chromoblastomycosis differed significantly from that of C. carrionii and revealed deeper divergence among three differentiated species with smaller numbers of haplotypes, indicating a longer evolutionary history.

An outbreak of an unusual strain of Listeria monocytogenes infection in North-East Scotland.

PubMed

Okpo, Emmanuel; Leith, Jayne; Smith-Palmer, Alison; Bell, John; Parks, Duncan; Browning, Fiona; Byers, Lynn; Corrigan, Helen; Webster, Diana; Karcher, Anne M; Murray, Andrew; Storey, Tom

2015-01-01

Listeria monocytogenes infection is an important cause of illness and hospitalization in vulnerable individuals. In the present study, we describe a community outbreak of Listeria monocytogenes in the North-East region of Scotland, which was epidemiologically, environmentally and microbiologically linked to a local meat product and ready-to-eat product manufacturer. Infected individuals were interviewed, and an environmental investigation was conducted. Clinical and environmental samples were tested by culture, and isolates were typed by fluorescent amplified fragment length polymorphism (fAFLP). Three cases of Listeria monocytogenes were linked geographically, had the same serotype (1/2a) and were indistinguishable by fAFLP type XII.6. The human, food and environmental isolates were of the same serotype and were indistinguishable by molecular typing. This is the first community outbreak of L. monocytogenes reported in Scotland since the current outbreak surveillance was established in 1996. Epidemiological and laboratory evidence indicated poor hand hygiene, unhygienic practices and cross-contamination throughout the manufacturing process of ready-to-eat foods as a possible cause of the outbreak. More stringent control of commercial food establishments that provide ready-to-eat food and the need to advise specifically vulnerable groups, e.g., pregnant women, of the risk of L. monocytogenes in ready-to-eat food is urgently needed. Copyright © 2015 King Saud Bin Abdulaziz University for Health Sciences. Published by Elsevier Ltd. All rights reserved.

Multilocus sequence typing reveals a novel subspeciation of Lactobacillus delbrueckii.

PubMed

Tanigawa, Kana; Watanabe, Koichi

2011-03-01

Currently, the species Lactobacillus delbrueckii is divided into four subspecies, L. delbrueckii subsp. delbrueckii, L. delbrueckii subsp. bulgaricus, L. delbrueckii subsp. indicus and L. delbrueckii subsp. lactis. These classifications were based mainly on phenotypic identification methods and few studies have used genotypic identification methods. As a result, these subspecies have not yet been reliably delineated. In this study, the four subspecies of L. delbrueckii were discriminated by phenotype and by genotypic identification [amplified-fragment length polymorphism (AFLP) and multilocus sequence typing (MLST)] methods. The MLST method developed here was based on the analysis of seven housekeeping genes (fusA, gyrB, hsp60, ileS, pyrG, recA and recG). The MLST method had good discriminatory ability: the 41 strains of L. delbrueckii examined were divided into 34 sequence types, with 29 sequence types represented by only a single strain. The sequence types were divided into eight groups. These groups could be discriminated as representing different subspecies. The results of the AFLP and MLST analyses were consistent. The type strain of L. delbrueckii subsp. delbrueckii, YIT 0080(T), was clearly discriminated from the other strains currently classified as members of this subspecies, which were located close to strains of L. delbrueckii subsp. lactis. The MLST scheme developed in this study should be a useful tool for the identification of strains of L. delbrueckii to the subspecies level.

Infection and colonisation due to Scedosporium in Northern Spain. An in vitro antifungal susceptibility and molecular epidemiology study of 60 isolates.

PubMed

Lackner, M; Rezusta, A; Villuendas, M C; Palacian, M P; Meis, J F; Klaassen, C H

2011-10-01

Since the latest taxonomical changes in the genus Scedosporium by Gilgado et al. in 2010, no species-specific studies on epidemiology and antifungal susceptibility patterns (AFSP) have so far been published. This study aimed to provide qualitative epidemiological data of Scedosporium spp. isolated from cystic fibrosis (CF) patients and immunocompromised patients from Northern Spain. Isolates were identified by using amplified fragment length polymorphism (AFLP), and species-specific AFSP were generated for all currently available antifungal compounds. AFLP was a useful tool for identification to species-level and for the discrimination of inter- and intra-patient isolates. Scedosporium prolificans represents the most prevalent species in the respiratory tract of CF patients and immunocompromised patients in Northern-Spain, followed by Pseudallescheria boydii, P. apiosperma, and P. ellipsoidea. CF patients were exclusively colonised with either P. boydii or S. prolificans. Patients were colonised over years exclusively with isolates affiliated to one species, but some patients were colonised with multiple strains with different AFSP. The sum of those co-colonising strains in one patient, may appear in vitro and in vivo as a multi-resistant S. prolificans isolate, as strains are morphologically identical and might therefore be regarded as only one strain. A majority of Scedosporium strains (with exception of S. prolificans) were found susceptible for voriconazole and micafungin. © 2011 Blackwell Verlag GmbH.

First report of Candida auris in America: Clinical and microbiological aspects of 18 episodes of candidemia.

PubMed

Calvo, Belinda; Melo, Analy S A; Perozo-Mena, Armindo; Hernandez, Martin; Francisco, Elaine Cristina; Hagen, Ferry; Meis, Jacques F; Colombo, Arnaldo Lopes

2016-10-01

Characterization of a hospital outbreak of Candida auris candidemia that involved 18 critically ill patients in Venezuela. Bloodstream isolates of C. auris obtained from 18 patients admitted at a medical center in Maracaibo, between March, 2012 and July, 2013 were included. Species identification was confirmed by ITS rDNA sequencing. Isolates were subsequently typed by amplified fragment length polymorphism fingerprinting (AFLP). Susceptibility testing was performed according to CLSI. Clinical data were collected from all cases by using a standard clinical form. A total of 13 critically ill pediatric and 5 adult patients, with a median age of 26 days, were included. All were previously exposed to antibiotics and multiple invasive medical procedures. Clinical management included prompt catheter removal and antifungal therapy. Thirteen patients (72%) survived up to 30 days after onset of candidemia. AFLP fingerprinting of all C. auris isolates suggested a clonal outbreak. The isolates were considered resistant to azoles, but susceptible to anidulafungin and 50% of isolates exhibited amphotericin B MIC values of >1 μg/ml. The study demonstrated that C. auris is a multiresistant yeast pathogen that can be a source of health-care associated infections in tertiary care hospitals with a high potential for nosocomial horizontal transmission. Copyright © 2016 The British Infection Association. Published by Elsevier Ltd. All rights reserved.

Molecular and antimicrobial susceptibility characterization of Globicatella sulfidifaciens isolated from sow's urinary tract infection.

PubMed

Matajira, Carlos E C; Moreno, Luisa Z; Gomes, Vasco T M; Silva, Ana Paula S; Mesquita, Renan E; Amigo, Cristina R; Christ, Ana Paula G; Sato, Maria Inês Z; Moreno, Andrea M

2017-12-01

The Globicatella genus comprises Gram-positive, facultative anaerobic, α-hemolytic and catalase negative cocci morphologically and phenotypically very similar to Streptococcus and Aerococcus genus which can lead to misidentification and underestimation of this pathogen. Globicatella species have already been isolated from human and animals with heart and brain disorders. Their clinical relevance in animals, and its zoonotic potential, remains unknown due to the difficulty in their identification. To present the isolation, phenotypic and molecular characterization of G. sulfidifaciens from urinary tract infection in sows. Urine samples from 140 sows of two swine herds located in São Paulo State (Brazil) yielded the isolation of three presumptive G. sulfidifaciens strains. Identification and species confirmation were done by MALDI-TOF MS and 16S rRNA sequencing. Strains were further characterized by single enzyme amplified fragments length polymorphism (SE-AFLP) and broth microdilution techniques. All three isolates were confirmed as G. sulfidifaciens. The SE-AFLP genotyping resulted in distinct fingerprint patterns for each strain. All isolates presented high MIC values to tetracycline, sulphonamides, aminoglycosides and tylosin tartrate, which present high usage in human and animal medicine. Globicatella sulfidifaciens could be related to sporadic urinary tract infections in swine and appear to present alarming antimicrobial susceptibility profile. It is necessary to differentiate Streptococcus-like microorganisms in routine laboratory diagnostics for the correct identification of underestimated species potentially pathogenic to animals.

Genetic variation in the endangered Southwestern Willow Flycatcher

USGS Publications Warehouse

Busch, Joseph; Miller, Mark P.; Paxton, E.H.; Sogge, M.K.; Keim, Paul

2000-01-01

The Southwestern Willow Flycatcher (Empidonax traillii extimus) is an endangered Neotropical migrant that breeds in isolated remnants of dense riparian habitat in the southwestern United States. We estimated genetic variation at 20 breeding sites of the Southwestern Willow Flycatcher (290 individuals) using 38 amplified fragment length polymorphisms (AFLPs). Our results suggest that considerable genetic diversity exists within the subspecies and within local breeding sites. Statistical analyses of genetic variation revealed only slight, although significant, differentiation among breeding sites (Mantel's r = 0.0705, P < 0.0005; θ = 0.0816, 95% CI = 0.0608 to 0.1034; ΦST = 0.0458, P < 0.001). UPGMA cluster analysis of the AFLP markers indicates that extensive gene flow has occurred among breeding sites. No one site stood out as being genetically unique or isolated. Therefore, the small level of genetic structure that we detected may not be biologically significant. Ongoing field studies are consistent with this conclusion. Of the banded birds that were resighted or recaptured in Arizona during the 1996 to 1998 breeding seasons, one-third moved between breeding sites and two-thirds were philopatric. Low differentiation may be the result of historically high rangewide diversity followed by recent geographic isolation of breeding sites, although observational data indicate that gene flow is a current phenomenon. Our data suggest that breeding groups of E. t. extimus act as a metapopulation.

Tracking genes of ecological relevance using a genome scan in two independent regional population samples of Arabis alpina.

PubMed

Poncet, Bénédicte N; Herrmann, Doris; Gugerli, Felix; Taberlet, Pierre; Holderegger, Rolf; Gielly, Ludovic; Rioux, Delphine; Thuiller, Wilfried; Aubert, Serge; Manel, Stéphanie

2010-07-01

Understanding the genetic basis of adaptation in response to environmental variation is fundamental as adaptation plays a key role in the extension of ecological niches to marginal habitats and in ecological speciation. Based on the assumption that some genomic markers are correlated to environmental variables, we aimed to detect loci of ecological relevance in the alpine plant Arabis alpina L. sampled in two regions, the French (99 locations) and the Swiss (109 locations) Alps. We used an unusually large genome scan [825 amplified fragment length polymorphism loci (AFLPs)] and four environmental variables related to temperature, precipitation and topography. We detected linkage disequilibrium among only 3.5% of the considered AFLP loci. A population structure analysis identified no admixture in the study regions, and the French and Swiss Alps were differentiated and therefore could be considered as two independent regions. We applied generalized estimating equations (GEE) to detect ecologically relevant loci separately in the French and Swiss Alps. We identified 78 loci of ecological relevance (9%), which were mainly related to mean annual minimum temperature. Only four of these loci were common across the French and Swiss Alps. Finally, we discuss that the genomic characterization of these ecologically relevant loci, as identified in this study, opens up new perspectives for studying functional ecology in A. alpina, its relatives and other alpine plant species.

Dynamics in the tomato root transcriptome on infection with the potato cyst nematode Globodera rostochiensis.

PubMed

Swiecicka, Magdalena; Filipecki, Marcin; Lont, Dieuwertje; Van Vliet, Joke; Qin, Ling; Goverse, Aska; Bakker, Jaap; Helder, Johannes

2009-07-01

Plant parasitic nematodes infect roots and trigger the formation of specialized feeding sites by substantial reprogramming of the developmental process of root cells. In this article, we describe the dynamic changes in the tomato root transcriptome during early interactions with the potato cyst nematode Globodera rostochiensis. Using amplified fragment length polymorphism-based mRNA fingerprinting (cDNA-AFLP), we monitored 17 600 transcript-derived fragments (TDFs) in infected and uninfected tomato roots, 1-14 days after inoculation with nematode larvae. Six hundred and twenty-four TDFs (3.5%) showed significant differential expression on nematode infection. We employed GenEST, a computer program which links gene expression profiles generated by cDNA-AFLP and databases of cDNA sequences, to identify 135 tomato sequences. These sequences were grouped into eight functional categories based on the presence of genes involved in hormone regulation, plant pathogen defence response, cell cycle and cytoskeleton regulation, cell wall modification, cellular signalling, transcriptional regulation, primary metabolism and allocation. The presence of unclassified genes was also taken into consideration. This article describes the responsiveness of numerous tomato genes hitherto uncharacterized during infection with endoparasitic cyst nematodes. The analysis of transcriptome profiles allowed the sequential order of expression to be dissected for many groups of genes and the genes to be connected with the biological processes involved in compatible interactions between the plant and nematode.

CHOICE OF METHODOLOGY FOR ASSESSING GENETIC IMPACTS OF ENVIRONMENTAL STRESSORS: POLYMORPHISM AND REPRODUCIBILITY OF RAPD AND AFLP FINGERPRINTS. (R826603)

EPA Science Inventory

The perspectives, information and conclusions conveyed in research project abstracts, progress reports, final reports, journal abstracts and journal publications convey the viewpoints of the principal investigator and may not represent the views and policies of ORD and EPA. Concl...

Identification and characterization of the WRKY transcription factor family in Pinus monticola.

PubMed

Liu, Jun-Jun; Ekramoddoullah, Abul K M

2009-01-01

The WRKY gene family represents an ancient and highly complex group of transcription factors involved in signal transduction pathways of numerous plant developmental processes and host defense response. Up to now, most WRKY proteins have been identified in a few angiosperm species. Identification of WRKY genes in a conifer species would facilitate a comprehensive understanding of the evolutionary and function-adaptive process of this superfamily in plants. We performed PCR on genomic DNA to clone WRKY sequences from western white pine (Pinus monticola), one of the most valuable conifer species endangered by white pine blister rust (Cronartium ribicola). In total, 83 P. monticola WRKY (PmWRKY) sequences were identified using degenerate primers targeted to the WRKY domain. A phylogenetic analysis revealed that PmWRKY members fell into four major groups (1, 2a+2b, 2c, and 2d+2e) described in Arabidopsis and rice. Because of high genetic diversity of the PmWRKY family, a modified AFLP method was used to detect DNA polymorphism of this gene family. Polymorphic fragments accounted for 17%-35% of total PCR products in the AFLP profiles. Among them, one WRKY AFLP marker was linked to the major resistance gene (Cr2) against C. ribicola. The results of this study provide basic genomic information for a conifer WRKY gene family, which will pave the way for elucidating gene evolutionary mechanisms in plants and unveiling the precise roles of PmWRKY in conifer development and defense response.

A preliminary study for identification of candidate AFLP markers under artificial selection for shell color in pearl oyster Pinctada fucata.

PubMed

Zou, Keshu; Zhang, Dianchang; Guo, Huayang; Zhu, Caiyan; Li, Min; Jiang, Shigui

2014-05-25

Pearl oyster Pinctada fucata is widely cultured to produce seawater pearl in South China, and the quality of pearl is significantly affected by its shell color. Thus the Pearl Oyster Selective Breeding Program (POSBP) was carried out for the shell color and growth traits. The black (B), gold (G), red (R) and white (W) shell strains with fast growth trait were achieved after five successive generation selection. In this study, AFLP technique was used to scan genome of four strains with different shell colors to identify the candidate markers under artificial selection. Eight AFLP primer combinations were screened and yielded 688 loci, 676 (98.26%) of which were polymorphic. In black, gold, red and white strains, the percentage of polymorphic loci was 90.41%, 87.79%, 93.60% and 93.31%, respectively, Nei's gene diversity was 0.3225, 0.2829, 0.3221 and 0.3292, Shannon's information index was 0.4801, 0.4271, 0.4825 and 0.4923, and the value of FST was 0.1805. These results suggested that the four different shell color strains had high genetic diversity and great genetic differentiation among strains, which had been subjected to the continuous selective pressures during the artificial selective breeding. Furthermore, six outlier loci were considered as the candidate markers under artificial selection for shell color. This study provides a molecular evidence for the inheritance of shell color of P. fucata. Copyright © 2014 Elsevier B.V. All rights reserved.

High-density linkage mapping revealed suppression of recombination at the sex determination locus in papaya.

PubMed Central

Ma, Hao; Moore, Paul H; Liu, Zhiyong; Kim, Minna S; Yu, Qingyi; Fitch, Maureen M M; Sekioka, Terry; Paterson, Andrew H; Ming, Ray

2004-01-01

A high-density genetic map of papaya (Carica papaya L.) was constructed using 54 F(2) plants derived from cultivars Kapoho and SunUp with 1501 markers, including 1498 amplified fragment length polymorphism (AFLP) markers, the papaya ringspot virus coat protein marker, morphological sex type, and fruit flesh color. These markers were mapped into 12 linkage groups at a LOD score of 5.0 and recombination frequency of 0.25. The 12 major linkage groups covered a total length of 3294.2 cM, with an average distance of 2.2 cM between adjacent markers. This map revealed severe suppression of recombination around the sex determination locus with a total of 225 markers cosegregating with sex types. The cytosine bases were highly methylated in this region on the basis of the distribution of methylation-sensitive and -insensitive markers. This high-density genetic map is essential for cloning of specific genes of interest such as the sex determination gene and for the integration of genetic and physical maps of papaya. PMID:15020433

Identification of AFLP markers linked to fertility restorer genes for tournefortii cytoplasmic male-sterility system in Brassica napus.

PubMed

Janeja, H S; Banga, S S; Lakshmikumaran, M

2003-06-01

The tournefortii cytoplasmic male-sterility system is being used as a method of pollination control to develop hybrids in Brassica napus. Genetic analyses have indicated that two dominant genes, one major ( Rft1) and another minor ( Rft2), were required to achieve complete fertility restoration. Though the major gene ( Rft1) can cause complete fertility restoration on its own, its expression was significantly enhanced in the presence of the minor gene ( Rft2). In the absence of Rft1, Rft2 caused only partial fertility restoration. We used a pair of near-isogenic lines (NILs), differing for the presence/absence of Rf genes, to identify AFLP markers linked to fertility restorer genes. A total of 64 EcoRI/ MseI primer combinations were surveyed which produced 3,225 bands, of which 19 (0.006%) were polymorphic between parental NILs. Primer combinations which led to the identification of polymorphic bands present in fertile parental NILs were used for assaying a mapping population of 70 F(2) plants for determining the segregation pattern of markers. Initial screening resulted in the identification of five AFLP markers. The recombination analyses of these AFLP markers revealed that at least two (EACC/MCTT(105), EAAG/MCTC(80)) were present in the same linkage group along with the Rf loci. Marker EACC/MCTT(105) was separated from the major gene ( Rft1) by a distance of 18.1 cM, while it was 33.2 cM away from the minor fertility restorer gene ( Rft2). Another marker EAAG/MCTC(80) was also located adjacent to Rft1 at a distance of 18.1 cM, but on other side. Identification of flanking markers (EACC/MCTT(105), EAAG/MCTC(80)) for the major fertility restorer gene ( Rft1) provides a crucial component for marker-assisted selection and map-based cloning of the restorer genes, and can hence be used to construct elite restorer genotypes.

Suitability of Cryopreservation for the Long‐term Storage of Rare and Endangered Plant Species: a Case History for Cosmos atrosanguineus

PubMed Central

WILKINSON, TIM; WETTEN, ANDREW; PRYCHID, CHRISSIE; FAY, MICHAEL F.

2003-01-01

The suitability of cryopreservation for the secure, long‐term storage of the rare and endangered species Cosmos atrosanguineus was investigated. Using encapsulation/dehydration of shoot tips in alginate strips, survival rates of up to 100 % and shoot regeneration of up to 35 % were achieved. Light and electron microscopy studies indicated that cellular damage to some regions of the shoot tip during the freeze/thaw procedure was high, although cell survival in and around the meristematic region allowed shoot tip regeneration. The genetic fingerprinting technique, amplified fragment length polymorphisms (AFLPs), showed that no detectable genetic variation was present between material of C. atrosanguineus at the time of initiation into tissue culture and that which had been cryopreserved, stored in liquid nitrogen for 12 months and regenerated. Weaned plantlets that were grown under glasshouse conditions exhibited no morphological variation from non‐frozen controls. PMID:12495921

Genetic Diversity and Genetic Relationships of Purple Willow (Salix purpurea L.) from Natural Locations

PubMed Central

Prinz, Kathleen; Przyborowski, Jerzy A.

2017-01-01

In this study, the genetic diversity and structure of 13 natural locations of Salix purpurea were determined with the use of AFLP (amplified length polymorphism), RAPD (randomly amplified polymorphic DNA) and ISSR (inter-simple sequence repeats). The genetic relationships between 91 examined S. purpurea genotypes were evaluated by analyses of molecular variance (AMOVA), principal coordinates analyses (PCoA) and UPGMA (unweighted pair group method with arithmetic mean) dendrograms for both single marker types and a combination of all marker systems. The locations were assigned to distinct regions and the analysis of AMOVA (analysis of molecular variance) revealed a high genetic diversity within locations. The genetic diversity between both regions and locations was relatively low, but typical for many woody plant species. The results noted for the analyzed marker types were generally comparable with few differences in the genetic relationships among S. purpurea locations. A combination of several marker systems could thus be ideally suited to understand genetic diversity patterns of the species. This study makes the first attempt to broaden our knowledge of the genetic parameters of the purple willow (S. purpurea) from natural location for research and several applications, inter alia breeding purposes. PMID:29301207

High-resolution melt analysis to identify and map sequence-tagged site anchor points onto linkage maps: a white lupin (Lupinus albus) map as an exemplar.

PubMed

Croxford, Adam E; Rogers, Tom; Caligari, Peter D S; Wilkinson, Michael J

2008-01-01

* The provision of sequence-tagged site (STS) anchor points allows meaningful comparisons between mapping studies but can be a time-consuming process for nonmodel species or orphan crops. * Here, the first use of high-resolution melt analysis (HRM) to generate STS markers for use in linkage mapping is described. This strategy is rapid and low-cost, and circumvents the need for labelled primers or amplicon fractionation. * Using white lupin (Lupinus albus, x = 25) as a case study, HRM analysis was applied to identify 91 polymorphic markers from expressed sequence tag (EST)-derived and genomic libraries. Of these, 77 generated STS anchor points in the first fully resolved linkage map of the species. The map also included 230 amplified fragment length polymorphisms (AFLP) loci, spanned 1916 cM (84.2% coverage) and divided into the expected 25 linkage groups. * Quantitative trait loci (QTL) analyses performed on the population revealed genomic regions associated with several traits, including the agronomically important time to flowering (tf), alkaloid synthesis and stem height (Ph). Use of HRM-STS markers also allowed us to make direct comparisons between our map and that of the related crop, Lupinus angustifolius, based on the conversion of RFLP, microsatellite and single nucleotide polymorphism (SNP) markers into HRM markers.

Spaceflight-induced genetic and epigenetic changes in the rice (Oryza sativa L.) genome are independent of each other.

PubMed

Ou, Xiufang; Long, Likun; Wu, Ying; Yu, Yingjie; Lin, Xiuyun; Qi, Xin; Liu, Bao

2010-07-01

An array of studies have reported that the spaceflight environment is mutagenic and may induce phenotypic and genetic changes in diverse organisms. We reported recently that in at least some plant species (e.g., rice) the spaceflight environment can be particularly potent in generating heritable epigenetic changes in the form of altered cytosine methylation patterns and activation of transposable elements. To further study the issue of spaceflight-induced genomic instability, and in particular to test whether the incurred genetic and epigenetic changes are connected or independent of each other, we performed the present study. We subjected seeds of the standard laboratory rice (Oryza sativa L.) cultivar Nipponbare to a spaceflight in the spaceship Long March 2 for 18 days. We then investigated the genetic and DNA methylation stabilities of 11 randomly selected plants germinated from the spaceflown seeds by using two kinds of DNA markers, amplified fragment length polymorphism (AFLP) and methylation sensitive amplified polymorphism (MSAP). For AFLP, by using 15 primer combinations, we assessed 460 genomic loci and found that the frequencies of genetic changes across the 11 plants ranged from 0.7% to 6.7% with an average frequency of 3.5%. For MSAP, by using 14 primer combinations, we assessed 467 loci and detected the occurrence of four major types of cytosine methylation alterations at the CCGG sites, namely CG or CNG hypomethylation and CG or CNG hypermethylation. Collectively, the frequencies of the two kinds of hypermethylation, CG (1.95%) and CNG (1.44%), are about two times higher than those of the two kinds of hypomethylation, CG (0.76%) and CNG (0.80%), though different plants showed variable frequencies for each type of alteration. Further analysis suggested that both the genetic and cytosine methylation changes manifested apparent mutational bias towards specific genomic regions, but the two kinds of instabilities are independent of each other based on correlation analysis.

Evidence for progenitor–derivative speciation in sexually deceptive orchids

PubMed Central

Schlüter, Philipp M.; Ruas, Paulo M.; Kohl, Gudrun; Ruas, Claudete F.; Stuessy, Tod F.; Paulus, Hannes F.

2011-01-01

Background and Aims Sexually deceptive orchids of the genus Ophrys use mimicry of pollinator females to attract specific pollinators. Pollinator shifts may drive speciation in Ophrys, since novel pollinators may in principle act as isolating factors immediately. It is thus possible that evolution of novel species occurs rapidly and with a progenitor–derivative pattern. The aims of this study are to compare genetic structure and diversity among widespread and geographically restricted Ophrys taxa, to test whether genetic structure is associated with specific pollinators, and to investigate whether any widespread species may have acted as a progenitor for the evolution of more restricted taxa. Methods Genetic differentiation and diversity were investigated in O. leucadica and O. cinereophila, the two taxa of the Ophrys fusca sensu lato complex widespread in the Aegean, and three geographically restricted taxa from Rhodes, O. attaviria, O. parvula and O. persephonae, all differing in their specific pollinators. This was done using amplified fragment length polymorphism (AFLP) DNA fingerprinting, and sequencing of the low-copy nuclear gene LEAFY (LFY). Key Results All taxa were found to be separate genetic entities, with O. leucadica forming two geographic groups from the west and east of the Aegean. Genetic structure was significantly shaped by pollinators and geography, and comparison of sequence and AFLP data revealed ancestral polymorphisms shared among several taxa. Among the sampled taxa, O. leucadica harbours the greatest genetic differentiation and geographic structure, and the highest genetic diversity. Part of the genome of O. parvula, endemic to Rhodes, may be derived from O. leucadica. Conclusions Pollinators probably influence the genetic structure of the investigated Ophrys species. The genetic pattern identified is consistent with O. leucadica being the oldest of the sampled taxa, making O. leucadica a candidate progenitor species from which more restricted taxa such as O. parvula may have evolved. PMID:21890487

On the origins of Balkan endemics: the complex evolutionary history of the Cyanus napulifer group (Asteraceae).

PubMed

Olšavská, Katarína; Slovák, Marek; Marhold, Karol; Štubňová, Eliška; Kučera, Jaromír

2016-11-01

The Balkan Peninsula is one of the most important centres of plant diversity in Europe. Here we aim to fill the gap in the current knowledge of the evolutionary processes and factors modelling this astonishing biological richness by applying multiple approaches to the Cyanus napulifer group. To reconstruct the mode of diversification within the C. napulifer group and to uncover its relationships with potential relatives with x = 10 from Europe and Northern Africa, we examined variation in genetic markers (amplified fragment length polymorphisms [AFLPs]; 460 individuals), relative DNA content (4',6-diamidino-2-phenylindole [DAPI] flow cytometry, 330 individuals) and morphology (multivariate morphometrics, 40 morphological characters, 710 individuals). To elucidate its evolutionary history, we analysed chloroplast DNA (cpDNA) sequences of the genus Cyanus deposited in the GenBank database. The AFLPs revealed a suite of closely related entities with variable levels of differentiation. The C. napulifer group formed a genetically well-defined unit. Samples outside the group formed strongly diversified and mostly species-specific genetic lineages with no further geographical patterns, often characterized also by a different DNA content. AFLP analysis of the C. napulifer group revealed extensive radiation and split it into nine allopatric (sub)lineages with varying degrees of congruence among genetic, DNA-content and morphological patterns. Genetic admixture was usually detected in contact zones between genetic lineages. Plastid data indicated extensive maintenance of ancestral variation across Cyanus perennials. The C. napulifer group is an example of a rapidly and recently diversified plant group whose genetic lineages have evolved in spatio-temporal isolation on the topographically complex Balkan Peninsula. Adaptive radiation, accompanied in some cases by long-term isolation and hybridization, has contributed to the formation of this species complex and its mosaic pattern. © The Author 2016. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Genetic and epigenetic divergence between disturbed and undisturbed subpopulations of a Mediterranean shrub: a 20-year field experiment.

PubMed

Herrera, Carlos M; Bazaga, Pilar

2016-06-01

Little is known on the potential of ecological disturbance to cause genetic and epigenetic changes in plant populations. We take advantage of a long-term field experiment initiated in 1986 to study the demography of the shrub Lavandula latifolia , and compare genetic and epigenetic characteristics of plants in two adjacent subplots, one experimentally disturbed and one left undisturbed, 20 years after disturbance. Experimental setup was comparable to an unreplicated 'Before-After-Control-Impact' (BACI) design where a single pair of perturbed and control areas were compared. When sampled in 2005, plants in the two subplots had roughly similar ages, but they had established in contrasting environments: dense conspecific population ('Undisturbed' subpopulation) versus open area with all conspecifics removed ('Disturbed' subpopulation). Plants were characterized genetically and epigenetically using amplified fragment length polymorphism (AFLP) and two classes of methylation-sensitive AFLP (MSAP) markers. Subpopulations were similar in genetic diversity but differed in epigenetic diversity and multilocus genetic and epigenetic characteristics. Epigenetic divergence between subpopulations was statistically unrelated to genetic divergence. Bayesian clustering revealed an abrupt linear boundary between subpopulations closely coincident with the arbitrary demarcation line between subplots drawn 20 years back, which supports that genetic and epigenetic divergence between subpopulations was caused by artificial disturbance. There was significant fine-scale spatial structuring of MSAP markers in both subpopulations, which in the Undisturbed one was indistinguishable from that of AFLP markers. Genetic differences between subpopulations could be explained by divergent selection alone, while the concerted action of divergent selection and disturbance-driven appearance of new methylation variants in the Disturbed subpopulation is proposed to explain epigenetic differences. This study provides the first empirical evidence to date suggesting that relatively mild disturbances could leave genetic and epigenetic signatures on the next adult generation of long-lived plants.

Breakthrough candidemia after the introduction of broad spectrum antifungal agents: A 5-year retrospective study.

PubMed

Breda, Giovanni L; Tuon, Felipe F; Meis, Jacques F; Herkert, Patricia F; Hagen, Ferry; de Oliveira, Letícia Z; Dias, Viviane de Carvalho; da Cunha, Clóvis Arns; Queiroz-Telles, Flávio

2018-06-01

Candidemia is the main invasive fungal disease among hospitalized patients. Several breakthrough candidemia (BrC) cases have been reported, but few studies evaluate the epidemiology, risk factors, molecular characterization, antifungal susceptibility profile and outcome of those patients, especially in developing countries and including patients using broad spectrum antifungals. We conducted a retrospective study from 2011 to 2016, including patients aged 12 years or older with candidemia. Epidemiological characteristics and risk factors for candidemia were evaluated and compared with patients with BrC using univariate and multivariate analysis. Sequential Candida isolates from BrC were identified by internal transcribed spacer sequencing, genotyped with amplified fragment length polymorphism fingerprinting (AFLP), and tested for antifungal susceptibility. From 148 candidemia episodes, 27 breakthrough episodes (18%) were identified, with neutropenia and mucositis being independent risk factors for BrC. Candida non-albicans was more frequent in the BrC group (P < .001). AFLP showed high correlation with conventional methods of identification among breakthrough isolates and a high genetic similarity among isolates from the same patient was observed. C. albicans was the most susceptible species with low MIC values for all antifungal agents tested. In contrast, we found isolates of C. glabrata, C. parapsilosis and C. tropicalis resistant to triazoles and echinocandins. In conclusion, BrC occurred mainly in severely immunosuppressed patients, with neutropenia and mucositis. Mortality did not differ between the groups. Candida non-albicans species were more recovered from BrC, with C. albicans being the most susceptible to antifungals.

Outbreak of Bacillus cereus infections in a neonatal intensive care unit traced to balloons used in manual ventilation.

PubMed

Van Der Zwet, W C; Parlevliet, G A; Savelkoul, P H; Stoof, J; Kaiser, A M; Van Furth, A M; Vandenbroucke-Grauls, C M

2000-11-01

In 1998, an outbreak of systemic infections caused by Bacillus cereus occurred in the Neonatal Intensive Care Unit of the University Hospital Vrije Universiteit, Amsterdam, The Netherlands. Three neonates developed sepsis with positive blood cultures. One neonate died, and the other two neonates recovered. An environmental survey, a prospective surveillance study of neonates, and a case control study were performed, in combination with molecular typing, in order to identify potential sources and transmission routes of infection. Genotypic fingerprinting by amplified-fragment length polymorphism (AFLP) showed that the three infections were caused by a single clonal type of B. cereus. The same strain was found in trachea aspirate specimens of 35 other neonates. The case control study showed mechanical ventilation with a Sensormedics ventilation machine to be a risk factor for colonization and/or infection (odds ratio, 9.8; 95% confidence interval, 1.1 to 88.2). Prospective surveillance showed that colonization with B. cereus occurred exclusively in the respiratory tract of mechanically ventilated neonates. The epidemic strain of B. cereus was found on the hands of nursing staff and in balloons used for manual ventilation. Sterilization of these balloons ended the outbreak. We conclude that B. cereus can cause outbreaks of severe opportunistic infection in neonates. Typing by AFLP proved very useful in the identification of the outbreak and in the analysis of strains recovered from the environment to trace the cause of the epidemic.

Outbreak of Bacillus cereus Infections in a Neonatal Intensive Care Unit Traced to Balloons Used in Manual Ventilation

PubMed Central

Van Der Zwet, Wil C.; Parlevliet, Gerard A.; Savelkoul, Paul H.; Stoof, Jeroen; Kaiser, Annie M.; Van Furth, A. Marceline; Vandenbroucke-Grauls, Christina M.

2000-01-01

In 1998, an outbreak of systemic infections caused by Bacillus cereus occurred in the Neonatal Intensive Care Unit of the University Hospital Vrije Universiteit, Amsterdam, The Netherlands. Three neonates developed sepsis with positive blood cultures. One neonate died, and the other two neonates recovered. An environmental survey, a prospective surveillance study of neonates, and a case control study were performed, in combination with molecular typing, in order to identify potential sources and transmission routes of infection. Genotypic fingerprinting by amplified-fragment length polymorphism (AFLP) showed that the three infections were caused by a single clonal type of B. cereus. The same strain was found in trachea aspirate specimens of 35 other neonates. The case control study showed mechanical ventilation with a Sensormedics ventilation machine to be a risk factor for colonization and/or infection (odds ratio, 9.8; 95% confidence interval, 1.1 to 88.2). Prospective surveillance showed that colonization with B. cereus occurred exclusively in the respiratory tract of mechanically ventilated neonates. The epidemic strain of B. cereus was found on the hands of nursing staff and in balloons used for manual ventilation. Sterilization of these balloons ended the outbreak. We conclude that B. cereus can cause outbreaks of severe opportunistic infection in neonates. Typing by AFLP proved very useful in the identification of the outbreak and in the analysis of strains recovered from the environment to trace the cause of the epidemic. PMID:11060080

Viability, purity, and genetic stability of entomopathogenic fungi species using different preservation methods.

PubMed

Ayala-Zermeño, Miguel A; Gallou, Adrien; Berlanga-Padilla, Angélica M; Andrade-Michel, Gilda Y; Rodríguez-Rodríguez, José C; Arredondo-Bernal, Hugo C; Montesinos-Matías, Roberto

2017-11-01

Preservation methods for entomopathogenic fungi (EPF) require effective protocols to ensure uniform processes and to avoid alterations during storage. The aim of this study was to preserve Beauveria bassiana, Metarhizium acridum, M. anisopliae, M. rileyi, Isaria javanica, Hirsutella thompsonii, H. citriformis and Lecanicillium lecanii in mineral oil (MO), sterile water (SW), silica gel (SG), lyophilisation (L), ultracold-freezing at -70 °C, and cryopreservation at -196 °C. The viability and purity of the fungi were then verified: phenotypic characteristics were evaluated qualitatively at 6, 12 and 24 m. Genetic stability was tested by amplified fragment length polymorphisms (AFLP) analysis at 24 m. Of the eight species of EPF, three remained viable in SW, five in MO and L, six at -70 °C, seven in SG, and eight at -196 °C. No significant changes were observed in AFLP patterns at 24 m of storage. The most effective preservation methods for EPF were SG, L, -70 and -196 °C. Beauveria bassiana, M. acridum, M. anisopliae, M. rileyi and I. javanica remained stable with all methods, while the remaining species were less compatible. The optimisation of preservation methods for EPF facilitates the development of reliable protocols to ensure their inherent characteristics in culture collections. Copyright © 2017 British Mycological Society. Published by Elsevier Ltd. All rights reserved.

An efficient cDNA-AFLP-based strategy for the identification of putative pathogenicity factors from the potato cyst nematode Globodera rostochiensis.

PubMed

Qin, L; Overmars, H; Helder, J; Popeijus, H; van der Voort, J R; Groenink, W; van Koert, P; Schots, A; Bakker, J; Smant, G

2000-08-01

A new strategy has been designed to identify putative pathogenicity factors from the dorsal or subventral esophageal glands of the potato cyst nematode Globodera rostochiensis. Three independent criteria were used for selection. First, genes of interest should predominantly be expressed in infective second-stage juveniles, and not, or to a far lesser extent, in younger developmental stages. For this, gene expression profiles from five different developmental stages were generated with cDNA-AFLP (amplified fragment length polymorphism). Secondly, the mRNA corresponding to such a putative pathogenicity factor should predominantly be present in the esophageal glands of pre-parasitic juveniles. This was checked by in situ hybridization. As a third criterion, these proteinaceous factors should be preceded by a signal peptide for secretion. Expression profiles of more than 4,000 genes were generated and three up-regulated, dorsal gland-specific proteins preceded by signal peptide for secretion were identified. No dorsal gland genes have been cloned before from plant-parasitic nematodes. The partial sequence of these three factors, A4, A18, and A41, showed no significant homology to any known gene. Their presence in the dorsal glands of infective juveniles suggests that these proteins could be involved in feeding cell initiation, and not in migration in the plant root or in protection against plant defense responses. Finally, the applicability of this new strategy in other plant-microbe interactions is discussed.

An annotated genetic map of loblolly pine based on microsatellite and cDNA markers

USDA-ARS?s Scientific Manuscript database

Previous loblolly pine (Pinus taeda L.) genetic linkage maps have been based on a variety of DNA polymorphisms, such as AFLPs, RAPDs, RFLPs, and ESTPs, but only a few SSRs (simple sequence repeats), also known as simple tandem repeats or microsatellites, have been mapped in P. taeda. The objective o...

Alterations of leaf cell ultrastructures and AFLP DNA profiles in Earth-grown tomato plants propagated from long-term six years Mir-flown seeds

NASA Astrophysics Data System (ADS)

Liu, Min; Xue, Huai; Pan, Yi; Zhang, Chunhua; Lu, Jinying

Leaf cell ultrastructures and DNA variations in the firstand the second-generation of Earthgrown tomato (Lycopersicon esculentun Mill) plants that had been endured a long-term six years spaceflight in the Mir were compared to their ground-based control plants, under observations with a Transmission Electron Microscope and the Amplification Fragment Length Polymorphism (AFLP) analysis. For alterations in the morphological ultrastructures, one plant among the 11 first-generation plants generated from 30 Mir-flown seeds had a three-layered palisade cell structure, while other 10 first-generation plants and all ground-based controls had one-layered palisade cell structure in leaves. Starch grains were larger and in clusters, numbers of starch grains increased in the chloroplasts in the Mir-flown plants. Leaf cells became contracted and deformed, and cell shape patterns were different in the Mir-flown plants. For the leaf genomic DNA alterations, 34 DNA bands were polymorphic with a 1.32% polymorphism among 2582 DNA bands in the first-generation Mir-flown plants. Band types in the spaceflight treated plants were also different from those in the ground-based control. Of 11 survived first-generation plants, 7 spaceflight treated plants (Plant Nos. 1-6 and No. 9) had a same 7 polymorphic bands and a same 0.27%DNA mutation. The DNA mutation rate was greatest in Plants No.10 and No.7 (0.90% and 0.94%), less in Plant No.11 (0.31%) and least in Plant No.8 (0.20%). For the 38 send-generation plants propagated from the No. 5 Mir-flown seed, 6 DNA bands were polymorphic with a 0.23% polymorphism among 2564 amplified DNA bands. Among those 38 second-generation plants amplified by primer pair (E4: ACC, M8: CTT), one DNA band disappeared in 29 second-generation plants and in the original Mir-flown No. 5 plant, compared to the ground-base controls. Among the 38 second-generation plants generated from the Mir-flown No. 5 seed, the DNA band types of 29 second-generation plants were different from that of the ground-base controls and had a same 6 polymorphic bands and a same 0.23% DNA mutation. For the 49 second-generation plants derived from the Mir-flown No. 6 seed, 7 DNA bands were polymorphic with 0.27% polymorphism among 2564 amplified DNA bands. With only one exception among those 49 second-generation plants amplified by primer pair (E3: ACA, M3: CAG), one DNA band disappeared in 48 second-generation plants and in the original Mir-flown No. 6 plant, compared to the ground-based controls. Among the 49 second-generation plants generated from the Mir-flown No. 6 seed, the DNA band types of 48 second-generation plants were different from that of the ground-base controls and had a same 7 polymorphic bands and a same 0.27% DNA mutation. Our results indicated that leaf cell ultrastructures had been altered and heredity variations had been induced by seeds being exposed to a long-term outer-space environment. Further research is needed to elucidate the dynamics and mechanisms resulting in such variations. Plant biology studies in the space environment may open potential approaches to induce mutations and to screen new plant varieties by ground-based selections among spaceflight treated seeds or seedlings.

Inverse Correlation of Population Similarity and Introduction Date for Invasive Ascidians

PubMed Central

Silva, Nathan; Smith, William C.

2008-01-01

The genomes of many marine invertebrates, including the purple sea urchin and the solitary ascidians Ciona intestinalis and Ciona savignyi, show exceptionally high levels of heterozygosity, implying that these populations are highly polymorphic. Analysis of the C. savignyi genome found little evidence to support an elevated mutation rate, but rather points to a large population size contributing to the polymorphism level. In the present study, the relative genetic polymorphism levels in sampled populations of ten different ascidian species were determined using a similarity index generated by AFLP analysis. The goal was to determine the range of polymorphism within the populations of different species, and to uncover factors that may contribute to the high level of polymorphism. We observe that, surprisingly, the levels of polymorphism within these species show a negative correlation with the reported age of invasive populations, and that closely related species show substantially different levels of genetic polymorphism. These findings show exceptions to the assumptions that invasive species start with a low level of genetic polymorphism that increases over time and that closely related species have similar levels of genetic polymorphism. PMID:18575620

ASSESSMENT OF MOLECULAR GENETIC STABILITY BETWEEN LONG-TERM CRYOPRESERVED AND TISSUE CULTURED WASABI (Wasabia japonica) PLANTS.

PubMed

Maki, S; Hirai, Y; Niino, T; Matsumoto, T

2015-01-01

Maintaining the genetic integrity in long-term tissue cultured and cryopreserved plants is important for the conservation of plant genetic resources. In this study, the genetic stability of cryopreserved wasabi shoot tips stored for 10 years at -150 degree C was visualized using Amplified Fragment Length Polymorphism (AFLP) and Methylation Sensitive Amplified Polymorphism (MSAP). The study included plants derived from cryopreserved shoot tips after 10.5 years storage at -150 degree C (LN10yr), after 2 h storage at -196 degree C (LN2hr), cryopreservation controls (No LN cooling (TC)) and non-treated controls without LN cooling (LC). The donor plants for LN2hr, TC and LC were also maintained in vitro at 20 degree C for the same period. Neither technique detected genetic variations in either control or cryopreserved plants. Some mutations were noted in plants maintained in tissue culture for 10 years. Comparison of genome stability for TC and LN2hr plants showed only a minor change in DNA. However, when comparing the LC and Ln10yr, many differences were found. We conclude that cryopreservation is a superior conservation method compared to tissue culture in maintaining genetic stability for a long-term storage of wasabi germplasm.

Methods for Genome-Wide Analysis of Gene Expression Changes in Polyploids

PubMed Central

Wang, Jianlin; Lee, Jinsuk J.; Tian, Lu; Lee, Hyeon-Se; Chen, Meng; Rao, Sheetal; Wei, Edward N.; Doerge, R. W.; Comai, Luca; Jeffrey Chen, Z.

2007-01-01

Polyploidy is an evolutionary innovation, providing extra sets of genetic material for phenotypic variation and adaptation. It is predicted that changes of gene expression by genetic and epigenetic mechanisms are responsible for novel variation in nascent and established polyploids (Liu and Wendel, 2002; Osborn et al., 2003; Pikaard, 2001). Studying gene expression changes in allopolyploids is more complicated than in autopolyploids, because allopolyploids contain more than two sets of genomes originating from divergent, but related, species. Here we describe two methods that are applicable to the genome-wide analysis of gene expression differences resulting from genome duplication in autopolyploids or interactions between homoeologous genomes in allopolyploids. First, we describe an amplified fragment length polymorphism (AFLP)–complementary DNA (cDNA) display method that allows the discrimination of homoeologous loci based on restriction polymorphisms between the progenitors. Second, we describe microarray analyses that can be used to compare gene expression differences between the allopolyploids and respective progenitors using appropriate experimental design and statistical analysis. We demonstrate the utility of these two complementary methods and discuss the pros and cons of using the methods to analyze gene expression changes in autopolyploids and allopolyploids. Furthermore, we describe these methods in general terms to be of wider applicability for comparative gene expression in a variety of evolutionary, genetic, biological, and physiological contexts. PMID:15865985

Characterization of the multiple resistance traits of somatic hybrids between Solanum cardiophyllum Lindl. and two commercial potato cultivars.

PubMed

Thieme, Ramona; Rakosy-Tican, Elena; Nachtigall, Marion; Schubert, Jörg; Hammann, Thilo; Antonova, Olga; Gavrilenko, Tatjana; Heimbach, Udo; Thieme, Thomas

2010-10-01

Interspecific somatic hybrids between commercial cultivars of potato Solanum tuberosum L. Agave and Delikat and the wild diploid species Solanum cardiophyllum Lindl. (cph) were produced by protoplast electrofusion. The hybrid nature of the regenerated plants was confirmed by flow cytometry, simple sequence repeat (SSR), amplified fragment length polymorphism (AFLP), microsatellite-anchored fragment length polymorphism (MFLP) markers and morphological analysis. Somatic hybrids were assessed for their resistance to Colorado potato beetle (CPB) using a laboratory bioassay, to Potato virus Y (PVY) by mechanical inoculation and field trials, and foliage blight in a greenhouse and by field trials. Twenty-four and 26 somatic hybrids of cph + cv. Agave or cph + cv. Delikat, respectively, showed no symptoms of infection with PVY, of which 3 and 12, respectively, were also resistant to foliage blight. One hybrid of cph + Agave performed best in CPB and PVY resistance tests. Of the somatic hybrids that were evaluated for their morphology and tuber yield in the field for 3 years, four did not differ significantly in tuber yield from the parental and standard cultivars. Progeny of hybrids was obtained by pollinating them with pollen from a cultivar, selfing or cross-pollination. The results confirm that protoplast electrofusion can be used to transfer the CPB, PVY and late blight resistance of cph into somatic hybrids. These resistant somatic hybrids can be used in pre-breeding studies, molecular characterization and for increasing the genetic diversity available for potato breeding by marker-assisted combinatorial introgression into the potato gene pool.

[AFLP Analysis of Genetic Diversity in the Genus Mallus Mill. (Apple)].

