Science.gov

Sample records for repeated multilocus microsatellite

  1. Novel Polymorphic Multilocus Microsatellite Markers to Distinguish Candida tropicalis Isolates

    PubMed Central

    Chen, Sharon; Kong, Fanrong; Wang, He; Zhang, Li; Hou, Xin; Xu, Ying-Chun

    2016-01-01

    Candida tropicalis is an important pathogen. Here we developed and evaluated a polymorphic multilocus microsatellite scheme employing novel genetic markers for genotyping of C. tropicalis. Using 10 isolates from 10 unique (separate) patients to screen over 4000 tandem repeats from the C. tropicalis genome (strain MYA-3404), six new candidate microsatellite loci (ctm1, ctm3, ctm8, ctm18, ctm24 and ctm26) were selected according to amplification success, observed polymorphisms and stability of flanking regions by preliminary testing. Two known microsatellite loci CT14 and URA3 were also studied. The 6-locus scheme was then tested against a set of 82 different isolates from 32 patients. Microsatellite genotypes of isolates from the same patient (two to five isolates per patient) were identical. The six loci produced eight to 17 allele types and identified 11 to 24 genotypes amongst 32 patients’ isolates, achieving a discriminatory power (DP) of 0.76 to 0.97 (versus 0.78 for both CT14 and URA3 loci, respectively). Testing of a combination of only three loci, ctm1, ctm3 and ctm24, also achieved maximum typing efficiency (DP = 0.99, 29 genotypes). The microsatellite typing scheme had good correlation compared with pulsed-field gel electrophoresis, although was slightly less discriminatory. The new six-locus microsatellite typing scheme is a potentially valuable tool for genotyping and investigating microevolution of C. tropicalis. PMID:27820850

  2. Extensive polymorphism in Cryptosporidium parvum identified by multilocus microsatellite analysis.

    PubMed

    Feng, X; Rich, S M; Akiyoshi, D; Tumwine, J K; Kekitiinwa, A; Nabukeera, N; Tzipori, S; Widmer, G

    2000-08-01

    Restriction fragment length polymorphism and DNA sequence analysis discern two main types of Cryptosporidium parvum. We present a survey of length polymorphism at several microsatellite loci for type 1 and type 2 isolates. A total of 14 microsatellite loci were identified from C. parvum DNA sequences deposited in public databases. All repeats were mono-, di-, and trinucleotide repeats of A, AT, and AAT, reflecting the high AT content of the C. parvum genome. Several of these loci showed significant length polymorphism, with as many as seven alleles identified for a single locus. Differences between alleles ranged from 1 to 27 bp. Karyotype analysis using probes flanking three microsatellites localized each marker to an individual chromosomal band, suggesting that these markers are single copy. In a sample of 19 isolates for which at least three microsatellites were typed, a majority of isolates displayed a unique multilocus fingerprint. Microsatellite analysis of isolates passaged between different host species identified genotypic changes consistent with changes in parasite populations.

  3. Multilocus microsatellite typing of Leishmania and clinical applications: a review

    PubMed Central

    Aluru, Srikanth; Hide, Mallorie; Michel, Gregory; Bañuls, Anne-Laure; Marty, Pierre; Pomares, Christelle

    2015-01-01

    Microsatellite markers have been used for Leishmania genetic studies worldwide, giving useful insight into leishmaniasis epidemiology. Understanding the geographic distribution, dynamics of Leishmania populations, and disease epidemiology improved markedly with this tool. In endemic foci, the origins of antimony-resistant strains and multidrug treatment failures were explored with multilocus microsatellite typing (MLMT). High genetic variability was detected but no association between parasite genotypes and drug resistance was established. An association between MLMT profiles and clinical disease manifestations was highlighted in only three studies and this data needs further confirmation. At the individual level, MLMT provided information on relapse and reinfection when multiple leishmaniasis episodes occurred. This information could improve knowledge of epidemiology and guide therapeutic choices for active chronic visceral leishmaniasis, the disease form in some HIV-positive patients. PMID:25950900

  4. Typing Candida Species Using Microsatellite Length Polymorphism and Multilocus Sequence Typing.

    PubMed

    Garcia-Hermoso, Dea; Desnos-Ollivier, Marie; Bretagne, Stéphane

    2016-01-01

    To gain more insight into the epidemiological relationships between isolates of Candida spp. obtained from various origins, several molecular typing techniques have been developed. Two methods have emerged in the 2000s as soon as enough knowledge of the Candida spp. genomes was available to choose adequate loci and primers, namely microsatellite length polymorphism (MLP) and multilocus sequence typing (MLST). To contrast with previous PCR-based methods, specific amplifications with stringent conditions easily reproducible are the basis of MLP and MLST. MLST relies on Sanger sequencing to detect single-nucleotide polymorphisms within housekeeping genes. MLP needs a first in silico step to select tandemly repeated stretches of two to five nucleotides. One of the two primers used to amplify a microsatellite locus is labeled and fragment sizing is automatically performed using high-resolution electrophoresis platforms. MLST provides results easily comparable between laboratories and active MLST schemes are publicly available for the main Candida species. For comparative studies, MLP needs standards to compensate for the electrophoretic variations depending on the platforms used. Both methods can help us gain insight into the genetic relatedness of fungal isolates, both with advantages and drawbacks, and the choice of one method rather than the other depends on the task in question.