PubMed

Savelyeva, E N; Kudryavtsev, A M

2015-10-01

The first molecular genetic analysis of the apple species and varieties from Russian collections with the AFLP marker system was performed in order to study the genetic diversity of the genus Malus, as well as to clarify the phylogeny and solve some systematic issues of the genus. Nienty-one apple accessions, including species from five sections of the genus Malus and hybrid species, were examined. The level of polymorphism constituted 90.2%. It was demonstrated that the classical taxonomy of the genus Malus, which identifies five sections based on differences in their morphological characters, is valid and may be used to classify apple species. The species assignment of the Antonovka landraces was established. All of them belonged to the species M. domestica. It was demonstrated that the Yakutskaya apple variety was a domesticated species of the section Gymnomeles, presumably, M. baccata. AFLP analysis confirmed the hybrid nature of many species. The relationships between apple varieties of the Golden group with American wild species were demonstrated. The data suggest that the species M. sieversii was the ancestor of not only the domestic apple but also of other species of the Malus sections.

Cloning and expression analysis of Zmglp1, a new germin-like protein gene in maize.

PubMed

Fan, Zhanmin; Gu, Hongya; Chen, Xiaowei; Song, Hui; Wang, Qian; Liu, Meihua; Qu, Li-Jia; Chen, Zhangliang

2005-06-17

The cDNA and genomic DNA of a green tissue-specific gene were cloned from maize (Zea mays L.) using cDNA-amplified fragment length polymorphism (cDNA-AFLP) and library screening. The deduced protein was highly similar to Hordeum vulgare germin-like protein 1 (HvGLP1), and the maize gene was therefore designated Zmglp1. Northern blot specifically detected the mRNA of Zmglp1 in young whorl leaves at the early-whorl stage. However, at the late-whorl, tassel, and silk stages, Zmglp1 transcripts were highly abundant in young whorl leaves; less abundant in mature leaves, young tassels, and cobs; and not detectable in roots, immature kernels, and stalks. RNA in situ hybridization revealed that Zmglp1 expressed only in mesophyllous, phloem, and guard cells in the young whorl leaves. Deletion analysis of the promoter in transgenic Arabidopsis resulted in the identification of several regions containing important regulatory cis-elements controlling the expression levels and circadian rhythm-oscillated patterns of Zmglp1.

[Clonal variability in longevity of the cercariae of Himasthla elongata (Trematoda: Echinostomatidae)].

PubMed

Levakin, I A; Losev, E A; Zavirskiĭ, Ia V; Galaktionov, K V

2013-01-01

The study was carried out on Himasthla elongata cercariae shed by infected Littorina littorea snails. The infected periwinkles were collected from the settlement with the low prevalence of H. elongata. As shown earlier with the use of AFLP (Amplified Fragment Length Polymorphisms) method, rediae groups in all the infected periwinkles of this settlement arise from the infection of a mollusc with a single miracidium. Therefore, the cercariae shed by an infected mollusc have the same genotype or, in other words, represent a clone. The LT50 (the time during which 50% of cercariae perish in the experimental dish) were measured experimentally for cercariae Himasthla elongata belong to different clones. The investigated parameter demonstrated a high level of interclonal variability. Two groups of cercarial clones were identified: one of them was characterized by the high level of intraclonal variability in LT50 and the second, by the low one. It is assumed that the observed heterogeneity may be stipulated by different degrees of mitotic recombinations during formation of different cercarial clones.

Applying genotoxicology tools to identify environmental stressors in support of river management.

PubMed

Oberholster, Paul J; Hill, Liesl; Jappie, Shaamiela; Truter, Johannes C; Botha, Anna-Maria

2016-02-01

Although bioassay approaches are useful for identifying chemicals of potential concern, they provide little understanding of the mechanisms of chemical toxicity. Without this understanding, it is difficult to address some of the key challenges that currently face aquatic ecotoxicology. To overcome this, the toxicity potential of the water samples was assessed and surviving organisms (Physa acuta) were used for protein activity measurements and gene expression profiling by making use of complementary DNA amplified fragment length polymorphism (cDNA-AFLP) analysis. From the data it was evident that the impacts of specific pollutants (e.g. sewage) on organisms at the cellular level could be identified, and that the expressed stressor genes can be used as bioindicators/markers/genetic signatures or fingerprints during identification of point source pollution. From an ecosystem management point of view these insights could assist with the forecasting and reduction of environmental risks on catchment level by implementing suitable management interventions. Copyright © 2015 Elsevier Ltd. All rights reserved.

Several Pleistocene refugia detected in the high alpine plant Phyteuma globulariifolium sternb & hoppe (Campanulaceae) in the European Alps.

PubMed

Schönswetter, P; Tribsch, A; Barfuss, M; Niklfeld, H

2002-12-01

Phyteuma globulariifolium is a high alpine plant species growing in the European Alps and the Pyrenees. In order to elucidate its glacial history, 325 individuals from 69 populations were analysed using the amplified fragment length polymorphism (AFLP) technique. A strongly hierarchical phylogeographical pattern was detected: Two major east-west vicariant groups can be separated along a gap in the distributional area. A further subdivision into at least four populational groups is in congruence with presumed peripheral glacial refugia. There is no indication for survival on unglaciated mountain tops (nunataks) in the interior of the Pleistocene ice shield covering the Alps. Our results favour glacial survival in peripheral, unglaciated or not fully glaciated areas. Populations of P. globulariifolium in the Pyrenees are the result of relatively recent long-distance dispersal. Within the Alps, there is strong differentiation among groups of populations, whereas within them the differentiation is weak. This suggests high levels of gene-flow over short to middle distances.

The voyage of an invasive species across continents: genetic diversity of North American and European Colorado potato beetle populations.

PubMed

Grapputo, Alessandro; Boman, Sanna; Lindström, Leena; Lyytinen, Anne; Mappes, Johanna

2005-12-01

The paradox of successful invading species is that they are likely to be genetically depauperate compared to their source population. This study on Colorado potato beetles is one of the few studies of the genetic consequences of continent-scale invasion in an insect pest. Understanding gene flow, population structure and the potential for rapid evolution in native and invasive populations offers insights both into the dynamics of small populations that become successful invaders and for their management as pests. We used this approach to investigate the invasion of the Colorado potato beetle (Leptinotarsa decemlineata) from North America to Europe. The beetles invaded Europe at the beginning of the 20th century and expanded almost throughout the continent in about 30 years. From the analysis of mitochondrial DNA (mtDNA) and amplified fragment length polymorphism (AFLP) markers, we found the highest genetic diversity in beetle populations from the central United States. The European populations clearly contained only a fraction of the genetic variability observed in North American populations. European populations show a significant reduction at nuclear markers (AFLPs) and are fixed for one mitochondrial haplotype, suggesting a single successful founder event. Despite the high vagility of the species and the reduction of genetic diversity in Europe, we found a similar, high level of population structure and low gene flow among populations on both continents. Founder events during range expansion, agricultural management with crop rotation, and selection due to insecticide applications are most likely the causes partitioning genetic diversity in this species.

Cluster of cases of Salmonella enterica serotype Rissen infection in a general hospital, Italy, 2007.

PubMed

Boschi, T; Aquilini, D; Degl'Innocenti, R; Aleo, A; Romani, C; Nicoletti, P; Buonomini, M I; Marconi, P; Bilei, S; Mammina, C; Nastasi, A

2010-12-01

In 2007, three strains of Salmonella enterica serotype Rissen (S. Rissen) were isolated in the laboratory of diagnostic microbiology of the General Hospital of Prato, Tuscany, Italy, over a 1 month and half interval of time. The first isolate was recovered on January 26 from an outpatient with enteritis. Then, two strains were isolated on February 16 and March 11 respectively, from central venous catheters of patients who were being hospitalized in two departments of the Hospital. An epidemiologically linked cluster of cases of salmonellosis was suspected. The three strains were submitted to single enzyme-amplified fragment length polymorphism (SE-AFLP) and XbaI macrorestriction and pulsed-field gel electrophoresis (PFGE) that yielded undistinguishable profiles. Epidemiological investigations failed to identify a common source of infection within the Hospital. Moreover, the third patient had been exclusively total parenteral nutrition fed since his admission with a stomach cancer diagnosis. The first patient had a community-acquired infection, but the source of her illness was uncertain. Twenty-five further isolates identified in the years 2004-2007 in the same geographical area showed distinctly different PFGE and SE-AFLP patterns. The three patients seemed to represent a cluster of epidemiologically unrelated cases caused by a previously never recognized S. Rissen strain. Rapid subtyping of isolates is essential in the early investigation of potential outbreaks, but synthesis of conventional and molecular epidemiological investigation and availability of surveillance data is often critical to prevent the initiation of time-consuming, expensive and ineffective further investigations and control interventions. © 2009 Blackwell Verlag GmbH.

Genetic Diversity Among Botulinum Neurotoxin Producing Clostridial Strains

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hill, K K; Smith, T J; Helma, C H

2006-07-06

Clostridium botulinum is a taxonomic designation for many diverse anaerobic spore forming rod-shaped bacteria which have the common property of producing botulinum neurotoxins (BoNTs). The BoNTs are exoneurotoxins that can cause severe paralysis and even death in humans and various other animal species. A collection of 174 C. botulinum strains were examined by amplified fragment length polymorphism (AFLP) analysis and by sequencing of the 16S rRNA gene and BoNT genes to examine genetic diversity within this species. This collection contained representatives of each of the seven different serotypes of botulinum neurotoxins (BoNT A-G). Analysis of the16S rRNA sequences confirmed earliermore » reports of at least four distinct genomic backgrounds (Groups I-IV) each of which has independently acquired one or more BoNT serotypes through horizontal gene transfer. AFLP analysis provided higher resolution, and can be used to further subdivide the four groups into sub-groups. Sequencing of the BoNT genes from serotypes A, B and E in multiple strains confirmed significant sequence variation within each serotype. Four distinct lineages within each of the BoNT A and B serotypes, and five distinct lineages of serotype E strains were identified. The nucleotide sequences of the seven serotypes of BoNT were compared and show varying degrees of interrelatedness and recombination as has been previously noted for the NTNH gene which is linked to BoNT. These analyses contribute to the understanding of the evolution and phylogeny within this species and assist in the development of improved diagnostics and therapeutics for treatment of botulism.« less

Effects of Sublethal Fungicides on Mutation Rates and Genomic Variation in Fungal Plant Pathogen, Sclerotinia sclerotiorum.

PubMed

Amaradasa, B Sajeewa; Everhart, Sydney E

2016-01-01

Pathogen exposure to sublethal doses of fungicides may result in mutations that may represent an important and largely overlooked mechanism of introducing new genetic variation into strictly clonal populations, including acquisition of fungicide resistance. We tested this hypothesis using the clonal plant pathogen, Sclerotinia sclerotiorum. Nine susceptible isolates were exposed independently to five commercial fungicides with different modes of action: boscalid (respiration inhibitor), iprodione (unclear mode of action), thiophanate methyl (inhibition of microtubulin synthesis) and azoxystrobin and pyraclostrobin (quinone outside inhibitors). Mycelium of each isolate was inoculated onto a fungicide gradient and sub-cultured from the 50-100% inhibition zone for 12 generations and experiment repeated. Mutational changes were assessed for all isolates at six neutral microsatellite (SSR) loci and for a subset of isolates using amplified fragment length polymorphisms (AFLPs). SSR analysis showed 12 of 85 fungicide-exposed isolates had a total of 127 stepwise mutations with 42 insertions and 85 deletions. Most stepwise deletions were in iprodione- and azoxystrobin-exposed isolates (n = 40/85 each). Estimated mutation rates were 1.7 to 60-fold higher for mutated loci compared to that expected under neutral conditions. AFLP genotyping of 33 isolates (16 non-exposed control and 17 fungicide exposed) generated 602 polymorphic alleles. Cluster analysis with principal coordinate analysis (PCoA) and discriminant analysis of principal components (DAPC) identified fungicide-exposed isolates as a distinct group from non-exposed control isolates (PhiPT = 0.15, P = 0.001). Dendrograms based on neighbor-joining also supported allelic variation associated with fungicide-exposure. Fungicide sensitivity of isolates measured throughout both experiments did not show consistent trends. For example, eight isolates exposed to boscalid had higher EC50 values at the end of the experiment, and when repeated, only one isolate had higher EC50 while most isolates showed no difference. Results of this support the hypothesis that sublethal fungicide stress increases mutation rates in a largely clonal plant pathogen under in vitro conditions. Collectively, this work will aid our understanding how non-lethal fungicide exposure may affect genomic variation, which may be an important mechanism of novel trait emergence, adaptation, and evolution for clonal organisms.

Effects of Sublethal Fungicides on Mutation Rates and Genomic Variation in Fungal Plant Pathogen, Sclerotinia sclerotiorum

PubMed Central

Amaradasa, B. Sajeewa

2016-01-01

Pathogen exposure to sublethal doses of fungicides may result in mutations that may represent an important and largely overlooked mechanism of introducing new genetic variation into strictly clonal populations, including acquisition of fungicide resistance. We tested this hypothesis using the clonal plant pathogen, Sclerotinia sclerotiorum. Nine susceptible isolates were exposed independently to five commercial fungicides with different modes of action: boscalid (respiration inhibitor), iprodione (unclear mode of action), thiophanate methyl (inhibition of microtubulin synthesis) and azoxystrobin and pyraclostrobin (quinone outside inhibitors). Mycelium of each isolate was inoculated onto a fungicide gradient and sub-cultured from the 50–100% inhibition zone for 12 generations and experiment repeated. Mutational changes were assessed for all isolates at six neutral microsatellite (SSR) loci and for a subset of isolates using amplified fragment length polymorphisms (AFLPs). SSR analysis showed 12 of 85 fungicide-exposed isolates had a total of 127 stepwise mutations with 42 insertions and 85 deletions. Most stepwise deletions were in iprodione- and azoxystrobin-exposed isolates (n = 40/85 each). Estimated mutation rates were 1.7 to 60-fold higher for mutated loci compared to that expected under neutral conditions. AFLP genotyping of 33 isolates (16 non-exposed control and 17 fungicide exposed) generated 602 polymorphic alleles. Cluster analysis with principal coordinate analysis (PCoA) and discriminant analysis of principal components (DAPC) identified fungicide-exposed isolates as a distinct group from non-exposed control isolates (PhiPT = 0.15, P = 0.001). Dendrograms based on neighbor-joining also supported allelic variation associated with fungicide-exposure. Fungicide sensitivity of isolates measured throughout both experiments did not show consistent trends. For example, eight isolates exposed to boscalid had higher EC50 values at the end of the experiment, and when repeated, only one isolate had higher EC50 while most isolates showed no difference. Results of this support the hypothesis that sublethal fungicide stress increases mutation rates in a largely clonal plant pathogen under in vitro conditions. Collectively, this work will aid our understanding how non-lethal fungicide exposure may affect genomic variation, which may be an important mechanism of novel trait emergence, adaptation, and evolution for clonal organisms. PMID:27959950

Genetic diversity of clinical and environmental isolates of Vibrio cholerae determined by amplified fragment length polymorphism fingerprinting.

PubMed

Jiang, S C; Matte, M; Matte, G; Huq, A; Colwell, R R

2000-01-01

Vibrio cholerae, the causative agent of major epidemics of diarrheal disease in Bangladesh, South America, Southeastern Asia, and Africa, was isolated from clinical samples and from aquatic environments during and between epidemics over the past 20 years. To determine the evolutionary relationships and molecular diversity of these strains, in order to understand sources, origin, and epidemiology, a novel DNA fingerprinting technique, amplified fragment length polymorphism (AFLP), was employed. Two sets of restriction enzyme-primer combinations were tested for fingerprinting of V. cholerae serogroup O1, O139, and non-O1, O139 isolates. Amplification of HindIII- and TaqI-digested genomic DNA produced 30 to 50 bands for each strain. However, this combination, although capable of separating environmental isolates of O1 and non-O1 strains, was unable to distinguish between O1 and O139 clinical strains. This result confirmed that clinical O1 and O139 strains are genetically closely related. On the other hand, AFLP analyses of restriction enzyme ApaI- and TaqI-digested genomic DNA yielded 20 to 30 bands for each strain, but were able to separate O1 from O139 strains. Of the 74 strains examined with the latter combination, 26 serogroup O1 strains showed identical banding patterns and were represented by the O1 El Tor strain of the seventh pandemic. A second group, represented by O139 Bengal, included 12 strains of O139 clinical isolates, with 7 from Thailand, 3 from Bangladesh, and 2 from India. Interestingly, an O1 clinical isolate from Africa also grouped with the O139 clinical isolates. Eight clinical O1 isolates from Mexico grouped separately from the O1 El Tor of the seventh pandemic, suggesting an independent origin of these isolates. Identical fingerprints were observed between an O1 environmental isolate from a river in Chile and an O1 clinical strain from Kenya, both isolated more than 10 years apart. Both strains were distinct from the O1 seventh pandemic strain. Two O139 clinical isolates from Africa clustered with environmental non-O1 isolates, independent of other O139 strains included in the study. These results suggest that although a single clone of pathogenic V. cholerae appears responsible for many cases of cholera in Asia, Africa, and Latin America during the seventh pandemic, other cases of clinical cholera were caused by toxigenic V. cholerae strains that appear to have been derived locally from environmental O1 or non-O1 strains.

Epidemiological Investigation of Legionella pneumophila Serogroup 2 to 14 Isolates from Water Samples by Amplified Fragment Length Polymorphism and Sequence-Based Typing and Detection of Virulence Traits

PubMed Central

Katsiaflaka, Anna; Pournaras, Spyros; Kristo, Ioulia; Mouchtouri, Varvara A.; Kyritsi, Maria; Velonakis, Emmanuel; Vatopoulos, Alkiviadis C.

2016-01-01

ABSTRACT The aim of this study is to explore the dispersion, clonality, and virulence of Legionella pneumophila serogroups 2 to 14 in the Greek environment. Eighty L. pneumophila serogroup 2 to 14 strains isolated from water distribution systems of hotels, hospitals, athletic venues, and ferries in Greece were tested by monoclonal antibodies (MAbs) for serogroup discrimination and molecularly by amplified fragment length polymorphism (AFLP) for genetic diversity. Fifty-six of 80 strains were also typed by the sequence-based typing (SBT) method. Αll strains were further analyzed for detection of two pathogenicity loci: Legionella vir homologue (lvh) and repeats in structural toxin (rtxA). Thirty-seven strains (46.2%) belonged to serogroup 6, 26 strains (32.5%) to serogroup 3, and 7 (8.8%) to other serogroups (4, 5, 8, and 10). Ten strains (12.5%) were nontypeable (NT) into the known serogroups. Thirty-nine different AFLP types were found among the 80 L. pneumophila serogroup 2 to 14 strains, and 24 different SBT types were found among the 56 strains tested. Among the 80 strains, the lvh locus was present in 75 (93.8%), the rtxA locus was found in 76 (95%), and both loci were found in 73 (91.3%) strains. This study showed that there is genetic variability of L. pneumophila serogroups 2 to 14 in the Greek environment as well as a high percentage of the pathogenicity loci. Ιntroducing an effective diagnostic test for L. pneumophila serogroups 2 to 14 in urine and promoting the examination of respiratory specimens from patients hospitalized for pneumonia in Greek hospitals are essential. IMPORTANCE In this study, the dispersion, clonality, and virulence of environmental isolates of Legionella pneumophila serogroups 2 to 14 (Lp2–14) in Greece were investigated. Genetic variability of Lp2–14 in the Greek environment was identified together with the presence of the pathogenicity loci in a high percentage of the isolates. Despite the high prevalence of Lp2–14 in the Greek environment, no clinical cases were reported, which may be due to underdiagnosis of the disease. Almost all the legionellosis cases are diagnosed in Greece by using the urine antigen test, which is specific for Lp1. There is an urgent need to improve the clinical diagnosis of legionellosis by introducing an effective diagnostic test for Lp2–14 in urine and by promoting the PCR examination of respiratory specimens from patients with compatible clinical symptoms. PMID:27496776

An annotated genetic map of loblolly pine based on microsatellite and cDNA markers

Treesearch

Craig S. Echt; Surya Saha; Konstantin V. Krutovsky; Kokulapalan Wimalanathan; John E. Erpelding; Chun Liang; C Dana Nelson

2011-01-01

Previous loblolly pine (Pinus taeda L.) genetic linkage maps have been based on a variety of DNA polymorphisms, such as AFLPs, RAPDs, RFLPs, and ESTPs, but only a few SSRs (simple sequence repeats), also known as simple tandem repeats or microsatellites, have been mapped in P. taeda. The objective of this study was to integrate a large set of SSR markers from a variety...

Genetic differentiation in Pyrenophora teres f. teres populations from Syria and Tunisia as assessed by AFLP markers.

PubMed

Bouajila, A; Zoghlami, N; Murad, S; Baum, M; Ghorbel, A; Nazari, K

2013-06-01

To investigate the level of genetic differentiation and diversity among Pyrenophora teres isolate populations originating from different agro-ecological areas of Syria and Tunisia and to determine the potential of AFLP profiling in genotyping Pyrenophora teres f. teres. In this study, AFLP markers have been employed to identify patterns of population structure in 20 Pyrenophora teres f. teres populations from Syria and Tunisia. Ninety-four isolates were studied by the use of a protocol that involved stringent PCR amplification of fragments derived from digestion of genomic DNA with restriction enzymes EcoRI and MesI. Based on 401 amplified polymorphic DNA markers (AFLP), variance analyses indicated that most of the variation was partitioned within rather than between populations. Genotypic diversity (GD) was high for populations from Rihane, local landraces and different agro-ecological zones (GD = 0·75-0·86). There was high genetic differentiation among pathogen populations from different host populations in Syria (Gst  = 0·31, ht = 0·190) and Tunisia (Gst  = 0·39, ht = 0·263), which may be partly explained by the low gene flow around the areas sampled. A phenetic tree revealed three groups with high bootstrap values (55, 68, 76) and reflected the grouping of isolates based on host, or agro-ecological areas. AFLP profiling is an effective method for typing the genetically diverse pathogen Pyrenophora teres f. teres. The study represents a comparative analysis of the genetic diversity in P. teres isolates from two countries spanning two continents and also shows that several distinct P. teres genotypes may be found in a given environment. The implications of these findings for Pyrenophora teres f. teres evolutionary potential and net blotch-resistance breeding in Syria and Tunisia were also discussed. © 2012 The Society for Applied Microbiology.

Direct isolation of differentially expressed genes from a specific chromosome region of common wheat: application of the amplified fragment length polymorphism-based mRNA fingerprinting (AMF) method in combination with a deletion line of wheat.

PubMed

Kojima, T; Habu, Y; Iida, S; Ogihara, Y

2000-05-01

The amplified restriction fragment length polymorphism (AFLP)-based mRNA fingerprinting (AMF) method makes it possible systematically and conveniently to identify differentially expressed cDNAs with high reproducibility. We have applied the AMF method to the cloning of the Q gene of common wheat, which is located on the long arm of chromosome 5A and pleiotropically controls the spike morphology and the threshing character of seeds. Using the AMF method, we compared the fingerprints of mRNA samples extracted from the young spikes of Triticum aestivum cv. Chinese Spring (CS) carrying the Q gene to those of a chromosome deletion line of CS, namely, q5, which lacks 15% of 5AL including the Q gene. Approximately 12,200 fragments were produced after PCR with 256 primer combinations. Of these, 92 fragments were differentially expressed between CS and q5. Northern and Southern analyses showed that 16 fragments gave specific or relatively stronger transcript signals in CS, and these clones were present in single copy or in low copy numbers in the wheat genome. Four clones were genetically mapped to the region deleted in q5. Subsequently, one clone, pTaQ22, was mapped at the same locus as the Q gene, indicating that pTaQ22 corresponds to the Q gene or is tightly linked to it. DNA sequence data showed that pTaQ22 had no homology to any known genes, thus suggesting a novel function for this gene in flower morphogenesis. This AMF method might provide a straightforward method for isolating genes in the hexaploid background of common wheat.

A High-Density Genetic Map Identifies a Novel Major QTL for Boron Efficiency in Oilseed Rape (Brassica napus L.)

PubMed Central

Wang, Xiaohua; Zhao, Hua; Shi, Lei; Xu, Fangsen

2014-01-01

Low boron (B) seriously limits the growth of oilseed rape (Brassica napus L.), a high B demand species that is sensitive to low B conditions. Significant genotypic variations in response to B deficiency have been observed among B. napus cultivars. To reveal the genetic basis for B efficiency in B. napus, quantitative trait loci (QTLs) for the plant growth traits, B uptake traits and the B efficiency coefficient (BEC) were analyzed using a doubled haploid (DH) population derived from a cross between a B-efficient parent, Qingyou 10, and a B-inefficient parent, Westar 10. A high-density genetic map was constructed based on single nucleotide polymorphisms (SNPs) assayed using Brassica 60 K Infinium BeadChip Array, simple sequence repeats (SSRs) and amplified fragment length polymorphisms (AFLPs). The linkage map covered a total length of 2139.5 cM, with 19 linkage groups (LGs) and an average distance of 1.6 cM between adjacent markers. Based on hydroponic evaluation of six B efficiency traits measured in three separate repeated trials, a total of 52 QTLs were identified, accounting for 6.14–46.27% of the phenotypic variation. A major QTL for BEC, qBEC-A3a, was co-located on A3 with other QTLs for plant growth and B uptake traits under low B stress. Using a subset of substitution lines, qBEC-A3a was validated and narrowed down to the interval between CNU384 and BnGMS436. The results of this study provide a novel major locus located on A3 for B efficiency in B. napus that will be suitable for fine mapping and marker-assisted selection breeding for B efficiency in B. napus. PMID:25375356

A novel method to infer the origin of polyploids from Amplified Fragment Length Polymorphism data reveals that the alpine polyploid complex of Senecio carniolicus (Asteraceae) evolved mainly via autopolyploidy.

PubMed

Winkler, Manuela; Escobar García, Pedro; Gattringer, Andreas; Sonnleitner, Michaela; Hülber, Karl; Schönswetter, Peter; Schneeweiss, Gerald M

2017-09-01

Despite its evolutionary and ecological relevance, the mode of polyploid origin has been notoriously difficult to be reconstructed from molecular data. Here, we present a method to identify the putative parents of polyploids and thus to infer the mode of their origin (auto- vs. allopolyploidy) from Amplified Fragment Length Polymorphism (AFLP) data. To this end, we use Cohen's d of distances between in silico polyploids, generated within a priori defined scenarios of origin from a priori delimited putative parental entities (e.g. taxa, genetic lineages), and natural polyploids. Simulations show that the discriminatory power of the proposed method increases mainly with increasing divergence between the lower-ploid putative ancestors and less so with increasing delay of polyploidization relative to the time of divergence. We apply the new method to the Senecio carniolicus aggregate, distributed in the European Alps and comprising two diploid, one tetraploid and one hexaploid species. In the eastern part of its distribution, the S. carniolicus aggregate was inferred to comprise an autopolyploid series, whereas for western populations of the tetraploid species, an allopolyploid origin involving the two diploid species was the most likely scenario. Although this suggests that the tetraploid species has two independent origins, other evidence (ribotype distribution, morphology) is consistent with the hypothesis of an autopolyploid origin with subsequent introgression by the second diploid species. Altogether, identifying the best among alternative scenarios using Cohen's d can be straightforward, but particular scenarios, such as allopolyploid origin vs. autopolyploid origin with subsequent introgression, remain difficult to be distinguished. © 2016 John Wiley & Sons Ltd.

Morphological, chemical and molecular characterization of Centella asiatica germplasms for commercial cultivation in the Indo-Gangetic plains.

PubMed

Prasad, Archana; Dhawan, Sunita S; Mathur, Ajay K; Prakash, Om; Gupta, Madan M; Verma, Ram K; Lal, Raj K; Mathur, Archana

2014-06-01

Centella asiatica germplasm collected from north, north-eastern and southern parts of India was compared for biomass and centellosides productivity under uniform agro-climatic conditions of the Indo-Gangetic plains at Lucknow. The highest biomass accumulation (411.9 g FW/m2 area) was recorded in accession A from north India, followed by 284.0, 135.7 and 29.2 g FW/m2 in accessions M, B and E from southern, eastern and north-eastern regions, respectively. Accession M possessed the highest asiaticoside content (52.1 mg/gDW) that was 1.58, 2.34 and 21.7 folds more than accessions A, B and E, respectively. The madecassoside level in leaves of accessions B and M was comparable (28.9 and 25.7 mg/gDW) and two folds more than accession A (13.9 mg/gDW). The madecassic and asiatic acid content in leaf tissue of all four accessions remained low in Lucknow. Amplified fragment length polymorphism (AFLP) analysis with 23 primers yielded 696 fragments, 563 of which were polymorphic. Accession M out-grouped with genetic dissimilarity indices of 83, 85 and 95% from accessions A, E and B, respectively. Commercial cultivation of accessions M and A through a four months growth cycle (June to September) in agro-climatic conditions of the Indo-Gangetic plains is suggested.

Genetic variation and structure in remnant population of critically endangered Melicope zahlbruckneri

USGS Publications Warehouse

Raji, J. A.; Atkinson, Carter T.

2016-01-01

The distribution and amount of genetic variation within and between populations of plant species are important for their adaptability to future habitat changes and also critical for their restoration and overall management. This study was initiated to assess the genetic status of the remnant population of Melicope zahlbruckneri–a critically endangered species in Hawaii, and determine the extent of genetic variation and diversity in order to propose valuable conservation approaches. Estimated genetic structure of individuals based on molecular marker allele frequencies identified genetic groups with low overall differentiation but identified the most genetically diverse individuals within the population. Analysis of Amplified Fragment Length Polymorphic (AFLP) marker loci in the population based on Bayesian model and multivariate statistics classified the population into four subgroups. We inferred a mixed species population structure based on Bayesian clustering and frequency of unique alleles. The percentage of Polymorphic Fragment (PPF) ranged from 18.8 to 64.6% for all marker loci with an average of 54.9% within the population. Inclusion of all surviving M. zahlbruckneri trees in future restorative planting at new sites are suggested, and approaches for longer term maintenance of genetic variability are discussed. To our knowledge, this study represents the first report of molecular genetic analysis of the remaining population of M. zahlbruckneri and also illustrates the importance of genetic variability for conservation of a small endangered population.

Degree of Hybridization in Seed Stands of Pinus engelmannii Carr. In the Sierra Madre Occidental, Durango, Mexico

PubMed Central

Ávila-Flores, Israel Jaime; Hernández-Díaz, José Ciro; González-Elizondo, Maria Socorro; Prieto-Ruíz, José Ángel; Wehenkel, Christian

2016-01-01

Hybridization is an important evolutionary force, because interspecific gene transfer can introduce more new genetic material than is directly generated by mutations. Pinus engelmannii Carr. is one of the nine most common pine species in the pine-oak forest ecoregion in the state of Durango, Mexico. This species is widely harvested for lumber and is also used in reforestation programmes. Interspecific hybrids between P.engelmannii and Pinus arizonica Engelm. have been detected by morphological analysis. The presence of hybrids in P. engelmannii seed stands may affect seed quality and reforestation success. Therefore, the goals of this research were to identify introgressive hybridization between P. engelmannii and other pine species in eight seed stands of this species in Durango, Mexico, and to examine how hybrid proportion is related to mean genetic dissimilarity between trees in these stands, using Amplified Fragment Length Polymorphism (AFLP) markers and morphological traits. Differences in the average current annual increment of putative hybrids and pure trees were also tested for statistical significance. Morphological and genetic analyses of 280 adult trees were carried out. Putative hybrids were found in all the seed stands studied. The hybrids did not differ from the pure trees in vigour or robustness. All stands with putative P. engelmannii hybrids detected by both AFLPs and morphological traits showed the highest average values of the Tanimoto distance, which indicates: i) more heterogeneous genetic material, ii) higher genetic variation and therefore iii) the higher evolutionary potential of these stands, and iv) that the morphological differentiation (hybrid/not hybrid) is strongly associated with the Tanimoto distance per stand. We conclude that natural pairwise hybrids are very common in the studied stands. Both morphological and molecular approaches are necessary to confirm the genetic identity of forest reproductive material. PMID:27064490

Genetic Variation in Fusarium Section Liseola from No-Till Maize in Argentina†

PubMed Central

Chulze, S. N.; Ramirez, M. L.; Torres, A.; Leslie, J. F.

2000-01-01

Strains of Fusarium species belonging to section Liseola cause stalk and ear rot of maize and produce important mycotoxins, such as fumonisins. We isolated two species, Fusarium verticillioides (Gibberella fujikuroi mating population A) and Fusarium proliferatum (G. fujikuroi mating population D) from maize cultivated under no-till conditions at five locations in the Córdoba province of Argentina. We determined the effective population number for mating population A (Ne) and found that the Ne for mating type was 89% of the count (total population) and that the Ne for male or hermaphrodite status was 36%. Thus, the number of strains that can function as the female parent limits Ne, and sexual reproduction needs to occur only once every 54 to 220 asexual generations to maintain this level of sexual fertility. Our results indicate that the fungal populations isolated from no-till maize are similar to those recovered from maize managed with conventional tillage. We placed 36 strains from mating population A into 28 vegetative compatibility groups (VCGs). Of the 13 strains belonging to five multimember VCGs, only 2 isolates belonging to one VCG were clones based on amplified fragment length polymorphism (AFLP) fingerprints. Members of the other four multimember VCGs had an average similarity index of 0.89, and members of one VCG were no more closely related to other members of the same VCG than they were to other members of the population as a whole. This finding suggests that the common assumption that strains in the same VCG are either clonal or very closely related needs to be examined in more detail. The variability observed with AFLPs and VCGs suggests that sexual reproduction may occur more frequently than estimated by Ne. PMID:11097907

The attractiveness fragment—AFLP analysis of local adaptation and sexual selection in a caeliferan grasshopper, Chorthippus biguttulus

NASA Astrophysics Data System (ADS)

Klappert, Kirsten; Butlin, Roger K.; Reinhold, Klaus

2007-08-01

Genetic variability among males is a necessary precondition for the evolution of female choice based on indirect genetic benefits. In addition to mutations and host parasite cycles, migration of locally adapted individuals offers an explanation for the maintenance of genetic variability. In a previous study, conducting a reciprocal transplant experiment on a grasshopper, Chorthippus biguttulus, we found that environmental conditions significantly influenced not only body condition but also an important trait of male calling song, the amplitude of song. Although not significant, all other analysed physical and courtship song traits and attractiveness were superior in native than in transferred males. Thus, we concluded that local adaptation has a slight but consistent influence on a range of traits in our study populations, including male acoustic attractiveness. In our present study, we scanned male grasshoppers from the same two populations for amplification fragment length polymorphism (AFLP) loci connected with acoustic attractiveness to conspecific females. We found greater differences in allele frequencies between the two populations, for some loci, than are expected from a balance between drift and gene flow. These loci are potentially connected with locally adapted traits. We examined whether these alleles show the proposed genotype environment interaction by having different associations with attractiveness in the two populations. One locus was significantly related to sexual attractiveness; however, this was independent of the males’ population affiliation. Future research on the evolution of female choice will benefit from knowledge of the underlying genetic architecture of male traits under intraspecific sexual selection, and the ‘population genomics’ approach can be a powerful tool for revealing this structure.

High levels of effective long-distance dispersal may blur ecotypic divergence in a rare terrestrial orchid.

PubMed

Vanden Broeck, An; Van Landuyt, Wouter; Cox, Karen; De Bruyn, Luc; Gyselings, Ralf; Oostermeijer, Gerard; Valentin, Bertille; Bozic, Gregor; Dolinar, Branko; Illyés, Zoltán; Mergeay, Joachim

2014-07-07

Gene flow and adaptive divergence are key aspects of metapopulation dynamics and ecological speciation. Long-distance dispersal is hard to detect and few studies estimate dispersal in combination with adaptive divergence. The aim of this study was to investigate effective long-distance dispersal and adaptive divergence in the fen orchid (Liparis loeselii (L.) Rich.). We used amplified fragment length polymorphism (AFLP)-based assignment tests to quantify effective long-distance dispersal at two different regions in Northwest Europe. In addition, genomic divergence between fen orchid populations occupying two distinguishable habitats, wet dune slacks and alkaline fens, was investigated by a genome scan approach at different spatial scales (continental, landscape and regional) and based on 451 AFLP loci. We expected that different habitats would contribute to strong divergence and restricted gene flow resulting in isolation-by-adaptation. Instead, we found remarkably high levels of effective long-distance seed dispersal and low levels of adaptive divergence. At least 15% of the assigned individuals likely originated from among-population dispersal events with dispersal distances up to 220 km. Six (1.3%) 'outlier' loci, potentially reflecting local adaptation to habitat-type, were identified with high statistical support. Of these, only one (0.22%) was a replicated outlier in multiple independent dune-fen population comparisons and thus possibly reflecting truly parallel divergence. Signals of adaptation in response to habitat type were most evident at the scale of individual populations. The findings of this study suggest that the homogenizing effect of effective long-distance seed dispersal may overwhelm divergent selection associated to habitat type in fen orchids in Northwest Europe.

Molecular Phylogenetics of the Genus Neoconocephalus (Orthoptera, Tettigoniidae) and the Evolution of Temperate Life Histories

PubMed Central

Snyder, Robert L.; Frederick-Hudson, Katy H.; Schul, Johannes

2009-01-01

Background The katydid genus Neoconocephalus (25+ species) has a prominent acoustic communication system and occurs in large parts of the Neotropics and Nearctic. This group has been subject of numerous behavioral, physiological, and evolutionary studies of its acoustic communication system. Two distinct life histories occur in this group: The tropical life history incorporates multiple generations/year and direct egg development without environmental triggers. Temperate life history is characterized by overwintering in the egg stage, cold trigger of egg development, and one generation/year. This study reconstructs the phylogenetic relationships within the genus to (1) determine the evolutionary history of the temperate life history, and (2) to support comparative studies of evolutionary and physiological problems in this genus. Methodology/Principal Findings We used Amplified Fragment Length Polymorphisms (AFLP), and sequences of two nuclear loci and one mitochondrial locus to reconstruct phylogenetic relationships. The analysis included 17 ingroup and two outgroup species. AFLP and mitochondrial data provided resolution at the species level while the two nuclear loci revealed only deeper nodes. The data sets were combined in a super-matrix to estimate a total evidence tree. Seven of the temperate species form a monophyletic group; however, three more temperate species were placed as siblings of tropical species. Conclusions/Significance Our analyses support the reliability of the current taxonomic treatment of the Neoconocephalus fauna of Caribbean, Central, and North America. Ancestral state reconstruction of life history traits was not conclusive, however at least four transitions between life histories occurred among our sample of species. The proposed phylogeny will strengthen conclusions from comparative work in this group. PMID:19779617

AFLP-based transcript profiling for cassava genome-wide expression analysis in the onset of storage root formation.

PubMed

Sojikul, Punchapat; Kongsawadworakul, Panida; Viboonjun, Unchera; Thaiprasit, Jittrawan; Intawong, Burapat; Narangajavana, Jarunya; Svasti, Mom Rajawong Jisnuson

2010-10-01

Cassava (Manihot esculenta Crantz) is a root crop that accumulates large quantities of starch, and it is an important source of carbohydrate. Study on gene expressions during storage root development provides important information on storage root formation and starch accumulation as well as unlock new traits for improving of starch yield. cDNA-Amplified Fragment Length Polymorphism (AFLP) was used to compare gene expression profiles in fibrous and storage roots of cassava cultivar Kasetsart 50. Total of 155 differentially expressed transcript-derived fragments with undetectable or low expression in leaves were characterized and classified into 11 groups regarding to their functions. The four major groups were no similarity (20%), hypothetical or unknown proteins (17%), cellular metabolism and biosynthesis (17%) and cellular communication and signaling (14%). Interestingly, sulfite reductase (MeKD82), calcium-dependent protein kinase (CDPK) (MeKD83), ent-kaurene synthase (KS) (MeKD106) and hexose transporter (HT) (MeKD154) showed root-specific expression patterns. This finding is consistent with previously reported genes involved in the initiation of potato tuber. Semi-quantitative reverse transcription polymerase chain reaction of early-developed root samples confirmed that those four genes exhibited significant expression with similar pattern in the storage root initiation and early developmental stages. We proposed that KS and HT may involve in transient induction of CDPK expression, which may play an important role in the signaling pathway of storage root initiation. Sulfite reductase, on the other hand, may involve in storage root development by facilitating sulfur-containing protein biosynthesis or detoxifying the cyanogenic glucoside content through aspartate biosynthesis. Copyright © Physiologia Plantarum 2010.

Ecology and genetic diversity of the dense-flowered orchid, Neotinea maculata, at the centre and edge of its range

PubMed Central

Duffy, Karl J.; Scopece, Giovanni; Cozzolino, Salvatore; Fay, Michael F.; Smith, Rhian J.; Stout, Jane C.

2009-01-01

Background and Aims Species may occur over a wide geographical range within which populations can display large variation in reproductive success and genetic diversity. Neotinea maculata is a rare orchid of conservation concern at the edge of its range in Ireland, where it occurs in small populations. However, it is relatively common throughout the Mediterranean region. Here, factors that affect rarity of N. maculata in Ireland are investigated by comparing Irish populations with those found in Italy, where it is more common. Methods Vegetation communities, breeding system and genetic diversity were compared using three amplified fragment length polymorphism (AFLP) primer pairs in populations in Ireland and Italy. Vegetation was quantified using quadrats taken along transects in study populations, and hand pollination experiments were performed to assess reliance of N. maculata on pollinators in both Irish and Italian populations. Key Results Neotinea maculata occupies different vegetation communities in Italian and Irish populations. Breeding system experiments show that N. maculata is 100 % autogamous, and there are no differences in fruit and seed production in selfed, outcrossed and unmanipulated plants. AFLP markers revealed that Irish and Italian populations have similar genetic diversity and are distinct from each other. Conclusions Neotinea maculata does not suffer any negative effects of autogamous reproduction; it self-pollinates and sets seed readily in the absence of pollinators. It occupies a variety of habitats in both Ireland and Italy; however, Irish populations are small and rare and should be conserved. This could be due to climatic factors and the absence of suitable soil mycorrhizas to allow recruitment from seed. PMID:18940852

Degree of Hybridization in Seed Stands of Pinus engelmannii Carr. In the Sierra Madre Occidental, Durango, Mexico.

PubMed

Ávila-Flores, Israel Jaime; Hernández-Díaz, José Ciro; González-Elizondo, Maria Socorro; Prieto-Ruíz, José Ángel; Wehenkel, Christian

2016-01-01

Hybridization is an important evolutionary force, because interspecific gene transfer can introduce more new genetic material than is directly generated by mutations. Pinus engelmannii Carr. is one of the nine most common pine species in the pine-oak forest ecoregion in the state of Durango, Mexico. This species is widely harvested for lumber and is also used in reforestation programmes. Interspecific hybrids between P.engelmannii and Pinus arizonica Engelm. have been detected by morphological analysis. The presence of hybrids in P. engelmannii seed stands may affect seed quality and reforestation success. Therefore, the goals of this research were to identify introgressive hybridization between P. engelmannii and other pine species in eight seed stands of this species in Durango, Mexico, and to examine how hybrid proportion is related to mean genetic dissimilarity between trees in these stands, using Amplified Fragment Length Polymorphism (AFLP) markers and morphological traits. Differences in the average current annual increment of putative hybrids and pure trees were also tested for statistical significance. Morphological and genetic analyses of 280 adult trees were carried out. Putative hybrids were found in all the seed stands studied. The hybrids did not differ from the pure trees in vigour or robustness. All stands with putative P. engelmannii hybrids detected by both AFLPs and morphological traits showed the highest average values of the Tanimoto distance, which indicates: i) more heterogeneous genetic material, ii) higher genetic variation and therefore iii) the higher evolutionary potential of these stands, and iv) that the morphological differentiation (hybrid/not hybrid) is strongly associated with the Tanimoto distance per stand. We conclude that natural pairwise hybrids are very common in the studied stands. Both morphological and molecular approaches are necessary to confirm the genetic identity of forest reproductive material.

Genotypic and Antimicrobial Susceptibility of Carbapenem-resistant Acinetobacter baumannii: Analysis of ISAba Elements and blaOXA-23-like Genes Including a New Variant

PubMed Central

Bahador, Abbas; Raoofian, Reza; Pourakbari, Babak; Taheri, Mohammad; Hashemizadeh, Zahra; Hashemi, Farhad B.

2015-01-01

Carbapenem-resistant Acinetobacter baumannii (CR-AB) causes serious nosocomial infections, especially in ICU wards of hospitals, worldwide. Expression of blaOXA genes is the chief mechanism of conferring carbapenem resistance among CR-AB. Although some blaOXA genes have been studied among CR-AB isolates from Iran, their blaOXA-23-like genes have not been investigated. We used a multiplex-PCR to detect Ambler class A, B, and D carbapenemases of 85 isolates, and determined that 34 harbored blaOXA-23-like genes. Amplified fragment length polymorphism (AFLP) genotyping, followed by DNA sequencing of blaOXA-23-like amplicons of CR-AB from each AFLP group was used to characterize their blaOXA-23-like genes. We also assessed the antimicrobial susceptibility pattern of CR-AB isolates, and tested whether they harbored insertion sequences ISAba1 and ISAba4. Sequence comparison with reference strain A. baumannii (NCTC12156) revealed five types of mutations in blaOXA-23-like genes; including one novel variant and four mutants that were already reported from China and the USA. All of the blaOXA-23-like genes mutations were associated with increased minimum inhibitory concentrations (MICs) against imipenem. ISAba1 and ISAba4 sequences were detected upstream of blaOXA-23 genes in 19 and 7% of isolates, respectively. The isolation of CR-AB with new blaOXA-23 mutations including some that have been reported from the USA and China highlights CR-AB pervasive distribution, which underscores the importance of concerted national and global efforts to control the spread of CR-AB isolates worldwide. PMID:26617588

High levels of effective long-distance dispersal may blur ecotypic divergence in a rare terrestrial orchid

PubMed Central

2014-01-01

Background Gene flow and adaptive divergence are key aspects of metapopulation dynamics and ecological speciation. Long-distance dispersal is hard to detect and few studies estimate dispersal in combination with adaptive divergence. The aim of this study was to investigate effective long-distance dispersal and adaptive divergence in the fen orchid (Liparis loeselii (L.) Rich.). We used amplified fragment length polymorphism (AFLP)-based assignment tests to quantify effective long-distance dispersal at two different regions in Northwest Europe. In addition, genomic divergence between fen orchid populations occupying two distinguishable habitats, wet dune slacks and alkaline fens, was investigated by a genome scan approach at different spatial scales (continental, landscape and regional) and based on 451 AFLP loci. Results We expected that different habitats would contribute to strong divergence and restricted gene flow resulting in isolation-by-adaptation. Instead, we found remarkably high levels of effective long-distance seed dispersal and low levels of adaptive divergence. At least 15% of the assigned individuals likely originated from among-population dispersal events with dispersal distances up to 220 km. Six (1.3%) ‘outlier’ loci, potentially reflecting local adaptation to habitat-type, were identified with high statistical support. Of these, only one (0.22%) was a replicated outlier in multiple independent dune-fen population comparisons and thus possibly reflecting truly parallel divergence. Signals of adaptation in response to habitat type were most evident at the scale of individual populations. Conclusions The findings of this study suggest that the homogenizing effect of effective long-distance seed dispersal may overwhelm divergent selection associated to habitat type in fen orchids in Northwest Europe. PMID:24998243

Molecular cloning, sequence analysis and homology modeling of the first caudata amphibian antifreeze-like protein in axolotl (Ambystoma mexicanum).

PubMed

Zhang, Songyan; Gao, Jiuxiang; Lu, Yiling; Cai, Shasha; Qiao, Xue; Wang, Yipeng; Yu, Haining

2013-08-01

Antifreeze proteins (AFPs) refer to a class of polypeptides that are produced by certain vertebrates, plants, fungi, and bacteria and which permit their survival in subzero environments. In this study, we report the molecular cloning, sequence analysis and three-dimensional structure of the axolotl antifreeze-like protein (AFLP) by homology modeling of the first caudate amphibian AFLP. We constructed a full-length spleen cDNA library of axolotl (Ambystoma mexicanum). An EST having highest similarity (∼42%) with freeze-responsive liver protein Li16 from Rana sylvatica was identified, and the full-length cDNA was subsequently obtained by RACE-PCR. The axolotl antifreeze-like protein sequence represents an open reading frame for a putative signal peptide and the mature protein composed of 93 amino acids. The calculated molecular mass and the theoretical isoelectric point (pl) of this mature protein were 10128.6 Da and 8.97, respectively. The molecular characterization of this gene and its deduced protein were further performed by detailed bioinformatics analysis. The three-dimensional structure of current AFLP was predicted by homology modeling, and the conserved residues required for functionality were identified. The homology model constructed could be of use for effective drug design. This is the first report of an antifreeze-like protein identified from a caudate amphibian.

Screening interspecific hybrids of Populus (P. ciliata x maximowiczii) using AFLP markers.

PubMed

Chauhan, N; Negi, M S; Sabharwal, V; Khurana, D K; Lakshmikumaran, M

2004-03-01

Hybrids of Populus ciliata x maximowiczii are very vigorous and outperform both the parents in growth performance and yield. Genetic evaluation of 24 of these interspecific hybrids along with the two mother trees ( Populus ciliata), and five male-parent ( Populus maximowiczii) genotypes was carried out using the AFLP marker assay. Eight AFLP primer combinations detected 428 markers, of which 280 (66%) were polymorphic. Genetic relationships within the samples were evaluated by generating the similarity matrix based on Jaccard's coefficient. The phenetic dendrograms, as well as the PCO plots, separated the hybrids and the two parent species into three distinct clusters. The hybrids grouped closer to the P. ciliata (female parent) cluster as compared to the P. maximowiczii (male parent) cluster. The hybrid cluster contained internal groupings, which correlated to some extent with growth performance. The four best performing hybrids (42m1, 65m1, 23m2, Cm2-5-20/91) formed a distinct sub-cluster. Data from a single primer combination was sufficient for distinguishing the hybrids from the parents and assigning paternity. The hybrids showed 22 markers that were absent in P. ciliata but were monomorphically present in all the hybrids, suggesting outcrossing and common paternity. Further, these 22 markers were found in all the P. maximowiczii genotypes confirming it as the male parent. These male-specific markers can be converted to SCAR markers and used for rapid screening of the P.ciliata x maximowiczii hybrids. The primer combination E-AAC x M-CAA was identified as most suitable for ascertaining true hybridity. AFLP proves to be a useful tool for screening of P. ciliata x maximowiczii hybrids at the early stages of development.

Introgression Threatens the Genetic Diversity of Quercus austrocochinchinensis (Fagaceae), an Endangered Oak: A Case Inferred by Molecular Markers

PubMed Central

An, Miao; Deng, Min; Zheng, Si-Si; Jiang, Xiao-Long; Song, Yi-Gang

2017-01-01

Natural introgression can cause negative effects where rare species experience genetic assimilation and invade by their abundant congeners. Quercus austrocochinchinensis and Q. kerrii (subgenus Cyclobalanopsis) are a pair of closely related species in the Indo-China area. Morphological intermediates of the two species have been reported in this region. In this study, we used AFLP, SSR and two key leaf morphological diagnostic traits to study the two Q. austrocochinchinensis populations, two pure Q. kerrii and two putative hybrid populations in China. Rates of individual admixture were examined using the Bayesian clustering programs STRUCTURE and NewHybrids, with no a priori species assignment. In total, we obtained 151 SSR alleles and 781 polymorphic loci of AFLP markers. Population differentiation inferred by SSR and AFLP was incoherent with recognized species boundaries. Bayesian admixture analyses and principal coordinate analysis identified more hybrids and backcrossed individuals than morphological intermediates in the populations. SSR inferred a wide genetic assimilation in Q. austrocochinchinensis, except for subpopulation D2 in the core area of Xi-Shuang-Ban-Na Nature Reserve (XSBN). However, AFLP recognized more Q. austrocochinchinensis purebreds than SSR. Analysis using NewHybrids on AFLP data indicated that these hybridized individuals were few F2 and predominantly backcrosses with both parental species. All these evidences indicate the formation of a hybrid swarm at XSBN where the two species co-exist. Both AFLP and SSR recognized that the core protected area of XSBN (D2) has a high percentage of Q. austrocochinchinensis purebreds and a unique germplasm. The Hainan population and the other subpopulations of XSBN of the species might have lost their genetic integrity. Our results revealed a clear genetic differentiation in the populations and subpopulations of Q. austrocochinchinensis and ongoing introgression between Q. austrocochinchinensis and Q. kerrii at the disturbed contact areas. Combining the results from genetic and morphological analyses, the conservation of subpopulation D2 should be prioritized. Conservation and restoration of the integrity of tropical ravine rainforest is an important long-term goal for the successful conservation of Q. austrocochinchinensis. The fine-scale landscape might play an essential role in shaping the spatial patterns of hybridization. Further studies are needed to evaluate these patterns and dynamics. PMID:28270827

Spatial and temporal aspects of the genetic structure of Juniperus communis populations.

PubMed

Merwe, M V; Winfield, M O; Arnold, G M; Parker, J S

2000-04-01

Juniperus communis is a dioecious, wind pollinated shrub or small tree that produces 'berries' (female cones) containing a small number of seeds that are thought to be dispersed by birds. The expectation, therefore, would be that populations of Juniper are genetically diverse with little structuring between them. In Britain, the species has two main centres of distribution: a highland zone in the north and west, in which populations are still large and sexually reproducing, and a southern zone on chalk downlands in which populations are small and fragmented and individuals suffer from a decline in fertility. Thus, one would expect the large sexually viable populations in the north to exhibit high levels of within-population genetic variation, while the declining southern populations would be genetically depauperate. The analysis of amplified fragment length polymorphisms (AFLPs) was used to test this hypothesis. Surprisingly, all populations studied showed high levels of genetic variation although there was clear structuring between populations. On the basis of the geographical structuring of the populations it was hypothesized that J. communis colonized Britain via three separate routes.

Iceberg Scour and Shell Damage in the Antarctic Bivalve Laternula elliptica

PubMed Central

Harper, Elizabeth M.; Clark, Melody S.; Hoffman, Joseph I.; Philipp, Eva E. R.; Peck, Lloyd S.; Morley, Simon A.

2012-01-01

We document differences in shell damage and shell thickness in a bivalve mollusc (Laternula elliptica) from seven sites around Antarctica with differing exposures to ice movement. These range from 60% of the sea bed impacted by ice per year (Hangar Cove, Antarctic Peninsula) to those protected by virtually permanent sea ice cover (McMurdo Sound). Patterns of shell damage consistent with blunt force trauma were observed in populations where ice scour frequently occurs; damage repair frequencies and the thickness of shells correlated positively with the frequency of iceberg scour at the different sites with the highest repair rates and thicker shells at Hangar Cove (74.2% of animals damaged) compared to the other less impacted sites (less than 10% at McMurdo Sound). Genetic analysis of population structure using Amplified Fragment Length Polymorphisms (AFLPs) revealed no genetic differences between the two sites showing the greatest difference in shell morphology and repair rates. Taken together, our results suggest that L. elliptica exhibits considerable phenotypic plasticity in response to geographic variation in physical disturbance. PMID:23029484

Biogeography and evolution of Thermococcus isolates from hydrothermal vent systems of the Pacific

PubMed Central

Price, Mark T.; Fullerton, Heather; Moyer, Craig L.

2015-01-01

Thermococcus is a genus of hyperthermophilic archaea that is ubiquitous in marine hydrothermal environments growing in anaerobic subsurface habitats but able to survive in cold oxygenated seawater. DNA analyses of Thermococcus isolates were applied to determine the relationship between geographic distribution and relatedness focusing primarily on isolates from the Juan de Fuca Ridge and South East Pacific Rise. Amplified fragment length polymorphism (AFLP) analysis and multilocus sequence typing (MLST) were used to resolve genomic differences in 90 isolates of Thermococcus, making biogeographic patterns and evolutionary relationships apparent. Isolates were differentiated into regionally endemic populations however there was also evidence in some lineages of cosmopolitan distribution. The biodiversity identified in Thermococcus isolates and presence of distinct lineages within the same vent site suggests the utilization of varying ecological niches in this genus. In addition to resolving biogeographic patterns in Thermococcus, this study has raised new questions about the closely related Pyrococcus genus. The phylogenetic placement of Pyrococcus type strains shows the close relationship between Thermococcus and Pyrococcus and the unresolved divergence of these two genera. PMID:26441901

Population Structure of Pythium irregulare, P. ultimum, and P. sylvaticum in Forest Nursery Soils of Oregon and Washington.

PubMed

Weiland, Jerry E; Garrido, Patricia; Kamvar, Zhian N; Espíndola, Andrés S; Marek, Stephen M; Grünwald, Niklaus J; Garzón, Carla D

2015-05-01

Pythium species are important soilborne pathogens occurring in the forest nursery industry of the Pacific Northwest. However, little is known about their genetic diversity or population structure and it is suspected that isolates are moved among forest nurseries on seedling stock and shared field equipment. In order to address these concerns, a total of 115 isolates of three Pythium species (P. irregulare, P. sylvaticum, and P. ultimum) were examined at three forest nurseries using simple sequence repeat (SSR) and amplified fragment length polymorphism (AFLP) markers. Analyses revealed distinct patterns of intraspecific variation for the three species. P. sylvaticum exhibited the most diversity, followed by P. irregulare, while substantial clonality was found in P. ultimum. For both P. irregulare and P. sylvaticum, but not P. ultimum, there was evidence for significant variation among nurseries. However, all three species also exhibited at least two distinct lineages not associated with the nursery of origin. Finally, evidence was found that certain lineages and clonal genotypes, including fungicide-resistant isolates, are shared among nurseries, indicating that pathogen movement has occurred.

Cytosine Methylation Alteration in Natural Populations of Leymus chinensis Induced by Multiple Abiotic Stresses

PubMed Central

Yu, Yingjie; Yang, Xuejiao; Wang, Huaying; Shi, Fengxue; Liu, Ying; Liu, Jushan; Li, Linfeng; Wang, Deli; Liu, Bao

2013-01-01

Background Human activity has a profound effect on the global environment and caused frequent occurrence of climatic fluctuations. To survive, plants need to adapt to the changing environmental conditions through altering their morphological and physiological traits. One known mechanism for phenotypic innovation to be achieved is environment-induced rapid yet inheritable epigenetic changes. Therefore, the use of molecular techniques to address the epigenetic mechanisms underpinning stress adaptation in plants is an important and challenging topic in biological research. In this study, we investigated the impact of warming, nitrogen (N) addition, and warming+nitrogen (N) addition stresses on the cytosine methylation status of Leymus chinensis Tzvel. at the population level by using the amplified fragment length polymorphism (AFLP), methylation-sensitive amplified polymorphism (MSAP) and retrotransposon based sequence-specific amplification polymorphism (SSAP) techniques. Methodology/Principal Findings Our results showed that, although the percentages of cytosine methylation changes in SSAP are significantly higher than those in MSAP, all the treatment groups showed similar alteration patterns of hypermethylation and hypomethylation. It meant that the abiotic stresses have induced the alterations in cytosine methylation patterns, and the levels of cytosine methylation changes around the transposable element are higher than the other genomic regions. In addition, the identification and analysis of differentially methylated loci (DML) indicated that the abiotic stresses have also caused targeted methylation changes at specific loci and these DML might have contributed to the capability of plants in adaptation to the abiotic stresses. Conclusions/Significance Our results demonstrated that abiotic stresses related to global warming and nitrogen deposition readily evoke alterations of cytosine methylation, and which may provide a molecular basis for rapid adaptation by the affected plant populations to the changed environments. PMID:23418457

AFLPs reveal different population genetic structure under contrasting environments in the marine snail Nucella lapillus L.

PubMed

Carro, Belén; Quintela, María; Ruiz, José Miguel; Barreiro, Rodolfo

2012-01-01

Dispersal has received growing attention in marine ecology, particularly since evidence obtained with up-to-date techniques challenged the traditional view. The dogwhelk Nucella lapillus L., a sedentary gastropod with direct development, is a good example: dispersal was traditionally assumed to be limited until studies with microsatellites disputed this idea. To shed some light on this controversy, the genetic structure of dogwhelk populations in northwest Spain was investigated with highly polymorphic AFLP markers giving special attention to the influence of hydrodynamic stress. In agreement with the expectations for a poor disperser, our results show a significant genetic structure at regional (<200 km) and areal scales (<15 km). However, the spatial genetic structure varied with wave-exposure in the present case study: IBD was evident under sheltered conditions but absent from the exposed area where genetic differentiation was stronger. Our results provide evidence that differences in wave-exposure can exert a detectable influence on the genetic structure of coastal organisms, even in species without a planktonic larva.

An evaluation of a new approach to the regeneration of Helichrysum italicum (Roth) G. Don, and the molecular characterization of the variation among sets of differently derived regenerants.

PubMed

Perrini, Rosaria; Alba, Vittorio; Ruta, Claudia; Morone-Fortunato, Irene; Blanco, Antonio; Montemurro, Cinzia

2009-01-01

A protocol for the induction of regeneration from leaves of Helichrysum italicum was established. Calli were found to form on the basal medium only when it was supplemented with thidiazuron (TDZ) alone or in combination with naphthalene acetic acid (NAA), with a percentage ranking of at least 80%. The hormone-free medium showed the highest percentage of shoot regeneration (62%) even though no callus formed. AFLP markers were employed to verify tissue culture-induced variation in the regenerated plantlets obtained by direct shoot regeneration or the indirect shoot regeneration process (callus formation). Seven out of the eleven AFLP primer pairs yielded polymorphic patterns. The average number of fragments per primer pair was 64.1. Singletons were represented by 12 (2.7%) fragments. Student's T-test was performed both on the average number of shared fragments and on the nucleotide diversity, and no significant statistical difference was observed between the two regeneration treatments.

Somatic Embryogenesis in Peach Palm Using the Thin Cell Layer Technique: Induction, Morpho-histological Aspects and AFLP Analysis of Somaclonal Variation

PubMed Central

Steinmacher, D. A.; Krohn, N. G.; Dantas, A. C. M.; Stefenon, V. M.; Clement, C. R.; Guerra, M. P.

2007-01-01

Background and Aims The thin cell layer (TCL) technique is based on the use of very small explants and has allowed enhanced in vitro morphogenesis in several plant species. The present study evaluated the TCL technique as a procedure for somatic embryo production and plantlet regeneration of peach palm. Methods TCL explants from different positions in the shoot apex and leaf sheath of peach palm were cultivated in MS culture medium supplemented with 0–600 µm Picloram in the presence of activated charcoal. The production of primary calli and embryogenic calli was evaluated in these different conditions. Histological and amplified fragment length polymorphism (AFLP) analyses were conducted to study in vitro morphogenetic responses and genetic stability, respectively, of the regenerated plantlets. Key Results Abundant primary callus induction was observed from TCLs of the shoot meristem in culture media supplemented with 150–600 µm Picloram (83–97 %, respectively). The production of embryogenic calli depends on Picloram concentration and explant position. The best response observed was 43 % embryogenic callus production from shoot meristem TCL on 300 µm Picloram. In maturation conditions, 34 ± 4 somatic embryos per embryogenic callus were obtained, and 45·0 ± 3·4 % of these fully developed somatic embryos were converted, resulting in plantlets ready for acclimatization, of which 80 % survived. Histological studies revealed that the first cellular division events occurred in cells adjacent to vascular tissue, resulting in primary calli, whose growth was ensured by a meristematic zone. A multicellular origin of the resulting somatic embryos arising from the meristematic zone is suggested. During maturation, histological analyses revealed bipolarization of the somatic embryos, as well as the development of new somatic embryos. AFLP analyses revealed that 92 % of the regenerated plantlets were true to type. The use of TCL explants considerably improves the number of calli and somatic embryos produced in comparison with previously described protocols for in vitro regeneration of peach palm. Conclusions The present study suggests that the TCL somatic embryogenesis protocol developed is feasible, although it still requires further optimization for in vitro multiplication of peach palm, especially the use of similar explants obtained from adult palm trees. PMID:17670751

Novel genomic resources for a climate change sensitive mammal: characterization of the American pika transcriptome.

PubMed

Lemay, Matthew A; Henry, Philippe; Lamb, Clayton T; Robson, Kelsey M; Russello, Michael A

2013-05-10

When faced with climate change, species must either shift their home range or adapt in situ in order to maintain optimal physiological balance with their environment. The American pika (Ochotona princeps) is a small alpine mammal with limited dispersal capacity and low tolerance for thermal stress. As a result, pikas have become an important system for examining biotic responses to changing climatic conditions. Previous research using amplified fragment length polymorphisms (AFLPs) has revealed evidence for environmental-mediated selection in O. princeps populations distributed along elevation gradients, yet the anonymity of AFLP loci and lack of available genomic resources precluded the identification of associated gene regions. Here, we harnessed next-generation sequencing technology in order to characterize the American pika transcriptome and identify a large suite of single nucleotide polymorphisms (SNPs), which can be used to elucidate elevation- and site-specific patterns of sequence variation. We constructed pooled cDNA libraries of O. princeps from high (1400 m) and low (300 m) elevation sites along a previously established transect in British Columbia. Transcriptome sequencing using the Roche 454 GS FLX titanium platform generated 780 million base pairs of data, which were assembled into 7,325 high coverage contigs. These contigs were used to identify 24,261 novel SNP loci. Using high resolution melt analysis, we developed 17 of these SNPs into genotyping assays, which were validated with independent DNA samples from British Columbia Canada and Oregon State USA. In addition, we detected haplotypes in the NADH dehydrogenase subunit 5 of the mitochondrial genome that were fixed and different among elevations, suggesting that this may be an informative target gene for studying the role of cellular respiration in local adaptation. We also identified contigs that were unique to each elevation, including a high elevation-specific contig that was a positive match with the hemoglobin alpha chain from the plateau pika, a species restricted to high elevation steppes in Asia. Elevation-specific contigs may represent candidate regions subject to differential levels of gene expression along this elevation gradient. To our knowledge, this is the first broad-scale, transcriptome-level study conducted within the Ochotonidae, providing novel genomic resources for studying pika ecology, behaviour and population history.

Using soil seed banks to assess temporal patterns of genetic variation in invasive plant populations.

PubMed

Fennell, Mark; Gallagher, Tommy; Vintro, Luis Leon; Osborne, Bruce

2014-05-01

Most research on the genetics of invasive plant species has focused on analyzing spatial differences among existing populations. Using a long-established Gunnera tinctoria population from Ireland, we evaluated the potential of using plants derived from seeds associated with different soil layers to track genetic variation through time. This species and site were chosen because (1) G. tinctoria produces a large and persistent seed bank; (2) it has been present in this locality, Sraheens, for ∼90 years; (3) the soil is largely undisturbed; and (4) the soil's age can be reliably determined radiometrically at different depths. Amplified fragment length polymorphic markers (AFLPs) were used to assess differences in the genetic structure of 75 individuals sampled from both the standing population and from four soil layers, which spanned 18 cm (estimated at ∼90 years based on (210)Pb and (137)Cs dating). While there are difficulties in interpreting such data, including accounting for the effects of selection, seed loss, and seed migration, a clear pattern of lower total allele counts, percentage polymorphic loci, and genetic diversity was observed in deeper soils. The greatest percentage increase in the measured genetic variables occurred prior to the shift from the lag to the exponential range expansion phases and may be of adaptive significance. These findings highlight that seed banks in areas with long-established invasive populations can contain valuable genetic information relating to invasion processes and as such, should not be overlooked.

Using soil seed banks to assess temporal patterns of genetic variation in invasive plant populations

PubMed Central

Fennell, Mark; Gallagher, Tommy; Vintro, Luis Leon; Osborne, Bruce

2014-01-01

Most research on the genetics of invasive plant species has focused on analyzing spatial differences among existing populations. Using a long-established Gunnera tinctoria population from Ireland, we evaluated the potential of using plants derived from seeds associated with different soil layers to track genetic variation through time. This species and site were chosen because (1) G. tinctoria produces a large and persistent seed bank; (2) it has been present in this locality, Sraheens, for ∼90 years; (3) the soil is largely undisturbed; and (4) the soil's age can be reliably determined radiometrically at different depths. Amplified fragment length polymorphic markers (AFLPs) were used to assess differences in the genetic structure of 75 individuals sampled from both the standing population and from four soil layers, which spanned 18 cm (estimated at ∼90 years based on 210Pb and 137Cs dating). While there are difficulties in interpreting such data, including accounting for the effects of selection, seed loss, and seed migration, a clear pattern of lower total allele counts, percentage polymorphic loci, and genetic diversity was observed in deeper soils. The greatest percentage increase in the measured genetic variables occurred prior to the shift from the lag to the exponential range expansion phases and may be of adaptive significance. These findings highlight that seed banks in areas with long-established invasive populations can contain valuable genetic information relating to invasion processes and as such, should not be overlooked. PMID:24967082

Identification, genetic localization, and allelic diversity of selectively amplified microsatellite polymorphic loci in lettuce and wild relatives (Lactuca spp.).

PubMed

Witsenboer, H; Michelmore, R W; Vogel, J

1997-12-01

Selectively amplified microsatellite polymorphic locus (SAMPL) analysis is a method of amplifying microsatellite loci using generic PCR primers. SAMPL analysis uses one AFLP primer in combination with a primer complementary to microsatellite sequences. SAMPL primers based on compound microsatellite sequences provided the clearest amplification patterns. We explored the potential of SAMPL analysis in lettuce to detect PCR-based codominant microsatellite markers. Fifty-eight SAMPLs were identified and placed on the genetic map. Seventeen were codominant. SAMPLs were dispersed with RFLP markers on 11 of the 12 main linkage groups in lettuce, indicating that they have a similar genomic distribution. Some but not all fragments amplified by SAMPL analysis were confirmed to contain microsatellite sequences by Southern hybridization. Forty-five cultivars of lettuce and five wild species of Lactuca were analyzed to determine the allelic diversity for codominant SAMPLs. From 3 to 11 putative alleles were found for each SAMPL; 2-6 alleles were found within Lactuca sativa and 1-3 alleles were found among the crisphead genotypes, the most genetically homogeneous plant type of L. sativa. This allelic diversity is greater than that found for RFLP markers. Numerous new alleles were observed in the wild species; however, there were frequent null alleles. Therefore, SAMPL analysis is more applicable to intraspecific than to interspecific comparisons. A phenetic analysis based on SAMPLs resulted in a dendrogram similar to those based on RFLP and AFLP markers.

Key processes for Cheirolophus (Asteraceae) diversification on oceanic islands inferred from AFLP data.

PubMed

Vitales, Daniel; García-Fernández, Alfredo; Pellicer, Jaume; Vallès, Joan; Santos-Guerra, Arnoldo; Cowan, Robyn S; Fay, Michael F; Hidalgo, Oriane; Garnatje, Teresa

2014-01-01

The radiation of the genus Cheirolophus (Asteraceae) in Macaronesia constitutes a spectacular case of rapid diversification on oceanic islands. Twenty species - nine of them included in the IUCN Red List of Threatened Species - have been described to date inhabiting the Madeiran and Canarian archipelagos. A previous phylogenetic study revealed that the diversification of Cheirolophus in Macaronesia started less than 2 Ma. As a result of such an explosive speciation process, limited phylogenetic resolution was reported, mainly due to the low variability of the employed molecular markers. In the present study, we used highly polymorphic AFLP markers to i) evaluate species' boundaries, ii) infer their evolutionary relationships and iii) investigate the patterns of genetic diversity in relation to the potential processes likely involved in the radiation of Cheirolophus. One hundred and seventy-two individuals representing all Macaronesian Cheirolophus species were analysed using 249 AFLP loci. Our results suggest that geographic isolation played an important role in this radiation process. This was likely driven by the combination of poor gene flow capacity and a good ability for sporadic long-distance colonisations. In addition, we also found some traces of introgression and incipient ecological adaptation, which could have further enhanced the extraordinary diversification of Cheirolophus in Macaronesia. Last, we hypothesize that current threat categories assigned to Macaronesian Cheirolophus species do not reflect their respective evolutionary relevance, so future evaluations of their conservation status should take into account the results presented here.

Key Processes for Cheirolophus (Asteraceae) Diversification on Oceanic Islands Inferred from AFLP Data

PubMed Central

Vitales, Daniel; García-Fernández, Alfredo; Pellicer, Jaume; Vallès, Joan; Santos-Guerra, Arnoldo; Cowan, Robyn S.; Fay, Michael F.; Hidalgo, Oriane; Garnatje, Teresa

2014-01-01

The radiation of the genus Cheirolophus (Asteraceae) in Macaronesia constitutes a spectacular case of rapid diversification on oceanic islands. Twenty species – nine of them included in the IUCN Red List of Threatened Species – have been described to date inhabiting the Madeiran and Canarian archipelagos. A previous phylogenetic study revealed that the diversification of Cheirolophus in Macaronesia started less than 2 Ma. As a result of such an explosive speciation process, limited phylogenetic resolution was reported, mainly due to the low variability of the employed molecular markers. In the present study, we used highly polymorphic AFLP markers to i) evaluate species' boundaries, ii) infer their evolutionary relationships and iii) investigate the patterns of genetic diversity in relation to the potential processes likely involved in the radiation of Cheirolophus. One hundred and seventy-two individuals representing all Macaronesian Cheirolophus species were analysed using 249 AFLP loci. Our results suggest that geographic isolation played an important role in this radiation process. This was likely driven by the combination of poor gene flow capacity and a good ability for sporadic long-distance colonisations. In addition, we also found some traces of introgression and incipient ecological adaptation, which could have further enhanced the extraordinary diversification of Cheirolophus in Macaronesia. Last, we hypothesize that current threat categories assigned to Macaronesian Cheirolophus species do not reflect their respective evolutionary relevance, so future evaluations of their conservation status should take into account the results presented here. PMID:25412495

AFLPs Reveal Different Population Genetic Structure under Contrasting Environments in the Marine Snail Nucella lapillus L.

PubMed Central

Carro, Belén; Quintela, María; Ruiz, José Miguel; Barreiro, Rodolfo

2012-01-01

Dispersal has received growing attention in marine ecology, particularly since evidence obtained with up-to-date techniques challenged the traditional view. The dogwhelk Nucella lapillus L., a sedentary gastropod with direct development, is a good example: dispersal was traditionally assumed to be limited until studies with microsatellites disputed this idea. To shed some light on this controversy, the genetic structure of dogwhelk populations in northwest Spain was investigated with highly polymorphic AFLP markers giving special attention to the influence of hydrodynamic stress. In agreement with the expectations for a poor disperser, our results show a significant genetic structure at regional (<200 km) and areal scales (<15 km). However, the spatial genetic structure varied with wave-exposure in the present case study: IBD was evident under sheltered conditions but absent from the exposed area where genetic differentiation was stronger. Our results provide evidence that differences in wave-exposure can exert a detectable influence on the genetic structure of coastal organisms, even in species without a planktonic larva. PMID:23185435

Co-occurrence and genotypic distribution of Phytophthora species recovered from watersheds and plant nurseries of eastern Tennessee.

PubMed

Hulvey, Jon; Gobena, Daniel; Finley, Ledare; Lamour, Kurt

2010-01-01

In 2008 statewide surveys of symptomatic foliage of nursery plants from Tennessee resulted in isolation of 43 isolates of Phytophthora spp. This sample set includes four described species (P. citrophthora, P. citricola, P. nicotianae, P. syringae), and a provisional species of Phytophthora ('P. hydropathica'). At the same time a stream-baiting survey was initiated to recover Phytophthora from eight watersheds in eastern Tennessee, some of which are near plant nurseries. Baiting was accomplished by submerging healthy Rhododendron leaves approximately 1 wk and isolation onto selective media. Six baiting periods were completed, and in total 98 Phytophthora isolates and 45 isolates of Pythium spp. were recovered. Three described species (P. citrophthora, P. citricola and P. irrigata) and the provisional species 'P. hydropathica' were obtained as well as three undescribed Phytophthora taxa and Pythium litorale. Isolates from both surveys were identified to species with morphology and the internal transcribed spacer (ITS) sequence. Isolates from species co-occurring in streams and nurseries (P. citricola, P. citrophthora and 'P. hydropathica') were characterized further with amplified fragment length polymorphism (AFLP) analyses and mefenoxam tolerance assays. Isolates representing a putative clonal genotype of P. citricola were obtained from both environmental and nursery sample sets.

Technological properties of bakers' yeasts in durum wheat semolina dough.

PubMed

Giannone, Virgilio; Longo, Chiara; Damigella, Arcangelo; Raspagliesi, Domenico; Spina, Alfio; Palumbo, Massimo

2010-04-01

Properties of 13 Saccharomyces cerevisiae strains isolated from different sources (traditional sourdoughs, industrial baking yeasts etc.) were studied in dough produced with durum wheat (Sicilian semolina, variety Mongibello). Durum wheat semolina and durum wheat flour are products prepared from grain of durum wheat (Triticum durum Desf.) by grinding or milling processes in which the bran and germ are essentially removed and the remainder is comminuted to a suitable degree of fineness. Acidification and leavening properties of the dough were evaluated. Strains isolated from traditional sourdoughs (DSM PST18864, DSM PST18865 and DSM PST18866) showed higher leavening power, valuable after the first and second hours of fermentation, than commercial baking yeasts. In particular the strain DSM PST 18865 has also been successfully tested in bakery companies for the improvement of production processes. Baking and staling tests were carried out on five yeast strains to evaluate their fermentation ability directly and their resistance to the staling process. Amplified fragment length polymorphism (fAFLP) was used to investigate genetic variations in the yeast strains. This study showed an appreciable biodiversity in the microbial populations of both wild and commercial yeast strains.

Susceptibility to Phytophthora ramorum in a key infectious host: landscape variation in host genotype, host phenotype, and environmental factors.

PubMed

Anacker, Brian L; Rank, Nathan E; Hüberli, Daniel; Garbelotto, Matteo; Gordon, Sarah; Harnik, Tami; Whitkus, Richard; Meentemeyer, Ross

2008-01-01

Sudden oak death is an emerging forest disease caused by the invasive pathogen Phytophthora ramorum. Genetic and environmental factors affecting susceptibility to P. ramorum in the key inoculum-producing host tree Umbellularia californica (bay laurel) were examined across a heterogeneous landscape in California, USA. Laboratory susceptibility trials were conducted on detached leaves and assessed field disease levels for 97 host trees from 12 225-m(2) plots. Genotype and phenotype characteristics were assessed for each tree. Effects of plot-level environmental conditions (understory microclimate, amount of solar radiation and topographic moisture potential) on disease expression were also evaluated. Susceptibility varied significantly among U. californica trees, with a fivefold difference in leaf lesion size. Lesion size was positively related to leaf area, but not to other phenotypic traits or to field disease level. Genetic diversity was structured at three spatial scales, but primarily among individuals within plots. Lesion size was significantly related to amplified fragment length polymorphism (AFLP) markers, but local environment explained most variation in field disease level. Thus, substantial genetic variation in susceptibility to P. ramorum occurs in its principal foliar host U. californica, but local environment mediates expression of susceptibility in nature.

Outbreak of encephalitic listeriosis in red-legged partridges (Alectoris rufa).

PubMed

Jeckel, S; Wood, A; Grant, K; Amar, C; King, S A; Whatmore, A M; Koylass, M; Anjum, M; James, J; Welchman, D de B

2015-01-01

An outbreak of neurological disease was investigated in red-legged partridges between 8 and 28 days of age. Clinical signs included torticollis, head tilt and incoordination and over an initial eight day period approximately 30-40 fatalities occurred per day. No significant gross post mortem findings were detected. Histopathological examination of the brain and bacterial cultures followed by partial sequencing confirmed a diagnosis of encephalitis due to Listeria monocytogenes. Further isolates were obtained from follow-up carcasses, environmental samples and pooled tissue samples of newly imported day-old chicks prior to placement on farm. These isolates had the same antibiotic resistance pattern as the isolate of the initial post mortem submission and belonged to the same fluorescent amplified fragment length polymorphism (fAFLP) subtype. This suggested that the isolates were very closely related or identical and that the pathogen had entered the farm with the imported day-old chicks, resulting in disease manifestation in partridges between 8 and 28 days of age. Reports of outbreaks of encephalitic listeriosis in avian species are rare and this is to the best of our knowledge the first reported outbreak in red-legged partridges.

High resolution linkage maps of the model organism Petunia reveal substantial synteny decay with the related genome of tomato.

PubMed

Bossolini, Eligio; Klahre, Ulrich; Brandenburg, Anna; Reinhardt, Didier; Kuhlemeier, Cris

2011-04-01

Two linkage maps were constructed for the model plant Petunia. Mapping populations were obtained by crossing the wild species Petunia axillaris subsp. axillaris with Petunia inflata, and Petunia axillaris subsp. parodii with Petunia exserta. Both maps cover the seven chromosomes of Petunia, and span 970 centimorgans (cM) and 700 cM of the genomes, respectively. In total, 207 markers were mapped. Of these, 28 are multilocus amplified fragment length polymorphism (AFLP) markers and 179 are gene-derived markers. For the first time we report on the development and mapping of 83 Petunia microsatellites. The two maps retain the same marker order, but display significant differences of recombination frequencies at orthologous mapping intervals. A complex pattern of genomic rearrangements was detected with the related genome of tomato (Solanum lycopersicum), indicating that synteny between Petunia and other Solanaceae crops has been considerably disrupted. The newly developed markers will facilitate the genetic characterization of mutants and ecological studies on genetic diversity and speciation within the genus Petunia. The maps will provide a powerful tool to link genetic and genomic information and will be useful to support sequence assembly of the Petunia genome.

Genetic Structure in the Seabuckthorn Carpenter Moth (Holcocerus hippophaecolus) in China: The Role of Outbreak Events, Geographical and Host Factors

PubMed Central

Tao, Jing; Chen, Min; Zong, Shi-Xiang; Luo, You-Qing

2012-01-01

Understanding factors responsible for structuring genetic diversity is of fundamental importance in evolutionary biology. The seabuckthorn carpenter moth (Holcocerus hippophaecolus Hua) is a native species throughout the north of China and is considered the main threat to seabuckthorn, Hippophae rhamnoides L. We assessed the influence of outbreaks, environmental factors and host species in shaping the genetic variation and structure of H. hippophaecolus by using Amplified Fragment Length Polymorphism (AFLP) markers. We rejected the hypothesis that outbreak-associated genetic divergence exist, as evidenced by genetic clusters containing a combination of populations from historical outbreak areas, as well as non-outbreak areas. Although a small number of markers (4 of 933 loci) were identified as candidates under selection in response to population densities. H. hippophaecolus also did not follow an isolation-by-distance pattern. We rejected the hypothesis that outbreak and drought events were driving the genetic structure of H. hippophaecolus. Rather, the genetic structure appears to be influenced by various confounding bio-geographical factors. There were detectable genetic differences between H. hippophaecolus occupying different host trees from within the same geographic location. Host-associated genetic divergence should be confirmed by further investigation. PMID:22291983

Cephalosporium maydis is a distinct species in the Gaeumannomyces-Harpophora species complex.

PubMed

Saleh, Amgad A; Leslie, John F

2004-01-01

Cephalosporium maydis is an important plant pathogen whose phylogenetic position relative to other fungi has not been established clearly. We compared strains of C. maydis, strains from several other plant-pathogenic Cephalosporium spp. and several possible relatives within the Gaeumannomyces-Harpophora species complex, to which C. maydis has been suggested to belong based on previous preliminary DNA sequence analyses. DNA sequences of the nuclear genes encoding the rDNA ITS region, β-tubulin, histone H3, and MAT-2 support the hypothesis that C. maydis is a distinct taxon within the Gaeumannomyces-Harpophora species complex. Based on amplified fragment length polymorphism (AFLP) profiles, C. maydis also is distinct from the other tested species of Cephalosporium, Phialophora sensu lato and members of Gaeumannomyces-Harpophora species complex, which supports its classification as Harpophora maydis. Oligonucleotide primers for H. maydis were developed that can be used in a PCR diagnostic protocol to rapidly and reliably detect and identify this pathogen. These diagnostic PCR primers will aid the detection of H. maydis in diseased maize because this fungus can be difficult to detect and isolate, and the movement of authentic cultures may be limited by quarantine restrictions.

Lactobacillus crustorum sp. nov., isolated from two traditional Belgian wheat sourdoughs.

PubMed

Scheirlinck, Ilse; Van der Meulen, Roel; Van Schoor, Ann; Huys, Geert; Vandamme, Peter; De Vuyst, Luc; Vancanneyt, Marc

2007-07-01

A polyphasic taxonomic study of the lactic acid bacteria (LAB) population in three traditional Belgian sourdoughs, sampled between 2002 and 2004, revealed a group of isolates that could not be assigned to any recognized LAB species. Initially, sourdough isolates were screened by means of (GTG)(5)-PCR fingerprinting. Four isolates displaying unique (GTG)(5)-PCR patterns were further investigated by means of phenylalanyl-tRNA synthase (pheS) gene sequence analysis and represented a bifurcated branch that could not be allocated to any LAB species present in the in-house pheS database. Their phylogenetic affiliation was determined using 16S rRNA gene sequence analysis and showed that the four sourdough isolates belong to the Lactobacillus plantarum group with Lactobacillus mindensis, Lactobacillus farciminis and Lactobacillus nantensis as closest relatives. Further genotypic and phenotypic studies, including whole-cell protein analysis (SDS-PAGE), amplified fragment length polymorphism (AFLP) fingerprinting, DNA-DNA hybridization, DNA G+C content analysis, growth characteristics and biochemical features, demonstrated that the new sourdough isolates represent a novel Lactobacillus species for which the name Lactobacillus crustorum sp. nov. is proposed. The type strain of the new species is LMG 23699(T) (=CCUG 53174(T)).

Universal Plant DNA Barcode Loci May Not Work in Complex Groups: A Case Study with Indian Berberis Species

PubMed Central

Roy, Sribash; Tyagi, Antariksh; Shukla, Virendra; Kumar, Anil; Singh, Uma M.; Chaudhary, Lal Babu; Datt, Bhaskar; Bag, Sumit K.; Singh, Pradhyumna K.; Nair, Narayanan K.; Husain, Tariq; Tuli, Rakesh

2010-01-01

Background The concept of DNA barcoding for species identification has gained considerable momentum in animals because of fairly successful species identification using cytochrome oxidase I (COI). In plants, matK and rbcL have been proposed as standard barcodes. However, barcoding in complex genera is a challenging task. Methodology and Principal Findings We investigated the species discriminatory power of four reportedly most promising plant DNA barcoding loci (one from nuclear genome- ITS, and three from plastid genome- trnH-psbA, rbcL and matK) in species of Indian Berberis L. (Berberidaceae) and two other genera, Ficus L. (Moraceae) and Gossypium L. (Malvaceae). Berberis species were delineated using morphological characters. These characters resulted in a well resolved species tree. Applying both nucleotide distance and nucleotide character-based approaches, we found that none of the loci, either singly or in combinations, could discriminate the species of Berberis. ITS resolved all the tested species of Ficus and Gossypium and trnH-psbA resolved 82% of the tested species in Ficus. The highly regarded matK and rbcL could not resolve all the species. Finally, we employed amplified fragment length polymorphism test in species of Berberis to determine their relationships. Using ten primer pair combinations in AFLP, the data demonstrated incomplete species resolution. Further, AFLP analysis showed that there was a tendency of the Berberis accessions to cluster according to their geographic origin rather than species affiliation. Conclusions/Significance We reconfirm the earlier reports that the concept of universal barcode in plants may not work in a number of genera. Our results also suggest that the matK and rbcL, recommended as universal barcode loci for plants, may not work in all the genera of land plants. Morphological, geographical and molecular data analyses of Indian species of Berberis suggest probable reticulate evolution and thus barcode markers may not work in this case. PMID:21060687

Population genetic structure and phylogeographical pattern of rice grasshopper, Oxya hyla intricata, across Southeast Asia.

PubMed

Li, Tao; Zhang, Min; Qu, Yanhua; Ren, Zhumei; Zhang, Jianzhen; Guo, Yaping; Heong, K L; Villareal, Bong; Zhong, Yang; Ma, Enbo

2011-04-01

The rice grasshopper, Oxya hyla intricata, is a rice pest in Southeast Asia. In this study, population genetic diversity and structure of this Oxya species was examined using both DNA sequences and AFLP technology. The samples of 12 populations were collected from four Southeast Asian countries, among which 175 individuals were analysed using mitochondrial DNA cytochrome c oxidase subunit I (COI) sequences, and 232 individuals were examined using amplified fragment length polymorphisms (AFLP) to test whether the phylogeographical pattern and population genetics of this species are related to past geological events and/or climatic oscillations. No obvious trend of genetic diversity was found along a latitude/longitude gradient among different geographical groups. Phylogenetic analysis indicated three deep monophyletic clades that approximately correspond to three geographical regions separated by high mountains and a deep strait, and TCS analysis also revealed three disconnected networks, suggesting that spatial and temporal separations by vicariance, which were also supported by AMOVA as a source of the molecular variance presented among groups. Gene flow analysis showed that there had been frequent historical gene flow among local populations in different regions, but the networks exhibited no shared haplotype among populations. In conclusion, the past geological events and climatic fluctuations are the most important factor on the phylogeographical structure and genetic patterns of O. hyla intricata in Southeast Asia. Habitat, vegetation, and anthropogenic effect may also contribute to gene flow and introgression of this species. Moreover, temperature, abundant rainfall and a diversity of graminaceous species are beneficial for the migration of O. hyla intricata. High haplotype diversity, deep phylogenetic division, negative Fu's F (s) values and unimodal and multimodal distribution shapes all suggest a complicated demographic expansion pattern of these O. hyla intricata populations, which might have been caused by climatic oscillations during glacial periods in the Quaternary.

Molecular characterization of Cryptococcus neoformans isolated from the environment in Beijing, China.

PubMed

Dou, Hongtao; Wang, Huizhu; Xie, Shaowei; Chen, Xinxin; Xu, Zhipeng; Xu, Yingchun

2017-10-01

The molecular type of environmental Cryptococcus neoformans in Beijing was not clear. Our study aims to reveal the molecular characterization of C. neoformans complex from environment in Beijing, China. A total of 435 samples of pigeon droppings from 11 different homes in Beijing were collected from August to November in 2015. Pigeon droppings were inoculated onto caffeic acid cornmeal agar (CACA) to screen C. neoformans complex. Bruker Biotyper matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) was performed for species identification. Serotype and mating type was determined by specific primers. Restriction fragment length polymorphisms of URA5 (URA5-RFLP) were applied to genotype. Multi-locus sequence typing (MLST) was done for further identification and sequence type (ST) determination. Altogether, 81 isolates of C. neoformans AFLP1/VNI were recognized from 435 pigeon droppings in this study. The positive rate for C. neoformans AFLP1/VNI from pigeon droppings in different homes varied from 5.0% to 52.6%, the average was 20.2%. All of these cryptococcal strains were serotype A, MATα. They were genotyped as VNI by URA5-RFLP and were confirmed by MLST. No other molecular types of C. neoformans and Cryptococcus gattii isolates were isolated. Their STs were identified as ST 31 (n = 54, 66.7%), followed by ST 53 (n = 10), ST 191 (n = 8), ST 5 (n = 5), ST 57 (n = 3), and ST 38 (n = 1). We concluded that not only clinical but also environmental isolates of C. neoformans need to be investigated more deeply and more extensively. The virulence difference between ST 5 and ST 31 need to be explored in the future. © The Author 2017. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Transcriptional responses in Honey Bee larvae infected with chalkbrood fungus

PubMed Central

2010-01-01

Background Diseases and other stress factors working synergistically weaken honey bee health and may play a major role in the losses of bee populations in recent years. Among a large number of bee diseases, chalkbrood has been on the rise. We present here the experimental identification of honey bee genes that are differentially expressed in response to infection of honey bee larvae with the chalkbrood fungus, Ascosphaera apis. Results We used cDNA-AFLP ®Technology to profile transcripts in infected and uninfected bee larvae. From 64 primer combinations, over 7,400 transcriptionally-derived fragments were obtained A total of 98 reproducible polymorphic cDNA-AFLP fragments were excised and sequenced, followed by quantitative real-time RT-PCR (qRT-PCR) analysis of these and additional samples. We have identified a number of differentially-regulated transcripts that are implicated in general mechanisms of stress adaptation, including energy metabolism and protein transport. One of the most interesting differentially-regulated transcripts is for a chitinase-like enzyme that may be linked to anti-fungal activities in the honey bee larvae, similarly to gut and fat-body specific chitinases found in mosquitoes and the red flour beetle. Surprisingly, we did not find many components of the well-characterized NF-κB intracellular signaling pathways to be differentially-regulated using the cDNA-AFLP approach. Therefore, utilizing qRT-PCR, we probed some of the immune related genes to determine whether the lack of up-regulation of their transcripts in our analysis can be attributed to lack of immune activation or to limitations of the cDNA-AFLP approach. Conclusions Using a combination of cDNA-AFLP and qRT-PCR analyses, we were able to determine several key transcriptional events that constitute the overall effort in the honey bee larvae to fight natural fungal infection. Honey bee transcripts identified in this study are involved in critical functions related to transcriptional regulation, apoptotic degradation of ubiquitinated proteins, nutritional regulation, and RNA processing. We found that immune regulation of the anti-fungal responses in honey bee involves highly coordinated activation of both NF-κB signaling pathways, leading to production of anti-microbial peptides. Significantly, activation of immune responses in the infected bee larvae was associated with down-regulation of major storage proteins, leading to depletion of nutritional resources. PMID:20565973

Transcriptional responses in honey bee larvae infected with chalkbrood fungus.

PubMed

Aronstein, Katherine A; Murray, Keith D; Saldivar, Eduardo

2010-06-21

Diseases and other stress factors working synergistically weaken honey bee health and may play a major role in the losses of bee populations in recent years. Among a large number of bee diseases, chalkbrood has been on the rise. We present here the experimental identification of honey bee genes that are differentially expressed in response to infection of honey bee larvae with the chalkbrood fungus, Ascosphaera apis. We used cDNA-AFLP Technology to profile transcripts in infected and uninfected bee larvae. From 64 primer combinations, over 7,400 transcriptionally-derived fragments were obtained A total of 98 reproducible polymorphic cDNA-AFLP fragments were excised and sequenced, followed by quantitative real-time RT-PCR (qRT-PCR) analysis of these and additional samples.We have identified a number of differentially-regulated transcripts that are implicated in general mechanisms of stress adaptation, including energy metabolism and protein transport. One of the most interesting differentially-regulated transcripts is for a chitinase-like enzyme that may be linked to anti-fungal activities in the honey bee larvae, similarly to gut and fat-body specific chitinases found in mosquitoes and the red flour beetle. Surprisingly, we did not find many components of the well-characterized NF-kappaB intracellular signaling pathways to be differentially-regulated using the cDNA-AFLP approach. Therefore, utilizing qRT-PCR, we probed some of the immune related genes to determine whether the lack of up-regulation of their transcripts in our analysis can be attributed to lack of immune activation or to limitations of the cDNA-AFLP approach. Using a combination of cDNA-AFLP and qRT-PCR analyses, we were able to determine several key transcriptional events that constitute the overall effort in the honey bee larvae to fight natural fungal infection. Honey bee transcripts identified in this study are involved in critical functions related to transcriptional regulation, apoptotic degradation of ubiquitinated proteins, nutritional regulation, and RNA processing. We found that immune regulation of the anti-fungal responses in honey bee involves highly coordinated activation of both NF-kappaB signaling pathways, leading to production of anti-microbial peptides. Significantly, activation of immune responses in the infected bee larvae was associated with down-regulation of major storage proteins, leading to depletion of nutritional resources.

Microsatellites for the genus Cucurbita and an SSR-based genetic linkage map of Cucurbita pepo L.

PubMed Central

Gong, L.; Stift, G.; Kofler, R.; Pachner, M.

2008-01-01

Until recently, only a few microsatellites have been available for Cucurbita, thus their development is highly desirable. The Austrian oil-pumpkin variety Gleisdorfer Ölkürbis (C. pepo subsp. pepo) and the C. moschata cultivar Soler (Puerto Rico) were used for SSR development. SSR-enriched partial genomic libraries were established and 2,400 clones were sequenced. Of these 1,058 (44%) contained an SSR at least four repeats long. Primers were designed for 532 SSRs; 500 primer pairs produced fragments of expected size. Of these, 405 (81%) amplified polymorphic fragments in a set of 12 genotypes: three C. moschata, one C. ecuadorensis, and eight C. pepo representing all eight cultivar groups. On an average, C. pepo and C. moschata produced 3.3 alleles per primer pair, showing high inter-species transferability. There were 187 SSR markers detecting polymorphism between the USA oil-pumpkin variety “Lady Godiva” (O5) and the Italian crookneck variety “Bianco Friulano” (CN), which are the parents of our previous F2 mapping population. It has been used to construct the first published C. pepo map, containing mainly RAPD and AFLP markers. Now the updated map comprises 178 SSRs, 244 AFLPs, 230 RAPDs, five SCARs, and two morphological traits (h and B). It contains 20 linkage groups with a map density of 2.9 cM. The observed genome coverage (Co) is 86.8%. Electronic supplementary material The online version of this article (doi:10.1007/s00122-008-0750-2) contains supplementary material, which is available to authorized users. PMID:18379753

Effect of latitudinal gradient and impact of logging on genetic diversity of Cedrela lilloi along the Argentine Yungas Rainforest

PubMed Central

Inza, Maria V; Zelener, Noga; Fornes, Luis; Gallo, Leonardo A

2012-01-01

Cedrela lilloi C. DC. (cedro coya, Meliaceae), an important south American timber species, has been historically overexploited through selective logging in Argentine Yungas Rainforest. Management and conservation programs of the species require knowledge of its genetic variation patterns; however, no information is available. Molecular genetic variability of the species was characterized to identify high-priority populations for conservation and domestication purposes. Fourteen native populations (160 individuals) along a latitudinal gradient and with different logging's intensities were assessed by 293 polymorphic AFLP (amplified fragment length polymorphism) markers. Genetic diversity was low (Ht = 0.135), according to marginal location of the species in Argentina. Most of the diversity was distributed within populations (87%). Northern populations showed significant higher genetic diversity (R2= 0.69) that agreed with latitudinal pattern of distribution of taxonomic diversity in the Yungas. Three clusters were identified by Bayesian analysis in correspondence with northern, central, and southern Yungas. An analysis of molecular variance (AMOVA) revealed significant genetic differences among latitudinal clusters even when logging (ΦRT = 0.07) and unlogging populations (ΦPT = 0.10) were separately analyzed. Loss of genetic diversity with increasing logging intensity was observed between neighboring populations with different disturbance (ΦPT = 0.03–0.10). Bottlenecks in disturbed populations are suggested as the main cause. Our results emphasize both: the necessity of maintaining the genetic diversity in protected areas that appear as possible long-term refuges of the species; and to rescue for the national system of protected areas some high genetic diversity populations that are on private fields. PMID:23170208

Effects of long-term space condition on cell ultrastructure and the molecular level change of the tomato

NASA Astrophysics Data System (ADS)

Jinying, L.; Min, L.; Huai, X.; Yi, P.; Chunhua, Z.; Nechitalo, G.

Effects of long-term exposure to physical factors of space flight on dormant seeds were studied on plants derived from tomato seeds flown for 6 years on board of the space station MIR Upon return to the Earth the seeds were germinated and grown to maturity Samples of plants were compared to plants from parallel ground-based controls Various differences of ultrastructure of the tomato leaf cell were observed with an electron microscope One plant carried by space station has the anatomy of leaves with a three-layered palisade tissue and other plants similar with ground controls have the anatomy of leaves with a one-layered palisade tissue The number of starch grains per chloroplast of every space-treated tomato leaf increased significantly compared with that of the ground control The leaf cell walls of two plants carried by space station became contracted and deformed The size of chloroplast in some space-treated plants was larger and the lamellae s structure of some chloroplasts turned curvature and loose The results obtained point out to significant changes occurring on the molecular level among the space-flight treated seedlings and the ground control The leaves of plants were used for AFLP Amplification Fragment Length Polymorphism analysis For the first generation space-flight treated tomato plants among 64 pairs of primers used in this experiment 43 primers generated the same DNA bands type and 21 primers generated a different DNA band type 2582 DNA bands were produced among which 34 DNA bands were polymorphic with the percentage

A Single, Plastic Population of Mycosphaerella pinodes Causes Ascochyta Blight on Winter and Spring Peas (Pisum sativum) in France

PubMed Central

Guibert, Michèle; Leclerc, Aurélie; Andrivon, Didier; Tivoli, Bernard

2012-01-01

Plant diseases are caused by pathogen populations continuously subjected to evolutionary forces (genetic flow, selection, and recombination). Ascochyta blight, caused by Mycosphaerella pinodes, is one of the most damaging necrotrophic pathogens of field peas worldwide. In France, both winter and spring peas are cultivated. Although these crops overlap by about 4 months (March to June), primary Ascochyta blight infections are not synchronous on the two crops. This suggests that the disease could be due to two different M. pinodes populations, specialized on either winter or spring pea. To test this hypothesis, 144 pathogen isolates were collected in the field during the winter and spring growing seasons in Rennes (western France), and all the isolates were genotyped using amplified fragment length polymorphism (AFLP) markers. Furthermore, the pathogenicities of 33 isolates randomly chosen within the collection were tested on four pea genotypes (2 winter and 2 spring types) grown under three climatic regimes, simulating winter, late winter, and spring conditions. M. pinodes isolates from winter and spring peas were genetically polymorphic but not differentiated according to the type of cultivars. Isolates from winter pea were more pathogenic than isolates from spring pea on hosts raised under winter conditions, while isolates from spring pea were more pathogenic than those from winter pea on plants raised under spring conditions. These results show that disease developed on winter and spring peas was initiated by a single population of M. pinodes whose pathogenicity is a plastic trait modulated by the physiological status of the host plant. PMID:23023742

Functional Multi-Locus QTL Mapping of Temporal Trends in Scots Pine Wood Traits

PubMed Central

Li, Zitong; Hallingbäck, Henrik R.; Abrahamsson, Sara; Fries, Anders; Gull, Bengt Andersson; Sillanpää, Mikko J.; García-Gil, M. Rosario

2014-01-01

Quantitative trait loci (QTL) mapping of wood properties in conifer species has focused on single time point measurements or on trait means based on heterogeneous wood samples (e.g., increment cores), thus ignoring systematic within-tree trends. In this study, functional QTL mapping was performed for a set of important wood properties in increment cores from a 17-yr-old Scots pine (Pinus sylvestris L.) full-sib family with the aim of detecting wood trait QTL for general intercepts (means) and for linear slopes by increasing cambial age. Two multi-locus functional QTL analysis approaches were proposed and their performances were compared on trait datasets comprising 2 to 9 time points, 91 to 455 individual tree measurements and genotype datasets of amplified length polymorphisms (AFLP), and single nucleotide polymorphism (SNP) markers. The first method was a multilevel LASSO analysis whereby trend parameter estimation and QTL mapping were conducted consecutively; the second method was our Bayesian linear mixed model whereby trends and underlying genetic effects were estimated simultaneously. We also compared several different hypothesis testing methods under either the LASSO or the Bayesian framework to perform QTL inference. In total, five and four significant QTL were observed for the intercepts and slopes, respectively, across wood traits such as earlywood percentage, wood density, radial fiberwidth, and spiral grain angle. Four of these QTL were represented by candidate gene SNPs, thus providing promising targets for future research in QTL mapping and molecular function. Bayesian and LASSO methods both detected similar sets of QTL given datasets that comprised large numbers of individuals. PMID:25305041

Functional multi-locus QTL mapping of temporal trends in Scots pine wood traits.

PubMed

Li, Zitong; Hallingbäck, Henrik R; Abrahamsson, Sara; Fries, Anders; Gull, Bengt Andersson; Sillanpää, Mikko J; García-Gil, M Rosario

2014-10-09

Quantitative trait loci (QTL) mapping of wood properties in conifer species has focused on single time point measurements or on trait means based on heterogeneous wood samples (e.g., increment cores), thus ignoring systematic within-tree trends. In this study, functional QTL mapping was performed for a set of important wood properties in increment cores from a 17-yr-old Scots pine (Pinus sylvestris L.) full-sib family with the aim of detecting wood trait QTL for general intercepts (means) and for linear slopes by increasing cambial age. Two multi-locus functional QTL analysis approaches were proposed and their performances were compared on trait datasets comprising 2 to 9 time points, 91 to 455 individual tree measurements and genotype datasets of amplified length polymorphisms (AFLP), and single nucleotide polymorphism (SNP) markers. The first method was a multilevel LASSO analysis whereby trend parameter estimation and QTL mapping were conducted consecutively; the second method was our Bayesian linear mixed model whereby trends and underlying genetic effects were estimated simultaneously. We also compared several different hypothesis testing methods under either the LASSO or the Bayesian framework to perform QTL inference. In total, five and four significant QTL were observed for the intercepts and slopes, respectively, across wood traits such as earlywood percentage, wood density, radial fiberwidth, and spiral grain angle. Four of these QTL were represented by candidate gene SNPs, thus providing promising targets for future research in QTL mapping and molecular function. Bayesian and LASSO methods both detected similar sets of QTL given datasets that comprised large numbers of individuals. Copyright © 2014 Li et al.

Epigenetic patterns newly established after interspecific hybridization in natural populations of Solanum

PubMed Central

Cara, Nicolás; Marfil, Carlos F; Masuelli, Ricardo W

2013-01-01

Interspecific hybridization is known for triggering genetic and epigenetic changes, such as modifications on DNA methylation patterns and impact on phenotypic plasticity and ecological adaptation. Wild potatoes (Solanum, section Petota) are adapted to multiple habitats along the Andes, and natural hybridizations have proven to be a common feature among species of this group. Solanum × rechei, a recently formed hybrid that grows sympatrically with the parental species S. kurtzianum and S. microdontum, represents an ideal model for studying the ecologically and evolutionary importance of hybridization in generating of epigenetic variability. Genetic and epigenetic variability and their correlation with morphological variation were investigated in wild and ex situ conserved populations of these three wild potato species using amplified fragment length polymorphism (AFLP) and methylation-sensitive amplified polymorphism (MSAP) techniques. We observed that novel methylation patterns doubled the number of novel genetic patterns in the hybrid and that the morphological variability measured on 30 characters had a higher correlation with the epigenetic than with the genetic variability. Statistical comparison of methylation levels suggested that the interspecific hybridization induces genome demethylation in the hybrids. A Bayesian analysis of the genetic data reveled the hybrid nature of S. × rechei, with genotypes displaying high levels of admixture with the parental species, while the epigenetic information assigned S. × rechei to its own cluster with low admixture. These findings suggested that after the hybridization event, a novel epigenetic pattern was rapidly established, which might influence the phenotypic plasticity and adaptation of the hybrid to new environments. PMID:24198938

Multi-scale variation in spatial heterogeneity for microbial community structure in an eastern Virginia agricultural field

NASA Technical Reports Server (NTRS)

Franklin, Rima B.; Mills, Aaron L.

2003-01-01

To better understand the distribution of soil microbial communities at multiple spatial scales, a survey was conducted to examine the spatial organization of community structure in a wheat field in eastern Virginia (USA). Nearly 200 soil samples were collected at a variety of separation distances ranging from 2.5 cm to 11 m. Whole-community DNA was extracted from each sample, and community structure was compared using amplified fragment length polymorphism (AFLP) DNA fingerprinting. Relative similarity was calculated between each pair of samples and compared using geostatistical variogram analysis to study autocorrelation as a function of separation distance. Spatial autocorrelation was found at scales ranging from 30 cm to more than 6 m, depending on the sampling extent considered. In some locations, up to four different correlation length scales were detected. The presence of nested scales of variability suggests that the environmental factors regulating the development of the communities in this soil may operate at different scales. Kriging was used to generate maps of the spatial organization of communities across the plot, and the results demonstrated that bacterial distributions can be highly structured, even within a habitat that appears relatively homogeneous at the plot and field scale. Different subsets of the microbial community were distributed differently across the plot, and this is thought to be due to the variable response of individual populations to spatial heterogeneity associated with soil properties. c2003 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.

Clonal plasticity and diversity facilitates the adaptation of Rhododendron aureum Georgi to alpine environment

PubMed Central

Wang, Xiaolong; Zhao, Wei; Li, Lin; You, Jian; Ni, Biao

2018-01-01

Four small oval populations and five large intensive populations of Rhododendron aureum growing at the alpine in Changbai Mountain (China) were studied in two types of habitat (in the tundra and in Betula ermanii forest). Identification and delimitation of genets were inferred from excavation in small populations and from amplified fragment length polymorphism (AFLP) markers by the standardized sampling design in large populations. Clonal architecture and clonal diversity were then estimated. For the four small populations, they were monoclonal, the spacer length (18.6 ± 5.6 in tundra, 29.7 ± 9.7 in Betula ermanii forest, P < 0.05) was shorter and branching intensity (136.7 ± 32.9 in tundra, 43.4 ± 12.3 in Betula ermanii forest, P < 0.05) was higher in the tundra than that in Betula ermanii forest. For the five large populations, they were composed of multiple genets with high level of clonal diversity (Simpson’s index D = 0.84, clonal richness R = 0.25, Fager's evenness E = 0.85); the spatial distribution of genets showed that the clonal growth strategy of R. aureum exhibits both guerilla and phalanx. Our results indicate that the clonal plasticity of R. aureum could enhance exploitation of resource heterogeneity and in turn greatly contribute to maintenance or improvement of fitness and the high clonal diversity of R. aureum increase the evolutionary rates to adapt the harsh alpine environment in Changbai Mountain. PMID:29746526

Comparative assessment of genetic and epigenetic variation among regenerants of potato (Solanum tuberosum) derived from long-term nodal tissue-culture and cell selection.

PubMed

Dann, Alison L; Wilson, Calum R

2011-04-01

Three long-term nodal tissued cultured Russet Burbank potato clones and nine thaxtomin A-treated regenerant lines, derived from the nodal lines, were assessed for genetic and epigenetic (in the form of DNA methylation) differences by AFLP and MSAP. The treated regenerant lines were originally selected for superior resistance to common scab disease and acceptable tuber yield in pot and field trials. The long-term, tissue culture clone lines exhibited genetic (8.75-15.63% polymorphisms) and epigenetic (12.56-26.13% polymorphisms) differences between them and may represent a stress response induced by normal plant growth disruption. The thaxtomin A-treated regenerant lines exhibited much higher significant (p < 0.05) genetic (2-29.38%) and epigenetic (45.22-51.76%) polymorphisms than the nodal cultured parent clones. Methylation-sensitive mutations accumulated within the regenerant lines are significantly correlated (p < 0.05) to disease resistance. However, linking phenotypic differences that could be of benefit to potato growers, to single gene sequence polymorphisms in a tetraploid plant such as the potato would be extremely difficult since it is assumed many desirable traits are under polygenic control.

Development and characterization of 16 polymorphic microsatellite markers from Taiwan cow-tail fir, Keteleeria davidiana var. formosana (Pinaceae) and cross-species amplification in other Keteleeria taxa

PubMed Central

2014-01-01

Background Keteleeria davidiana var. formosana (Pinaceae), Taiwan cow-tail fir, is an endangered species listed on the IUCN Red List of Threatened Species and only two populations remain, both on the Taiwan Island. Sixteen polymorphic microsatellite loci were developed in an endangered and endemic gymnosperm species, Keteleeria davidiana var. formosana, and were tested in an additional 6 taxa, K. davidiana var. calcarea, K. davidiana var. chienpeii, K. evelyniana, K. fortunei, K. fortunei var. cyclolepis, and K. pubescens, to evaluate the genetic variation available for conservation management and to reconstruct the phylogeographic patterns of this ancient lineage. Findings Polymorphic primer sets were developed from K. davidiana var. formosana using the modified AFLP and magnetic bead enrichment method. The number of alleles ranged from 3 to 16, with the observed heterozygosity ranging from 0.28 to 1.00. All of the loci were found to be interspecifically amplifiable. Conclusions These polymorphic and transferable loci will be potentially useful for future studies that will focus on identifying distinct evolutionary units within species and establishing the phylogeographic patterns and the process of speciation among closely related species. PMID:24755442

Linkage mapping in tetraploid willows: segregation of molecular markers and estimation of linkage phases support an allotetraploid structure for Salix alba x Salix fragilis interspecific hybrids.

PubMed

Barcaccia, G; Meneghetti, S; Albertini, E; Triest, L; Lucchin, M

2003-02-01

Salix alba-Salix fragilis complex includes closely related dioecious polyploid species, which are obligate outcrossers. Natural populations of these willows and their hybrids are represented by a mixture of highly heterozygous genotypes sharing a common gene pool. Since nothing is known about their genomic constitution, tetraploidy (2n=4x=76) in willow species makes basic and applied genetic studies difficult. We have used a two-way pseudotestcross strategy and single-dose markers (SDMs) to construct the first linkage maps for both pistillate and staminate willows. A total of 242 amplified fragment length polymorphisms (AFLPs) and 50 selective amplifications of microsatellite polymorphic loci (SAMPL) markers, which showed 1:1 segregation in the F(1) mapping populations, were used in linkage analysis. In S. alba, 73 maternal and 48 paternal SDMs were mapped to 19 and 16 linkage groups covering 708 and 339 cM, respectively. In S. fragilis, 13 maternal and 33 paternal SDMs were mapped in six and 14 linkage groups covering 98 and 321 cM, respectively. For most cosegregation groups, a comparable number of markers linked in coupling and repulsion was identified. This finding suggests that most of chromosomes pair preferentially as occurs in allotetraploid species exhibiting disomic inheritance. The detection of 10 pairs of marker alleles from single parents showing codominant inheritance strengthens this hypothesis. The fact that, of the 1122 marker loci identified in the two male and female parents, the vast majority (77.5%) were polymorphic and as few as 22.5% were shared between parental species highlight that S. alba and S. fragilis genotypes are differentiated. The highly difference between S. alba- and S. fragilis-specific markers found in both parental combinations (on average, 65.3 vs 34.7%, respectively) supports the (phylogenetic) hypothesis that S. fragilis is derived from S. alba-like progenitors.

Patterns of cytosine methylation in an elite rice hybrid and its parental lines, detected by a methylation-sensitive amplification polymorphism technique.

PubMed

Xiong, L Z; Xu, C G; Saghai Maroof, M A; Zhang, Q

1999-04-01

DNA methylation is known to play an important role in the regulation of gene expression in eukaryotes. In this study, we assessed the extent and pattern of cytosine methylation in the rice genome, using the technique of methylation-sensitive amplified polymorphism (MSAP), which is a modification of the amplified fragment length polymorphism (AFLP) method that makes use of the differential sensitivity of a pair of isoschizomers to cytosine methylation. The tissues assayed included seedlings and flag leaves of an elite rice hybrid, Shanyou 63, and the parental lines Zhenshan 97 and Minghui 63. In all, 1076 fragments, each representing a recognition site cleaved by either or both of the isoschizomers, were amplified using 16 pairs of selective primers. A total of 195 sites were found to be methylated at cytosines in one or both parents, and the two parents showed approximately the same overall degree of methylation (16.3%), as revealed by the incidence of differential digestion by the isoschizomers. Four classes of patterns were identified in a comparative assay of cytosine methylation in the parents and hybrid; increased methylation was detected in the hybrid compared to the parents at some of the recognition sites, while decreased methylation in the hybrid was detected at other sites. A small proportion of the sites was found to be differentially methylated in seedlings and flag leaves; DNA from young seedlings was methylated to a greater extent than that from flag leaves. Almost all of the methylation patterns detected by MSAP could be confirmed by Southern analysis using the isolated amplified fragments as probes. The results clearly demonstrate that the MSAP technique is highly efficient for large-scale detection of cytosine methylation in the rice genome. We believe that the technique can be adapted for use in other plant species.

Genetic diversity, population structure, and traditional culture of Camellia reticulata.

PubMed

Xin, Tong; Huang, Weijuan; De Riek, Jan; Zhang, Shuang; Ahmed, Selena; Van Huylenbroeck, Johan; Long, Chunlin

2017-11-01

Camellia reticulata is an arbor tree that has been cultivated in southwestern China by various sociolinguistic groups for esthetic purposes as well as to derive an edible seed oil. This study examined the influence of management, socio-economic factors, and religion on the genetic diversity patterns of Camellia reticulata utilizing a combination of ethnobotanical and molecular genetic approaches. Semi-structured interviews and key informant interviews were carried out with local communities in China's Yunnan Province. We collected plant material ( n  = 190 individuals) from five populations at study sites using single-dose AFLP markers in order to access the genetic diversity within and between populations. A total of 387 DNA fragments were produced by four AFLP primer sets. All DNA fragments were found to be polymorphic (100%). A relatively high level of genetic diversity was revealed in C. reticulata samples at both the species ( H sp  = 0.3397, I sp  = 0.5236) and population (percentage of polymorphic loci = 85.63%, H pop  = 0.2937, I pop  = 0.4421) levels. Findings further revealed a relatively high degree of genetic diversity within C. reticulata populations (Analysis of Molecular Variance = 96.31%). The higher genetic diversity within populations than among populations of C. reticulata from different geographies is likely due to the cultural and social influences associated with its long cultivation history for esthetic and culinary purposes by diverse sociolinguistic groups. This study highlights the influence of human management, socio-economic factors, and other cultural variables on the genetic and morphological diversity of C. reticulata at a regional level. Findings emphasize the important role of traditional culture on the conservation and utilization of plant genetic diversity.

Evidence for nonallopatric speciation among closely related sympatric Heliotropium species in the Atacama Desert

PubMed Central

Luebert, Federico; Jacobs, Pit; Hilger, Hartmut H; Muller, Ludo A H

2014-01-01

The genetic structure of populations of closely related, sympatric species may hold the signature of the geographical mode of the speciation process. In fully allopatric speciation, it is expected that genetic differentiation between species is homogeneously distributed across the genome. In nonallopatric speciation, the genomes may remain undifferentiated to a large extent. In this article, we analyzed the genetic structure of five sympatric species from the plant genus Heliotropium in the Atacama Desert. We used amplified fragment length polymorphisms (AFLPs) to characterize the genetic structure of these species and evaluate their genetic differentiation as well as the number of loci subject to positive selection using divergence outlier analysis (DOA). The five species form distinguishable groups in the genetic space, with zones of overlap, indicating that they are possibly not completely isolated. Among-species differentiation accounts for 35% of the total genetic differentiation (FST = 0.35), and FST between species pairs is positively correlated with phylogenetic distance. DOA suggests that few loci are subject to positive selection, which is in line with a scenario of nonallopatric speciation. These results support the idea that sympatric species of Heliotropium sect. Cochranea are under an ongoing speciation process, characterized by a fluctuation of population ranges in response to pulses of arid and humid periods during Quaternary times. PMID:24558582

Evidence for nonallopatric speciation among closely related sympatric Heliotropium species in the Atacama Desert.

PubMed

Luebert, Federico; Jacobs, Pit; Hilger, Hartmut H; Muller, Ludo A H

2014-02-01

The genetic structure of populations of closely related, sympatric species may hold the signature of the geographical mode of the speciation process. In fully allopatric speciation, it is expected that genetic differentiation between species is homogeneously distributed across the genome. In nonallopatric speciation, the genomes may remain undifferentiated to a large extent. In this article, we analyzed the genetic structure of five sympatric species from the plant genus Heliotropium in the Atacama Desert. We used amplified fragment length polymorphisms (AFLPs) to characterize the genetic structure of these species and evaluate their genetic differentiation as well as the number of loci subject to positive selection using divergence outlier analysis (DOA). The five species form distinguishable groups in the genetic space, with zones of overlap, indicating that they are possibly not completely isolated. Among-species differentiation accounts for 35% of the total genetic differentiation (F ST = 0.35), and F ST between species pairs is positively correlated with phylogenetic distance. DOA suggests that few loci are subject to positive selection, which is in line with a scenario of nonallopatric speciation. These results support the idea that sympatric species of Heliotropium sect. Cochranea are under an ongoing speciation process, characterized by a fluctuation of population ranges in response to pulses of arid and humid periods during Quaternary times.

Testing the link between population genetic differentiation and clade diversification in Costa Rican orchids.

PubMed

Kisel, Yael; Moreno-Letelier, Alejandra C; Bogarín, Diego; Powell, Martyn P; Chase, Mark W; Barraclough, Timothy G

2012-10-01

Species population genetics could be an important factor explaining variation in clade species richness. Here, we use newly generated amplified fragment length polymorphism (AFLP) data to test whether five pairs of sister clades of Costa Rican orchids that differ greatly in species richness also differ in average neutral genetic differentiation within species, expecting that if the strength of processes promoting differentiation within species is phylogenetically heritable, then clades with greater genetic differentiation should diversify more. Contrary to expectation, neutral genetic differentiation does not correlate directly with total diversification in the clades studied. Neutral genetic differentiation varies greatly among species and shows no heritability within clades. Half of the variation in neutral genetic differentiation among populations can be explained by ecological variables, and species-level traits explain the most variation. Unexpectedly, we find no isolation by distance in any species, but genetic differentiation is greater between populations occupying different niches. This pattern corresponds with those observed for microscopic eukaryotes and could reflect effective widespread dispersal of tiny and numerous orchid seeds. Although not providing a definitive answer to whether population genetics processes affect clade diversification, this work highlights the potential for addressing new macroevolutionary questions using a comparative population genetic approach. © 2012 The Author(s). Evolution© 2012 The Society for the Study of Evolution.

Sewage treatment plant associated genetic differentiation in the blue mussel from the Baltic Sea and Swedish west coast

PubMed Central

Lönn, Mikael; Lind, Emma E.; Świeżak, Justyna; Smolarz, Katarzyna; Grahn, Mats

2016-01-01

Human-derived environmental pollutants and nutrients that reach the aquatic environment through sewage effluents, agricultural and industrial processes are constantly contributing to environmental changes that serve as drivers for adaptive responses and evolutionary changes in many taxa. In this study, we examined how two types of point sources of aquatic environmental pollution, harbors and sewage treatment plants, affect gene diversity and genetic differentiation in the blue mussel in the Baltic Sea area and off the Swedish west coast (Skagerrak). Reference sites (REF) were geographically paired with sites from sewage treatments plant (STP) and harbors (HAR) with a nested sampling scheme, and genetic differentiation was evaluated using a high-resolution marker amplified fragment length polymorphism (AFLP). This study showed that genetic composition in the Baltic Sea blue mussel was associated with exposure to sewage treatment plant effluents. In addition, mussel populations from harbors were genetically divergent, in contrast to the sewage treatment plant populations, suggesting that there is an effect of pollution from harbors but that the direction is divergent and site specific, while the pollution effect from sewage treatment plants on the genetic composition of blue mussel populations acts in the same direction in the investigated sites. PMID:27812424

Incidence, size and spatial structure of clones in second-growth stands of coast redwood, Sequoia sempervirens (Cupressaceae).

PubMed

Douhovnikoff, Vladimir; Cheng, Adelaide M; Dodd, Richard S

2004-07-01

The ecology and evolutionary potential of coast redwood (Sequoia sempervirens) is significantly influenced by the important role clonal spread plays in its reproduction and site persistence. In nine second-growth stands, amplified fragment length polymorphisms (AFLPs) were used to identify redwood clonal architecture. Clones (multistem genets) dominated sites by representing an average of 70% of stems measured, ranging in size from two to 20 stems. As a result, a relatively small number of genets can monopolize a disproportionate amount of site resources, are more likely to persist over time, and have greater on-site genetic representation. Clones were not limited to fairy-ring structures, but consisted of a wide range of shapes including concentric rings, ring chains, disjunct, and linear structures. Between-ramet distances of up to 40 m were measured, indicating that clonal reproduction is not limited to basal stump resprouting. Clonal structure in second-growth stands was similar to earlier reports from old growth, emphasizing the importance of site persistence and long-term, gradual site development. Smaller ramet numbers per genet in old growth is probably due to local within-genet self thinning. Management and conservation of redwoods will benefit from a better understanding of the dynamics and structure of clonal spread in these forests.

Confirmation of Single-Locus Sex Determination and Female Heterogamety in Willow Based on Linkage Analysis.

PubMed

Chen, Yingnan; Wang, Tiantian; Fang, Lecheng; Li, Xiaoping; Yin, Tongming

2016-01-01

In this study, we constructed high-density genetic maps of Salix suchowensis and mapped the gender locus with an F1 pedigree. Genetic maps were separately constructed for the maternal and paternal parents by using amplified fragment length polymorphism (AFLP) markers and the pseudo-testcross strategy. The maternal map consisted of 20 linkage groups that spanned a genetic distance of 2333.3 cM; whereas the paternal map contained 21 linkage groups that covered 2260 cM. Based on the established genetic maps, it was found that the gender of willow was determined by a single locus on linkage group LG_03, and the female was the heterogametic gender. Aligned with mapped SSR markers, linkage group LG_03 was found to be associated with chromosome XV in willow. It is noteworthy that marker density in the vicinity of the gender locus was significantly higher than that expected by chance alone, which indicates severe recombination suppression around the gender locus. In conclusion, this study confirmed the findings on the single-locus sex determination and female heterogamety in willow. It also provided additional evidence that validated the previous studies, which found that different autosomes evolved into sex chromosomes between the sister genera of Salix (willow) and Populus (poplar).

Confirmation of Single-Locus Sex Determination and Female Heterogamety in Willow Based on Linkage Analysis

PubMed Central

Fang, Lecheng; Li, Xiaoping; Yin, Tongming

2016-01-01

In this study, we constructed high-density genetic maps of Salix suchowensis and mapped the gender locus with an F1 pedigree. Genetic maps were separately constructed for the maternal and paternal parents by using amplified fragment length polymorphism (AFLP) markers and the pseudo-testcross strategy. The maternal map consisted of 20 linkage groups that spanned a genetic distance of 2333.3 cM; whereas the paternal map contained 21 linkage groups that covered 2260 cM. Based on the established genetic maps, it was found that the gender of willow was determined by a single locus on linkage group LG_03, and the female was the heterogametic gender. Aligned with mapped SSR markers, linkage group LG_03 was found to be associated with chromosome XV in willow. It is noteworthy that marker density in the vicinity of the gender locus was significantly higher than that expected by chance alone, which indicates severe recombination suppression around the gender locus. In conclusion, this study confirmed the findings on the single-locus sex determination and female heterogamety in willow. It also provided additional evidence that validated the previous studies, which found that different autosomes evolved into sex chromosomes between the sister genera of Salix (willow) and Populus (poplar). PMID:26828940

Intraspecific genotypic variability determines concentrations of key truffle volatiles

PubMed Central

Splivallo, Richard; Valdez, Nayuf; Kirchhoff, Nina; Ona, Marta Castiella; Schmidt, Jean-Pierre; Feussner, Ivo; Karlovsky, Petr

2012-01-01

Aroma variability in truffles has been attributed to maturation (Tuber borchii), linked to environmental factors (Tuber magnatum), but the involvement of genetic factors has been ignored. We investigated aroma variability in Tuber uncinatum, a species with wide distribution. Our aim was to assess aroma variability at different spatial scales (i.e. trees, countries) and to quantify how aroma was affected by genotype, fruiting body maturity, and geographical origin. A volatile fingerprinting method was used to analyze the aroma of 223 T. uncinatum fruiting bodies from seven European countries. Maturity was estimated from spore melanization. Genotypic fingerprinting was performed by amplified fragment length polymorphism (AFLP). Discriminant analysis revealed that, regardless of the geographical origin of the truffles, most of the aroma variability was caused by eight-carbon-containing volatiles (C8-VOCs). In an orchard of T. uncinatum, truffles producing different concentrations of C8-VOCs clustered around distinct host trees. This clustering was not associated with maturity, but was associated with fungal genotype. These results indicate that the variation in C8-VOCs in truffles is most likely under genetic control. They exemplify that understanding the factors behind aroma variability requires a holistic approach. Furthermore, they also raise new questions regarding the ecological role of 1-octen-3-ol in truffles. PMID:22394027

Population Explosion in the Yellow-Spined Bamboo Locust Ceracris kiangsu and Inferences for the Impact of Human Activity

PubMed Central

Fan, Zhou; Jiang, Guo-Fang; Liu, Yu-Xiang; He, Qi-Xin; Blanchard, Benjamin

2014-01-01

Geographic distance and geographical barriers likely play a considerable role in structuring genetic variation in species, although some migratory species may have less phylogeographic structure on a smaller spatial scale. Here, genetic diversity and the phylogenetic structure among geographical populations of the yellow-spined bamboo locust, Ceracris kiangsu, were examined with 16S rDNA and amplified fragment length polymorphisms (AFLPs). In this study, no conspicuous phylogeographical structure was discovered from either Maximum parsimony (MP) and Neighbor-joining (NJ) phylogenetic analyses. The effect of geographical isolation was not conspicuous on a large spatial scale.At smaller spatial scales local diversity of some populations within mountainous areas were detected using Nei's genetic distance and AMOVA. There is a high level of genetic diversity and a low genetic differentiation among populations in the C. kiangsu of South and Southeast China. Our analyses indicate that C. kiangsu is a monophyletic group. Our results also support the hypothesis that the C. kiangsu population is in a primary differentiation stage. Given the mismatch distribution, it is likely that a population expansion in C. kiangsu occurred about 0.242 Ma during the Quaternary interglaciation. Based on historical reports, we conjecture that human activities had significant impacts on the C. kiangsu gene flow. PMID:24603526

Genetic architecture of conspecific sperm precedence in Allonemobius fasciatus and A. socius.

PubMed

Britch, Seth C; Swartout, Emma J; Hampton, Daniel D; Draney, Michael L; Chu, Jiming; Marshall, Jeremy L; Howard, Daniel J

2007-06-01

The evolution of barriers to gene exchange is centrally important to speciation. We used the crickets Allonemobius fasciatus and A. socius to investigate the genetic architecture of conspecific sperm precedence (CSP), a postinsemination prezygotic reproductive barrier. With amplified fragment-length polymorphism (AFLP) markers and controlled crosses we constructed linkage maps and estimated positions of QTL associated with CSP. The majority of QTL have low to moderate effects, although a few QTL exist in A. socius with large effects, and the numbers of QTL are comparable to numbers of genes accounting for species differences in other studies. The QTL are spread across many unlinked markers, yet QTL placed with linked markers are on a small number of linkage groups that could reflect the role of the large Allonemobius sex chromosome in prezygotic isolation. Although many QTL had positive effects on conspecific sperm utilization several QTL also exerted negative effects, which could be explained by intraspecific sexual conflict, sperm competition, or epistasis of introgressed genes on novel backgrounds. One unexpected outcome was that A. socius CSP alleles have a stronger effect than those from A. fasciatus in hybrid females, causing hybrids to behave like A. socius with regard to sperm utilization. Implications of this asymmetry in the Allonemobius hybrid zone are discussed.

Isolation of genes from female sterile flowers in Medicago sativa.

PubMed

Capomaccio, Stefano; Barone, Pierluigi; Reale, Lara; Veronesi, Fabio; Rosellini, Daniele

2009-06-01

A better knowledge of female sporogenesis and gametogenesis could have several practical applications, from commercial hybrid seed production to gene containment in GM crops. With the purpose of isolating genes involved in the megasporogenesis process, the cDNA-AFLP technique was employed to isolate transcript-derived fragments (TDF) differentially expressed between female-fertile and female-sterile full-sib alfalfa plants. This female sterility trait involves female-specific arrest of sporogenesis at early prophase associated with ectopic, massive callose deposition within the nucellus. Ninety-six TDFs were generated and BLAST analyses revealed similarities with genes involved in different Gene Ontology categories. Three TDFs were selected based on their putative functions: showing high similarity to a soybean flower-expressed beta 1,3-glucanase, to an Arabidopsis thaliana MAPKKK, and to an A. thaliana eukaryotic initiation translation factor eIF4G III, respectively. The full length mRNA sequences were obtained. RT-PCR and in situ hybridizations were performed to confirm differential expression during flower development. The genomic organization of the three genes was assessed through sequencing and Southern experiments. Sequence polymorphisms were found between sterile and fertile plants. Our approach based on differential display and bulked segregant analysis was successful in isolating genes that were differentially expressed between fertile and sterile alfalfa plants.

Identification of differentially expressed genes in brown planthopper Nilaparvata lugens (Hemiptera: Delphacidae) responding to host plant resistance.

PubMed

Yang, Zhifan; Zhang, Futie; Zhu, Lili; He, Guangcun

2006-02-01

The brown planthopper Nilaparvata lugens Stål is one of the major insect pests of rice Oryza sativa L. The host resistance exhibits profound effects on growth, development and propagation of N. lugens. To investigate the molecular response of N. lugens to host resistance, a cDNA-amplified fragment length polymorphism (cDNA-AFLP) technique was employed to identify the differentially expressed genes in the nymphs feeding on three rice varieties. Of the 2,800 cDNA bands analysed, 54 were up-regulated and seven down-regulated qualitatively in N. lugens when the ingestion sources were changed from susceptible rice plants to resistant ones. Sequence analysis of the differential transcript-derived fragments showed that the genes involved in signalling, stress response, gene expression regulation, detoxification and metabolism were regulated by host resistance. Four of the transcript-derived fragments corresponding to genes encoding for a putative B subunit of phosphatase PP2A, a nemo kinase, a cytochrome P450 monooxygenase and a prolyl endopeptidase were further characterized in detail. Northern blot analysis confirmed that the expression of the four genes was enhanced in N. lugens feeding on resistant rice plants. The roles of these genes in the defensive response of N. lugens to host plant resistance were discussed.

The Pattern of Genetic Variability in Apomictic Clones of Taraxacum officinale Indicates the Alternation of Asexual and Sexual Histories of Apomicts

PubMed Central

Majeský, Ľuboš; Vašut, Radim J.; Kitner, Miloslav; Trávníček, Bohumil

2012-01-01

Dandelions (genus Taraxacum) comprise a group of sexual diploids and apomictic polyploids with a complicated reticular evolution. Apomixis (clonal reproduction through seeds) in this genus is considered to be obligate, and therefore represent a good model for studying the role of asexual reproduction in microevolutionary processes of apomictic genera. In our study, a total of 187 apomictic individuals composing a set of nine microspecies (sampled across wide geographic area in Europe) were genotyped for six microsatellite loci and for 162 amplified fragment length polymorphism (AFLP) markers. Our results indicated that significant genetic similarity existed within accessions with low numbers of genotypes. Genotypic variability was high among accessions but low within accessions. Clustering methods discriminated individuals into nine groups corresponding to their phenotypes. Furthermore, two groups of apomictic genotypes were observed, which suggests that they had different asexual histories. A matrix compatibility test suggests that most of the variability within accession groups was mutational in origin. However, the presence of recombination was also detected. The accumulation of mutations in asexual clones leads to the establishment of a network of clone mates. However, this study suggests that the clones primarily originated from the hybridisation between sexual and apomicts. PMID:22870257

[Psilocybin - public available psychodysleptic].

PubMed

Dydak, Karolina; Śliwińska-Mossoń, Mariola; Milnerowicz, Halina

2015-09-07

Substances of plant origin have been used to induce hallucinations for a long time, in religious ceremonies and rituals as well as in pain relief. Psilocybin and psilocin naturally occur in the fungal genus Psilocybe. Due to the psychedelic effects and relative harmlessness of these substances and the fact that they do not cause physical addiction, psilocybin and psilocin recently have been increasingly replacing synthetic psychodysleptics, such as diethylamide D-lysergic acid. Both compounds as psychoactive substances are illegal, but psilocybin, in addition to psychotropic action, also shows positive effects, which from a medical point of view indicate its therapeutic potential and capacity for use in therapy. However, poisoning by psilocin and its derivatives is still a major clinical and social problem, mainly among young people, which is why quick and reliable identification of these substances is very important. Traditional ways of assigning the sample to a particular taxon, such as morphological and biochemical analysis or palynological and sporological studies, are not very universal and often do not provide clear results. Credibility, high speed and lower cost of DNA analysis make genetic methods more often used to determine the species of fungi. These methods are random amplification of polymorphic DNA (RAPD), amplified fragment length polymorphism (AFLP) and high resolution melting (HRM). Moreover, analysis of the regions ITS1 and nLSU was suggested as a valid method for application in the molecular taxonomy of fungi for forensic purposes. Modern methods of identifying psilocybin and psilocin in fungi and biological material are: zone capillary electrophoresis, high performance liquid chromatography, gas chromatography and liquid chromatography coupled with mass spectrometry. The mentioned methods are successfully used for the identification of psychoactive substances in fungi as well as in blood and urine samples.

Yalongjiang River Has Had an Important Role in the Dispersal and Divergence of Rosa soulieana in the Hengduan Mountains of China.

PubMed

Jian, Hongying; Zhang, Yonghong; Qiu, Xianqin; Yan, Huijun; Wang, Qigang; Zhang, Hao; Sun, Hang

2016-01-01

The Hengduan Mountains are the core of the Sino-Himalayan Floristic Region. Rosa soulieana Crép. is an important wild rose species that is widely distributed in the Hengduan Mountains. To provide better future utilization of this wild rose, and also to add some possible proof of the effect of geomorphological and ecological characteristics of the Hengduan Mountains on the current spatial distribution and genetic diversity of local species, the genetic diversity and genetic structure of 556 individuals from 37 populations of R. soulieana were studied using fluorescent amplified fragment length polymorphisms (AFLPs). R. soulieana showed a moderately high level of genetic diversity and a high level of genetic differentiation at the species level. The total percentage of polymorphic loci, total heterozygosity (Ht), Shannon index (I), and heterozygosity value within populations (Hs) were 97.8%, 0.253, 0.339, and 0.139, respectively. More than half of the total genetic variation (54.0%) occurred within populations, and the overall gene differentiation coefficient (Gst) was 0.451. The genetic differentiation among populations was positively and significantly correlated with geographic distance. The neighbor-joining cluster and the Bayesian analysis divided all the populations and individuals into 3 groups, and did not support the morphology based intraspecific varieties. The results confirmed that the ancient R. soulieana of the third group survived in northwestern Yunnan and Yalongjiang valley and then moved upnorth along the valley. The spatial distribution of the other two groups was the result of allopatric divergence due to long period of adaptation to the different climatic conditions of its distribution at either side of the Yalongjiang River.

Comparison of space flight and heavy ion radiation induced genomic/epigenomic mutations in rice (Oryza sativa)

NASA Astrophysics Data System (ADS)

Shi, Jinming; Lu, Weihong; Sun, Yeqing

2014-04-01

Rice seeds, after space flight and low dose heavy ion radiation treatment were cultured on ground. Leaves of the mature plants were obtained for examination of genomic/epigenomic mutations by using amplified fragment length polymorphism (AFLP) and methylation sensitive amplification polymorphism (MSAP) method, respectively. The mutation sites were identified by fragment recovery and sequencing. The heritability of the mutations was detected in the next generation. Results showed that both space flight and low dose heavy ion radiation can induce significant alterations on rice genome and epigenome (P < 0.05). For both genetic and epigenetic assays, while there was no significant difference in mutation rates and their ability to be inherited to the next generation, the site of mutations differed between the space flight and radiation treated groups. More than 50% of the mutation sites were shared by two radiation treated groups, radiated with different LET value and dose, while only about 20% of the mutation sites were shared by space flight group and radiation treated group. Moreover, in space flight group, we found that DNA methylation changes were more prone to occur on CNG sequence than CG sequence. Sequencing results proved that both space flight and heavy ion radiation induced mutations were widely spread on rice genome including coding region and repeated region. Our study described and compared the characters of space flight and low dose heavy ion radiation induced genomic/epigenomic mutations. Our data revealed the mechanisms of application of space environment for mutagenesis and crop breeding. Furthermore, this work implicated that the nature of mutations induced under space flight conditions may involve factors beyond ion radiation.

Development and Characterization of Somatic Hybrids of Ulva reticulata Forsskål (×) Monostroma oxyspermum (Kutz.)Doty

PubMed Central

Gupta, Vishal; Kumari, Puja; Reddy, CRK

2015-01-01

Ulvophycean species with diverse trait characteristics provide an opportunity to create novel allelic recombinant variants. The present study reports the development of seaweed variants with improved agronomic traits through protoplast fusion between Monostroma oxyspermum (Kutz.) Doty and Ulva reticulata Forsskål. A total of 12 putative hybrids were screened based on the variations in morphology and total DNA content over the fusion partners. DNA-fingerprinting by inter simple sequence repeat (ISSR) and amplified fragment length polymorphism (AFLP) analysis confirmed genomic introgression in the hybrids. The DNA fingerprint revealed sharing of parental alleles in regenerated hybrids and a few alleles that were unique to hybrids. The epigenetic variations in hybrids estimated in terms of DNA methylation polymorphism also revealed sharing of methylation loci with both the fusion partners. The functional trait analysis for growth showed a hybrid with heterotic trait (DGR% = 36.7 ± 1.55%) over the fusion partners U. reticulata (33.2 ± 2.6%) and M. oxyspermum (17.8 ± 1.77%), while others were superior to the mid-parental value (25.2 ± 2.2%) (p < 0.05). The fatty acid (FA) analysis of hybrids showed notable variations over fusion partners. Most hybrids showed increased polyunsaturated FAs (PUFAs) compared to saturated FAs (SFAs) and mainly includes the nutritionally important linoleic acid, α-linolenic acid, oleic acid, stearidonic acid, and docosahexaenoic acid. The other differences observed include superior cellulose content and antioxidative potential in hybrids over fusion partners. The hybrid varieties with superior traits developed in this study unequivocally demonstrate the significance of protoplast fusion technique in developing improved varients of macroalgae. PMID:25688248

Vibrio communis sp. nov., isolated from the marine animals Mussismilia hispida, Phyllogorgia dilatata, Palythoa caribaeorum, Palythoa variabilis and Litopenaeus vannamei.

PubMed

Chimetto, Luciane A; Cleenwerck, Ilse; Alves, Nelson; Silva, Bruno Sergio; Brocchi, Marcelo; Willems, Anne; De Vos, Paul; Thompson, Fabiano L

2011-02-01

Eight Vibrio isolates originating from the marine corals Mussismilia hispida and Phyllogorgia dilatata and the zoanthids Palythoa caribaeorum and Palythoa variabilis in Brazil and the Pacific white shrimp (Litopenaeus vannamei) in Ecuador were studied by means of a polyphasic approach. The novel isolates formed a tight monophyletic group in the genus Vibrio and were closely related to species of the Vibrio harveyi group, to which they showed more than 99 % 16S rRNA gene sequence similarity. Analysis based on concatenated sequences of the following seven genes, 16S rRNA, gyrB, recA, rpoA, topA, pyrH and mreB (5633 bp in length), showed clear separation between the isolates and species of the V. harveyi group. Amplified fragment length polymorphism (AFLP) analysis, performed previously, revealed that a representative isolate of this group, LMG 20370, was clearly separate from known Vibrio species (it belonged to the so-called AFLP cluster A31). DNA-DNA hybridization (DDH) experiments with representative isolates and type strains of the V. harveyi species group revealed high DDH between the novel isolates (more than 74 %) and less than 70 % DDH towards type strains of related Vibrio species, proving the novel species status of the isolates. Phenotypically, the novel species belongs to the arginine dihydrolase (A)-negative, lysine decarboxylase (L)-positive and ornithine decarboxylase (O)-positive (A-/L+/O+) cluster reported previously. Most species of the V. harveyi group (i.e. Vibrio rotiferianus, V. harveyi, V. parahaemolyticus and V. alginolyticus) also belong to this A-/L+/O+ cluster. However, several phenotypic features can be used for the identification of the novel species. In contrast to its closest phylogenetic neighbours, the novel species exhibits esterase (C4) and N-acetyl-β-glucosaminidase activities, but it does not produce acetoin, does not use citrate, α-ketoglutaric acid or propionic acid and does not ferment melibiose. The novel species can also be differentiated on the basis of the presence of the fatty acids C(17 : 0,) C(17 : 1)ω8c, iso-C(17 : 0) and iso-C(13 : 0) and the absence of the fatty acid C(18 : 0). The name Vibrio communis sp. nov. is proposed for this taxon. Strain R-40496(T) (=LMG 25430(T) =CAIM 1816(T)) is the type strain.

Identification of genes differentially expressed during interaction of Mexican lime tree infected with "Candidatus Phytoplasma aurantifolia"

PubMed Central

2011-01-01

Background "Candidatus Phytoplasma aurantifolia", is the causative agent of witches' broom disease in Mexican lime trees (Citrus aurantifolia L.), and is responsible for major losses of Mexican lime trees in Southern Iran and Oman. The pathogen is strictly biotrophic, and thus is completely dependent on living host cells for its survival. The molecular basis of compatibility and disease development in this system is poorly understood. Therefore, we have applied a cDNA- amplified fragment length polymorphism (AFLP) approach to analyze gene expression in Mexican lime trees infected by "Ca. Phytoplasma aurantifolia". Results We carried out cDNA-AFLP analysis on grafted infected Mexican lime trees of the susceptible cultivar at the representative symptoms stage. Selective amplifications with 43 primer combinations allowed the visualisation of 55 transcript-derived fragments that were expressed differentially between infected and non-infected leaves. We sequenced 51 fragments, 36 of which were identified as lime tree transcripts after homology searching. Of the 36 genes, 70.5% were down-regulated during infection and could be classified into various functional groups. We showed that Mexican lime tree genes that were homologous to known resistance genes tended to be repressed in response to infection. These included the genes for modifier of snc1 and autophagy protein 5. Furthermore, down-regulation of genes involved in metabolism, transcription, transport and cytoskeleton was observed, which included the genes for formin, importin β 3, transducin, L-asparaginase, glycerophosphoryl diester phosphodiesterase, and RNA polymerase β. In contrast, genes that encoded a proline-rich protein, ubiquitin-protein ligase, phosphatidyl glycerol specific phospholipase C-like, and serine/threonine-protein kinase were up-regulated during the infection. Conclusion The present study identifies a number of candidate genes that might be involved in the interaction of Mexican lime trees with "Candidatus Phytoplasma aurantifolia". These results should help to elucidate the molecular basis of the infection process and to identify genes that could be targeted to increase plant resistance and inhibit the growth and reproduction of the pathogen. PMID:21194490

Persistence, Transmission, and Virulence Characteristics of Aeromonas Strains in a Duckweed Aquaculture-Based Hospital Sewage Water Recycling Plant in Bangladesh▿

PubMed Central

Rahman, Mokhlasur; Huys, Geert; Rahman, Motiur; Albert, M. John; Kühn, Inger; Möllby, Roland

2007-01-01

The persistence and transmission of Aeromonas in a duckweed aquaculture-based hospital sewage water treatment plant in Bangladesh was studied. A total of 670 samples from different sites of the hospital sewage water treatment plant, from feces of hospitalized children suffering from diarrhea, from environmental control ponds, and from feces of healthy humans were collected over a period of three years. In total, 1,315 presumptive Aeromonas isolates were biochemically typed by the PhenePlate rapid screening system (PhP-AE). A selection of 90 representative isolates was further analyzed with PhenePlate (PhP) extended typing (PhP-48), fatty acid methyl ester analysis, and amplified fragment length polymorphism (AFLP) fingerprinting. In addition, the prevalence of the putative virulence factors hemolysin and cytotoxin and the presence of the cytolytic enterotoxin gene (AHCYTOEN) were analyzed. Aeromonas was found at all sites of the treatment plant, in 40% of the samples from environmental control ponds, in 8.5% of the samples from hospitalized children suffering from diarrhea, and in 3.5% of samples from healthy humans. A significantly high number of Aeromonas bacteria was found in duckweed, which indicates that duckweed may serve as a reservoir for these bacteria. PhP-AE typing allowed identification of more than 192 distinct PhP types, of which 18 major PhP types (MTs) were found in multiple sites and during several occasions. AFLP fingerprinting revealed the prevalence of genotypically indistinguishable Aeromonas isolates among certain PhP MTs recovered from different sampling occasions and/or at multiple sites. Hemolytic and cytotoxic activities were observed in 43% of the tested strains, whereas 29% possessed the cytolytic enterotoxin gene AHCYTOEN. Collectively, two specific MTs associated with diarrhea were shown to exhibit high cytotoxicity. Furthermore, all tested isolates of these major types were positive for the cytolytic enterotoxin gene. In conclusion, our data indicate that certain phenotypically and genotypically stable clonal lineages of Aeromonas have persisted in the treatment system for a prolonged period and might spread from the hospitalized children suffering from diarrhea to fish produced for human consumption through the sewage water treatment system. PMID:17194839

Chromosome elimination, addition and introgression in intertribal partial hybrids between Brassica rapa and Isatis indigotica

PubMed Central

Tu, Yuqin; Sun, Jian; Ge, Xianhong; Li, Zaiyun

2009-01-01

Background and Aims Partial hybrids with female-parent-type phenotypes and chromosome numbers but altered genomic compositions have been reported in wide crosses of several plants. In order to introgress desirable genes from a wild relative, Isatis indigotica (a dye and medicinal plant; 2n = 14), into Brassica crops, intertribal sexual hybridizations were carried out with B. rapa (2n = 20), and the resulting hybrids and their progenies were characterized. Methods Using genomic in situ hybridization (GISH) and amplified fragment length polymorphism (AFLP), chromosomal/genomic components of the hybrids and their progenies were analysed. Key Results Many hybrid plants were obtained from the mature seeds harvested from the B. rapa × I. indigotica cross, and these exhibited different morphological traits. However, the majority of them did not survive and only three plants grew to maturity. These three hybrids showed poor growth and much smaller stature than the two parents, but had some morphological traits and chemical composition of I. indigotica. One plant had 2n = 10, the haploid chromosome number of B. rapa, and was absolutely sterile. The other two plants had 20 and 22 somatic chromosomes and were male sterile but produced seeds following pollinations with B. rapa. All back-cross progenies over several generations maintained a B. rapa-type phenotype and also displayed some variations in morphological characters and fatty acid compositions. They were all 2n = 20 and showed good seed-set. The hybrid with 2n = 22 produced some progeny plants with 2n = 21 and 2n = 22. GISH detected two chromosomes of I. indigotica in the hybrid with 2n = 22 but none in the one with 2n = 20. AFLP bands specific for I. indigotica, novel for two parents or absent in B. rapa, were detected in the two hybrids and their progenies. These progeny plants were novel B. rapa types with an altered genomic constitution or alien additions. Conclusions Complete or partial chromosome elimination and diploidization with genomic rearrangements were considered to lead to the formation of partial hybrids in this cross. PMID:19258339

Asymmetric hybridization in Rhododendron agastum: a hybrid taxon comprising mainly F1s in Yunnan, China

PubMed Central

Zha, Hong-Guang; Milne, Richard I.; Sun, Hang

2010-01-01

Background and Aims Rhododendron (Ericaceae) is a large woody genus in which hybridization is thought to play an important role in evolution and speciation, particularly in the Sino-Himalaya region where many interfertile species often occur sympatrically. Rhododendron agastum, a putative hybrid species, occurs in China, western Yunnan Province, in mixed populations with R. irroratum and R. delavayi. Methods Material of these taxa from two sites 400 km apart (ZhuJianYuan, ZJY and HuaDianBa, HDB) was examined using cpDNA and internal transcribed spacer (ITS) sequences, and amplified fragment length polymorphism (AFLP) loci, to test the possibility that R. agastum was in fact a hybrid between two of the other species. Chloroplast trnL-F and trnS-trnG sequences together distinguished R. irroratum, R. delavayi and some material of R. decorum, which is also considered a putative parent of R. agastum. Key Results All 14 R. agastum plants from the HDB site had the delavayi cpDNA haplotype, whereas at the ZJY site 17 R. agastum plants had this haplotype and four had the R. irroratum haplotype. R. irroratum and R. delavayi are distinguished by five unequivocal point mutations in their ITS sequences; every R. agastum accession had an additive pattern (double peaks) at each of these sites. Data from AFLP loci were acquired for between ten and 21 plants of each taxon from each site, and were analysed using a Bayesian approach implemented by the program NewHybrids. The program confirmed the identity of all accessions of R. delavayi, and all R. irroratum except one, which was probably a backcross. All R. agastum from HDB and 19 of 21 from ZJY were classified as F1 hybrids; the other two could not be assigned a class. Conclusions Rhododendron agastum represents populations of hybrids between R. irroratum and R. delavayi, which comprise mostly or only F1s, at the two sites examined. The sites differ in that at HDB there was no detected variation in cpDNA type or hybrid class, whereas at ZJY there was variation in both. PMID:19887474

Diversification in continental island archipelagos: new evidence on the roles of fragmentation, colonization and gene flow on the genetic divergence of Aegean Nigella (Ranunculaceae).

PubMed

Jaros, Ursula; Tribsch, Andreas; Comes, Hans Peter

2018-02-12

Disentangling the relative roles of past fragmentation (vicariance), colonization (dispersal) and post-divergence gene flow in the genetic divergence of continental island organisms remains a formidable challenge. Amplified fragment length polymorphisms (AFLPs) were used to (1) gain further insights into the biogeographical processes underlying the Pleistocene diversification of the Aegean Nigella arvensis complex; (2) evaluate the role of potential key factors driving patterns of population genetic variability (mating system, geographical isolation and historical contingencies); and (3) test the robustness of conclusions previously drawn from chloroplast (cp) DNA. Genetic diversity was analysed for 235 AFLP markers from 48 populations (497 individuals) representing 11 taxa of the complex using population genetic methods and Bayesian assignment tests. Most designated taxa are identifiable as genetically distinct units. Both fragmentation and dispersal-driven diversification processes occurred at different geological time scales, from Early to Late Pleistocene, specifically (1) sea barrier-induced vicariant speciation in the Cyclades, the Western Cretan Strait and Ikaria; and (2) bi-regional colonizations of the 'Southern Aegean Island Arc' from the Western vs. Eastern Aegean mainland, followed by allopatric divergences in Crete vs. Rhodos and Karpathos/Kasos. Outcrossing island taxa experienced drift-related demographic processes that are magnified in the two insular selfing species. Population genetic differentiation on the mainland seems largely driven by dispersal limitation, while in the Central Aegean it may still be influenced by historical events (island fragmentation and sporadic long-distance colonization). The biogeographical history of Aegean Nigella is more complex than expected for a strictly allopatric vicariant model of divergence. Nonetheless, the major phylogeographical boundaries of this radiation are largely congruent with the geography and history of islands, with little evidence for ongoing gene exchange between divergent taxa. The present results emphasize the need to investigate further biological and landscape features and contemporary vs. historical processes in driving population divergence and taxon diversification in Aegean plant radiations. © The Author(s) 2017. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Development and characterization of microsatellite markers for rice leaffolder, Cnaphalocrocis medinalis (Guenée) and cross-species amplification in other Pyralididae.

PubMed

An, Baoguang; Deng, Xiaolong; Shi, Huiyun; Ding, Meng; Lan, Jie; Yang, Jing; Li, Yangsheng

2014-02-01

Rice leaffolder, Cnaphalocrocis medinalis (Guenée), is a destructive and widespread pest on rice. In this study, 20 microsatellite markers were isolated and characterized from C. medinalis partial genomic libraries using the method of fast isolation by AFLP of sequence containing repeats. Of these markers, 18 markers displayed polymorphisms. Polymorphisms were evaluated in 48 individuals from two natural populations. The number of alleles per locus ranged from 2 to 15, and the expected and observed heterozygosities ranged from 0.324 to 0.934 and from 0.304 to 0.917, respectively. Cross-species amplification was also performed to test the transferability of the 20 microsatellite markers and a moderate level of cross amplication was observed across the three species of Pyralididae (26.67 %). These microsatellite loci would facilitate the future study on population genetics and molecular genetics of rice leaffolder and would also be useful for study in Chilo suppressalis, Scirpophaga incertulas and Pyrausta nubilalis.

Viability and Biological Properties of Barley Seeds Expose to Outside of International Space Station

NASA Astrophysics Data System (ADS)

Sugimoto, Manabu; Ishii, Makoto; Mori, Izumi; Shagimardanova, Elena; Gusev, Oleg; Sychev, Vladimir; Levinskikh, Margarita; Novikova, Nataliya; Grigoriev, Anatoly

Plants play an important role in supplying nutrients and oxygen to human under material recycle system in space as well as on earth, therefore, seed storage in space should be necessary to self-supply foods when number of astronauts would stay and investigate for a long-term habitation of orbit and the bases of the Moon and Mars. In order to understand the effect of real space environment on the preservation of seeds, the seeds of malting barley, Haruna Nijo, were exposed to outside of the Pier docking station of International Space Station in the framework of the Biorisk-MSN program. After exposure to outside of International Space Station for 13 months, the seeds (SP) were transported to Earth, soaked in water, and germinated on the filter paper filled with water. The germination ratio of SP was 82%, while that of the ground control was 96%, showing that the barley seeds survived cosmic radiation, vacuum, and temperature excursion in space. The germinated seeds of SP and ground control were transplanted to the Wagner pots filled with soil and grown for 5 months in the greenhouse. The agronomic character, such as number of main stem leaf and ear, straw weight, culm length, ear length, thousand kernel weight, and percentage of ripening, were not different significantly between SP and ground control. The germination ratio of the harvested SP was 96% as same as that of the harvested ground control. Genomic DNA and protein were extracted from leaves of the barleys and analyzed by AFLP and 2-DE, respectively. The results demonstrated no significant difference in genetic polymorphism and protein production in these barleys. From our results, barley seeds could survive real space environment for the long-term habitation without phenotypic and genotypic damages.

Disentangling the role of hybridization in the evolution of the endangered Arizona cliffrose (Purshia subintegra; Rosaceae): A molecular and morphological analysis

USGS Publications Warehouse

Travis, S.E.; Baggs, J.E.; Maschinski, J.

2008-01-01

Hybridization may threaten the conservation status of rare species through genetic assimilation and may confound the ability to distinguish among taxa. We studied these issues in an endangered shrub, Purshia subintegra (Rosaceae), known from four populations growing on limestone outcrops in central Arizona (USA). Using amplified fragment length polymorphisms (AFLP) and the Bayesian clustering algorithm implemented in STRUCTURE, we identified three distinct genetic lineages among Arizona Purshia subintegra and P. stansburiana. An initial split divided San Carlos Basin P. subintegra (considered P. pinkavae by Schaack) from northern P. stansburiana populations (FST = 0.394). A subsequent split separated northern P. stansburiana from two P. subintegra populations at Horseshoe Lake and Burro Creek (FST = 0.207), which comprised a nearly perfect admixture of the two lineages identified in the initial analysis. In the Verde River Valley P. subintegra is sympatric with P. stansburiana and exhibited an average 27% P. stansburiana genes for 5 of 6 stands analyzed, indicating ongoing hybridization and backcrossing with P. subintegra. Individuals carrying >90% P. subintegra markers are identifiable 68% of the time based on morphology, with leaf lobing, leaf size, and leaf length acting as the most reliable indicators of taxonomic status. However, the genetic and morphological distance correlation among individuals was low (r = 0.17, P = 0.0002), indicating that morphology cannot always accurately predict genetic admixture or taxonomy. Overall, our study confirmed the genetic distinctiveness of the San Carlos Basin population, an ancient natural hybrid origin of P. subintegra, and the presence of a hybrid swarm in the Verde Valley, whose conservation value may lie in its heightened genetic diversity. ?? 2007 Springer Science+Business Media B.V.

Histological and transcript analyses of intact somatic embryos in an elite maize (Zea mays L.) inbred line Y423.

PubMed

Liu, Beibei; Su, Shengzhong; Wu, Ying; Li, Ying; Shan, Xiaohui; Li, Shipeng; Liu, Hongkui; Dong, Haixiao; Ding, Meiqi; Han, Junyou; Yuan, Yaping

2015-07-01

Intact somatic embryos were obtained from an elite maize inbred line Y423, bred in our laboratory. Using 13-day immature embryos after self-pollination as explants, and after 4-5 times subculture, a large number of somatic embryos were detected on the surface of the embryonic calli on the medium. The intact somatic embryos were transferred into the differential medium, where the plantlets regenerated with shoots and roots forming simultaneously. Histological analysis and scanning electron micrographs confirmed the different developmental stages of somatic embryogenesis, including globular-shaped embryo, pear-shaped embryo, scutiform embryo, and mature embryo. cDNA-amplified fragment length polymorphism (cDNA-AFLP) was used for comparative transcript profiling between embryogenic and non-embryogenic calli of a new elite maize inbred line Y423 during somatic embryogenesis. Differentially expressed genes were cloned and sequenced. Gene Ontology analysis of 117 candidate genes indicated their involvement in cellular component, biological process and molecular function. Nine of the candidate genes were selected. The changes in their expression levels during embryo induction and regeneration were analyzed in detail using quantitative real-time PCR. Two full-length cDNA sequences, encoding ZmSUF4 (suppressor of fir 4-like protein) and ZmDRP3A (dynamin-related protein), were cloned successfully from intact somatic embryos of the elite inbred maize line Y423. Here, a procedure for maize plant regeneration from somatic embryos is described. Additionally, the possible roles of some of these genes during the somatic embryogenesis has been discussed. This study is a systematic analysis of the cellular and molecular mechanism during the formation of intact somatic embryos in maize. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Patterns of Post-Glacial Genetic Differentiation in Marginal Populations of a Marine Microalga

PubMed Central

Tahvanainen, Pia; Alpermann, Tilman J.; Figueroa, Rosa Isabel; John, Uwe; Hakanen, Päivi; Nagai, Satoshi; Blomster, Jaanika; Kremp, Anke

2012-01-01

This study investigates the genetic structure of an eukaryotic microorganism, the toxic dinoflagellate Alexandrium ostenfeldii, from the Baltic Sea, a geologically young and ecologically marginal brackish water estuary which is predicted to support evolution of distinct, genetically impoverished lineages of marine macroorganisms. Analyses of the internal transcribed spacer (ITS) sequences and Amplified Fragment Length Polymorphism (AFLP) of 84 A. ostenfeldii isolates from five different Baltic locations and multiple external sites revealed that Baltic A. ostenfeldii is phylogenetically differentiated from other lineages of the species and micro-geographically fragmented within the Baltic Sea. Significant genetic differentiation (F ST) between northern and southern locations was correlated to geographical distance. However, instead of discrete genetic units or continuous genetic differentiation, the analysis of population structure suggests a complex and partially hierarchic pattern of genetic differentiation. The observed pattern suggests that initial colonization was followed by local differentiation and varying degrees of dispersal, most likely depending on local habitat conditions and prevailing current systems separating the Baltic Sea populations. Local subpopulations generally exhibited low levels of overall gene diversity. Association analysis suggests predominately asexual reproduction most likely accompanied by frequency shifts of clonal lineages during planktonic growth. Our results indicate that the general pattern of genetic differentiation and reduced genetic diversity of Baltic populations found in large organisms also applies to microscopic eukaryotic organisms. PMID:23300940

Hybridization and adaptation to introduced balloon vines in an Australian soapberry bug.

PubMed

Andres, J A; Thampy, P R; Mathieson, M T; Loye, J; Zalucki, M P; Dingle, H; Carroll, S P

2013-12-01

Contemporary adaptation of plant feeding insects to introduced hosts provides clear cases of ecologically based population divergence. In most cases the mechanisms permitting rapid differentiation are not well known. Here we study morphological and genetic variation associated with recent shifts by the Australian soapberry bug Leptocoris tagalicus onto two naturalized Neotropical balloon vines, Cardiospermum halicacabum and C. grandiflorum that differ in time since introduction. Our results show that these vines have much larger fruits than the native hosts (Whitewood tree -Atalaya hemiglauca- and Woolly Rambutan -Alectryon tomentosus-) and that bugs living on them have evolved significantly longer beaks and new allometries. Genetic analyses of mitochondrial haplotypes and amplified fragment length polymorphic (AFLP) markers indicate that the lineage of bugs on the annual vine C. halicacabum, the older introduction, is intermediate between the two subspecies of L. tagalicus found on native hosts. Moreover, where the annual vine and Whitewood tree co-occur, the morphology and genomic composition of the bugs are similar to those occurring in allopatry. These results show that hybridization provided the genetic elements underlying the strongly differentiated 'Halicacabum bugs'. In contrast, the bugs feeding on the recently introduced perennial balloon vine (C. grandiflorum) showed no evidence of admixture, and are genetically indistinguishable from the nearby populations on a native host. © 2013 John Wiley & Sons Ltd.

Seven cases of Saccharomyces fungaemia related to use of probiotics.

PubMed

Roy, Ujjwayini; Jessani, Laxman G; Rudramurthy, Shivaprakash M; Gopalakrishnan, Ram; Dutta, Soma; Chakravarty, Chandrashish; Jillwin, Joseph; Chakrabarti, Arunaloke

2017-06-01

Probiotics are increasingly used in critically ill patients without enough safety data. The aim of the present study was to determine the association of probiotics with Saccharomyces cerevisiae fungaemia. Seven patients with S. cerevisiae fungaemia were reported at two hospitals in India between July 2014 and September 2015. Detailed clinical history of patients was recorded. Besides the seven patient isolates, three probiotics sachets used in those patients and five unrelated clinical isolates were used for association study by Fluorescent amplified fragment length polymorphism (FAFLP). Antifungal susceptibility testing was performed by broth microdilution technique of CLSI (M27-A3) and interpreted according to CLSI (M27S4). Two patients were premature neonates and five were adults. They were admitted in intensive care unit and were on probiotics containing S. boulardii (except one adult patient). FAFLP analysis showed 96.4-99.7% similarity between blood and corresponding probiotic isolates. Of the three AFLP types (group I, II, II) identified, all the probiotic isolates clustered in group I (major cluster) including majority of the blood isolates. The isolates were susceptible to all antifungal agents tested. Five patients, who could be evaluated, responded promptly to echinocandins or voriconazole. As the prescription of probiotic containing S. boulardii in critically ill patient's leads to the fungaemia, we recommend avoiding this probiotic in those patients. © 2017 Blackwell Verlag GmbH.

Differential gene expression revealed with RNA-Seq and parallel genotype selection of the ornithine decarboxylase gene in fish inhabiting polluted areas.

PubMed

Vega-Retter, C; Rojas-Hernandez, N; Vila, I; Espejo, R; Loyola, D E; Copaja, S; Briones, M; Nolte, A W; Véliz, D

2018-03-19

How organisms adapt to unfavorable environmental conditions by means of plasticity or selection of favorable genetic variants is a central issue in evolutionary biology. In the Maipo River basin, the fish Basilichthys microlepidotus inhabits polluted and non-polluted areas. Previous studies have suggested that directional selection drives genomic divergence between these areas in 4% of Amplified Fragment Length Polymorphism (AFLP) loci, but the underlying genes and functions remain unknown. We hypothesized that B. microlepidotus in this basin has plastic and/or genetic responses to these conditions. Using RNA-Seq, we identified differentially expressed genes in individuals from two polluted sites compared with fish inhabiting non-polluted sites. In one polluted site, the main upregulated genes were related to cellular proliferation as well as suppression and progression of tumors, while biological processes and molecular functions involved in apoptotic processes were overrepresented in the upregulated genes of the second polluted site. The ornithine decarboxylase gene (related to tumor promotion and progression), which was overexpressed in both polluted sites, was sequenced, and a parallel pattern of a heterozygote deficiency and increase of the same homozygote genotype in both polluted sites compared with fish inhabiting the non-polluted sites was detected. These results suggest the occurrence of both a plastic response in gene expression and an interplay between phenotypic change and genotypic selection in the face of anthropogenic pollution.

Genome Shuffling and Gentamicin-Resistance to Improve ε-Poly-L-Lysine Productivity of Streptomyces albulus W-156.

PubMed

Wang, Liang; Chen, Xusheng; Wu, Guangyao; Zeng, Xin; Ren, Xidong; Li, Shu; Tang, Lei; Mao, Zhonggui

2016-12-01

Genome shuffling has been a recently effective method for screening the desirable phenotypes of industrial strains. Here, we combined genome shuffling and gentamicin resistance to improve the production of ε-poly-L-lysine in Streptomyces albulus W-156. Five starting mutants with higher ε-poly-L-lysine (ε-PL) productivities were firstly obtained by atmospheric and room temperature plasma (ARTP) mutagenesis. After three rounds of genome shuffling with increasing concentration of gentamicin for selection, S. albulus AG3-28, was finally got with a production of 3.43 g/L in shaking flask. In a 5-L fermenter, AG3-28 exhibited a higher ε-PL productivity (56.5 g/L) than the initial strain W-156 (37.5 g/L). Key enzyme activities in primary and secondary metabolic pathways were analyzed, and the transcription levels of hrdD and pls were determined by quantitative real time-polymerase chain reaction (qRT-PCR). Increase of key enzyme activities and the upregulation of the gene transcriptional levels demonstrated that ε-PL synthetic pathway in AG3-28 was obviously strengthened, which might be responsible for the high productivity. Moreover, hyper-yield strain AG3-28 was found to produce a slightly lower ε-PL polymerization degree than the parent strain. Amplified fragment length polymorphism (AFLP) analysis reflects the genetic diversity among the derivates after genome shuffling.

Identification of desiccation tolerance transcripts potentially involved in rape (Brassica napus L.) seeds development and germination.

PubMed

Lang, Sirui; Liu, Xiaoxia; Ma, Gang; Lan, QinYing; Wang, Xiaofeng

2014-10-01

To investigate regulatory processes and protective mechanisms leading to desiccation tolerance (DT) in seeds, cDNA amplified fragment length polymorphism (cDNA-AFLP) in conjunction with 128 primer combinations was used to detect differential gene expression in rape seeds in response to DT during seed development and germination. We obtained approximately 8000 transcript-derived fragments (TDFs), of which 394 TDFs with differential expression patterns ("sustained expression", "up-regulated", "couple with seed DT", and "down-regulated") were excised from gels and re-amplified by polymerase chain reaction (PCR). After sequencing and comparison with the National Center for Biotechnology Information database, 176 TDFs presented significant similarity with known genes that could be classified into the following categories: metabolism and energy, stress resistance and defense, storage, signal transduction, and other functional categories. Using semiquantitative reverse-transcription PCR and real-time PCR approaches, the significance of the differences was further confirmed in fresh seeds and dehydrated seeds. The genes that encode superoxide dismutase, peroxiredoxin, caleosin, oleosin S3, steroleosin, late embryogenesis abundant protein, glutathione reductase, β-glucosidase, S23 transcriptional repressor, and some heat-shock proteins could be associated with DT. The results of this study will aid in the identification of candidate genes for future experiments that seek to understand seed DT. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

Sweet vernal grasses (Anthoxanthum) colonized African mountains along two fronts in the Late Pliocene, followed by secondary contact, polyploidization and local extinction in the Pleistocene.

PubMed

Tusiime, Felly Mugizi; Gizaw, Abel; Wondimu, Tigist; Masao, Catherine Aloyce; Abdi, Ahmed Abdikadir; Muwanika, Vincent; Trávníček, Pavel; Nemomissa, Sileshi; Popp, Magnus; Eilu, Gerald; Brochmann, Christian; Pimentel, Manuel

2017-07-01

High tropical mountains harbour remarkable and fragmented biodiversity thought to a large degree to have been shaped by multiple dispersals of cold-adapted lineages from remote areas. Few dated phylogenetic/phylogeographic analyses are however available. Here, we address the hypotheses that the sub-Saharan African sweet vernal grasses have a dual colonization history and that lineages of independent origins have established secondary contact. We carried out rangewide sampling across the eastern African high mountains, inferred dated phylogenies from nuclear ribosomal and plastid DNA using Bayesian methods, and performed flow cytometry and AFLP (amplified fragment length polymorphism) analyses. We inferred a single Late Pliocene western Eurasian origin of the eastern African taxa, whose high-ploid populations in one mountain group formed a distinct phylogeographic group and carried plastids that diverged from those of the currently allopatric southern African lineage in the Mid- to Late Pleistocene. We show that Anthoxanthum has an intriguing history in sub-Saharan Africa, including Late Pliocene colonization from southeast and north, followed by secondary contact, hybridization, allopolyploidization and local extinction during one of the last glacial cycles. Our results add to a growing body of evidence showing that isolated tropical high mountain habitats have a dynamic recent history involving niche conservatism and recruitment from remote sources, repeated dispersals, diversification, hybridization and local extinction. © 2017 John Wiley & Sons Ltd.

Genetic Divergence and Chemotype Diversity in the Fusarium Head Blight Pathogen Fusarium poae.

PubMed

Vanheule, Adriaan; De Boevre, Marthe; Moretti, Antonio; Scauflaire, Jonathan; Munaut, Françoise; De Saeger, Sarah; Bekaert, Boris; Haesaert, Geert; Waalwijk, Cees; van der Lee, Theo; Audenaert, Kris

2017-08-23

Fusarium head blight is a disease caused by a complex of Fusarium species. F. poae is omnipresent throughout Europe in spite of its low virulence. In this study, we assessed a geographically diverse collection of F. poae isolates for its genetic diversity using AFLP (Amplified Fragment Length Polymorphism). Furthermore, studying the mating type locus and chromosomal insertions, we identified hallmarks of both sexual recombination and clonal spread of successful genotypes in the population. Despite the large genetic variation found, all F. poae isolates possess the nivalenol chemotype based on Tri7 sequence analysis. Nevertheless, Tri gene clusters showed two layers of genetic variability. Firstly, the Tri1 locus was highly variable with mostly synonymous mutations and mutations in introns pointing to a strong purifying selection pressure. Secondly, in a subset of isolates, the main trichothecene gene cluster was invaded by a transposable element between Tri5 and Tri6 . To investigate the impact of these variations on the phenotypic chemotype, mycotoxin production was assessed on artificial medium. Complex blends of type A and type B trichothecenes were produced but neither genetic variability in the Tri genes nor variability in the genome or geography accounted for the divergence in trichothecene production. In view of its complex chemotype, it will be of utmost interest to uncover the role of trichothecenes in virulence, spread and survival of F. poae .

Patterns of post-glacial genetic differentiation in marginal populations of a marine microalga.

PubMed

Tahvanainen, Pia; Alpermann, Tilman J; Figueroa, Rosa Isabel; John, Uwe; Hakanen, Päivi; Nagai, Satoshi; Blomster, Jaanika; Kremp, Anke

2012-01-01

This study investigates the genetic structure of an eukaryotic microorganism, the toxic dinoflagellate Alexandrium ostenfeldii, from the Baltic Sea, a geologically young and ecologically marginal brackish water estuary which is predicted to support evolution of distinct, genetically impoverished lineages of marine macroorganisms. Analyses of the internal transcribed spacer (ITS) sequences and Amplified Fragment Length Polymorphism (AFLP) of 84 A. ostenfeldii isolates from five different Baltic locations and multiple external sites revealed that Baltic A. ostenfeldii is phylogenetically differentiated from other lineages of the species and micro-geographically fragmented within the Baltic Sea. Significant genetic differentiation (F(ST)) between northern and southern locations was correlated to geographical distance. However, instead of discrete genetic units or continuous genetic differentiation, the analysis of population structure suggests a complex and partially hierarchic pattern of genetic differentiation. The observed pattern suggests that initial colonization was followed by local differentiation and varying degrees of dispersal, most likely depending on local habitat conditions and prevailing current systems separating the Baltic Sea populations. Local subpopulations generally exhibited low levels of overall gene diversity. Association analysis suggests predominately asexual reproduction most likely accompanied by frequency shifts of clonal lineages during planktonic growth. Our results indicate that the general pattern of genetic differentiation and reduced genetic diversity of Baltic populations found in large organisms also applies to microscopic eukaryotic organisms.

Drifting propagules and receding swamps: genetic footprints of mangrove recolonization and dispersal along tropical coasts.

PubMed

Nettel, Alejandro; Dodd, Richard S

2007-04-01

Two issues that have captured the attention of tropical plant evolutionary biologists in recent years are the relative role of long distance dispersal (LDD) over vicariance in determining plant distributions and debate about the extent that Quaternary climatic changes affected tropical species. Propagules of some mangrove species are assumed to be capable of LDD due to their ability to float and survive for long periods of time in salt water. Mangrove species responded to glaciations with a contraction of their range. Thus, widespread mangrove species are an ideal system to study LDD and recolonization in the tropics. We present phylogenetic and phylogeographic analyses based on internal transcribed spacers region (ITS) sequences, chloroplast DNA (cpDNA), and amplified fragment length polymorphisms (AFLPs) of genomic DNA that demonstrate recent LDD across the Atlantic, rejecting the hypothesis of vicariance for the widespread distribution of the black mangrove (Avicennia germinans). Northern latitude populations likely became extinct during the late Quaternary due to frosts and aridification; these locations were recolonized afterward from southern populations. In some low latitude regions populations went extinct or were drastically reduced during the Quaternary because of lack of suitable habitat as sea levels changed. Our analyses show that low latitude Pacific populations of A. germinans harbor more diversity and reveal deeper divergence than Atlantic populations. Implications for our understanding of phylogeography of tropical species are discussed.

Pleistocene refugia and genetic diversity patterns in West Africa: Insights from the liana Chasmanthera dependens (Menispermaceae)

PubMed Central

Schmidt, Marco; Muellner-Riehl, Alexandra Nora; Ogundipe, Oluwatoyin Temitayo; Paule, Juraj

2017-01-01

Processes shaping the African Guineo-Congolian rain forest, especially in the West African part, are not well understood. Recent molecular studies, based mainly on forest tree species, confirmed the previously proposed division of the western African Guineo-Congolian rain forest into Upper Guinea (UG) and Lower Guinea (LG) separated by the Dahomey Gap (DG). Here we studied nine populations in the area of the DG and the borders of LG and UG of the widespread liana species, Chasmanthera dependens (Menispermaceae) by amplified fragment length polymorphism (AFLP), a chloroplast DNA sequence marker, and modelled the distribution based on current as well as paleoclimatic data (Holocene Climate Optimum, ca. 6 kyr BP and Last Glacial Maximum, ca. 22 kyr BP). Current population genetic structure and geographical pattern of cpDNA was related to present as well as historical modelled distributions. Results from this study show that past historical factors played an important role in shaping the distribution of C. dependens across West Africa. The Cameroon Volcanic Line seems to represent a barrier for gene flow in the present as well as in the past. Distribution modelling proposed refugia in the Dahomey Gap, supported also by higher genetic diversity. This is in contrast with the phylogeographic patterns observed in several rainforest tree species and could be explained by either diverging or more relaxed ecological requirements of this liana species. PMID:28301470

Evolution of genomic diversity and sex at extreme environments: Fungal life under hypersaline Dead Sea stress

PubMed Central

Kis-Papo, Tamar; Kirzhner, Valery; Wasser, Solomon P.; Nevo, Eviatar

2003-01-01

We have found that genomic diversity is generally positively correlated with abiotic and biotic stress levels (1–3). However, beyond a high-threshold level of stress, the diversity declines to a few adapted genotypes. The Dead Sea is the harshest planetary hypersaline environment (340 g·liter–1 total dissolved salts, ≈10 times sea water). Hence, the Dead Sea is an excellent natural laboratory for testing the “rise and fall” pattern of genetic diversity with stress proposed in this article. Here, we examined genomic diversity of the ascomycete fungus Aspergillus versicolor from saline, nonsaline, and hypersaline Dead Sea environments. We screened the coding and noncoding genomes of A. versicolor isolates by using >600 AFLP (amplified fragment length polymorphism) markers (equal to loci). Genomic diversity was positively correlated with stress, culminating in the Dead Sea surface but dropped drastically in 50- to 280-m-deep seawater. The genomic diversity pattern paralleled the pattern of sexual reproduction of fungal species across the same southward gradient of increasing stress in Israel. This parallel may suggest that diversity and sex are intertwined intimately according to the rise and fall pattern and adaptively selected by natural selection in fungal genome evolution. Future large-scale verification in micromycetes will define further the trajectories of diversity and sex in the rise and fall pattern. PMID:14645702

Inheritance of restriction fragment length polymorphisms, random amplified polymorphic DNAs and isozymes in coastal Douglas-fir

Treesearch

K.D. Jermstad; A.M. Reem; J.R. Henifin; N.C. Wheeler; D.B Neale

1994-01-01

A total of 225 new genetic loci [151 restriction fragment length polymorphisms (RFLP) and 74 random amplified polymorphic DNAs (RAPD)] in coastal Douglas- fir [Pseudotsuga menziesii (Mirb.) Franco var. menziesii] have been identified using a three-generation outbred pedigree. The Mendelian inheritance of 16 RFLP loci and 29...

Restriction fragment length polymorphism among Israeli Holstein-Friesian dairy bulls.

PubMed

Beckmann, J S; Kashi, Y; Hallerman, E M; Nave, A; Soller, M

1986-01-01

Israeli Holstein-Friesian dairy bulls were screened for restriction fragment length polymorphisms by hybridizing cloned DNA probes for bovine growth hormone, for chymosin, and for rat muscle beta-actin to restriction endonuclease-digested DNA immobilized on nitrocellulose filters. The population proved to be polymorphic at the growth hormone locus, with evidence consistent with the phenotypes being inherited in allelic fashion. A low level of polymorphism was also observed at one of the beta-actin gene family loci. The chymosin locus was monomorphic with the restriction enzymes utilized. The results illustrate the power of restriction fragment length polymorphism methodology in visualizing genetic variability in dairy cattle populations.

Acute fatty liver of pregnancy: a clinical study of 12 episodes in 11 patients.

PubMed Central

Reyes, H; Sandoval, L; Wainstein, A; Ribalta, J; Donoso, S; Smok, G; Rosenberg, H; Meneses, M

1994-01-01

Twelve episodes of acute fatty liver of pregnancy (AFLP) were diagnosed in 11 patients during the past 18 years in a general hospital in Santiago, Chile, with a prevalence of 1 per 15,900 deliveries. Acute fatty liver of pregnancy started between the 31st and 38th weeks of pregnancy, with malaise, vomiting, jaundice, and lethargy as the main clinical manifestations. Polydipsia (in nine episodes) and skin pruritus (in seven episodes) were unusual clinical findings. In two patients, pruritus started two and four weeks before AFLP, suggesting that an intrahepatic cholestasis of pregnancy preceded AFLP in those patients. Considering the current prevalence of both diseases in Chile, their association should be considered fortuitous. In another patient, two consecutive pregnancies were affected by AFLP, raising to three the number of reported patients with recurrent AFLP. In 11 episodes, liver biopsies supported the diagnosis of AFLP by showing small and midsized vacuolar cytoplasmic transformation as the most prominent histopathological feature. Positive intracellular fat staining was found in the four samples analysed. Studies by electron microscopy showed megamitochondria with paracrystalline inclusions in four samples. All the mothers survived, but fetal mortality was 58.3%. Several extrahepatic complications delayed maternal recovery for up to four weeks after delivery. This study confirms an improvement in maternal prognosis in AFLP, discusses the possibility of an epidemiological association with intrahepatic cholestasis of pregnancy, and increases the number of patients reported with recurrent AFLP. Images Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:8307428

Molecular characterization and antifungal susceptibility of Cryptococcus neoformans strains collected from a single institution in Lima, Peru.

PubMed

Bejar, Vilma; Tello, Mercedes; García, Ruth; Guevara, José M; Gonzales, Sofia; Vergaray, German; Valencia, Esther; Abanto, Enma; Ortega-Loayza, Alex G; Hagen, Ferry; Gutierrez, Ericson L

2015-01-01

Cryptococcosis is a fungal infection with a worldwide distribution, mainly caused by Cryptococcus neoformans and Cryptococcus gattii. To molecularly characterize the mating-types, serotypes, genotypes and antifungal susceptibility profiles of a set of retrospectively isolated C. neoformans strains from Lima, Peru. A set of 32 Cryptococcus spp. strains from the Institute of Tropical Medicine of the National University of San Marcos, Lima, Peru, were included in this retrospective study. Twenty-four strains were isolated from patients, while the remaining 8 were isolated from the environment. Using conventional PCR, 27 (84.4%) of the isolates were identified as C. neoformans var. grubii mating-type alpha and serotype A. Using the AFLP fingerprinting, it was shown that 16 (50%) of the C. neoformans strains were genotype AFLP1, 13 (40.6%) were genotype AFLP1B, 2 (6.3%) were genotype AFLP2, and 1 (3.1%) was found to be a hybrid between both C. neoformans varieties (genotype AFLP3). The antifungal susceptibility profiles for amphotericin B, fluconazole and voriconazole showed that all the 32 C. neoformans are sensitive to these antifungal compounds. In this study we observed that C. neoformans var. grubii (AFLP1 and AFLP1B) and C. neoformans var. neoformans (AFLP2) were the only cryptococcal varieties involved. All strains were found to be sensitive to the antifungals tested, results that are consistent with those found in the international literature. Copyright © 2013 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.

Determination of epigenetic inheritance, genetic inheritance, and estimation of genome DNA methylation in a full-sib family of Cupressus sempervirens L.

PubMed

Avramidou, Evangelia V; Doulis, Andreas G; Aravanopoulos, Filippos A

2015-05-15

Genetic inheritance and epigenetic inheritance are significant determinants of plant evolution, adaptation and plasticity. We studied inheritance of restriction site polymorphisms by the f-AFLP method and epigenetic DNA cytosine methylation inheritance by the f-MSAP technique. The study involved parents and 190 progeny of a Cupressus sempervirens L. full-sib family. Results from AFLP genetic data revealed that 71.8% of the fragments studied are under Mendelian genetic control, whereas faithful Mendelian inheritance for the MSAP fragments was low (4.29%). Further, MSAP fragment analysis showed that total methylation presented a mean of 28.2%, which was higher than the midparent value, while maternal inheritance was higher (5.65%) than paternal (3.01%). Interestingly de novo methylation in the progeny was high (19.65%) compared to parental methylation. Genetic and epigenetic distances for parents and offspring were not correlated (R(2)=0.0005). Furthermore, we studied correlation of total relative methylation and CG methylation with growth (height, diameter). We found CG/CNG methylation (N: A, C, T) to be positively correlated with height and diameter, while total relative methylation and CG methylation were positively correlated with height. Results are discussed in light of further research needed and of their potential application in breeding. Copyright © 2015 Elsevier B.V. All rights reserved.

Genetic Variability of Beauveria bassiana and a DNA Marker for Environmental Monitoring of a Highly Virulent Isolate Against Cosmopolites sordidus.

PubMed

Ferri, D V; Munhoz, C F; Neves, P M O; Ferracin, L M; Sartori, D; Vieira, M L C; Fungaro, M H P

2012-12-01

The banana weevil Cosmopolites sordidus (Germar) is one of a number of pests that attack banana crops. The use of the entomopathogenic fungus Beauveria bassiana as a biological control agent for this pest may contribute towards reducing the application of chemical insecticides on banana crops. In this study, the genetic variability of a collection of Brazilian isolates of B. bassiana was evaluated. Samples were obtained from various geographic regions of Brazil, and from different hosts of the Curculionidae family. Based on the DNA fingerprints generated by RAPD and AFLP, we found that 92 and 88 % of the loci were polymorphic, respectively. The B. bassiana isolates were attributed to two genotypic clusters based on the RAPD data, and to three genotypic clusters, when analyzed with AFLP. The nucleotide sequences of nuclear ribosomal DNA intergenic spacers confirmed that all isolates are in fact B. bassiana. Analysis of molecular variance showed that variability among the isolates was not correlated with geographic origin or hosts. A RAPD-specific marker for isolate CG 1024, which is highly virulent to C. sordidus, was cloned and sequenced. Based on the sequences obtained, specific PCR primers BbasCG1024F (5'-TGC GGC TGA GGA GGA CT-3') and BbasCG1024R (5'-TGC GGC TGA GTG TAG AAC-3') were designed for detecting and monitoring this isolate in the field.

Molecular basis of length polymorphism in the human zeta-globin gene complex.

PubMed Central

Goodbourn, S E; Higgs, D R; Clegg, J B; Weatherall, D J

1983-01-01

The length polymorphism between the human zeta-globin gene and its pseudogene is caused by an allele-specific variation in the copy number of a tandemly repeating 36-base-pair sequence. This sequence is related to a tandemly repeated 14-base-pair sequence in the 5' flanking region of the human insulin gene, which is known to cause length polymorphism, and to a repetitive sequence in intervening sequence (IVS) 1 of the pseudo-zeta-globin gene. Evidence is presented that the latter is also of variable length, probably because of differences in the copy number of the tandem repeat. The homology between the three length polymorphisms may be an indication of the presence of a more widespread group of related sequences in the human genome, which might be useful for generalized linkage studies. PMID:6308667

Fragment length polymorphisms among independent isolates of Epstein-Barr virus from immunocompromised and normal hosts.

PubMed

Katz, B Z; Niederman, J C; Olson, B A; Miller, G

1988-02-01

DNA restriction fragment length polymorphisms of Epstein-Barr virus (EBV) DNA were used as a molecular epidemiological tool to study multiple isolates of virus from the same and different individuals. We studied 35 EBV isolates: 19 from seven immunocompromised children and 16 from seven college students with mononucleosis. Analysis of the fragment length polymorphisms in this collection of isolates permitted several conclusions. Sites of polymorphism were most often encountered in regions with repetitive DNA. Epidemiologically unrelated patients harbored viruses that could be readily distinguished; by contrast, two infants and their mothers harbored similar viruses. Isolates from different sites in the same patient were similar. Variations between different clinical isolates of EBV mimic those found between different laboratory strains of the virus. Fragment length polymorphisms thus provide a useful marker for studying transmission and pathogenesis of EBV infections.

Temporal patterns of genetic variation across a 9-year-old aerial seed bank of the shrub Banksia hookeriana (Proteaceae).

PubMed

Barrett, Luke G; He, Tianhua; Lamont, Byron B; Krauss, Siegfried L

2005-11-01

The pattern of accumulation of genetic variation over time in seed banks is poorly understood. We examined the genetic structure of the aerial seed bank of Banksia hookeriana within a single 15-year-old population in fire-prone southwestern Australia, and compared genetic variation between adults and each year of a 9-year-old seed bank using amplified fragment length polymorphism (AFLP). B. hookeriana is well suited to the study of seed bank dynamics due to the canopy storage of its seeds, and because each annual crop can be identified. A total of 304 seeds from nine crop years and five maternal plants were genotyped, along with 113 plants from the adult population. Genetic variation, as assessed by the proportion of polymorphic markers (P(p)) and Shannon's index (I), increased slightly within the seed bank over time, while gene diversity (H(j)), did not change. P(p), I, and H(j) all indicated that genetic variation within the seed bank quickly approached the maximal level detected. Analysis of molecular variance revealed that less than 4% of variation could be accounted for by variation among seeds produced in different years, whereas there was greater differentiation among maternal plants (12.7%), and among individual seeds produced by different maternal plants (83.4%). With increasing population age, offspring generated each year were slightly more outbred, as indicated by an increase in the mean number of nonmaternal markers per offspring. There were no significant differences for H(j) or I between adults and the seed bank. Viability of seeds decreased with age, such that the viability of 9-year-old seeds was half that of 2-year-old seeds. These results suggest that variable fire frequencies have only limited potential to influence the amount of genetic variation stored within the seed bank of B. hookeriana.

Phylogeography and molecular diversity analysis of Jatropha curcas L. and the dispersal route revealed by RAPD, AFLP and nrDNA-ITS analysis.

PubMed

Sudheer Pamidimarri, D V N; Reddy, Muppala P

2014-05-01

Jatropha curcas L. (Euphorbiaceae) has acquired a great importance as a renewable source of energy with a number of environmental benefits. Very few attempts were made to understand the extent of genetic diversity and its distribution. This study was aimed to study the diversity and deduce the phylogeography of Jatropha curcas L. which is said to be the most primitive species of the genus Jatropha. Here we studied the intraspecific genetic diversity of the species distributed in different parts of the globe. The study also focused to understand the molecular diversity at reported probable center of origin (Mexico), and to reveal the dispersal route to other regions based on random amplified polymorphic DNA, amplified fragment length polymorphism and nrDNA-ITS sequences data. The overall genetic diversity of J. curcas found in the present study was narrow. The highest genetic diversity was observed in the germplasm collected from Mexico and supports the earlier hypothesis based on morphological data and natural distribution, it is the center for origin of the species. Least genetic diversity found in the Indian germplasm and clustering results revealed that the species was introduced simultaneously by two distinct germplasm and subsequently distributed in different parts of India. The present molecular data further revealed that J. curcas might have spread from the center of the origin to Cape Verde, than to Spain, Portuguese to other neighboring countries and simultaneously to Africa. The molecular evidence supports the Burkill et al. (A dictionary of the economic products of the Malay Peninsula, Governments of Malaysia and Singapore by the Ministry of Agriculture and Co-operatives. Kuala Lumpur, Malaysia, 1966) view of Portuguese might have introduced the species to India. The clustering pattern suggests that the distribution was interfered by human activity.

Diversity hotspots of the laurel forest on Tenerife, Canary Islands: a phylogeographic study of Laurus and Ixanthus.

PubMed

Betzin, Anja; Thiv, Mike; Koch, Marcus A

2016-09-01

Macaronesian laurel forest is among the worldwide hotspots of threatened biodiversity. With increasing evidence that woodland composition on the Canary Islands changed dramatically during the last few thousand years, the aim of this study was to find evidence for substantial recent population dynamics of two representative species from laurel forest. Amplified fragment length polymorphism (AFLP) was used to evaluate fine-scaled genetic variation of the paradigmatic tree Laurus novocanariensis (Lauraceae) and a long-lived herbaceous gentian from core laurel forest, Ixanthus viscosus (Gentianaceae), on Tenerife. Bioclimatic variables were analysed to study the respective climate niches. A chloroplast DNA screening was performed to evaluate additional genetic variation. Genetic diversity of the laurel tree showed severe geographic partitioning. On Tenerife, fine-scaled Bayesian clustering of genetic variation revealed a western and an eastern gene pool, separated by a zone of high admixture and with a third major gene pool. Compared with genetic clusters found on the other Canary Islands, the East-West differentiation on Tenerife seems to be more recent than differentiation between islands. This is substantiated by the finding of extremly low levels of chloroplast DNA-based polymorphisms. Ixanthus showed no geographic structuring of genetic variation. Genetic data from Tenerife indicate contemporary gene flow and dispersal on a micro/local scale rather than reflecting an old and relic woodland history. In particular for Laurus, it is shown that this species occupies a broad bioclimatic niche. This is not correlated with its respective distribution of genetic variation, therefore indicating its large potential for contemporary rapid and effective colonization. Ixanthus is more specialized to humid conditions and is mostly found in the natural Monteverde húmedo vegetation types, but even for this species indications for long-term persistence in the respective bioclimatically differentiated regions was not find. © The Author 2016. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Genetic structure in the coral, Montastraea cavernosa: assessing genetic differentiation among and within Mesophotic reefs.

PubMed

Brazeau, Daniel A; Lesser, Michael P; Slattery, Marc

2013-01-01

Mesophotic coral reefs (30-150 m) have recently received increased attention as a potential source of larvae (e.g., the refugia hypothesis) to repopulate a select subset of the shallow water (<30 m) coral fauna. To test the refugia hypothesis we used highly polymorphic Amplified Fragment Length Polymorphism (AFLP) markers as a means to assess small-scale genetic heterogeneity between geographic locations and across depth clines in the Caribbean coral, Montastraea cavernosa. Zooxanthellae-free DNA extracts of coral samples (N = 105) were analyzed from four depths, shallow (3-10 m), medium (15-25 m), deep (30-50 m) and very deep (60-90 m) from Little Cayman Island (LCI), Lee Stocking Island (LSI), Bahamas and San Salvador (SS), Bahamas which range in distance from 170 to 1,600 km apart. Using AMOVA analysis there were significant differences in ΦST values in pair wise comparisons between LCI and LSI. Among depths at LCI, there was significant genetic differentiation between shallow and medium versus deep and very deep depths in contrast there were no significant differences in ΦST values among depths at LSI. The assignment program AFLPOP, however, correctly assigned 95.7% of the LCI and LSI samples to the depths from which they were collected, differentiating among populations as little as 10 to 20 m in depth from one another. Discriminant function analysis of the data showed significant differentiation among samples when categorized by collection site as well as collection depth. FST outlier analyses identified 2 loci under positive selection and 3 under balancing selection at LCI. At LSI 2 loci were identified, both showing balancing selection. This data shows that adult populations of M. cavernosa separated by depths of tens of meters exhibits significant genetic structure, indicative of low population connectivity among and within sites and are not supplying successful recruits to adjacent coral reefs less than 30 m in depth.

Introgression of genomic components from Chinese Brassica rapa contributes to widening the genetic diversity in rapeseed (B. napus L.), with emphasis on the evolution of Chinese rapeseed.

PubMed

Qian, W; Meng, J; Li, M; Frauen, M; Sass, O; Noack, J; Jung, C

2006-06-01

In spite of its short history of being an oil crop in China, the Chinese semi-winter rapeseed (Brassica napus L., 2n = 38, AACC) has been improved rapidly by intentional introgression of genomic components from Chinese B. rapa (2n = 20, AA). As a result, the Chinese semi-winter rapeseed has diversified genetically from the spring and winter rapeseed grown in the other regions such as Europe and North America. The objectives of this study were to investigate the roles of the introgression of the genomic components from the Chinese B. rapa in widening the genetic diversity of rapeseed and to verify the role of this introgression in the evolution of the Chinese rapeseed. Ten lines of the new type of rapeseed, which were produced by introgression of Chinese B. rapa to Chinese normal rapeseed, were compared for genetic diversity using amplified fragment length polymorphism (AFLP) with three groups of 35 lines of the normal rapeseed, including 9 semi-winter rapeseed lines from China, 9 winter rapeseed lines from Europe and 17 spring rapeseed lines from Northern Europe, Canada and Australia. Analysis of 799 polymorphic fragments revealed that within the groups, the new type rapeseed had the highest genetic diversity, followed by the semi-winter normal rapeseed from China. Spring and winter rapeseed had the lowest genetic diversity. Among the groups, the new type rapeseed group had the largest average genetic distance to the other three groups. Principal component analysis and cluster analysis, however, could not separate the new type rapeseed group from Chinese normal rapeseed group. Our data suggested that the introgression of Chinese B. rapa could significantly diversify the genetic basis of the rapeseed and play an important role in the evolution of Chinese rapeseed. The use of new genetic variation for the exploitation of heterosis in Brassica hybrid breeding is discussed.

A hybrid BAC physical map of potato: a framework for sequencing a heterozygous genome

PubMed Central

2011-01-01

Background Potato is the world's third most important food crop, yet cultivar improvement and genomic research in general remain difficult because of the heterozygous and tetraploid nature of its genome. The development of physical map resources that can facilitate genomic analyses in potato has so far been very limited. Here we present the methods of construction and the general statistics of the first two genome-wide BAC physical maps of potato, which were made from the heterozygous diploid clone RH89-039-16 (RH). Results First, a gel electrophoresis-based physical map was made by AFLP fingerprinting of 64478 BAC clones, which were aligned into 4150 contigs with an estimated total length of 1361 Mb. Screening of BAC pools, followed by the KeyMaps in silico anchoring procedure, identified 1725 AFLP markers in the physical map, and 1252 BAC contigs were anchored the ultradense potato genetic map. A second, sequence-tag-based physical map was constructed from 65919 whole genome profiling (WGP) BAC fingerprints and these were aligned into 3601 BAC contigs spanning 1396 Mb. The 39733 BAC clones that overlap between both physical maps provided anchors to 1127 contigs in the WGP physical map, and reduced the number of contigs to around 2800 in each map separately. Both physical maps were 1.64 times longer than the 850 Mb potato genome. Genome heterozygosity and incomplete merging of BAC contigs are two factors that can explain this map inflation. The contig information of both physical maps was united in a single table that describes hybrid potato physical map. Conclusions The AFLP physical map has already been used by the Potato Genome Sequencing Consortium for sequencing 10% of the heterozygous genome of clone RH on a BAC-by-BAC basis. By layering a new WGP physical map on top of the AFLP physical map, a genetically anchored genome-wide framework of 322434 sequence tags has been created. This reference framework can be used for anchoring and ordering of genomic sequences of clone RH (and other potato genotypes), and opens the possibility to finish sequencing of the RH genome in a more efficient way via high throughput next generation approaches. PMID:22142254

Genetic admixture and lineage separation in a southern Andean plant

PubMed Central

Morello, Santiago; Sede, Silvana M.

2016-01-01

Mountain uplifts have generated new ecologic opportunities for plants, and triggered evolutionary processes, favouring an increase on the speciation rate in all continents. Moreover, mountain ranges may act as corridors or barriers for plant lineages and populations. In South America a high rate of diversification has been linked to Andean orogeny during Pliocene/Miocene. More recently, Pleistocene glacial cycles have also shaped species distribution and demography. The endemic genus Escallonia is known to have diversified in the Andes. Species with similar morphology obscure species delimitation and plants with intermediate characters occur naturally. The aim of this study is to characterize genetic variation and structure of two widespread species of Escallonia: E. alpina and E. rubra. We analyzed the genetic variation of populations of the entire distribution range of the species and we also included those with intermediate morphological characters; a total of 94 accessions from 14 populations were used for the Amplified Fragment Length Polymorphism (AFLP) analysis. Plastid DNA sequences (trnS-trnG, 3′trnV-ndhC intergenic spacers and the ndhF gene) from sixteen accessions of Escallonia species were used to construct a Statistical Parsimony network. Additionally, we performed a geometric morphometrics analysis on 88 leaves from 35 individuals of the two E. alpina varieties to further study their differences. Wright’s Fst and analysis of molecular variance tests performed on AFLP data showed a significant level of genetic structure at the species and population levels. Intermediate morphology populations showed a mixed genetic contribution from E. alpina var. alpina and E. rubra both in the Principal Coordinates Analysis (PCoA) and STRUCTURE. On the other hand, E. rubra and the two varieties of E. alpina are well differentiated and assigned to different genetic clusters. Moreover, the Statistical Parsimony network showed a high degree of divergence between the varieties of E. alpina: var. alpina is more closely related to E. rubra and other species than to its own counterpart E. alpina var. carmelitana. Geometric morphometrics analysis (Elliptic Fourier descriptors) revealed significant differences in leaf shape between varieties. We found that diversity in Escallonia species analyzed here is geographically structured and deep divergence between varieties of E. alpina could be associated to ancient evolutionary events like orogeny. Admixture in southern populations could be the result of hybridization at the margins of the parental species’ distribution range. PMID:27179539

Genetic fine-mapping of DIPLOSPOROUS in Taraxacum (dandelion; Asteraceae) indicates a duplicated DIP-gene

PubMed Central

2010-01-01

Background DIPLOSPOROUS (DIP) is the locus for diplospory in Taraxacum, associated to unreduced female gamete formation in apomicts. Apomicts reproduce clonally through seeds, including apomeiosis, parthenogenesis, and autonomous or pseudogamous endosperm formation. In Taraxacum, diplospory results in first division restitution (FDR) nuclei, and inherits as a dominant, monogenic trait, independent from the other apomixis elements. A preliminary genetic linkage map indicated that the DIP-locus lacks suppression of recombination, which is unique among all other map-based cloning efforts of apomeiosis to date. FDR as well as apomixis as a whole are of interest in plant breeding, allowing for polyploidization and fixation of hybrid vigor, respectively. No dominant FDR or apomixis genes have yet been isolated. Here, we zoom-in to the DIP-locus by largely extending our initial mapping population, and by analyzing (local) suppression of recombination and allele sequence divergence (ASD). Results We identified 24 recombinants between two most closely linked molecular markers to DIP in an F1-population of 2227 plants that segregates for diplospory and lacks parthenogenesis. Both markers segregated c. 1:1 in the entire population, indicating a 1:1 segregation rate of diplospory. Fine-mapping showed three amplified fragment length polymorphisms (AFLPs) closest to DIP at 0.2 cM at one flank and a single AFLP at 0.4 cM at the other flank. Our data lacked strong evidence for ASD at marker regions close to DIP. An unexpected bias towards diplosporous plants among the recombinants (20 out of 24) was found. One third of these diplosporous recombinants showed incomplete penetrance of 50-85% diplospory. Conclusions Our data give interesting new insights into the structure of the diplospory locus in Taraxacum. We postulate a locus with a minimum of two DIP-genes and possibly including one or two enhancers or cis-regulatory elements on the basis of the bias towards diplosporous recombinants and incomplete penetrance of diplospory in some of them. We define the DIP-locus to 0.6 cM, which is estimated to cover ~200-300 Kb, with the closest marker at 0.2 cM. Our results confirm the minor role of suppression of recombination and ASD around DIP, making it an excellent candidate to isolate via a chromosome-walking approach. PMID:20659311

Genetic fine-mapping of DIPLOSPOROUS in Taraxacum (dandelion; Asteraceae) indicates a duplicated DIP-gene.

PubMed

Vijverberg, Kitty; Milanovic-Ivanovic, Slavica; Bakx-Schotman, Tanja; van Dijk, Peter J

2010-07-26

DIPLOSPOROUS (DIP) is the locus for diplospory in Taraxacum, associated to unreduced female gamete formation in apomicts. Apomicts reproduce clonally through seeds, including apomeiosis, parthenogenesis, and autonomous or pseudogamous endosperm formation. In Taraxacum, diplospory results in first division restitution (FDR) nuclei, and inherits as a dominant, monogenic trait, independent from the other apomixis elements. A preliminary genetic linkage map indicated that the DIP-locus lacks suppression of recombination, which is unique among all other map-based cloning efforts of apomeiosis to date. FDR as well as apomixis as a whole are of interest in plant breeding, allowing for polyploidization and fixation of hybrid vigor, respectively. No dominant FDR or apomixis genes have yet been isolated. Here, we zoom-in to the DIP-locus by largely extending our initial mapping population, and by analyzing (local) suppression of recombination and allele sequence divergence (ASD). We identified 24 recombinants between two most closely linked molecular markers to DIP in an F1-population of 2227 plants that segregates for diplospory and lacks parthenogenesis. Both markers segregated c. 1:1 in the entire population, indicating a 1:1 segregation rate of diplospory. Fine-mapping showed three amplified fragment length polymorphisms (AFLPs) closest to DIP at 0.2 cM at one flank and a single AFLP at 0.4 cM at the other flank. Our data lacked strong evidence for ASD at marker regions close to DIP. An unexpected bias towards diplosporous plants among the recombinants (20 out of 24) was found. One third of these diplosporous recombinants showed incomplete penetrance of 50-85% diplospory. Our data give interesting new insights into the structure of the diplospory locus in Taraxacum. We postulate a locus with a minimum of two DIP-genes and possibly including one or two enhancers or cis-regulatory elements on the basis of the bias towards diplosporous recombinants and incomplete penetrance of diplospory in some of them. We define the DIP-locus to 0.6 cM, which is estimated to cover approximately 200-300 Kb, with the closest marker at 0.2 cM. Our results confirm the minor role of suppression of recombination and ASD around DIP, making it an excellent candidate to isolate via a chromosome-walking approach.

Boxer crabs induce asexual reproduction of their associated sea anemones by splitting and intraspecific theft.

PubMed

Schnytzer, Yisrael; Giman, Yaniv; Karplus, Ilan; Achituv, Yair

2017-01-01

Crabs of the genus Lybia have the remarkable habit of holding a sea anemone in each of their claws. This partnership appears to be obligate, at least on the part of the crab. The present study focuses on Lybia leptochelis from the Red Sea holding anemones of the genus Alicia (family Aliciidae). These anemones have not been found free living, only in association with L. leptochelis . In an attempt to understand how the crabs acquire them, we conducted a series of behavioral experiments and molecular analyses. Laboratory observations showed that the removal of one anemone from a crab induces a "splitting" behavior, whereby the crab tears the remaining anemone into two similar parts, resulting in a complete anemone in each claw after regeneration. Furthermore, when two crabs, one holding anemones and one lacking them, are confronted, the crabs fight, almost always leading to the "theft" of a complete anemone or anemone fragment by the crab without them. Following this, crabs "split" their lone anemone into two. Individuals of Alicia sp. removed from freshly collected L. leptochelis were used for DNA analysis. By employing AFLP (Fluorescence Amplified Fragments Length Polymorphism) it was shown that each pair of anemones from a given crab is genetically identical. Furthermore, there is genetic identity between most pairs of anemone held by different crabs, with the others showing slight genetic differences. This is a unique case in which one animal induces asexual reproduction of another, consequently also affecting its genetic diversity.

Geographic pattern of genetic variation in the European globeflower Trollius europaeus L. (Ranunculaceae) inferred from amplified fragment length polymorphism markers.

PubMed

Despres, Laurence; Loriot, Sandrine; Gaudeul, Myriam

2002-11-01

The distribution of genetic variation and the phylogenetic relationships between 18 populations of the arctic-alpine plant Trollius europaeus were analysed in three main regions (Alps, Pyrenees and Fennoscandia) by using dominant AFLP markers. Analysis of molecular variance revealed that most of the genetic variability was found within populations (64%), although variation among regions (17%) and among populations within regions (19%) was highly significant (P < 0.001). Accordingly, the global fixation index FST averaged over loci was high (0.39). The among-population differentiation indicates restricted gene flow, congruent with limited dispersal of specific globeflower's pollinating flies (Chiastocheta spp.). Within-population diversity levels were significantly higher in the Alps (mean Nei's expected heterozygosity HE = 0.229) than in the Pyrenees (HE= 0.197) or in Fennoscandia (HE = 0.158). This finding is congruent with the species-richness of the associated flies, which is maximum in the Alps. We discuss the processes involved in shaping observed patterns of genetic diversity within and among T. europaeus populations. Genetic drift is the major factor acting on the small Pyrenean populations at the southern edge of T. europaeus distribution, while large Fennoscandian populations result probably from a founder effect followed by demographic expansion. The Alpine populations represent moderately fragmented relics of large southern ancestral populations. The patterns of genetic variability observed in the host plant support the hypothesis of sympatric speciation in associated flies, rather than recurrent allopatric speciations.

Relatedness of medically important strains of Saccharomyces cerevisiae as revealed by phylogenetics and metabolomics.

PubMed

MacKenzie, Donald A; Defernez, Marianne; Dunn, Warwick B; Brown, Marie; Fuller, Linda J; de Herrera, Santiago R M Seco; Günther, Andreas; James, Steve A; Eagles, John; Philo, Mark; Goodacre, Royston; Roberts, Ian N

2008-07-01

Ten medically important Saccharomyces strains, comprising six clinical isolates of Saccharomyces cerevisiae and four probiotic strains of Saccharomyces boulardii, were characterized at the genetic and metabolic level and compared with non-medical, commercial yeast strains used in baking and wine-making. Strains were compared by genetic fingerprinting using amplified fragment length polymorphism (AFLP) analysis, by ribosomal DNA ITS1 sequencing and by metabolic footprinting using both direct injection mass spectrometry (DIMS) and gas chromatography-time of flight-mass spectrometry (GC-ToF-MS). Overall, the clinical isolates fell into different groupings when compared with the non-medical strains, with good but not perfect correlation amongst strains at both the genetic and metabolic levels. Probiotic strains of S. boulardii that are used therapeutically to treat human gastro-intestinal tract disorders showed tight clustering both genetically and metabolically. Metabolomics was found to be of value both as a taxonomic tool and as a means to investigate anomalous links between genotype and phenotype. Key discriminatory metabolites were identified when comparing the three main groups of clinical, probiotic and non-medical strains and included molecules such as trehalose, myo-inositol, lactic acid, fumaric acid and glycerol 3-phosphate. This study confirmed the link between a subset of clinical isolates and baking or probiotic strains but also highlighted that in general the clinical strains were more diverse at both the genomic and metabolic levels. Copyright 2008 John Wiley & Sons, Ltd.

A polygalacturonase-inhibiting protein with a role in pea defence against the cyst nematode Heterodera goettingiana.

PubMed

Veronico, Pasqua; Melillo, M Teresa; Saponaro, Concetta; Leonetti, Paola; Picardi, Ernesto; Jones, John T

2011-04-01

A cDNA of 312 bp, similar to polygalacturonase-inhibiting proteins (PGIPs), was isolated by cDNA-amplified fragment length polymorphism (cDNA-AFLP) from pea roots infected with the cyst nematode Heterodera goettingiana. The deduced amino acid sequence obtained from the complete Pspgip1 coding sequence was very similar to PGIPs described from several other plant species, and was identical in both MG103738 and Progress 9 genotypes, resistant and susceptible to H. goettingiana, respectively. Reverse transcription-polymerase chain reaction (RT-PCR) expression analysis revealed the differential regulation of the Pspgip1 gene in the two genotypes in response to wounding and nematode challenge. Mechanical wounding induced Pspgip1 expression in MG103738 within 8 h, but this response was delayed in Progress 9. In contrast, the response to nematode infection was more complex. The transcription of Pspgip1 was triggered rapidly in both genotypes, but the expression level returned to levels observed in uninfected plants more quickly in susceptible than in resistant roots. In addition, in situ hybridization showed that Pspgip1 was expressed in the cortical cells damaged as a result of nematode invasion in both genotypes. However, it was specifically localized in the cells bordering the nematode-induced syncytia in resistant roots. This suggests a role for this gene in counteracting nematode establishment inside the root. © 2010 THE AUTHORS. MOLECULAR PLANT PATHOLOGY © 2010 BSPP AND BLACKWELL PUBLISHING LTD.

From homothally to heterothally: Mating preferences and genetic variation within clones of the dinoflagellate Gymnodinium catenatum

NASA Astrophysics Data System (ADS)

Figueroa, Rosa Isabel; Rengefors, Karin; Bravo, Isabel; Bensch, Staffan

2010-02-01

The chain-forming dinoflagellate Gymnodinium catenatum Graham is responsible for outbreaks of paralytic shellfish poisoning (PSP), a human health threat in coastal waters. Sexuality in this species is of great importance in its bloom dynamics, and has been shown to be very complex but lacks an explanation. For this reason, we tested if unreported homothallic behavior and rapid genetic changes may clarify the sexual system of this alga. To achieve this objective, 12 clonal strains collected from the Spanish coast were analyzed for the presence of sexual reproduction. Mating affinity results, self-compatibility studies, and genetic fingerprinting (amplified fragment length polymorphism, AFLP) analysis on clonal strains, showed three facts not previously described for this species: (i) That there is a continuous mating system within G. catenatum, with either self-compatible strains (homothallic), or strains that needed to be outcrossed (heterothallic), and with a range of differences in cyst production among the crosses. (ii) There was intraclonal genetic variation, i.e. genetic variation within an asexual lineage. Moreover, the variability among homothallic clones was smaller than among the heterothallic ones. (iii) Sibling strains (the two strains established by the germination of one cyst) increased their intra- and inter-sexual compatibility with time. To summarize, we have found that G. catenatum's sexual system is much more complex than previously described, including complex homothallic/heterothallic behaviors. Additionally, high rates of genetic variability may arise in clonal strains, although explanations for the mechanisms responsible are still lacking.

Molecular characterization of natural orchid in South slopes of Mount Merapi, Sleman regency, Yogyakarta

NASA Astrophysics Data System (ADS)

Ferdiani, Defika I.; Devi, Fera L.; Koentjana, Johan P.; Milasari, Asri F.; Nur'aini, Indah; Semiarti, Endang

2015-09-01

Natural orchid is one of the most important tropical biodiversity. In Indonesia there are ± 6000 species out of 30000 orchids species in the world, of which there are ± 60 species at Mount Merapi. Repetitive eruption of Merapi have wiped out the biodiversity of orchids, therefore the efforts to conserve the orchids and to establish the database of natural orchids in Mount Merapi are needed. The orchid's database can be created based on DNA analysis, and establish barcoding DNA. DNA-barcodes can be used as molecular markers. The different character of morphology usually shows different pattern in DNA fragments. This research aims to characterize the phenotype and genotype of natural orchids of Mt. Merapi based on morphology and the structure of DNA in trnL-F intergenic region of chloroplasts DNA of orchid. Amplified Fragment Length Polymorphism (AFLP) technique was used to characterize the molecular types of orchids in silico of intergenic space area of orchid chloroplast. In this study, 11 species of orchids were characterized based on morphological and molecular characters. The molecular characters were obtained from trnL-F intergenic region of leaves chloroplasts. The data indicates that there is a conserve DNA pattern in all orchids and the distinctive characters of some orchids. In this study, based on trnL-F intergenic region of chloroplast genome, the phylogenetic tree revealed that 11 species of orchids at Mt. Merapi can be grouped into 2 clades, that matched with morphological characters.

Wide Distribution of Mitochondrial Genome Rearrangements in Wild Strains of the Cultivated Basidiomycete Agrocybe aegerita

PubMed Central

Barroso, G.; Blesa, S.; Labarere, J.

1995-01-01

We used restriction fragment length polymorphisms to examine mitochondrial genome rearrangements in 36 wild strains of the cultivated basidiomycete Agrocybe aegerita, collected from widely distributed locations in Europe. We identified two polymorphic regions within the mitochondrial DNA which varied independently: one carrying the Cox II coding sequence and the other carrying the Cox I, ATP6, and ATP8 coding sequences. Two types of mutations were responsible for the restriction fragment length polymorphisms that we observed and, accordingly, were involved in the A. aegerita mitochondrial genome evolution: (i) point mutations, which resulted in strain-specific mitochondrial markers, and (ii) length mutations due to genome rearrangements, such as deletions, insertions, or duplications. Within each polymorphic region, the length differences defined only two mitochondrial types, suggesting that these length mutations were not randomly generated but resulted from a precise rearrangement mechanism. For each of the two polymorphic regions, the two molecular types were distributed among the 36 strains without obvious correlation with their geographic origin. On the basis of these two polymorphisms, it is possible to define four mitochondrial haplotypes. The four mitochondrial haplotypes could be the result of intermolecular recombination between allelic forms present in the population long enough to reach linkage equilibrium. All of the 36 dikaryotic strains contained only a single mitochondrial type, confirming the previously described mitochondrial sorting out after cytoplasmic mixing in basidiomycetes. PMID:16534984

Genetic characterization of fig tree mutants with molecular markers.

PubMed

Rodrigues, M G F; Martins, A B G; Desidério, J A; Bertoni, B W; Alves, M C

2012-08-06

The fig (Ficus carica L.) is a fruit tree of great world importance and, therefore, the genetic improvement becomes an important field of research for better crops, being necessary to gather information on this species, mainly regarding its genetic variability so that appropriate propagation projects and management are made. The improvement programs of fig trees using conventional procedures in order to obtain new cultivars are rare in many countries, such as Brazil, especially due to the little genetic variability and to the difficulties in obtaining plants from gamete fusion once the wasp Blastophaga psenes, responsible for the natural pollinating, is not found in Brazil. In this way, the mutagenic genetic improvement becomes a solution of it. For this reason, in an experiment conducted earlier, fig plants formed by cuttings treated with gamma ray were selected based on their agronomic characteristics of interest. We determined the genetic variability in these fig tree selections, using RAPD and AFLP molecular markers, comparing them to each other and to the Roxo-de-Valinhos, used as the standard. For the reactions of DNA amplification, 140 RAPD primers and 12 primer combinations for AFLP analysis were used. The selections did not differ genetically between themselves and between them and the Roxo-de-Valinhos cultivar. Techniques that can detect polymorphism between treatments, such as DNA sequencing, must be tested. The phenotypic variation of plants may be due to epigenetic variation, necessitating the use of techniques with methylation-sensitive restriction enzymes.

AFLP markers provide insights into the evolutionary relationships and diversification of New Caledonian Araucaria species (Araucariaceae).

PubMed

Gaudeul, Myriam; Rouhan, Germinal; Gardner, Martin F; Hollingsworth, Peter M

2012-01-01

Despite its small size, New Caledonia is characterized by a very diverse flora and striking environmental gradients, which make it an ideal setting to study species diversification. Thirteen of the 19 Araucaria species are endemic to the territory and form a monophyletic group, but patterns and processes that lead to such a high species richness are largely unexplored. We used 142 polymorphic AFLP markers and performed analyses based on Bayesian clustering algorithms, genetic distances, and cladistics on 71 samples representing all New Caledonian Araucaria species. We examined correlations between the inferred evolutionary relationships and shared morphological, ecological, or geographic parameters among species, to investigate evolutionary processes that may have driven speciation. We showed that genetic divergence among the present New Caledonian Araucaria species is low, suggesting recent diversification rather than pre-existence on Gondwana. We identified three genetic groups that included small-leaved, large-leaved, and coastal species, but detected no association with soil preference, ecological habitat, or rainfall. The observed patterns suggested that speciation events resulted from both differential adaptation and vicariance. Last, we hypothesize that speciation is ongoing and/or there are cryptic species in some genetically (sometimes also morphologically) divergent populations. Further data are required to provide better resolution and understanding of the diversification of New Caledonian Araucaria species. Nevertheless, our study allowed insights into their evolutionary relationships and provides a framework for future investigations on the evolution of this emblematic group of plants in one of the world's biodiversity hotspots.

High level of molecular and phenotypic biodiversity in Jatropha curcas from Central America compared to Africa, Asia and South America

PubMed Central

2014-01-01

Background The main bottleneck to elevate jatropha (Jatropha curcas L.) from a wild species to a profitable biodiesel crop is the low genetic and phenotypic variation found in different regions of the world, hampering efficient plant breeding for productivity traits. In this study, 182 accessions from Asia (91), Africa (35), South America (9) and Central America (47) were evaluated at genetic and phenotypic level to find genetic variation and important traits for oilseed production. Results Genetic variation was assessed with SSR (Simple Sequence Repeat), TRAP (Target Region Amplification Polymorphism) and AFLP (Amplified fragment length polymorphism) techniques. Phenotypic variation included seed morphological characteristics, seed oil content and fatty acid composition and early growth traits. Jaccard’s similarity and cluster analysis by UPGM (Unweighted Paired Group Method) with arithmetic mean and PCA (Principle Component Analysis) indicated higher variability in Central American accessions compared to Asian, African and South American accessions. Polymorphism Information Content (PIC) values ranged from 0 to 0.65. In the set of Central American accessions. PIC values were higher than in other regions. Accessions from the Central American population contain alleles that were not found in the accessions from other populations. Analysis of Molecular Variance (AMOVA; P < 0.0001) indicated high genetic variation within regions (81.7%) and low variation across regions (18.3%). A high level of genetic variation was found on early growth traits and on components of the relative growth rate (specific leaf area, leaf weight, leaf weight ratio and net assimilation rate) as indicated by significant differences between accessions and by the high heritability values (50–88%). The fatty acid composition of jatropha oil significantly differed (P < 0.05) between regions. Conclusions The pool of Central American accessions showed very large genetic variation as assessed by DNA-marker variation compared to accessions from other regions. Central American accessions also showed the highest phenotypic variation and should be considered as the most important source for plant breeding. Some variation in early growth traits was found within a group of accessions from Asia and Africa, while these accessions did not differ in a single DNA-marker, possibly indicating epigenetic variation. PMID:24666927

Androgen receptor repeat length polymorphism associated with male-to-female transsexualism.

PubMed

Hare, Lauren; Bernard, Pascal; Sánchez, Francisco J; Baird, Paul N; Vilain, Eric; Kennedy, Trudy; Harley, Vincent R

2009-01-01

There is a likely genetic component to transsexualism, and genes involved in sex steroidogenesis are good candidates. We explored the specific hypothesis that male-to-female transsexualism is associated with gene variants responsible for undermasculinization and/or feminization. Specifically, we assessed the role of disease-associated repeat length polymorphisms in the androgen receptor (AR), estrogen receptor beta (ERbeta), and aromatase (CYP19) genes. Subject-control analysis included 112 male-to-female transsexuals and 258 non-transsexual males. Associations and interactions were investigated between CAG repeat length in the AR gene, CA repeat length in the ERbeta gene, and TTTA repeat length in the CYP19 gene and male-to-female transsexualism. A significant association was identified between transsexualism and the AR allele, with transsexuals having longer AR repeat lengths than non-transsexual male control subjects (p=.04). No associations for transsexualism were evident in repeat lengths for CYP19 or ERbeta genes. Individuals were then classified as short or long for each gene polymorphism on the basis of control median polymorphism lengths in order to further elucidate possible combined effects. No interaction associations between the three genes and transsexualism were identified. This study provides evidence that male gender identity might be partly mediated through the androgen receptor.

Androgen Receptor Repeat Length Polymorphism Associated with Male-to-Female Transsexualism

PubMed Central

Hare, Lauren; Bernard, Pascal; Sánchez, Francisco J.; Baird, Paul N.; Vilain, Eric; Kennedy, Trudy; Harley, Vincent R.

2012-01-01

Background There is a likely genetic component to transsexualism, and genes involved in sex steroidogenesis are good candidates. We explored the specific hypothesis that male-to-female transsexualism is associated with gene variants responsible for undermasculinization and/or feminization. Specifically, we assessed the role of disease-associated repeat length polymorphisms in the androgen receptor (AR), estrogen receptor β (ERβ), and aromatase (CYP19) genes. Methods Subject-control analysis included 112 male-to-female transsexuals and 258 non-transsexual males. Associations and interactions were investigated between CAG repeat length in the AR gene, CA repeat length in the ERβ gene, and TTTA repeat length in the CYP19 gene and male-to-female transsexualism. Results A significant association was identified between transsexualism and the AR allele, with transsexuals having longer AR repeat lengths than non-transsexual male control subjects (p = .04). No associations for transsexualism were evident in repeat lengths for CYP19 or ERβ genes. Individuals were then classified as short or long for each gene polymorphism on the basis of control median polymorphism lengths in order to further elucidate possible combined effects. No interaction associations between the three genes and transsexualism were identified. Conclusions This study provides evidence that male gender identity might be partly mediated through the androgen receptor. PMID:18962445

Screening of Israeli Holstein-Friesian cattle for restriction fragment length polymorphisms using homologous and heterologous deoxyribonucleic acid probes.

PubMed

Hallerman, E M; Nave, A; Soller, M; Beckmann, J S

1988-12-01

Genomic DNA of Israeli Holstein-Friesian dairy cattle were screened with a battery of 17 cloned or subcloned DNA probes in an attempt to document restriction fragment length polymorphisms at a number of genetic loci. Restriction fragment length polymorphisms were observed at the chymosin, oxytocin-neurophysin I, lutropin beta, keratin III, keratin VI, keratin VII, prolactin, and dihydrofolate reductase loci. Use of certain genomic DNA fragments as probes produced hybridization patterns indicative of satellite DNA at the respective loci. Means for distinguishing hybridizations to coding sequences for unique genes from those to satellite DNA were developed. Results of this study are discussed in terms of strategy for the systematic development of large numbers of bovine genomic polymorphisms.

First genetic linkage map of Taraxacum koksaghyz Rodin based on AFLP, SSR, COS and EST-SSR markers.

PubMed

Arias, Marina; Hernandez, Monica; Remondegui, Naroa; Huvenaars, Koen; van Dijk, Peter; Ritter, Enrique

2016-08-04

Taraxacum koksaghyz Rodin (TKS) has been studied in many occasions as a possible alternative source for natural rubber production of good quality and for inulin production. Some tire companies are already testing TKS tire prototypes. There are also many investigations on the production of bio-fuels from inulin and inulin applications for health improvement and in the food industry. A limited amount of genomic resources exist for TKS and particularly no genetic linkage map is available in this species. We have constructed the first TKS genetic linkage map based on AFLP, COS, SSR and EST-SSR markers. The integrated linkage map with eight linkage groups (LG), representing the eight chromosomes of Russian dandelion, has 185 individual AFLP markers from parent 1, 188 individual AFLP markers from parent 2, 75 common AFLP markers and 6 COS, 1 SSR and 63 EST-SSR loci. Blasting the EST-SSR sequences against known sequences from lettuce allowed a partial alignment of our TKS map with a lettuce map. Blast searches against plant gene databases revealed some homologies with useful genes for downstream applications in the future.

High Genetic and Epigenetic Stability in Coffea arabica Plants Derived from Embryogenic Suspensions and Secondary Embryogenesis as Revealed by AFLP, MSAP and the Phenotypic Variation Rate

PubMed Central

Bobadilla Landey, Roberto; Cenci, Alberto; Georget, Frédéric; Bertrand, Benoît; Camayo, Gloria; Dechamp, Eveline; Herrera, Juan Carlos; Santoni, Sylvain; Lashermes, Philippe; Simpson, June; Etienne, Hervé

2013-01-01

Embryogenic suspensions that involve extensive cell division are risky in respect to genome and epigenome instability. Elevated frequencies of somaclonal variation in embryogenic suspension-derived plants were reported in many species, including coffee. This problem could be overcome by using culture conditions that allow moderate cell proliferation. In view of true-to-type large-scale propagation of C. arabica hybrids, suspension protocols based on low 2,4-D concentrations and short proliferation periods were developed. As mechanisms leading to somaclonal variation are often complex, the phenotypic, genetic and epigenetic changes were jointly assessed so as to accurately evaluate the conformity of suspension-derived plants. The effects of embryogenic suspensions and secondary embryogenesis, used as proliferation systems, on the genetic conformity of somatic embryogenesis-derived plants (emblings) were assessed in two hybrids. When applied over a 6 month period, both systems ensured very low somaclonal variation rates, as observed through massive phenotypic observations in field plots (0.74% from 200 000 plant). Molecular AFLP and MSAP analyses performed on 145 three year-old emblings showed that polymorphism between mother plants and emblings was extremely low, i.e. ranges of 0–0.003% and 0.07–0.18% respectively, with no significant difference between the proliferation systems for the two hybrids. No embling was found to cumulate more than three methylation polymorphisms. No relation was established between the variant phenotype (27 variants studied) and a particular MSAP pattern. Chromosome counting showed that 7 of the 11 variant emblings analyzed were characterized by the loss of 1–3 chromosomes. This work showed that both embryogenic suspensions and secondary embryogenesis are reliable for true-to-type propagation of elite material. Molecular analyses revealed that genetic and epigenetic alterations are particularly limited during coffee somatic embryogenesis. The main change in most of the rare phenotypic variants was aneuploidy, indicating that mitotic aberrations play a major role in somaclonal variation in coffee. PMID:23418563

Identification of single nucleotide polymorphisms in the ASB15 gene and their associations with chicken growth and carcass traits.

PubMed

Wang, Y C; Jiang, R R; Kang, X T; Li, Z J; Han, R L; Geng, J; Fu, J X; Wang, J F; Wu, J P

2015-09-25

ASB15 is a member of the ankyrin repeat and suppressor of cytokine signaling box family, and is predominantly expressed in skeletal muscle. In the present study, an F2 resource population of Gushi chickens crossed with Anka broilers was used to investigate the genetic effects of the chicken ASB15 gene. Two single nucleotide polymorphisms (SNPs) (rs315759231 A>G and rs312619270 T>C) were identified in exon 7 of the ASB15 gene using forced chain reaction-restriction fragment length polymorphism and DNA sequencing. One was a missense SNP (rs315759231 A>G) and the other was a synonymous SNP (rs312619270 T>C). The rs315759231 A>G polymorphism was significantly associated with body weight at birth, 12-week body slanting length, semi-evisceration weight, evisceration weight, leg muscle weight, and carcass weight (P < 0.05). The rs312619270 T>C polymorphism was significantly associated with body weight at birth, 4, 8, and 12-week body weight, 8-week shank length, 12-week breast bone length, 8 and 12-week body slanting length, breast muscle weight, and carcass weight (P < 0.05). Our results suggest that the ASB15 gene profoundly affects chicken growth and carcass traits.

A second generation genetic linkage map of Japanese flounder (Paralichthys olivaceus)

PubMed Central

2010-01-01

Background Japanese flounder (Paralichthys olivaceus) is one of the most economically important marine species in Northeast Asia. Information on genetic markers associated with quantitative trait loci (QTL) can be used in breeding programs to identify and select individuals carrying desired traits. Commercial production of Japanese flounder could be increased by developing disease-resistant fish and improving commercially important traits. Previous maps have been constructed with AFLP markers and a limited number of microsatellite markers. In this study, improved genetic linkage maps are presented. In contrast with previous studies, these maps were built mainly with a large number of codominant markers so they can potentially be used to analyze different families and populations. Results Sex-specific genetic linkage maps were constructed for the Japanese flounder including a total of 1,375 markers [1,268 microsatellites, 105 single nucleotide polymorphisms (SNPs) and two genes]; 1,167 markers are linked to the male map and 1,067 markers are linked to the female map. The lengths of the male and female maps are 1,147.7 cM and 833.8 cM, respectively. Based on estimations of map lengths, the female and male maps covered 79 and 82% of the genome, respectively. Recombination ratio in the new maps revealed F:M of 1:0.7. All linkage groups in the maps presented large differences in the location of sex-specific recombination hot-spots. Conclusions The improved genetic linkage maps are very useful for QTL analyses and marker-assisted selection (MAS) breeding programs for economically important traits in Japanese flounder. In addition, SNP flanking sequences were blasted against Tetraodon nigroviridis (puffer fish) and Danio rerio (zebrafish), and synteny analysis has been carried out. The ability to detect synteny among species or genera based on homology analysis of SNP flanking sequences may provide opportunities to complement initial QTL experiments with candidate gene approaches from homologous chromosomal locations identified in related model organisms. PMID:20937088

Use of AFLP, plasmid typing and phenotyping in a comparative study to assess genetic diversity of Shigella flexneri strains.

PubMed Central

Herrera, S.; Cabrera, R.; Ramirez, M. M.; Usera, M. A.; Echeita, M. A.

2002-01-01

Shigella flexneri infections are one of the main causes of acute diarrhoea in Cuba. Twenty strains isolated from sporadic cases in nine different Cuban provinces were characterized. Serotyping, antibiotic-resistance typing, plasmid-typing and AFLP-typing were used to determine their suitability for use in epidemiological studies of S. flexneri. The predominant serotypes were serotype 6 (35%) and serotype 2 (35%). Eleven different plasmid profiles were detected (Diversity Index = 0.92). AFLP-typing discriminated 12 different patterns (DI = 0.95), these patterns were not coincident with plasmid-typing patterns. Both techniques combined distinguished 14 patterns among the 20 studied strains (DI = 0.99). There was no consistent relationship between plasmid-typing and AFLP-typing patterns or antibiotic-resistance typing patterns. Ninety-five percent of S. flexneri strains were multiresistant. PMID:12558326

First genetic linkage map of Taraxacum koksaghyz Rodin based on AFLP, SSR, COS and EST-SSR markers

PubMed Central

Arias, Marina; Hernandez, Monica; Remondegui, Naroa; Huvenaars, Koen; van Dijk, Peter; Ritter, Enrique

2016-01-01

Taraxacum koksaghyz Rodin (TKS) has been studied in many occasions as a possible alternative source for natural rubber production of good quality and for inulin production. Some tire companies are already testing TKS tire prototypes. There are also many investigations on the production of bio-fuels from inulin and inulin applications for health improvement and in the food industry. A limited amount of genomic resources exist for TKS and particularly no genetic linkage map is available in this species. We have constructed the first TKS genetic linkage map based on AFLP, COS, SSR and EST-SSR markers. The integrated linkage map with eight linkage groups (LG), representing the eight chromosomes of Russian dandelion, has 185 individual AFLP markers from parent 1, 188 individual AFLP markers from parent 2, 75 common AFLP markers and 6 COS, 1 SSR and 63 EST-SSR loci. Blasting the EST-SSR sequences against known sequences from lettuce allowed a partial alignment of our TKS map with a lettuce map. Blast searches against plant gene databases revealed some homologies with useful genes for downstream applications in the future. PMID:27488242

The effect of a promoter polymorphism on the transcription of nitric oxide synthase 1 and its relevance to Parkinson's disease.

PubMed

Rife, Terrie; Rasoul, Bareza; Pullen, Nicholas; Mitchell, David; Grathwol, Kristen; Kurth, Janice

2009-08-01

Transcriptional changes of the enzyme nitric oxide synthase I (NOS1) are believed to play a role in the development of many diseases. The gene for NOS1 has 12 alternative first exons (1A-1L). The 1F exon is one of the most highly utilized first exons in the brain and has a polymorphism ((TG)(m)TA(TG)(n)) located in its promoter region. The polymorphism's length has been suggested to affect NOS1 transcription and play a role in Parkinson's disease (PD); however, the actual influence of the polymorphism on NOS1 transcription has not been studied. To better characterize the links of the polymorphism with PD, a genotyping study was done comparing polymorphism length among 170 PD patients and 150 age-matched controls. The pattern of changes between the two group's allele frequencies shows statistical significance (P = 0.0359). The smallest polymorphism sizes are more predominant among PD patients than controls. To study the effects of this polymorphism on NOS1 gene transcription, reporter gene constructs were made by cloning the NOS1 1F promoter with polymorphism lengths of either 42, 54, or 62 bp in front of the luciferase gene and transfecting them into HeLa or Sk-N-MC cells. NOS1-directed reporter gene constructs with the 62-bp polymorphism increased transcription of luciferase 2.2-fold in HeLa and 1.8-fold in Sk-N-MC cells compared with reporter gene constructs with the 42-bp polymorphism. These data suggest that if smaller polymorphism size contributes to the higher NOS1 levels in PD patients, an as yet unknown transcriptional mechanism is required. Copyright 2009 Wiley-Liss, Inc.

Transgenerational epigenetics: Inheritance of global cytosine methylation and methylation-related epigenetic markers in the shrub Lavandula latifolia.

PubMed

Herrera, Carlos M; Alonso, Conchita; Medrano, Mónica; Pérez, Ricardo; Bazaga, Pilar

2018-04-01

The ecological and evolutionary significance of natural epigenetic variation (i.e., not based on DNA sequence variants) variation will depend critically on whether epigenetic states are transmitted from parents to offspring, but little is known on epigenetic inheritance in nonmodel plants. We present a quantitative analysis of transgenerational transmission of global DNA cytosine methylation (= proportion of all genomic cytosines that are methylated) and individual epigenetic markers (= methylation status of anonymous MSAP markers) in the shrub Lavandula latifolia. Methods based on parent-offspring correlations and parental variance component estimation were applied to epigenetic features of field-growing plants ('maternal parents') and greenhouse-grown progenies. Transmission of genetic markers (AFLP) was also assessed for reference. Maternal parents differed significantly in global DNA cytosine methylation (range = 21.7-36.7%). Greenhouse-grown maternal families differed significantly in global methylation, and their differences were significantly related to maternal origin. Methylation-sensitive amplified polymorphism (MSAP) markers exhibited significant transgenerational transmission, as denoted by significant maternal variance component of marker scores in greenhouse families and significant mother-offspring correlations of marker scores. Although transmission-related measurements for global methylation and MSAP markers were quantitatively lower than those for AFLP markers taken as reference, this study has revealed extensive transgenerational transmission of genome-wide global cytosine methylation and anonymous epigenetic markers in L. latifolia. Similarity of results for global cytosine methylation and epigenetic markers lends robustness to this conclusion, and stresses the value of considering both types of information in epigenetic studies of nonmodel plants. © 2018 Botanical Society of America.

Genetic mapping of clubroot resistance genes in oilseed rape.

PubMed

Werner, S; Diederichsen, E; Frauen, M; Schondelmaier, J; Jung, C

2008-02-01

Clubroot caused by the obligate biotrophic protist Plasmodiophora brassicae is a major disease of Brassica species. Clubroot resistances introduced from B. oleracea var. 'Böhmerwaldkohl' and resistance from B. rapa ECD-04 were genetically mapped in oilseed rape (B. napus L.). A doubled haploid (DH) population of rape seed was developed by crossing a resistant DH-line derived from a resynthesized B. napus with the susceptible cultivar 'Express'. The DH population was tested in the greenhouse against seven P. brassicae isolates showing low and high virulence toward B. oleracea or/and B. rapa. DH-lines with highest or lowest disease scores were used in a bulked segregant analysis (BSA), and 43 polymorphic AFLPs were identified. A genetic map of the whole genome was constructed using 338 AFLP and 156 anchored SSR markers. Nineteen QTL were detected on chromosomes N02, N03, N08, N13, N15, N16 and N19 giving resistance to seven different isolates. Race-specific effects were observed for all QTL, none of the QTL conferred resistance to all isolates. The phenotypic variance explained by the respective QTL ranged between 10.3 and 67.5%. All QTL could be assigned to both ancestral genomes of B. napus. In contrast to previous reports, a clear differentiation into major QTL from B. rapa and minor QTL from B. oleracea could not be found. Composite interval mapping confirmed the linkage relationships determined by BSA, thus demonstrating that markers for oligogenic traits can be selected by merely testing the distributional extremes of a segregating population.

Strong spatial genetic structure in five tropical Piper species: should the Baker–Fedorov hypothesis be revived for tropical shrubs?

PubMed Central

Lasso, E; Dalling, J W; Bermingham, E

2011-01-01

Fifty years ago, Baker and Fedorov proposed that the high species diversity of tropical forests could arise from the combined effects of inbreeding and genetic drift leading to population differentiation and eventually to sympatric speciation. Decades of research, however have failed to support the Baker–Fedorov hypothesis (BFH), and it has now been discarded in favor of a paradigm where most trees are self-incompatible or strongly outcrossing, and where long-distance pollen dispersal prevents population drift. Here, we propose that several hyper-diverse genera of tropical herbs and shrubs, including Piper (>1,000 species), may provide an exception. Species in this genus often have aggregated, high-density populations with self-compatible breeding systems; characteristics which the BFH would predict lead to high local genetic differentiation. We test this prediction for five Piper species on Barro Colorado Island, Panama, using Amplified Fragment Length Polymorphism (AFLP) markers. All species showed strong genetic structure at both fine- and large-spatial scales. Over short distances (200–750 m) populations showed significant genetic differentiation (Fst 0.11–0.46, P < 0.05), with values of spatial genetic structure that exceed those reported for other tropical tree species (Sp = 0.03–0.136). This genetic structure probably results from the combined effects of limited seed and pollen dispersal, clonal spread, and selfing. These processes are likely to have facilitated the diversification of populations in response to local natural selection or genetic drift and may explain the remarkable diversity of this rich genus. PMID:22393518

Aggressiveness Changes in Populations of Didymella pinodes over Winter and Spring Pea Cropping Seasons

PubMed Central

Laloi, G.; Montarry, J.; Guibert, M.; Andrivon, D.; Michot, D.

2016-01-01

ABSTRACT Ascochyta blight, caused by the necrotrophic ascomycete Didymella pinodes, is responsible for severe losses in winter and spring pea crops. Despite different climatic conditions, epidemics on winter and spring crops are due to a single population of D. pinodes, suggesting gene flow either between the two crops or from reservoir sources during the cropping season. This should lead to similar pathogenicity characteristics in isolates sampled from the two crops. However, these hypotheses have never been formally tested. We therefore sampled a total of 520 D. pinodes strains throughout a growing season from winter and spring pea plots (WP and SP, respectively) and from winter and spring trap plants (TWP and TSP). Amplified fragment length polymorphism (AFLP) markers revealed high genetic diversity within subpopulations, whereas pathogenicity tests showed that mean aggressiveness increases over the course of an epidemic. These results support the idea that alloinoculum contributes to the carryover of epidemics between winter and spring crops and that the most aggressive isolates are selected as an epidemic progresses. IMPORTANCE Ascochyta blight, caused by Didymella pinodes, is responsible for severe losses in pea crops. While previous studies have shown that ascochyta blight epidemics on winter and spring crops are due to a single population of D. pinodes, suggesting that isolates from the two crops present similar pathogenicity characteristics, that hypothesis have never been tested. Genetic analysis of subpopulations sampled throughout a growing season from winter and spring pea plots revealed high genetic diversity within subpopulations, whereas pathogenicity tests showed that mean aggressiveness increases over the course of an epidemic. PMID:27208102

Short communication: Subtyping of Staphylococcus haemolyticus isolates from milk and corresponding teat apices to verify the potential teat-skin origin of intramammary infections in dairy cows.

PubMed

Leroy, Frédéric; Van Coillie, Els; Braem, Gorik; Piessens, Veerle; Verbist, Bert; De Vuyst, Luc; De Vliegher, Sarne

2015-11-01

Coagulase-negative staphylococci (CNS) are a major cause of intramammary infections (IMI) in dairy cows and they colonize the teat skin. Staphylococcus haemolyticus, one of the more common CNS, has been identified as a highly versatile opportunistic species. The aim of the present study was to gain better insight into the adaptation of S. haemolyticus subtypes to the udder ecosystem with respect to IMI development. During a longitudinal observational study conducted over 13 mo on 6 Flemish dairy herds, S. haemolyticus isolates were recovered from milk and teat apices. A total of 44 S. haemolyticus isolates originating from milk (24 isolates) and teat apices (20 isolates) of 6 selected udder quarters were singled out and analyzed using a combined methodology of (GTG)5-PCR and amplified fragment length polymorphism (AFLP) fingerprinting to determine intraspecies differences. Combining both fingerprinting methods, 4 S. haemolyticus subtypes were obtained (I to IV). Subtypes I, II, and IV were recovered from both milk and teat apex samples and were found to be associated with persisting IMI. Subtype III, not apparently related to IMI, was isolated solely from teat apices and not from milk. In general, S. haemolyticus subtypes found in milk from infected quarters could be recovered from the corresponding teat apices, although the latter could be colonized with up to 3 different subtypes. Comparing subtypes from milk and teat apices indicates that the IMI-causing agent likely originates from the teat skin. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.

Genetic variation among interconnected populations of Catostomus occidentalis: Implications for distinguishing impacts of contaminants from biogeographical structuring

USGS Publications Warehouse

Whitehead, A.; Anderson, S.L.; Kuivila, K.M.; Roach, J.L.; May, B.

2003-01-01

Exposure to contaminants can affect survivorship, recruitment, reproductive success, mutation rates and migration, and may play a significant role in the partitioning of genetic variation among exposed and nonexposed populations. However, the application of molecular population genetic data to evaluate such influences has been uncommon and often flawed. We tested whether patterns of genetic variation among native fish populations (Sacramento sucker, Catostomus occidentalis) in the Central Valley of California were consistent with long-term pesticide exposure history, or primarily with expectations based on biogeography. Field sampling was designed to rigorously test for both geographical and contamination influences. Fine-scale structure of these interconnected populations was detected with both amplified fragment length polymorphisms (AFLP) and microsatellite markers, and patterns of variation elucidated by the two marker systems were highly concordant. Analyses indicated that biogeographical hypotheses described the data set better than hypotheses relating to common historical pesticide exposure. Downstream populations had higher genetic diversity than upstream populations, regardless of exposure history, and genetic distances showed that populations from the same river system tended to cluster together. Relatedness among populations reflected primarily directions of gene flow, rather than convergence among contaminant-exposed populations. Watershed geography accounted for significant partitioning of genetic variation among populations, whereas contaminant exposure history did not. Genetic patterns indicating contaminant-induced selection, increased mutation rates or recent bottlenecks were weak or absent. We stress the importance of testing contaminant-induced genetic change hypotheses within a biogeographical context. Strategic application of molecular markers for analysis of fine-scale structure, and for evaluating contaminant impacts on gene pools, is discussed.

Adaptive Genetic Divergence along Narrow Environmental Gradients in Four Stream Insects

PubMed Central

Watanabe, Kozo; Kazama, So; Omura, Tatsuo; Monaghan, Michael T.

2014-01-01

A central question linking ecology with evolutionary biology is how environmental heterogeneity can drive adaptive genetic divergence among populations. We examined adaptive divergence of four stream insects from six adjacent catchments in Japan by combining field measures of habitat and resource components with genome scans of non-neutral Amplified Fragment Length Polymorphism (AFLP) loci. Neutral genetic variation was used to measure gene flow and non-neutral genetic variation was used to test for adaptive divergence. We identified the environmental characteristics contributing to divergence by comparing genetic distances at non-neutral loci between sites with Euclidean distances for each of 15 environmental variables. Comparisons were made using partial Mantel tests to control for geographic distance. In all four species, we found strong evidence for non-neutral divergence along environmental gradients at between 6 and 21 loci per species. The relative contribution of these environmental variables to each species' ecological niche was quantified as the specialization index, S, based on ecological data. In each species, the variable most significantly correlated with genetic distance at non-neutral loci was the same variable along which each species was most narrowly distributed (i.e., highest S). These were gradients of elevation (two species), chlorophyll-a, and ammonia-nitrogen. This adaptive divergence occurred in the face of ongoing gene flow (F st = 0.01–0.04), indicating that selection was strong enough to overcome homogenization at the landscape scale. Our results suggest that adaptive divergence is pronounced, occurs along different environmental gradients for different species, and may consistently occur along the narrowest components of species' niche. PMID:24681871

Directional genetic selection by pulp mill effluent on multiple natural populations of three-spined stickleback (Gasterosteus aculeatus).

PubMed

Lind, Emma E; Grahn, Mats

2011-05-01

Contamination can cause a rapid environmental change which may require populations to respond with evolutionary changes. To evaluate the effects of pulp mill effluents on population genetics, we sampled three-spined sticklebacks (Gasterosteus aculeatus) near four pulp mills and four adjacent reference sites and analyzed Amplified Fragment Length Polymorphism (AFLP) to compare genetic variability. A fine scale genetic structure was detected and samples from polluted sites separated from reference sites in multidimensional scaling plots (P<0.005, 1000 permutations) and locus-by-locus Analysis of Molecular Variance (AMOVA) further confirmed that habitats are significantly separated (F(ST)=0.021, P<0.01, 1023 permutations). The amount of genetic variation between populations did not differ between habitats, and populations from both habitats had similar levels of heterozygosity (polluted sites Nei's Hs=0.11, reference sites Nei's Hs=0.11). Still, pairwise F(ST): s between three, out of four, pairs of polluted-reference sites were significant. A F(ST)-outlier analysis showed that 21 (8.4%) loci were statistically different from a neutral distribution at the P<0.05 level and therefore indicated to be under divergent selection. When removing 13 F(ST)-outlier loci, significant at the P<0.01 level, differentiation between habitats disappeared in a multidimensional scaling plot. In conclusion, pulp mill effluence has acted as a selective agent on natural populations of G. aculeatus, causing a convergence in genotype composition change at multiple sites in an open environment. © The Author(s) 2011. This article is published with open access at Springerlink.com

Geographic population structure in an outcrossing plant invasion after centuries of cultivation and recent founding events.

PubMed

Gaskin, John F; Schwarzländer, Mark; Gibson, Robert D; Simpson, Heather; Marshall, Diane L; Gerber, Esther; Hinz, Hariet

2018-04-01

Population structure and genetic diversity of invasions are the result of evolutionary processes such as natural selection, drift and founding events. Some invasions are also molded by specific human activities such as selection for cultivars and intentional introduction of desired phenotypes, which can lead to low genetic diversity in the resulting invasion. We investigated the population structure, diversity and origins of a species with both accidental and intentional introduction histories, as well as long-term selection as a cultivar. Dyer's woad ( Isatis tinctoria ; Brassicaceae) has been used as a dye source for at least eight centuries in Eurasia, was introduced to eastern USA in the 1600s, and is now considered invasive in the western USA. Our analyses of amplified fragment length polymorphisms (AFLPs) from 645 plants from the USA and Eurasia did not find significantly lower gene diversity ( H j ) in the invaded compared to the native range. This suggests that even though the species was under cultivation for many centuries, human selection of plants may not have had a strong influence on diversity in the invasion. We did find significantly lower genetic differentiation ( F st ) in the invasive range but our results still suggested that there are two distinct invasions in the western USA. Our data suggest that these invasions most likely originated from Switzerland, Ukraine and Germany, which correlates with initial biological control agent survey findings. Genetic information on population structure, diversity and origins assists in efforts to control invasive species, and continued combination of ecological and molecular analyses will help bring us closer to sustainable management of plant invasions.

Genomic scan as a tool for assessing the genetic component of phenotypic variance in wild populations.

PubMed

Herrera, Carlos M

2012-01-01

Methods for estimating quantitative trait heritability in wild populations have been developed in recent years which take advantage of the increased availability of genetic markers to reconstruct pedigrees or estimate relatedness between individuals, but their application to real-world data is not exempt from difficulties. This chapter describes a recent marker-based technique which, by adopting a genomic scan approach and focusing on the relationship between phenotypes and genotypes at the individual level, avoids the problems inherent to marker-based estimators of relatedness. This method allows the quantification of the genetic component of phenotypic variance ("degree of genetic determination" or "heritability in the broad sense") in wild populations and is applicable whenever phenotypic trait values and multilocus data for a large number of genetic markers (e.g., amplified fragment length polymorphisms, AFLPs) are simultaneously available for a sample of individuals from the same population. The method proceeds by first identifying those markers whose variation across individuals is significantly correlated with individual phenotypic differences ("adaptive loci"). The proportion of phenotypic variance in the sample that is statistically accounted for by individual differences in adaptive loci is then estimated by fitting a linear model to the data, with trait value as the dependent variable and scores of adaptive loci as independent ones. The method can be easily extended to accommodate quantitative or qualitative information on biologically relevant features of the environment experienced by each sampled individual, in which case estimates of the environmental and genotype × environment components of phenotypic variance can also be obtained.

Boxer crabs induce asexual reproduction of their associated sea anemones by splitting and intraspecific theft

PubMed Central

Karplus, Ilan; Achituv, Yair

2017-01-01

Crabs of the genus Lybia have the remarkable habit of holding a sea anemone in each of their claws. This partnership appears to be obligate, at least on the part of the crab. The present study focuses on Lybia leptochelis from the Red Sea holding anemones of the genus Alicia (family Aliciidae). These anemones have not been found free living, only in association with L. leptochelis. In an attempt to understand how the crabs acquire them, we conducted a series of behavioral experiments and molecular analyses. Laboratory observations showed that the removal of one anemone from a crab induces a “splitting” behavior, whereby the crab tears the remaining anemone into two similar parts, resulting in a complete anemone in each claw after regeneration. Furthermore, when two crabs, one holding anemones and one lacking them, are confronted, the crabs fight, almost always leading to the “theft” of a complete anemone or anemone fragment by the crab without them. Following this, crabs “split” their lone anemone into two. Individuals of Alicia sp. removed from freshly collected L. leptochelis were used for DNA analysis. By employing AFLP (Fluorescence Amplified Fragments Length Polymorphism) it was shown that each pair of anemones from a given crab is genetically identical. Furthermore, there is genetic identity between most pairs of anemone held by different crabs, with the others showing slight genetic differences. This is a unique case in which one animal induces asexual reproduction of another, consequently also affecting its genetic diversity. PMID:28168117

Testing the efficiency of nested barriers to dispersal in the Mediterranean high mountain plant Edraianthus graminifolius (Campanulaceae).

PubMed

Surina, Boštjan; Schneeweiss, Gerald M; Glasnović, Peter; Schönswetter, Peter

2014-06-01

Due to strong spatial heterogeneity and limited Pleistocene glaciation, the Balkan Peninsula is a major European biodiversity hot spot. Surprisingly little, however, is known about patterns and processes of intraspecific diversification of its biota in general and of high-altitude species in particular. A well-suited system to test hypotheses with respect to various isolating factors acting at different geographic scales and to explore full-range phylogeographic patterns on the Balkan Peninsula is Edraianthus graminifolius (Campanulaceae), distributed in the western Balkan mountain systems, the southwestern Carpathians and the Apennine Peninsula. To this end, we used a dense population sampling and employed amplified fragment length polymorphism (AFLP) markers and plastid DNA sequences supplemented by ecological niche modelling. The strongest splits were inferred to separate southern and northern Balkan populations from the central ones, from where range extension occurred to the Carpathians and, in more recent times, once or twice to the Apennine Peninsula. The three genetic groups in the western Balkan Peninsula were remarkably congruent among molecular markers, suggesting that the barriers to gene flow acted over long time periods facilitating allopatric differentiation. Each main group of Balkan populations contained genetically and geographically distinct subgroups, which likely are the result of local refugia during warmer periods. Evidently, the topographically highly complex and during the Last Glacial Maximum only locally glaciated Balkan Peninsula is a hot spot of species richness and endemism as well as a sanctuary of intraspecific genetic diversity, even if the underlying causes remain insufficiently understood. © 2014 John Wiley & Sons Ltd.

Mitochondrial respiratory pathways modulate nitrate sensing and nitrogen-dependent regulation of plant architecture in Nicotiana sylvestris

PubMed Central

Pellny, Till K; Van Aken, Olivier; Dutilleul, Christelle; Wolff, Tonja; Groten, Karin; Bor, Melike; De Paepe, Rosine; Reyss, Agnès; Van Breusegem, Frank; Noctor, Graham; Foyer, Christine H

2008-01-01

Mitochondrial electron transport pathways exert effects on carbon–nitrogen (C/N) relationships. To examine whether mitochondria–N interactions also influence plant growth and development, we explored the responses of roots and shoots to external N supply in wild-type (WT) Nicotiana sylvestris and the cytoplasmic male sterile II (CMSII) mutant, which has a N-rich phenotype. Root architecture in N. sylvestris seedlings showed classic responses to nitrate and sucrose availability. In contrast, CMSII showed an altered ‘nitrate-sensing’ phenotype with decreased sensitivity to C and N metabolites. The WT growth phenotype was restored in CMSII seedling roots by high nitrate plus sugars and in shoots by gibberellic acid (GA). Genome-wide cDNA-amplified fragment length polymorphism (AFLP) analysis of leaves from mature plants revealed that only a small subset of transcripts was altered in CMSII. Tissue abscisic acid content was similar in CMSII and WT roots and shoots, and growth responses to zeatin were comparable. However, the abundance of key transcripts associated with GA synthesis was modified both by the availability of N and by the CMSII mutation. The CMSII mutant maintained a much higher shoot/root ratio at low N than WT, whereas no difference was observed at high N. Shoot/root ratios were strikingly correlated with root amines/nitrate ratios, values of <1 being characteristic of high N status. We propose a model in which the amine/nitrate ratio interacts with GA signalling and respiratory pathways to regulate the partitioning of biomass between shoots and roots. PMID:18318685

Nitric oxide and gene regulation in plants.

PubMed

Grün, S; Lindermayr, C; Sell, S; Durner, J

2006-01-01

There is increasing evidence that nitric oxide (NO), which was first identified as a unique diffusible molecular messenger in animals, plays an important role in diverse physiological processes in plants. Recent progress that has deepened our understanding of NO signalling functions in plants, with special emphasis on defence signalling, is discussed here. Several studies, based on plants with altered NO-levels, have recently provided genetic evidence for the importance of NO in gene induction. For a general overview of which gene expression levels are altered by NO, two studies, involving large-scale transcriptional analyses of Arabidopsis thaliana using custom-made or commercial DNA-microarrays, were performed. Furthermore, a comprehensive transcript profiling by cDNA-amplification fragment length polymorphism (AFLP) revealed a number of Arabidopsis thaliana genes that are involved in signal transduction, disease resistance and stress response, photosynthesis, cellular transport, and basic metabolism. In addition, NO affects the expression of numerous genes in other plant species such as tobacco or soybean. The NO-dependent intracellular signalling pathway(s) that lead to the activation or suppression of these genes have not yet been defined. Several lines of evidence point to an interrelationship between NO and salicylic acid (SA) in plant defence. Recent evidence suggests that NO also plays a role in the wounding/jasmonic acid (JA) signalling pathway. NO donors affect both wounding-induced H2O2 synthesis and wounding- or JA-induced expression of defence genes. One of the major challenges ahead is to determine how the correct specific response is evoked, despite shared use of the NO signal and, in some cases, its downstream second messengers.

Pleistocene climatic oscillations rather than recent human disturbance influence genetic diversity in one of the world's highest treeline species.

PubMed

Peng, Yanling; Lachmuth, Susanne; Gallegos, Silvia C; Kessler, Michael; Ramsay, Paul M; Renison, Daniel; Suarez, Ricardo; Hensen, Isabell

2015-10-01

Biological responses to climatic change usually leave imprints on the genetic diversity and structure of plants. Information on the current genetic diversity and structure of dominant tree species has facilitated our general understanding of phylogeographical patterns. Using amplified fragment length polymorphism (AFLPs), we compared genetic diversity and structure of 384 adults of P. tarapacana with those of 384 seedlings across 32 forest sites spanning a latitudinal gradient of 600 km occurring between 4100 m and 5000 m a.s.l. in Polylepis tarapacana (Rosaceae), one of the world's highest treeline species endemic to the central Andes. Moderate to high levels of genetic diversity and low genetic differentiation were detected in both adults and seedlings, with levels of genetic diversity and differentiation being almost identical. Four slightly genetically divergent clusters were identified that accorded to differing geographical regions. Genetic diversity decreased from south to north and with increasing precipitation for adults and seedlings, but there was no relationship to elevation. Our study shows that, unlike the case for other Andean treeline species, recent human activities have not affected the genetic structure of P. tarapacana, possibly because its inhospitable habitat is unsuitable for agriculture. The current genetic pattern of P. tarapacana points to a historically more widespread distribution at lower altitudes, which allowed considerable gene flow possibly during the glacial periods of the Pleistocene epoch, and also suggests that the northern Argentinean Andes may have served as a refugium for historical populations. © 2015 Botanical Society of America.

Typing of Human Mycobacterium avium Isolates in Italy by IS1245-Based Restriction Fragment Length Polymorphism Analysis

PubMed Central

Lari, Nicoletta; Cavallini, Michela; Rindi, Laura; Iona, Elisabetta; Fattorini, Lanfranco; Garzelli, Carlo

1998-01-01

All but 2 of 63 Mycobacterium avium isolates from distinct geographic areas of Italy exhibited markedly polymorphic, multibanded IS1245 restriction fragment length polymorphism (RFLP) patterns; 2 isolates showed the low-number banding pattern typical of bird isolates. By computer analysis, 41 distinct IS1245 patterns and 10 clusters of essentially identical strains were detected; 40% of the 63 isolates showed genetic relatedness, suggesting the existence of a predominant AIDS-associated IS1245 RFLP pattern. PMID:9817900

Acetobacter sicerae sp. nov., isolated from cider and kefir, and identification of species of the genus Acetobacter by dnaK, groEL and rpoB sequence analysis.

PubMed

Li, Leilei; Wieme, Anneleen; Spitaels, Freek; Balzarini, Tom; Nunes, Olga C; Manaia, Célia M; Van Landschoot, Anita; De Vuyst, Luc; Cleenwerck, Ilse; Vandamme, Peter

2014-07-01

Five acetic acid bacteria isolates, awK9_3, awK9_4 ( = LMG 27543), awK9_5 ( = LMG 28092), awK9_6 and awK9_9, obtained during a study of micro-organisms present in traditionally produced kefir, were grouped on the basis of their MALDI-TOF MS profile with LMG 1530 and LMG 1531(T), two strains currently classified as members of the genus Acetobacter. Phylogenetic analysis based on nearly complete 16S rRNA gene sequences as well as on concatenated partial sequences of the housekeeping genes dnaK, groEL and rpoB indicated that these isolates were representatives of a single novel species together with LMG 1530 and LMG 1531(T) in the genus Acetobacter, with Acetobacter aceti, Acetobacter nitrogenifigens, Acetobacter oeni and Acetobacter estunensis as nearest phylogenetic neighbours. Pairwise similarity of 16S rRNA gene sequences between LMG 1531(T) and the type strains of the above-mentioned species were 99.7%, 99.1%, 98.4% and 98.2%, respectively. DNA-DNA hybridizations confirmed that status, while amplified fragment length polymorphism (AFLP) and random amplified polymorphic DNA (RAPD) data indicated that LMG 1531(T), LMG 1530, LMG 27543 and LMG 28092 represent at least two different strains of the novel species. The major fatty acid of LMG 1531(T) and LMG 27543 was C18 : 1ω7c. The major ubiquinone present was Q-9 and the DNA G+C contents of LMG 1531(T) and LMG 27543 were 58.3 and 56.7 mol%, respectively. The strains were able to grow on D-fructose and D-sorbitol as a single carbon source. They were also able to grow on yeast extract with 30% D-glucose and on standard medium with pH 3.6 or containing 1% NaCl. They had a weak ability to produce acid from d-arabinose. These features enabled their differentiation from their nearest phylogenetic neighbours. The name Acetobacter sicerae sp. nov. is proposed with LMG 1531(T) ( = NCIMB 8941(T)) as the type strain. © 2014 IUMS.

MULTIPLE ENZYME RESTRICTION FRAGMENT LENGTH POLYMORPHISM ANALYSIS FOR HIGH RESOLUTION DISTINCTION OF PSEUDOMONAS (SENSU STRICTO) 16S RRNA GENES

EPA Science Inventory

Pseudomonas specific 16S rDNA PCR amplification and multiple enzyme restriction fragment length polymorphism (MERFLP) analysis using a single digestion mixture of Alu I, Hinf I, Rsa I, and Tru 9I distinguished 150 published sequences and reference strains of authentic Pseudomonas...

Leptin and leptin receptor gene polymorphisms are correlated with production performance in the Arctic fox.

PubMed

Zhang, M; Bai, X J

2015-05-25

The polymerase chain reaction-single-strand conformation polymorphism technique was employed to measure mononucleotide diversity in the coding region of the leptin and leptin receptor genes in the Arctic fox. The relationships between specific genetic mutations and reproductive performance in Arctic foxes were determined to im-prove breeding strategies. We found that a leptin gene polymorphism was significantly associated with body weight (P < 0.01), abdominal circumference (P < 0.01), and fur length (P < 0.01). Furthermore, a polymorphism in the leptin receptor gene was associated with carcass weight and guard hair length (P < 0.01). Leptin and leptin receptor gene combinatorial genotypes were significantly associated with abdominal circumference, fur length (P < 0.01), and body weight (P < 0.05). The leptin gene is thus a key gene affecting body weight, abdominal circumference, and fur length in Arctic foxes, whereas variations in the leptin receptor mainly affect carcass weight and guard hair. The marker loci identified in this study can be used to assist in the selection of Arctic foxes for breeding to raise the production performance of this species.

avpr1a length polymorphism is not associated with either social or genetic monogamy in free-living prairie voles.

PubMed

Mabry, Karen E; Streatfeild, Craig A; Keane, Brian; Solomon, Nancy G

2011-01-01

Recent discoveries of single-gene influences on social behaviour have generated a great deal of interest in the proximate mechanisms underlying the expression of complex behaviours. Length polymorphism in a microsatellite in the regulatory region of the gene encoding the vasopressin 1a receptor (avpr1a) has been associated with both inter- and intra-specific variation in socially monogamous behaviour in voles (genus Microtus) under laboratory conditions. Here, we evaluate the relationship between avpr1a length polymorphism and social associations, genetic monogamy, and reproductive success in free-living prairie vole (M. ochrogaster) populations. We found no evidence of a relationship between avpr1a microsatellite length and any of our correlates of either social or genetic monogamy in the field. Our results, especially when taken in conjunction with those of recent experimental studies in semi-natural enclosures, suggest that avpr1a polymorphism is unlikely to have been a major influence in the evolution or maintenance of social monogamy in prairie voles under natural conditions.

avpr1a length polymorphism is not associated with either social or genetic monogamy in free-living prairie voles

PubMed Central

Mabry, Karen E.; Streatfeild, Craig A.; Keane, Brian; Solomon, Nancy G.

2010-01-01

Recent discoveries of single-gene influences on social behaviour have generated a great deal of interest in the proximate mechanisms underlying the expression of complex behaviours. Length polymorphism in a microsatellite in the regulatory region of the gene encoding the vasopressin 1a receptor (avpr1a) has been associated with both inter- and intra-specific variation in socially monogamous behaviour in voles (genus Microtus) under laboratory conditions. Here, we evaluate the relationship between avpr1a length polymorphism and social associations, genetic monogamy, and reproductive success in free-living prairie vole (M. ochrogaster) populations. We found no evidence of a relationship between avpr1a microsatellite length and any of our correlates of either social or genetic monogamy in the field. Our results, especially when taken in conjunction with those of recent experimental studies in semi-natural enclosures, suggest that avpr1a polymorphism is unlikely to have been a major influence in the evolution or maintenance of social monogamy in prairie voles under natural conditions. PMID:21442019

Increased apomixis expression concurrent with genetic and epigenetic variation in a newly synthesized Eragrostis curvula polyploid

NASA Astrophysics Data System (ADS)

Zappacosta, Diego C.; Ochogavía, Ana C.; Rodrigo, Juan M.; Romero, José R.; Meier, Mauro S.; Garbus, Ingrid; Pessino, Silvina C.; Echenique, Viviana C.

2014-04-01

Eragrostis curvula includes biotypes reproducing through obligate and facultative apomixis or, rarely, full sexuality. We previously generated a ``tetraploid-dihaploid-tetraploid'' series of plants consisting of a tetraploid apomictic plant (T), a sexual dihaploid plant (D) and a tetraploid artificial colchiploid (C). Initially, plant C was nearly 100% sexual. However, its capacity to form non-reduced embryo sacs dramatically increased over a four year period (2003-2007) to reach levels of 85-90%. Here, we confirmed high rates of apomixis in plant C, and used AFLPs and MSAPs to characterize the genetic and epigenetic variation observed in this plant in 2007 as compared to 2003. Of the polymorphic sequences, some had no coding potential whereas others were homologous to retrotransposons and/or protein-coding-like sequences. Our results suggest that in this particular plant system increased apomixis expression is concurrent with genetic and epigenetic modifications, possibly involving transposable elements.

msap: a tool for the statistical analysis of methylation-sensitive amplified polymorphism data.

PubMed

Pérez-Figueroa, A

2013-05-01

In this study msap, an R package which analyses methylation-sensitive amplified polymorphism (MSAP or MS-AFLP) data is presented. The program provides a deep analysis of epigenetic variation starting from a binary data matrix indicating the banding pattern between the isoesquizomeric endonucleases HpaII and MspI, with differential sensitivity to cytosine methylation. After comparing the restriction fragments, the program determines if each fragment is susceptible to methylation (representative of epigenetic variation) or if there is no evidence of methylation (representative of genetic variation). The package provides, in a user-friendly command line interface, a pipeline of different analyses of the variation (genetic and epigenetic) among user-defined groups of samples, as well as the classification of the methylation occurrences in those groups. Statistical testing provides support to the analyses. A comprehensive report of the analyses and several useful plots could help researchers to assess the epigenetic and genetic variation in their MSAP experiments. msap is downloadable from CRAN (http://cran.r-project.org/) and its own webpage (http://msap.r-forge.R-project.org/). The package is intended to be easy to use even for those people unfamiliar with the R command line environment. Advanced users may take advantage of the available source code to adapt msap to more complex analyses. © 2013 Blackwell Publishing Ltd.

DNA Polymorphism of Insulin-like Growth Factor-binding Protein-3 Gene and Its Association with Cashmere Traits in Cashmere Goats

PubMed Central

Liu, Haiying; Liu, Chao; Yang, Guiqin; Li, Hui; Dai, Jin; Cong, Yuyan; Li, Xuejian

2012-01-01

Insulin-like growth factor binding protein-3 (IGFBP-3) gene is important for regulation of growth and development in mammals. The present investigation was carried out to study DNA polymorphism by PCR-RFLP of IGFBP-3 gene and its effect on fibre traits of Chinese Inner Mongolian cashmere goats. The fibre traits data investigated were cashmere fibre diameter, combed cashmere weight, cashmere fibre length and guard hair length. Four hundred and forty-four animals were used to detect polymorphisms in the hircine IGFBP-3 gene. A 316-bp fragment of the IGFBP-3 gene in exon 2 was amplified and digested with HaeIII restriction enzyme. Three patterns of restriction fragments were observed in the populations. The frequency of AA, AB and BB genotypes was 0.58, 0.33 and 0.09 respectively. The allelic frequency of the A and B allele was 0.75 and 0.25 respectively. Nucleotide sequencing revealed a C>G transition in the exon 2 region of the IGFBP-3 gene resulting in R158G change which caused the polymorphism. Least squares analysis revealed a significant effect of genotypes on cashmere weight (p<0.0001), cashmere fibre length (p<0.001) and hair length (p<0.05) of the animals. The effect of genotypes on cashmere fibre diameter was not statistically significant (p>0.05). The animals of AB and BB genotypes showed higher cashmere weight, cashmere fibre length and hair length than the animals possessing AA genotype. These results suggested that polymorphisms in the hircine IGFBP-3 gene might be a potential molecular marker for cashmere weight in cashmere goats. PMID:25049511

AFLP Phylogeny of 36 Erythroxylum species- genetic relationships among Erythroxylum species inferred by AFLP analysis

USDA-ARS?s Scientific Manuscript database

The plant genus Erythroxylum is known for four cultivated taxa, Erythroxylum coca var. coca (Ecc), Erythroxylum coca var. ipadu (Eci), Erythroxylum novogranatense var. novogranatense (Enn) and Erythroxylum novogranatense var. truxillense (Ent) that are cultivated primarily for the illicit extraction...

AFLP Approach Reveals Variability in Phragmites australis: Implications for Its Die-Back and Evidence for Genotoxic Effects

PubMed Central

Coppi, Andrea; Lastrucci, Lorenzo; Cappelletti, David; Cerri, Martina; Ferranti, Francesco; Ferri, Valentina; Foggi, Bruno; Gigante, Daniela; Venanzoni, Roberto; Viciani, Daniele; Selvaggi, Roberta; Reale, Lara

2018-01-01

Phragmites australis is a subcosmopolitan species typical of wetlands being studied in Europe for its disappearance from natural stands, a phenomenon called reed die-back syndrome (RDBS). Although it is conjectured that low genetic variability contributes to RDBS, this aspect remains neglected to this day. Using a molecular fingerprinting approach and a sequence analysis of the trnT-trnL/rbcL-psaI regions of cpDNA, this study aimed to compare the genetic structure of stable vs. RDBS-affected P. australis stands from five wetlands of central Italy. Beforehand, in order to characterize the health condition of reed populations, the occurrence of the main macromorphological descriptors for RDBS was considered on 40 reed stands. Soil samples were also collected to examine the total content of heavy metals. The current study analyzed cpDNA in 19 samples and AFLP profiles in 381 samples to investigate the genetic structure of Phragmites populations. Based on the multinomial-Dirichlet model, an analysis of candidate loci under selective pressure was also performed. The relationships among AFLP data, RDBS descriptors and chemicals were evaluated with the use of Linear Mixed Models. The analysis of the cpDNA shows the occurrence of the haplotypes M (the most widespread), and K here recorded for the first time in Italy. Three new haplotypes were also described. The DNA fingerprinting analysis has produced a total of 322 loci (98% polymorphic) and shows the medium-to-high amount of genetic diversity. The significant genetic differentiation among wetlands (Fst = 0.337) suggests either low gene flow or small effective population size. Moreover, the low amount of outlier loci (only 5; l.5% of the total), seems to indicate the scarce occurrence of selective pressure upon the reed’s genome. Genetic diversity increased in relationship to the decrease in diameter and of flowering buds of the reed, two of the trends associated with the die-back. The current study rejects the hypothesis that genetic diversity massively contributed to RDBS. Moreover, significant relationships between genetic diversity and the total concentration of some heavy metals (Cr, Cu, and Zn) were highlighted, indicating possible genotoxic effects on P. australis. The current study represents a fact-finding background useful for the conservation of common reed. PMID:29632544

An AFLP estimation of the outcrossing rate of Spondias tuberosa (Anacardiaceae), an endemic species to the Brazilian semiarid region.

PubMed

Fernandes Santos, Carlos Antonio; de Souza Gama, Renata Natália Cândido

2013-06-01

The umbu tree (Spondias tuberosa) is one of the most important endemic species to the Brazilian tropical semiarid region. The umbu tree has edible fruits with a peculiar flavor that are consumed in natura or in a semi-industrialized form, such as jams, candies and juices. The majority of endemic species to Brazilian semiarid region have not been studied or sampled to form germ-plasm collections, which increases the risk of losing genetic variability of the adapted species to xerophytic conditions. The aim of this study was to estimate outcrossing rates in S. tuberosa using a multilocus mixed model in order to guide genetic resources and breeding programs of this species. DNA samples were extracted from 92 progenies of umbu trees, which were distributed among 12 families. These trees were planted by seed in 1991 in Petrolina, PE, Brazil. The experimental design was a randomized block, with a total of 42 progenies sampled in three regions. The experimental units were composed by five plants and five replications. The outcrossing rate was estimated by the multilocus model, which is available in the MLTR software, and was based on 17 polymorphic AFLP bands obtained from AAA_CTG and AAA_CTC primer combinations. The observed heterozygotes ranged from 0.147 to 0.499, with a maximum frequency estimated for the AAA_CTC 10 amplicon. The multilocus outcrossing estimation (t(m)) was 0.804 +/- 0.072, while the single-locus (t(s)) was 0.841 +/- 0.079, which suggests that S. tuberosa is predominantly an outcrossing species. The difference between t(m) and t(s) was -0.037 +/- 0.029, which indicates that biparental inbreeding was nearly absent. The mean inbreeding coefficient or fixation index (F) among maternal plants was--0.103 +/- 0.045, and the expected F was 0.108, which indicates that there was no excess of heterozygotes in the maternal population. The outcrossing estimates obtained in the present study indicate that S. tuberosa is an open-pollinated species. Biometrical models applied to this species should therefore take into account the deviation from random outcrossing to estimate genetic parameters and the constitution of broad germplasm samples to preserve the genetic variability of the species. Outcrossing rates based on AFLP and the mixed-mating model should be applied to other studies of plant species in the Brazilian semiarid region.

Infraspecific DNA methylation polymorphism in cotton (Gossypium hirsutum L.).

PubMed

Keyte, Anna L; Percifield, Ryan; Liu, Bao; Wendel, Jonathan F

2006-01-01

Cytosine methylation is important in the epigenetic regulation of gene expression and development in plants and has been implicated in silencing duplicate genes after polyploid formation in several plant groups. Relatively little information exists, however, on levels and patterns of methylation polymorphism (MP) at homologous loci within species. Here we explored the levels and patterns of methylation-polymorphism diversity at CCGG sites within allotetraploid cotton, Gossypium hirsutum, using a methylation-sensitive amplified fragment length polymorphism screen and a selected set of 20 G. hirsutum accessions for which we have information on genetic polymorphism levels and relationships. Methylation and MP exist at high levels within G. hirsutum: of 150 HpaII/MspI sites surveyed, 48 were methylated at the inner cytosine (32%) and 32 of these were polymorphic (67%). Both these values are higher than comparable measures of genetic diversity using restriction fragment length polymorphisms. The high percentage of methylation-polymorphic sites and potential relationship to gene expression underscore the potential significance of MP within and among populations. We speculate that biased correlation of methylation-polymorphic sites and genes in cotton may be a consequence of polyploidy and the attendant doubling of all genes.

Genetic diversity of sweet sorghum germplasm in Mexico using AFLP and SSR markers

USDA-ARS?s Scientific Manuscript database

The objective of this work was to evaluate the diversity and genetic relationships between lines and varieties of the sweet sorghum (Sorghum bicolor) germplasm bank of the National Institute for Forestry, Agriculture and Livestock Research, Mexico, using AFLP and SSR markers. The molecular markers ...

Relationship of European Persimmon (Diospyros kaki Thunb.) Cultivars to Asian Cultivars, Characterized Using AFLPs.

USDA-ARS?s Scientific Manuscript database

Sixty one persimmon (Diospyros kaki Thunb.) selections, including 17 Italian, 11 Spanish, 13 Japanese, six Korean, five Chinese, one Israeli, and eight of unknown origin, were evaluated for genetic differences by AFLP analysis. Relationships among cultivars were evaluated by UPGMA clustering, Neigh...

Distribution of a length polymorphism 5{prime} to exon 1 of the antithrombin III (ATIII) gene in the Chinese

DOE Office of Scientific and Technical Information (OSTI.GOV)

Low, P.S.; Liu, Y.; Saha, N.

A length polymorphism at the 5{prime} untranslated region of the ATIII gene has been described as having been detected by polymerase chain reaction (PCR) with a frequency of 0.75 for the short allele (S) in the Caucasian population. This length polymorphism of the ATIII gene has been studied in 251 Chinese healthy subjects. Genomic DNA was amplified by PCR with primers of published sequences. Fragments of the amplified DNA were separated by agarose gel electrophoresis (3% NuSieve and 1% Seakem GTG) and photographed on a UV transilluminator. The frequency of the short allele (S) was found to be significantly lowermore » (0.37) than that in the Caucasians (0.75). The distribution of genotypes of this polymorphism of the ATIII gene was at Hardy-Weinberg equilibrium. The large difference of allelic frequencies in the Mongoloid and Caucasian populations makes it a useful marker for population studies.« less

Essentials of Conservation Biotechnology: A mini review

NASA Astrophysics Data System (ADS)

Merlyn Keziah, S.; Subathra Devi, C.

2017-11-01

Equilibrium of biodiversity is essential for the maintenance of the ecosystem as they are interdependent on each other. The decline in biodiversity is a global problem and an inevitable threat to the mankind. Major threats include unsustainable exploitation, habitat destruction, fragmentation, transformation, genetic pollution, invasive exotic species and degradation. This review covers the management strategies of biotechnology which include sin situ, ex situ conservation, computerized taxonomic analysis through construction of phylogenetic trees, calculating genetic distance, prioritizing the group for conservation, digital preservation of biodiversities within the coding and decoding keys, molecular approaches to asses biodiversity like polymerase chain reaction, real time, randomly amplified polymorphic DNA, restriction fragment length polymorphism, amplified fragment length polymorphism, single sequence repeats, DNA finger printing, single nucleotide polymorphism, cryopreservation and vitrification.

Bootsie: estimation of coefficient of variation of AFLP data by bootstrap analysis

USDA-ARS?s Scientific Manuscript database

Bootsie is an English-native replacement for ASG Coelho’s “DBOOT” utility for estimating coefficient of variation of a population of AFLP marker data using bootstrapping. Bootsie improves on DBOOT by supporting batch processing, time-to-completion estimation, built-in graphs, and a suite of export t...

Characterization of Haemophilus parasuis isolated from Brazilian swine through serotyping, AFLP and PFGE.

PubMed

Castilla, Karina Salvagni; de Gobbi, Débora Dirani Sena; Moreno, Luisa Zanolli; Paixão, Renata; Coutinho, Tania Alen; dos Santos, José Lúcio; Moreno, Andrea Micke

2012-06-01

Haemophilus parasuis infection in pigs is characterized by fibrinous polyserositis, arthritis and meningitis. Despite the fact that traditional diagnosis is based on herd history, clinical signs, bacterial isolation and serotyping, molecular-based methods are alternatives for species-specific tests and epidemiological studies. The aim of this study was to characterize H. parasuis field strains from different states of Brazil, employing serotyping and genotyping methods. Serotyping revealed that serovar 4 was the most prevalent (26.1%), followed by serovars 5 (17.4%), 14 (8.7%), 13 (4.4%) and 2 (4.4%), whereas 39% of the strains were considered as untypeable. AFLP with a single enzyme and PFGE were able to type all isolates tested, generating 34 and 20 different profiles, respectively, including untypeable strains. Besides the slightly higher discrimination index presented by AFLP, PFGE with Not I restriction enzyme showed a better correlation with epidemiological data, grouping strains of the same serovar, animal or farm origin. The results indicated AFLP and PFGE as valuable tools for typing H. parasuis isolates collected in Brazil. Copyright © 2011 Elsevier Ltd. All rights reserved.

Mapped DNA probes from Ioblolly pine can be used for restriction fragment length polymorphism mapping in other conifers

Treesearch

M.R. Ahuja; M.E. Devey; A.T. Groover; K.D. Jermstad; D.B Neale

1994-01-01

A high-density genetic map based on restriction fragment length polymorphisms (RFLPs) is being constructed for loblolly pine (Pinus taeda L.). Consequently, a large number of DNA probes from loblolly pine are potentially available for use in other species. We have used some of these DNA probes to detect RFLPs in 12 conifers and an angiosperm....

Phylogenetic relationships in the genus Leontopodium (Asteraceae: Gnaphalieae) based on AFLP data

PubMed Central

SAFER, STEFAN; TREMETSBERGER, KARIN; GUO, YAN-PING; KOHL, GUDRUN; SAMUEL, MARY R.; STUESSY, TOD F.; STUPPNER, HERMANN

2012-01-01

The genus Leontopodium comprises 30–41 species. The centre of diversity is the Sino-Himalayan region in south-western China, where about 15 species occur. The two species native to Europe, L. alpinum (known as the common ‘Edelweiss’) and L. nivale, are part of the cultural heritage of the people living there. Despite its importance, very little is known about the systematics of the genus. Because recent molecular studies have shown that species within this genus are closely related and difficult to distinguish with rDNA and cpDNA data, we used AFLPs to obtain a more detailed understanding of the phylogeny of the genus. Our main aims were as follows: (1) to clarify species relationships within the genus; and (2) to reveal information about the biogeography of the genus. We used AFLPs with six primer combinations to investigate 216 individuals in 38 populations of 16 different species. With AFLPs, we were able to recognize 10 different groups, all of which had strong bootstrap support. These results were also congruent with the morphology-based taxonomy of the genus. Most private and rare fragments were found in the Yunnan region (south-western China) relative to Europe and Mongolia/central China, suggesting a long-lasting in situ history of populations in the centre of diversity of the genus. Our results illustrate the utility of AFLPs to resolve phylogenetic relationships between these closely related species. PMID:23258943

AFLP markers reveal high clonal diversity and extreme longevity in four key arctic-alpine species.

PubMed

de Witte, Lucienne C; Armbruster, Georg F J; Gielly, Ludovic; Taberlet, Pierre; Stöcklin, Jürg

2012-03-01

We investigated clonal diversity, genet size structure and genet longevity in populations of four arctic-alpine plants (Carex curvula, Dryas octopetala, Salix herbacea and Vaccinium uliginosum) to evaluate their persistence under past climatic oscillations and their potential resistance to future climate change. The size and number of genets were determined by an analysis of amplified fragment length polymorphisms and a standardized sampling design in several European arctic-alpine populations, where these species are dominant in the vegetation. Genet age was estimated by dividing the size by the annual horizontal size increment from in situ growth measurements. Clonal diversity was generally high but differed among species, and the frequency distribution of genet size was strongly left-skewed. The largest C. curvula genet had an estimated minimum age of c. 4100 years and a maximum age of c. 5000 years, although 84.8% of the genets in this species were <200 years old. The oldest genets of D. octopetala, S. herbacea and V. uliginosum were found to be at least 500, 450 and 1400 years old, respectively. These results indicate that individuals in the studied populations have survived pronounced climatic oscillations, including the Little Ice Age and the postindustrial warming. The presence of genets in all size classes and the dominance of presumably young individuals suggest repeated recruitment over time, a precondition for adaptation to changing environmental conditions. Together, persistence and continuous genet turnover may ensure maximum ecosystem resilience. Thus, our results indicate that long-lived clonal plants in arctic-alpine ecosystems can persist, despite considerable climatic change. © 2011 Blackwell Publishing Ltd.

Living on the edge: conservation genetics of seven thermophilous plant species in a high Arctic archipelago

PubMed Central

Brysting, Anne Krag; Elven, Reidar; Alsos, Inger Greve

2017-01-01

Abstract Small, isolated and/or peripheral populations are expected to harbour low levels of genetic variation and may therefore have reduced adaptability to environmental change, including climate warming. In the Arctic, global warming has already caused vegetation change across the region and is acting as a significant stressor on Arctic biodiversity. Many of the rare plants in the Arctic are relicts from early Holocene warm periods, but their ability to benefit from the current warming is dependent on the viability of their populations. We therefore examined Amplified Fragment Length Polymorphism (AFLP) data from regional red listed vascular plant species in the High Arctic archipelago of Svalbard and reference populations from the main distribution area of: (1) Botrychium lunaria, (2) Carex capillaris ssp. fuscidula, (3) Comastoma tenellum, (4) Kobresia simpliciuscula ssp. subholarctica, (5) Ranunculus wilanderi, (6) Sibbaldia procumbens and (7) Tofieldia pusilla. In addition, we gathered population size data in Svalbard. The Svalbard populations had low genetic diversity and distinctiveness and few or no private markers compared to populations outside the archipelago. This is similar to observations in other rare species in Svalbard and the genetic depletion may be due to an initial founder effect and/or a genetic bottleneck caused by late Holocene cooling. There seems to be limited gene flow from other areas and the Svalbard populations should therefore be considered as demographically independent management units. Overall, these management units have small and/or few populations and are therefore prone to stochastic events which may further increase vulnerability to inbreeding depression, loss of genetic variation, and reduced evolutionary potential. Our results support theory predicting lower levels of genetic diversity in small, isolated and/or peripheral populations and may be of importance for management of other rare plant species in the Arctic. PMID:28108432

AFLP diversity and spatial structure of Calycophyllum candidissimum (Rubiaceae), a dominant tree species of Nicaragua's critically endangered seasonally dry forest.

PubMed

Dávila-Lara, A; Affenzeller, M; Tribsch, A; Díaz, V; Comes, H P

2017-10-01

The Central American seasonally dry tropical (SDT) forest biome is one of the worlds' most endangered ecosystems, yet little is known about the genetic consequences of its recent fragmentation. A prominent constituent of this biome is Calycophyllum candidissimum, an insect-pollinated and wind-dispersed canopy tree of high socio-economic importance, particularly in Nicaragua. Here, we surveyed amplified fragment length polymorphisms across 13 populations of this species in Nicaragua to elucidate the relative roles of contemporary vs historical factors in shaping its genetic variation. Genetic diversity was low in all investigated populations (mean H E =0.125), and negatively correlated with latitude. Overall population differentiation was moderate (Φ ST =0.109, P<0.001), and Bayesian analysis of population structure revealed two major latitudinal clusters (I: 'Pacific North'+'Central Highland'; II: 'Pacific South'), along with a genetic cline between I and II. Population-based cluster analyses indicated a strong pattern of 'isolation by distance' as confirmed by Mantel's test. Our results suggest that (1) the low genetic diversity of these populations reflects biogeographic/population history (colonisation from South America, Pleistocene range contractions) rather than recent human impact; whereas (2) the underlying process of their isolation by distance pattern, which is best explained by 'isolation by dispersal limitation', implies contemporary gene flow between neighbouring populations as likely facilitated by the species' efficient seed dispersal capacity. Overall, these results underscore that even tree species from highly decimated forest regions may be genetically resilient to habitat fragmentation due to species-typical dispersal characteristics, the necessity of broad-scale measures for their conservation notwithstanding.

Expanded spectrum β-lactamase producing Escherichia coli isolated from chickens with colibacillosis in Egypt.

PubMed

El-Shazly, D A; Nasef, S A; Mahmoud, F F; Jonas, Daniel

2017-07-01

Throughout the world, expanded spectrum β-lactamases (ESBL) are increasing among clinical isolates of Enterobacteriaceae, both in humans and animals. Unfortunately, there is a paucity of data on ESBL or Ampicillin class C β-lactamase (AmpC) in Egypt, although antimicrobial consumption is high in this developing country. This study aims to characterize the resistance mechanisms to expanded spectrum cephalosporins among resistant veterinary Escherichia coli isolates in Egypt. We investigated 50 clinical multi-resistant E. coli strains isolated from 20 chicken farms for production of ESBL or AmpC. Antibiotic susceptibility was tested by Clinical and Laboratory Standards Institute (CLSI) disk diffusion and ESBL confirmatory tests. PCR and sequencing were performed to screen for plasmid mediated ESBL genes and genes encoding AmpC β-lactamases. All the isolates were phylogentically classified, investigated for harboring class 1 integrons, and genotyped by amplified fragment length polymorphism (AFLP). Three strains showed ESBL and 6 strains AmpC phenotypic patterns, respectively, with confirmed ESBL genes of blaTEM-57, blaSHV-12, blaCTX-M-14, and blaCMY-2 for AmpC producing strains. All ESBL strains belonged to phylogroup D with different clones isolated from different flocks, while most of the AmpC strains belonged to phylogroup B1 (4/6) and were assigned to the same genotype distributed in 2 different farms. Class 1 integrons were disseminated in 60% of all tested strains and in 100% of ESBL and AmpC strains. These results highlight the antimicrobial resistance problem in Egypt, caused in all probability by unwise use of antimicrobials in animal husbandry. The results call for a nationwide surveillance program to monitor antimicrobial resistance. © 2017 Poultry Science Association Inc.

Genetic connectivity in scleractinian corals across the Northern Gulf of Mexico: oil/gas platforms, and relationship to the Flower Garden Banks.

PubMed

Sammarco, Paul W; Brazeau, Daniel A; Sinclair, James

2012-01-01

The 3,000 oil/gas structures currently deployed in the northern Gulf of Mexico (GOM) provide hard substratum for marine organisms in a region where such has been rare since the Holocene. The major exception to this are the Flower Garden Banks (FGB). Corals are known to have colonized oil/gas platforms around the FGB, facilitating biogeographic expansion. We ask the question, what are the patterns of genetic affinity in these coral populations. We sampled coral tissue from populations of two species occurring on oil and gas platforms: Madracis decactis (hermatype) and Tubastraea coccinea (invasive ahermatype). We sampled 28 platforms along four transects from 20 km offshore to the continental shelf edge off 1) Matagorda Island, TX; 2) Lake Sabine, TX; 3) Terrebonne Bay, LA; and 4) Mobile, AL. The entire population of M. decactis was sampled between depths of 5 m and 37 m. T. coccinea populations were sub-sampled. Genetic variation was assessed using the PCR-based Amplified Fragment Length Polymorphisms (AFLPs). Data were analyzed via AFLPOP and STRUCTURE. Genetic connectivity among M. decactis platform populations was highest near the FGB and decreased to the east. Connectivity increased again in the eastern sector, indicating isolation between the populations from different sides of the Mississippi River (Transects 3 and 4). A point-drop in genetic affinity (relatedness) at the shelf edge south of Terrebonne Bay, LA indicated a population differing from all others in the northern GOM. Genetic affinities among T. coccinea were highest in the west and decreased to the east. Very low genetic affinities off Mobile, AL indicated a dramatic difference between those populations and those west of the Mississippi River, apparently a formidable barrier to larval dispersal.

Resistance of Cultivated Tomato to Cell Content-Feeding Herbivores Is Regulated by the Octadecanoid-Signaling Pathway1

PubMed Central

Li, Chuanyou; Williams, Mark M.; Loh, Ying-Tsu; Lee, Gyu In; Howe, Gregg A.

2002-01-01

The octadecanoid signaling pathway has been shown to play an important role in plant defense against various chewing insects and some pathogenic fungi. Here, we examined the interaction of a cell-content feeding arachnid herbivore, the two-spotted spider mite (Tetranychus urticae Koch), with cultivated tomato (Lycopersicon esculentum) and an isogenic mutant line (defenseless-1 [def-1]) that is deficient in the biosynthesis of the octadecanoid pathway-derived signal, jasmonic acid (JA). Spider mite feeding and fecundity on def-1 plants was significantly greater than on wild-type plants. Decreased resistance of def-1 plants was correlated with reduced JA accumulation and expression of defensive proteinase inhibitor (PI) genes, which were induced in mite-damaged wild-type leaves. Treatment of def-1 plants with methyl-JA restored resistance to spider mite feeding and reduced the fecundity of female mites. Plants expressing a 35S::prosystemin transgene that constitutively activates the octadecanoid pathway in a Def-1-dependent manner were highly resistant to attack by spider mites and western flower thrips (Frankliniella occidentalis), another cell-content feeder of economic importance. These findings indicate that activation of the octadecanoid signaling pathway promotes resistance of tomato to a broad spectrum of herbivores. The techniques of amplified fragment length polymorphism (AFLP) and bulk segregant analysis were used to map the Def-1 gene to a region on the long arm of chromosome 3 that is genetically separable from the map position of known JA biosynthetic genes. Tight linkage of Def-1 to a T-DNA insertion harboring the maize (Zea mays) Dissociation transposable element suggests a strategy for directed transposon tagging of the gene. PMID:12226528

The environmental source of emerging Apophysomyces variabilis infection in India.

PubMed

Prakash, Hariprasath; Ghosh, Anup Kumar; Rudramurthy, Shivaprakash Mandya; Paul, Raees Ahmad; Gupta, Sunita; Negi, Vishwanand; Chakrabarti, Arunaloke

2016-08-01

The rare mucoraceous fungus, Apophysomyces species complex ranks second after Rhizopus arrhizus causing mucormycosis in India. The source of this agent in the environment is not clearly known. We conducted an environmental study to find its presence in Indian soil. The soil samples from different geographical locations were analyzed for isolation of Mucorales. Rhizopus arrhizus (24.6%) was most commonly isolated from soil, followed by Lichtheimia spp. (23.2%), Cunninghamella spp. (21.7%), Rhizopus microsporus (14%) and Apophysomyces spp. (4.5%). The isolation of Apophysomyces species complex was significantly associated with low nitrogen content of the soil. Based on sequencing of internal transcribed spacer (ITS) and 28S (D1/D2) regions of ribosomal DNA, the Apophysomyces isolates were identified as Apophysomyces variabilis with 98 to 100% similarity to type strain A. variabilis (CBS658.93). The analysis of amplified fragment length polymorphism (AFLP) fingerprinting data demonstrated genomic diversity of A. variabilis isolates with multiple clades (similarity 40-90%). The minimum inhibitory concentrations (MIC), MIC50 and MIC90 for A. variabilis isolates were 1 and 4 μg/ml for amphotericin B, 0.25 and 0.5 μg/ml for itraconazole, 0.125 and 0.25 μg/ml for posaconazole, 0.06 and 0.12 μg/ml for terbinafine, respectively. The present study revealed abundant presence of A. variabilis in Indian soil with low nitrogen content, its genetic heterogeneity and relatively high MICs for amphotericin B. © The Author 2016. Published by Oxford University Press on behalf of The International Society for Human and Animal Mycology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Genetic structure and seed-mediated dispersal rates of an endangered shrub in a fragmented landscape: a case study for Juniperus communis in northwestern Europe

PubMed Central

2011-01-01

Background Population extinction risk in a fragmented landscape is related to the differential ability of the species to spread its genes across the landscape. The impact of landscape fragmentation on plant population dynamics will therefore vary across different spatial scales. We quantified successful seed-mediated dispersal of the dioecious shrub Juniperus communis in a fragmented landscape across northwestern Europe by using amplified fragment length polymorphism (AFLP) markers. Furthermore we investigated the genetic diversity and structure on two spatial scales: across northwestern Europe and across Flanders (northern Belgium). We also studied whether seed viability and populations size were correlated with genetic diversity. Results Unexpectedly, estimated seed-mediated dispersal rates were quite high and ranged between 3% and 14%. No population differentiation and no spatial genetic structure were detected on the local, Flemish scale. A significant low to moderate genetic differentiation between populations was detected at the regional, northwest European scale (PhiPT = 0.10). In general, geographically nearby populations were also genetically related. High levels of within-population genetic diversity were detected but no correlation was found between any genetic diversity parameter and population size or seed viability. Conclusions In northwestern Europe, landscape fragmentation has lead to a weak isolation-by-distance pattern but not to genetic impoverishment of common juniper. Substantial rates of successful migration by seed-mediated gene flow indicate a high dispersal ability which could enable Juniperus communis to naturally colonize suitable habitats. However, it is not clear whether the observed levels of migration will suffice to counterbalance the effects of genetic drift in small populations on the long run. PMID:21859457

Genetic structure and seed-mediated dispersal rates of an endangered shrub in a fragmented landscape: a case study for Juniperus communis in northwestern Europe.

PubMed

Vanden-Broeck, An; Gruwez, Robert; Cox, Karen; Adriaenssens, Sandy; Michalczyk, Inga M; Verheyen, Kris

2011-08-22

Population extinction risk in a fragmented landscape is related to the differential ability of the species to spread its genes across the landscape. The impact of landscape fragmentation on plant population dynamics will therefore vary across different spatial scales. We quantified successful seed-mediated dispersal of the dioecious shrub Juniperus communis in a fragmented landscape across northwestern Europe by using amplified fragment length polymorphism (AFLP) markers. Furthermore we investigated the genetic diversity and structure on two spatial scales: across northwestern Europe and across Flanders (northern Belgium). We also studied whether seed viability and populations size were correlated with genetic diversity. Unexpectedly, estimated seed-mediated dispersal rates were quite high and ranged between 3% and 14%. No population differentiation and no spatial genetic structure were detected on the local, Flemish scale. A significant low to moderate genetic differentiation between populations was detected at the regional, northwest European scale (PhiPT = 0.10). In general, geographically nearby populations were also genetically related. High levels of within-population genetic diversity were detected but no correlation was found between any genetic diversity parameter and population size or seed viability. In northwestern Europe, landscape fragmentation has lead to a weak isolation-by-distance pattern but not to genetic impoverishment of common juniper. Substantial rates of successful migration by seed-mediated gene flow indicate a high dispersal ability which could enable Juniperus communis to naturally colonize suitable habitats. However, it is not clear whether the observed levels of migration will suffice to counterbalance the effects of genetic drift in small populations on the long run.

Diversity and clinical impact of Acinetobacter baumannii colonization and infection at a military medical center.

PubMed

Petersen, Kyle; Cannegieter, Suzanne C; van der Reijden, Tanny J; van Strijen, Beppie; You, David M; Babel, Britta S; Philip, Andrew I; Dijkshoorn, Lenie

2011-01-01

The epidemiology of Acinetobacter baumannii emerging in combat casualties is poorly understood. We analyzed 65 (54 nonreplicate) Acinetobacter isolates from 48 patients (46 hospitalized and 2 outpatient trainees entering the military) from October 2004 to October 2005 for genotypic similarities, time-space relatedness, and antibiotic susceptibility. Clinical and surveillance cultures were compared by amplified fragment length polymorphism (AFLP) genomic fingerprinting to each other and to strains of a reference database. Antibiotic susceptibility was determined, and multiplex PCR was performed for OXA-23-like, -24-like, -51-like, and -58-like carbapenemases. Records were reviewed for overlapping hospital stays of the most frequent genotypes, and risk ratios were calculated for any association of genotype with severity of Acute Physiology and Chronic Health Evaluation II (APACHE II) score or injury severity score (ISS) and previous antibiotic use. Nineteen genotypes were identified; two predominated, one consistent with an emerging novel international clone and the other unique to our database. Both predominant genotypes were carbapenem resistant, were present at another hospital before patients' admission to our facility, and were associated with higher APACHE II scores, higher ISSs, and previous carbapenem antibiotics in comparison with other genotypes. One predominated in wound and respiratory isolates, and the other predominated in wound and skin surveillance samples. Several other genotypes were identified as European clones I to III. Acinetobacter genotypes from recruits upon entry to the military, unlike those in hospitalized patients, did not include carbapenem-resistant genotypes. Acinetobacter species isolated from battlefield casualties are diverse, including genotypes belonging to European clones I to III. Two carbapenem-resistant genotypes were epidemic, one of which appeared to belong to a novel international clone.

Integrating species distribution models (SDMs) and phylogeography for two species of Alpine Primula

PubMed Central

Schorr, G; Holstein, N; Pearman, P B; Guisan, A; Kadereit, J W

2012-01-01

The major intention of the present study was to investigate whether an approach combining the use of niche-based palaeodistribution modeling and phylo-geography would support or modify hypotheses about the Quaternary distributional history derived from phylogeographic methods alone. Our study system comprised two closely related species of Alpine Primula. We used species distribution models based on the extant distribution of the species and last glacial maximum (LGM) climate models to predict the distribution of the two species during the LGM. Phylogeographic data were generated using amplified fragment length polymorphisms (AFLPs). In Primula hirsuta, models of past distribution and phylogeographic data are partly congruent and support the hypothesis of widespread nunatak survival in the Central Alps. Species distribution models (SDMs) allowed us to differentiate between alpine regions that harbor potential nunatak areas and regions that have been colonized from other areas. SDMs revealed that diversity is a good indicator for nunataks, while rarity is a good indicator for peripheral relict populations that were not source for the recolonization of the inner Alps. In P. daonensis, palaeo-distribution models and phylogeographic data are incongruent. Besides the uncertainty inherent to this type of modeling approach (e.g., relatively coarse 1-km grain size), disagreement of models and data may partly be caused by shifts of ecological niche in both species. Nevertheless, we demonstrate that the combination of palaeo-distribution modeling with phylogeographical approaches provides a more differentiated picture of the distributional history of species and partly supports (P. hirsuta) and partly modifies (P. daonensis and P. hirsuta) hypotheses of Quaternary distributional history. Some of the refugial area indicated by palaeodistribution models could not have been identified with phylogeographic data. PMID:22833799

An ecological genetic delineation of local seed-source provenance for ecological restoration

PubMed Central

Krauss, Siegfried L; Sinclair, Elizabeth A; Bussell, John D; Hobbs, Richard J

2013-01-01

An increasingly important practical application of the analysis of spatial genetic structure within plant species is to help define the extent of local provenance seed collection zones that minimize negative impacts in ecological restoration programs. Here, we derive seed sourcing guidelines from a novel range-wide assessment of spatial genetic structure of 24 populations of Banksia menziesii (Proteaceae), a widely distributed Western Australian tree of significance in local ecological restoration programs. An analysis of molecular variance (AMOVA) of 100 amplified fragment length polymorphism (AFLP) markers revealed significant genetic differentiation among populations (ΦPT = 0.18). Pairwise population genetic dissimilarity was correlated with geographic distance, but not environmental distance derived from 15 climate variables, suggesting overall neutrality of these markers with regard to these climate variables. Nevertheless, Bayesian outlier analysis identified four markers potentially under selection, although these were not correlated with the climate variables. We calculated a global R-statistic using analysis of similarities (ANOSIM) to test the statistical significance of population differentiation and to infer a threshold seed collection zone distance of ∼60 km (all markers) and 100 km (outlier markers) when genetic distance was regressed against geographic distance. Population pairs separated by >60 km were, on average, twice as likely to be significantly genetically differentiated than population pairs separated by <60 km, suggesting that habitat-matched sites within a 30-km radius around a restoration site genetically defines a local provenance seed collection zone for B. menziesii. Our approach is a novel probability-based practical solution for the delineation of a local seed collection zone to minimize negative genetic impacts in ecological restoration. PMID:23919158

Linkage map of the peppered moth, Biston betularia (Lepidoptera, Geometridae): a model of industrial melanism

PubMed Central

Van't Hof, A E; Nguyen, P; Dalíková, M; Edmonds, N; Marec, F; Saccheri, I J

2013-01-01

We have constructed a linkage map for the peppered moth (Biston betularia), the classical ecological genetics model of industrial melanism, aimed both at localizing the network of loci controlling melanism and making inferences about chromosome dynamics. The linkage map, which is based primarily on amplified fragment length polymorphisms (AFLPs) and genes, consists of 31 linkage groups (LGs; consistent with the karyotype). Comparison with the evolutionarily distant Bombyx mori suggests that the gene content of chromosomes is highly conserved. Gene order is conserved on the autosomes, but noticeably less so on the Z chromosome, as confirmed by physical mapping using bacterial artificial chromosome fluorescence in situ hybridization (BAC-FISH). Synteny mapping identified three pairs of B. betularia LGs (11/29, 23/30 and 24/31) as being orthologous to three B. mori chromosomes (11, 23 and 24, respectively). A similar finding in an outgroup moth (Plutella xylostella) indicates that the B. mori karyotype (n=28) is a phylogenetically derived state resulting from three chromosome fusions. As with other Lepidoptera, the B. betularia W chromosome consists largely of repetitive sequence, but exceptionally we found a W homolog of a Z-linked gene (laminin A), possibly resulting from ectopic recombination between the sex chromosomes. The B. betularia linkage map, featuring the network of known melanization genes, serves as a resource for melanism research in Lepidoptera. Moreover, its close resemblance to the ancestral lepidopteran karyotype (n=31) makes it a useful reference point for reconstructing chromosome dynamic events and ancestral genome architectures. Our study highlights the unusual evolutionary stability of lepidopteran autosomes; in contrast, higher rates of intrachromosomal rearrangements support a special role of the Z chromosome in adaptive evolution and speciation. PMID:23211790

Genetic structure of natural and restored shoalgrass Halodule wrightii populations in the NW Gulf of Mexico

USGS Publications Warehouse

Travis, S.E.; Sheridan, P.

2006-01-01

The decline of seagrass communities worldwide has sparked an urgent need for effective restoration strategies, which require a working knowledge of population genetic structure. Halodule wrighti is a common seagrass of the Caribbean region that is being restored to areas of the Gulf of Mexico, yet little is known of its population genetics. This study provides an assessment of individual, clonal and population effects on the genetic structure of 4 natural H. wrightii populations occupying 170 km of coastline in and around Galveston Bay, Texas, for comparison with 7 restored populations ranging in age from 2 to 7 yr. By using molecular markers, in the form of amplified fragment length polymorphisms (AFLPs), we found considerable variation in clonal richness at the population scale (from 0.54 to 0.82), with the restored populations occupying an intermediate to high position within this range. Replicate sampling within individual seagrass beds of 3 to 5m diameter generally revealed higher levels of clonal richness, elevated by 4 to 22% over that at the population scale, suggesting that seed recruitment is more important at the local scale than at distances of >10 m. Genetic diversity was 2 to 3 times less than that expected for a widespread, outcrossing species like H. wrightii, although a 170% increase in the frequency of variable markers relative to the mean for all other populations was noted for a volunteer population that had recruited from a mixture of donor materials planted at a nearby restoration site. Within the spatial extent of this study, natural populations adhered to a model of isolation-by-distance, whereas donor materials from these same natural populations were undergoing a rapid genetic convergence within a restored site where they had been planted together. ?? Inter-Research 2006.

Clostridium botulinum Group I Strain Genotyping by 15-Locus Multilocus Variable-Number Tandem-Repeat Analysis ▿ †

PubMed Central

Fillo, Silvia; Giordani, Francesco; Anniballi, Fabrizio; Gorgé, Olivier; Ramisse, Vincent; Vergnaud, Gilles; Riehm, Julia M.; Scholz, Holger C.; Splettstoesser, Wolf D.; Kieboom, Jasper; Olsen, Jaran-Strand; Fenicia, Lucia; Lista, Florigio

2011-01-01

Clostridium botulinum is a taxonomic designation that encompasses a broad variety of spore-forming, Gram-positive bacteria producing the botulinum neurotoxin (BoNT). C. botulinum is the etiologic agent of botulism, a rare but severe neuroparalytic disease. Fine-resolution genetic characterization of C. botulinum isolates of any BoNT type is relevant for both epidemiological studies and forensic microbiology. A 10-locus multiple-locus variable-number tandem-repeat analysis (MLVA) was previously applied to isolates of C. botulinum type A. The present study includes five additional loci designed to better address proteolytic B and F serotypes. We investigated 79 C. botulinum group I strains isolated from human and food samples in several European countries, including types A (28), B (36), AB (4), and F (11) strains, and 5 nontoxic Clostridium sporogenes. Additional data were deduced from in silico analysis of 10 available fully sequenced genomes. This 15-locus MLVA (MLVA-15) scheme identified 86 distinct genotypes that clustered consistently with the results of amplified fragment length polymorphism (AFLP) and MLVA genotyping in previous reports. An MLVA-7 scheme, a subset of the MLVA-15, performed on a lab-on-a-chip device using a nonfluorescent subset of primers, is also proposed as a first-line assay. The phylogenetic grouping obtained with the MLVA-7 does not differ significantly from that generated by the MLVA-15. To our knowledge, this report is the first to analyze genetic variability among all of the C. botulinum group I serotypes by MLVA. Our data provide new insights into the genetic variability of group I C. botulinum isolates worldwide and demonstrate that this group is genetically highly diverse. PMID:22012011

Clostridium botulinum group I strain genotyping by 15-locus multilocus variable-number tandem-repeat analysis.

PubMed

Fillo, Silvia; Giordani, Francesco; Anniballi, Fabrizio; Gorgé, Olivier; Ramisse, Vincent; Vergnaud, Gilles; Riehm, Julia M; Scholz, Holger C; Splettstoesser, Wolf D; Kieboom, Jasper; Olsen, Jaran-Strand; Fenicia, Lucia; Lista, Florigio

2011-12-01

Clostridium botulinum is a taxonomic designation that encompasses a broad variety of spore-forming, Gram-positive bacteria producing the botulinum neurotoxin (BoNT). C. botulinum is the etiologic agent of botulism, a rare but severe neuroparalytic disease. Fine-resolution genetic characterization of C. botulinum isolates of any BoNT type is relevant for both epidemiological studies and forensic microbiology. A 10-locus multiple-locus variable-number tandem-repeat analysis (MLVA) was previously applied to isolates of C. botulinum type A. The present study includes five additional loci designed to better address proteolytic B and F serotypes. We investigated 79 C. botulinum group I strains isolated from human and food samples in several European countries, including types A (28), B (36), AB (4), and F (11) strains, and 5 nontoxic Clostridium sporogenes. Additional data were deduced from in silico analysis of 10 available fully sequenced genomes. This 15-locus MLVA (MLVA-15) scheme identified 86 distinct genotypes that clustered consistently with the results of amplified fragment length polymorphism (AFLP) and MLVA genotyping in previous reports. An MLVA-7 scheme, a subset of the MLVA-15, performed on a lab-on-a-chip device using a nonfluorescent subset of primers, is also proposed as a first-line assay. The phylogenetic grouping obtained with the MLVA-7 does not differ significantly from that generated by the MLVA-15. To our knowledge, this report is the first to analyze genetic variability among all of the C. botulinum group I serotypes by MLVA. Our data provide new insights into the genetic variability of group I C. botulinum isolates worldwide and demonstrate that this group is genetically highly diverse.

Genetic Connectivity in Scleractinian Corals across the Northern Gulf of Mexico: Oil/Gas Platforms, and Relationship to the Flower Garden Banks

PubMed Central

Sammarco, Paul W.; Brazeau, Daniel A.; Sinclair, James

2012-01-01

The 3,000 oil/gas structures currently deployed in the northern Gulf of Mexico (GOM) provide hard substratum for marine organisms in a region where such has been rare since the Holocene. The major exception to this are the Flower Garden Banks (FGB). Corals are known to have colonized oil/gas platforms around the FGB, facilitating biogeographic expansion. We ask the question, what are the patterns of genetic affinity in these coral populations. We sampled coral tissue from populations of two species occurring on oil and gas platforms: Madracis decactis (hermatype) and Tubastraea coccinea (invasive ahermatype). We sampled 28 platforms along four transects from 20 km offshore to the continental shelf edge off 1) Matagorda Island, TX; 2) Lake Sabine, TX; 3) Terrebonne Bay, LA; and 4) Mobile, AL. The entire population of M. decactis was sampled between depths of 5 m and 37 m. T. coccinea populations were sub-sampled. Genetic variation was assessed using the PCR-based Amplified Fragment Length Polymorphisms (AFLPs). Data were analyzed via AFLPOP and STRUCTURE. Genetic connectivity among M. decactis platform populations was highest near the FGB and decreased to the east. Connectivity increased again in the eastern sector, indicating isolation between the populations from different sides of the Mississippi River (Transects 3 and 4). A point-drop in genetic affinity (relatedness) at the shelf edge south of Terrebonne Bay, LA indicated a population differing from all others in the northern GOM. Genetic affinities among T. coccinea were highest in the west and decreased to the east. Very low genetic affinities off Mobile, AL indicated a dramatic difference between those populations and those west of the Mississippi River, apparently a formidable barrier to larval dispersal. PMID:22558066

Aggressiveness Changes in Populations of Didymella pinodes over Winter and Spring Pea Cropping Seasons.

PubMed

Laloi, G; Montarry, J; Guibert, M; Andrivon, D; Michot, D; Le May, C

2016-07-15

Ascochyta blight, caused by the necrotrophic ascomycete Didymella pinodes, is responsible for severe losses in winter and spring pea crops. Despite different climatic conditions, epidemics on winter and spring crops are due to a single population of D. pinodes, suggesting gene flow either between the two crops or from reservoir sources during the cropping season. This should lead to similar pathogenicity characteristics in isolates sampled from the two crops. However, these hypotheses have never been formally tested. We therefore sampled a total of 520 D. pinodes strains throughout a growing season from winter and spring pea plots (WP and SP, respectively) and from winter and spring trap plants (TWP and TSP). Amplified fragment length polymorphism (AFLP) markers revealed high genetic diversity within subpopulations, whereas pathogenicity tests showed that mean aggressiveness increases over the course of an epidemic. These results support the idea that alloinoculum contributes to the carryover of epidemics between winter and spring crops and that the most aggressive isolates are selected as an epidemic progresses. Ascochyta blight, caused by Didymella pinodes, is responsible for severe losses in pea crops. While previous studies have shown that ascochyta blight epidemics on winter and spring crops are due to a single population of D. pinodes, suggesting that isolates from the two crops present similar pathogenicity characteristics, that hypothesis have never been tested. Genetic analysis of subpopulations sampled throughout a growing season from winter and spring pea plots revealed high genetic diversity within subpopulations, whereas pathogenicity tests showed that mean aggressiveness increases over the course of an epidemic. Copyright © 2016, American Society for Microbiology. All Rights Reserved.

A sulphur deficiency-induced gene, sdi1, involved in the utilization of stored sulphate pools under sulphur-limiting conditions has potential as a diagnostic indicator of sulphur nutritional status.

PubMed

Howarth, Jonathan R; Parmar, Saroj; Barraclough, Peter B; Hawkesford, Malcolm J

2009-02-01

A sulphate deficiency-induced gene, sdi1, has been identified by cDNA-amplified fragment length polymorphism (AFLP) analysis utilizing field-grown, nutrient-deficient wheat (Triticum aestivum var. Hereward). The expression of sdi1 was specifically induced in leaf and root tissues in response to sulphate deficiency, but was not induced by nitrogen, phosphorus, potassium or magnesium deficiency. Expression was also shown to increase in plant tissues as the external sulphate concentration in hydroponically grown plants was reduced from 1.0 to 0.0 mm. On this basis, sdi1 gene expression has potential as a sensitive indicator of sulphur nutritional status in wheat. Genome-walking techniques were used to clone the 2.7-kb region upstream of sdi1 from genomic DNA, revealing several cis-element motifs previously identified as being associated with sulphur responses in plants. The Arabidopsis thaliana gene most highly homologous to sdi1 is At5g48850, which was also demonstrated to be induced by sulphur deficiency, an observation confirmed by the analysis of microarray data available in the public domain. The expression of Atsdi1 was induced more rapidly than previously characterized sulphur-responsive genes in the period immediately following the transfer of plants to sulphur-deficient medium. Atsdi1 T-DNA 'knockout' mutants were shown to maintain higher tissue sulphate concentrations than wild-type plants under sulphur-limiting conditions, indicating a role in the utilization of stored sulphate under sulphur-deficient conditions. The structural features of the sdi1 gene and its application in the genetic determination of the sulphur nutritional status of wheat crops are discussed.

Fruitlet abscission: A cDNA-AFLP approach to study genes differentially expressed during shedding of immature fruits reveals the involvement of a putative auxin hydrogen symporter in apple (Malus domestica L. Borkh).

PubMed

Dal Cin, Valeriano; Barbaro, Enrico; Danesin, Marcello; Murayama, Hideki; Velasco, Riccardo; Ramina, Angelo

2009-08-01

Apple Malus X domestica fruitlet abscission is preceded by a stimulation of ethylene biosynthesis and a gain in sensitivity to the hormone. This phase was studied by a differential screening carried out by cDNA-AFLP in abscising (AF) and non-abscising (NAF) fruitlet populations. Fifty-three primer combinations allowed for the isolation of 131, 66 and 30 differentially expressed bands from cortex, peduncle and seed, respectively. All sequences were then classified as up- or down-regulated by comparing the profile in AFs and NAFs. Almost all of these sequences showed significant homology to genes encoding proteins with known or putative function. The gene ontology analysis of the TDFs isolated indicated a deep change in metabolism, plastid and hormonal status, especially auxin. Furthermore, some common elements between abscission and senescence were identified. The isolation of the full length of one of these TDFs allowed for the identification of a gene encoding an auxin hydrogen symporter (MdAHS). Bioinformatic analysis indicated that the deduced protein shares some features with other auxin efflux carriers, which include PINs. Nevertheless the 3D structure pointed out substantial differences and a conformation largely dissimilar from canonical ion transporters. The expression analysis demonstrated that this gene is regulated by light and development but not affected by ethylene or auxin.

Polymorphic sequence-characterized codominant loci in the chestnut blight fungus, Cryphonectria parasitica

Treesearch

J. E. Davis; Thomas L. Kubisiak; M. G. Milgroom

2005-01-01

Studies on the population biology of the chestnut blight fungus, Cryphonectria parasitica, have previously been carried out with dominant restriction fragment length polymorphism (RFLP) fingerprinting markers. In this study, we described the development of 11 condominant markers from randomly amplified polymorphic DNAs (RAPDs). RAPD fragments were...

The Effect of Polymorphisms in DNA Repair Genes and Carcinogen Metabolizers on Leukocyte Telomere Length: A Cohort of Healthy Spanish Smokers.

PubMed

Verde, Zoraida; Reinoso-Barbero, Luis; Chicharro, Luis; Resano, Pilar; Sánchez-Hernández, Ignacio; Rodríguez González-Moro, Jose Miguel; Bandrés, Fernando; Gómez-Gallego, Félix; Santiago, Catalina

2016-04-01

Smoking implies exposure to carcinogenic agents that causes DNA damage, which could be suspected to enhance telomere attrition. To protect and deal with DNA damage, cells possess mechanisms that repair and neutralize harmful substances. Polymorphisms altering DNA repair capacity or carcinogen metabolism may lead to synergistic effects with tobacco carcinogen-induced shorter telomere length independently of cancer interaction. The aim of this study was to explore the association between leukocyte telomere length (LTL) and several genetic polymorphisms in DNA repair genes and carcinogen metabolizers in a cohort of healthy smokers. We evaluated the effect of six genetic polymorphisms in cytochrome P1A1 (Ile462Val), XRCC1 (Arg399Gln), APEX1 (Asp148Glu), XRCC3 (Thr241Met), and XPD (Asp312Asn; Lys751Gln) on LTL in a cohort of 145 healthy smokers in addition to smoking habits. Logistic regression analysis showed an association between XRCC1 399Gln allele and shorter telomere length (OR = 5.03, 95% CI = 1.08% to 23.36%). There were not association between the rest of polymorphisms analyzed and LTL. Continuous exposure to tobacco could overwhelm the DNA repair machinery, making the effect of the polymorphisms that reduce repair capacity more pronounced. Analyzing the function of smoking-induced DNA-repair genes and LTL is an important goal in order to identify therapeutic targets to treat smoking-induced diseases. © The Author 2015. Published by Oxford University Press on behalf of the Society for Research on Nicotine and Tobacco. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

Effects of vertebral number variations on carcass traits and genotyping of Vertnin candidate gene in Kazakh sheep.

PubMed

Zhang, Zhifeng; Sun, Yawei; Du, Wei; He, Sangang; Liu, Mingjun; Tian, Changyan

2017-09-01

The vertebral number is associated with body length and carcass traits, which represents an economically important trait in farm animals. The variation of vertebral number has been observed in a few mammalian species. However, the variation of vertebral number and quantitative trait loci in sheep breeds have not been well addressed. In our investigation, the information including gender, age, carcass weight, carcass length and the number of thoracic and lumbar vertebrae from 624 China Kazakh sheep was collected. The effect of vertebral number variation on carcass weight and carcass length was estimated by general linear model. Further, the polymorphic sites of Vertnin ( VRTN ) gene were identified by sequencing, and the association of the genotype and vertebral number variation was analyzed by the one-way analysis of variance model. The variation of thoracolumbar vertebrae number in Kazakh sheep (18 to 20) was smaller than that in Texel sheep (17 to 21). The individuals with 19 thoracolumbar vertebrae (T13L6) were dominant in Kazakh sheep (79.2%). The association study showed that the numbers of thoracolumbar vertebrae were positively correlated with the carcass length and carcass weight, statistically significant with carcass length. To investigate the association of thoracolumbar vertebrae number with VRTN gene, we genotyped the VRTN gene. A total of 9 polymorphic sites were detected and only a single nucleotide polymorphism (SNP) (rs426367238) was suggested to associate with thoracic vertebral number statistically. The variation of thoracolumbar vertebrae number positively associated with the carcass length and carcass weight, especially with the carcass length. VRTN gene polymorphism of the SNP (rs426367238) with significant effect on thoracic vertebral number could be as a candidate marker to further evaluate its role in influence of thoracolumbar vertebral number.

Detection and quantitation of single nucleotide polymorphisms, DNA sequence variations, DNA mutations, DNA damage and DNA mismatches

DOEpatents

McCutchen-Maloney, Sandra L.

2002-01-01

DNA mutation binding proteins alone and as chimeric proteins with nucleases are used with solid supports to detect DNA sequence variations, DNA mutations and single nucleotide polymorphisms. The solid supports may be flow cytometry beads, DNA chips, glass slides or DNA dips sticks. DNA molecules are coupled to solid supports to form DNA-support complexes. Labeled DNA is used with unlabeled DNA mutation binding proteins such at TthMutS to detect DNA sequence variations, DNA mutations and single nucleotide length polymorphisms by binding which gives an increase in signal. Unlabeled DNA is utilized with labeled chimeras to detect DNA sequence variations, DNA mutations and single nucleotide length polymorphisms by nuclease activity of the chimera which gives a decrease in signal.

Restriction fragment length polymorphism and allozyme linkage map of Cuphea lanceolata.

PubMed

Webb, D M; Knapp, S J; Tagliani, L A

1992-02-01

Cuphea lanceolata Ait. has had a significant role in the domestication of Cuphea and is a useful experimental organism for investigating how medium-chain lipids are synthesized in developing seeds. To expand the genetics of this species, a linkage map of the C. lanceolata genome was constructed using five allozyme and 32 restriction-fragment-length-polymorphism (RFLP) marker loci. These loci were assigned to six linkage groups that correspond to the six chromosomes of this species. Map length is 288 cM. Levels of polymorphism were estimated for three inbred lines of C. lanceolata and an inbred line of C. viscosissima using 84 random genomic clones and two restriction enzymes, EcoRI and HindIII. Of the probes 29% detected RFLPs between C. lanceolata and C. viscosissima lines. Crosses between these species can be exploited to expand the map.

A novel single-nucleotide polymorphism of the visfatin gene and its associations with performance traits in the chicken.

PubMed

Han, R-L; Lan, X-Y; Zhang, L-Z; Ren, G; Jing, Y-J; Li, M-J; Zhang, B; Zhao, M; Guo, Y-K; Kang, X-T; Chen, H

2010-01-01

Visfatin is a peptide that is predominantly expressed in visceral adipose tissue and is hypothesized to be related to obesity and insulin resistance. In this study, a novel silent single-nucleotide polymorphism (SNP) was found in exon 7 of the chicken visfatin gene (also known as PBEF1) by single-stranded conformation polymorphism (SSCP) and DNA sequencing. In total, 836 chickens forming an F2 resource population of Gushi chicken crossed with Anka broiler were genotyped by XbaI forced RFLP, and the associations of this polymorphism with chicken growth, carcass characteristics, and meat quality were analyzed. Significant associations were found between the polymorphism and 4-week body weight (BW4), 6-week body weight (BW6), 4-week body slanting length (BSL4), fat bandwidth (FBW), breast muscle water loss rate (BWLR) and breast muscle fiber density (BFD) (P < 0.05), as well as 4-week breastbone length (BBL4) (P < 0.01). These observations suggested that the polymorphism in exon7 of the visfatin gene had significant effects on the early growth traits of chicken.

Environmental sampling of Ceratonia siliqua (carob) trees in Spain reveals the presence of the rare Cryptococcus gattii genotype AFLP7/VGIV.

PubMed

Linares, Carlos; Colom, María Francisca; Torreblanca, Marina; Esteban, Violeta; Romera, Álvaro; Hagen, Ferry

2015-01-01

Cryptococcus gattii is a pathogenic basidiomycetous yeast that is emerging in temperate climate zones worldwide. C. gattii has repetitively been isolated from numerous tree species. Ongoing environmental sampling and molecular characterization is essential to understand the presence of this primary pathogenic microorganism in the Mediterranean environment. To report the first isolation of the rare C. gattii genotype AFLP7/VGIV from the environment in Europe. Samples were collected from woody debris of carob trees (Ceratonia siliqua) and olive trees (Olea europaea) in El Perelló, Tarragona, Spain. Cryptococcus species were further characterized by using URA5-RFLP, MALDI-TOF, AFLP and MLST. The antifungal susceptibility profile to amphotericin B, 5-fluorocytosine, fluconazole, itraconazole, posaconazole and voriconazole was determined using Sensititre Yeast One and E-test. Cultures from one carob tree revealed the presence of ten Cryptococcus-like colonies. One colony was identified as C. gattii, and subsequent molecular characterization showed that it was an α mating-type that belonged to the rare genotype AFLP7/VGIV. Antifungal susceptibility testing showed values within the range of sensitivity described for other isolates of the same genotype and within the epidemiological cutoff values for this species. The isolation of the rare C. gattii genotype AFLP7/VGIV in Spain is the first report in the European environment, implying the possible presence in other regions of the Mediterranean area, and underlines that clinicians must be aware for C. gattii infections in healthy individuals. Copyright © 2014 Revista Iberoamericana de Micología. Published by Elsevier Espana. All rights reserved.

Analysis of DNA methylation related to rice adult plant resistance to bacterial blight based on methylation-sensitive AFLP (MSAP) analysis.

PubMed

Sha, A H; Lin, X H; Huang, J B; Zhang, D P

2005-07-01

DNA methylation is known to play an important role in the regulation of gene expression in eukaryotes. The rice cultivar Wase Aikoku 3 becomes resistant to the blight pathogen Xanthomonas oryzae pv. oryzae at the adult stage. Using methylation-sensitive amplified polymorphism (MSAP) analysis, we compared the patterns of cytosine methylation in seedlings and adult plants of the rice cultivar Wase Aikoku 3 that had been inoculated with the pathogen Xanthomonas oryzae pv. oryzae, subjected to mock inoculation or left untreated. In all, 2000 DNA fragments, each representing a recognition site cleaved by either or both of two isoschizomers, were amplified using 60 pairs of selective primers. A total of 380 sites were found to be methylated. Of these, 45 showed differential cytosine methylation among the seedlings and adult plants subjected to different treatments, and overall levels of methylation were higher in adult plants than in seedlings. All polymorphic fragments were sequenced, and six showed homology to genes that code for products of known function. Northern analysis of three fragments indicated that their expression varied with methylation pattern, with hypermethylation being correlated with repression of transcription, as expected. The results suggest that significant differences in cytosine methylation exist between seedlings and adult plants, and that hypermethylation or hypomethylation of specific genes may be involved in the development of adult plant resistance (APR) in rice plants.

Differentially expressed genes in Populus simonii x P. nigra in respnse to NaCl stress using cDNA-AFLP

USDA-ARS?s Scientific Manuscript database

Salinity is an important environmental factor limiting growth and productivity of plants, and affects almost every aspect of the plant physiology and biochemistry. The objective of this study was to apply cDNA-AFLP and to identify differentially expressed genes in response to NaCl stress vs. no-stre...

Role of 3-Hydroxy Fatty Acid-Induced Hepatic Lipotoxicity in Acute Fatty Liver of Pregnancy

PubMed Central

Ibdah, Jamal A.

2018-01-01

Acute fatty liver of pregnancy (AFLP), a catastrophic illness for both the mother and the unborn offspring, develops in the last trimester of pregnancy with significant maternal and perinatal mortality. AFLP is also recognized as an obstetric and medical emergency. Maternal AFLP is highly associated with a fetal homozygous mutation (1528G>C) in the gene that encodes for mitochondrial long-chain hydroxy acyl-CoA dehydrogenase (LCHAD). The mutation in LCHAD results in the accumulation of 3-hydroxy fatty acids, such as 3-hydroxy myristic acid, 3-hydroxy palmitic acid and 3-hydroxy dicarboxylic acid in the placenta, which are then shunted to the maternal circulation leading to the development of acute liver injury observed in patients with AFLP. In this review, we will discuss the mechanistic role of increased 3-hydroxy fatty acid in causing lipotoxicity to the liver and in inducing oxidative stress, mitochondrial dysfunction and hepatocyte lipoapoptosis. Further, we also review the role of 3-hydroxy fatty acids in causing placental damage, pancreatic islet β-cell glucolipotoxicity, brain damage, and retinal epithelial cells lipoapoptosis in patients with LCHAD deficiency. PMID:29361796

Restriction site polymorphism-based candidate gene mapping for seedling drought tolerance in cowpea [Vigna unguiculata (L.) Walp.].

PubMed

Muchero, Wellington; Ehlers, Jeffrey D; Roberts, Philip A

2010-02-01

Quantitative trait loci (QTL) studies provide insight into the complexity of drought tolerance mechanisms. Molecular markers used in these studies also allow for marker-assisted selection (MAS) in breeding programs, enabling transfer of genetic factors between breeding lines without complete knowledge of their exact nature. However, potential for recombination between markers and target genes limit the utility of MAS-based strategies. Candidate gene mapping offers an alternative solution to identify trait determinants underlying QTL of interest. Here, we used restriction site polymorphisms to investigate co-location of candidate genes with QTL for seedling drought stress-induced premature senescence identified previously in cowpea. Genomic DNA isolated from 113 F(2:8) RILs of drought-tolerant IT93K503-1 and drought susceptible CB46 genotypes was digested with combinations of EcoR1 and HpaII, Mse1, or Msp1 restriction enzymes and amplified with primers designed from 13 drought-responsive cDNAs. JoinMap 3.0 and MapQTL 4.0 software were used to incorporate polymorphic markers onto the AFLP map and to analyze their association with the drought response QTL. Seven markers co-located with peaks of previously identified QTL. Isolation, sequencing, and blast analysis of these markers confirmed their significant homology with drought or other abiotic stress-induced expressed sequence tags (EST) from cowpea and other plant systems. Further, homology with coding sequences for a multidrug resistance protein 3 and a photosystem I assembly protein ycf3 was revealed in two of these candidates. These results provide a platform for the identification and characterization of genetic trait determinants underlying seedling drought tolerance in cowpea.

Restriction fragment length polymorphism of the human c-fms gene.

PubMed Central

Xu, D Q; Guilhot, S; Galibert, F

1985-01-01

By using blot hybridization with a v-fms probe, a polymorphism for EcoRI, HindIII, and BamHI restriction endonuclease sites associated with the human c-fms locus was observed in a random adult population. This restriction fragment length polymorphism can be explained on the basis of the existence of two alleles, a and b, and is due to a short (congruent to 500 base pairs) deletion characteristic of allele a. The distribution in the analyzed population (48 unrelated individuals) is 23% heterozygotes ab, 75% homozygotes bb, and 2% homozygotes aa. Though the inheritance of this polymorphism follows a Mendelian pattern, the children from couples ab X bb are of the following genotype: 74% ab and 26% bb. These deviations from the expected frequencies of 50% suggest a selective pressure in favor of heterozygotes. Images PMID:2986142

Production of polymorphic sperm by anopheline mosquitoes and their fate within the female genital tract.

PubMed

Klowden, Marc J; Chambers, Gail M

2004-12-01

The males of two mosquito species within the Anopheles gambiae complex, An. gambiae s.s. and An. quadriannulatus, as well as males of An. darlingi, produced sperm of significantly varying lengths, while a sperm polymorphism was absent in Aedes aegypti and other anophelines not suspected of belonging to species complexes. The polymorphic distribution of these sperm lengths was not significantly different in smaller adult males that were reared on a low larval diet. The reproductive tract of the female was more likely to contain larger sperm, but overall sperm retention varied depending on the size of the female and the volume of the spermatheca she contained. The presence of a sperm polymorphism may be a factor that has promoted speciation, as well as providing an indication that females may mate multiply.

Rapid identification of Campylobacter, Arcobacter, and Helicobacter isolates by PCR-restriction fragment length polymorphism analysis of the 16S rRNA gene.

PubMed

Marshall, S M; Melito, P L; Woodward, D L; Johnson, W M; Rodgers, F G; Mulvey, M R

1999-12-01

A rapid two-step identification scheme based on PCR-restriction fragment length polymorphism (PCR-RFLP) analysis of the 16S rRNA gene was developed in order to differentiate isolates belonging to the Campylobacter, Arcobacter, and Helicobacter genera. For 158 isolates (26 reference cultures and 132 clinical isolates), specific RFLP patterns were obtained and species were successfully identified by this assay.

Pathological and Molecular Characterization of Xanthomonas campestris Strains Causing Diseases of Cassava (Manihot esculenta)

PubMed Central

Verdier, Valérie; Boher, Bernard; Maraite, Henri; Geiger, Jean-Paul

1994-01-01

Fifty-one strains representing Xanthomonas campestris pv. manihotis and cassavae and different pathovars occurring on plants of the family Euphorbiaceae were characterized by ribotyping with a 16S+23S rRNA probe of Escherichia coli and by restriction fragment length polymorphism analysis with a plasmid probe from X. campestris pv. manihotis. Pathogenicity tests were performed on cassava (Manihot esculenta). Histological comparative studies were conducted on strains of two pathovars of X. campestris (vascular and mesophyllic) that attack cassava. Our results indicated that X. campestris pv. manihotis and cassavae have different modes of action in the host and supplemented the taxonomic data on restriction fragment length polymorphism that clearly separate the two pathovars. The plasmid probe could detect multiple restriction fragment length polymorphisms among strains of the pathovar studied. Ribotyping provides a useful tool for rapid identification of X. campestris pathovars on cassava. Images PMID:16349463

Short poly-glutamine repeat in the androgen receptor in New World monkeys.

PubMed

Hiramatsu, Chihiro; Paukner, Annika; Kuroshima, Hika; Fujita, Kazuo; Suomi, Stephen J; Inoue-Murayama, Miho

2017-12-01

The androgen receptor mediates various physiological and developmental functions and is highly conserved in mammals. Although great intraspecific length polymorphisms in poly glutamine (poly-Q) and poly glycine (poly-G) regions of the androgen receptor in humans, apes and several Old World monkeys have been reported, little is known about the characteristics of these regions in New World monkeys. In this study, we surveyed 17 species of New World monkeys and found length polymorphisms in these regions in three species (common squirrel monkeys, tufted capuchin monkeys and owl monkeys). We found that the poly-Q region in New World monkeys is relatively shorter than that in catarrhines (humans, apes and Old World monkeys). In addition, we observed that codon usage for poly-G region in New World monkeys is unique among primates. These results suggest that the length of polymorphic regions in androgen receptor genes have evolved uniquely in New World monkeys.

[Association analysis between SNPs of the growth hormone receptor gene and growth traits in arctic fox].

PubMed

DU, Zhi-Heng; Liu, Zong-Yue; Bai, Xiu-Juan

2010-06-01

Using single-strand conformation polymorphism (PCR-SSCP) and DNA sequencing, single nucleotide polymorphisms (SNPs) of growth hormone receptor (GHR) gene were detected in an arctic fox population. Correlation analysis between GHR polymorphisms and growth traits were carried out using the appropriate model. Four SNPs, G3A in the 5'UTR, C99T in the first exon, T59C and G65A in the fifth exon were identified on the arctic fox GHR gene. The G3A and C99T polymorphisms of GHR were associated with female fox body weight (Pamp;0.05) and the T59C and G65A polymorphisms of GHR were associated with male fox body weight (Pamp;0.05) and the skin length of the female fox (Pamp;0.01). Therefore, marker assistant selection on body weight and skin length of arctic foxes using these SNPs can be applied to get big and high quality arctic foxes.