  5. Triplet-repeat microsatellites shared among hard and soft pines.

    PubMed

    Kutil, B L; Williams, C G

    2001-01-01

    Vascular plant species have shown a low level of microsatellite conservation compared to many animal species. Finding trans-specific microsatellites for plants may be improved by using a priori knowledge of genome organization. Fifteen triplet-repeat microsatellites from hard pine (Pinus taeda L.) were tested for trans-specific amplification across seven hard pines (P. palustris Mill., P. echinata Mill., P. radiata D. Don., P. patula Schiede et Deppe, P. halepensis Mill., P. kesiya Royle), a soft pine (P. strobus L.), and Picea rubens Sargent. Seven of 15 microsatellites had trans-specific amplification in both hard and soft pine subgenera. Two P. taeda microsatellites had conserved flanking regions and repeat motifs in all seven hard pines, soft pine P. strobus, and P. rubens. Perfect triplet-repeat P. taeda microsatellites appear to be better candidates for trans-specific polymorphism than compound microsatellites. Not all perfect triplet-repeat microsatellites were conserved, but all conserved microsatellites had perfect repeat motifs. Persistent microsatellites PtTX2123 and PtTX3020 had highly conserved flanking regions and a conserved repeat motif composition with variable repeat unit numbers. Using trinucleotide microsatellites improved trans-specific microsatellite recovery among hard and soft pine species.

  6. Quasimonomorphic mononucleotide repeats for high-level microsatellite instability analysis.

    PubMed

    Buhard, Olivier; Suraweera, Nirosha; Lectard, Aude; Duval, Alex; Hamelin, Richard

    2004-01-01

    Microsatellite instability (MSI) analysis is becoming more and more important to detect sporadic primary tumors of the MSI phenotype as well as in helping to determine Hereditary Non-Polyposis Colorectal Cancer (HNPCC) cases. After some years of conflicting data due to the absence of consensus markers for the MSI phenotype, a meeting held in Bethesda to clarify the situation proposed a set of 5 microsatellites (2 mononucleotide repeats and 3 dinucleotide repeats) to determine MSI tumors. A second Bethesda consensus meeting was held at the end of 2002. It was discussed here that the 1998 microsatellite panel could underestimate high-level MSI tumors and overestimate low-level MSI tumors. Amongst the suggested changes was the exclusive use of mononucleotide repeats in place of dinucleotide repeats. We have already proposed a pentaplex MSI screening test comprising 5 quasimonomorphic mononucleotide repeats. This article compares the advantages of mono or dinucleotide repeats in determining microsatellite instability.

  7. Quasimonomorphic Mononucleotide Repeats for High-Level Microsatellite Instability Analysis

    PubMed Central

    Buhard, Olivier; Suraweera, Nirosha; Lectard, Aude; Duval, Alex; Hamelin, Richard

    2004-01-01

    Microsatellite instability (MSI) analysis is becoming more and more important to detect sporadic primary tumors of the MSI phenotype as well as in helping to determine Hereditary Non-Polyposis Colorectal Cancer (HNPCC) cases. After some years of conflicting data due to the absence of consensus markers for the MSI phenotype, a meeting held in Bethesda to clarify the situation proposed a set of 5 microsatellites (2 mononucleotide repeats and 3 dinucleotide repeats) to determine MSI tumors. A second Bethesda consensus meeting was held at the end of 2002. It was discussed here that the 1998 microsatellite panel could underestimate high-level MSI tumors and overestimate low-level MSI tumors. Amongst the suggested changes was the exclusive use of mononucleotide repeats in place of dinucleotide repeats. We have already proposed a pentaplex MSI screening test comprising 5 quasimonomorphic mononucleotide repeats. This article compares the advantages of mono or dinucleotide repeats in determining microsatellite instability. PMID:15528790

  8. Vectorette PCR isolation of microsatellite repeat sequences using anchored dinucleotide repeat primers.

    PubMed Central

    Lench, N J; Norris, A; Bailey, A; Booth, A; Markham, A F

    1996-01-01

    We have developed a vectorette PCR approach to provide an improved method for isolation of microsatellite repeats. The modified procedure relies on PCR amplification using a vectorette-specific primer in combination with one of a panel of anchored dinucleotide repeat primers. The target DNA to be screened for microsatellite sequences can be from YAC, P1, cosmid, bacteriophage or plasmid clones. We have used this technique to isolate novel, polymorphic microsatellite repeats from clones containing the amelogenin gene (AMGX) located on human chromosome Xp22.3. PMID:8668553

  9. Multilocus Microsatellite Typing as a New Tool for Discrimination of Leishmania infantum MON-1 Strains

    PubMed Central

    Ochsenreither, Sebastian; Kuhls, Katrin; Schaar, Matthias; Presber, Wolfgang; Schönian, Gabriele

    2006-01-01

    The Leishmania donovani complex, which consists of L. donovani, L. infantum-L. chagasi, and L. archibaldi, is responsible for visceral manifestations of leishmaniasis. Multilocus enzyme electrophoresis is the standard method for the characterization and identification of strains of Leishmania. For L. infantum, the predominance of zymodeme MON-1 significantly reduces the discriminative power of this approach. In the present study, we developed 17 independent polymorphic microsatellite markers for the typing of strains of L. infantum, with the main emphasis on zymodeme MON-1. The discriminative powers of 11 markers selected from among these markers were tested by using a panel of 63 isolates of the L. donovani complex. Unique multilocus genotypes were observed for the strains analyzed, with only three exceptions. Model-based and distance-based analyses of the data set showed comparable results. It was possible to discriminate between L. donovani sensu stricto, a non-MON-1 group of L. infantum isolates, and a MON-1 group of L. infantum isolates. Within MON-1, three clusters with geographical correlations became apparent. The frequency of heterozygosity in the alleles analyzed varied extremely between the different groups of isolates. The main clusters described are not consistent with species definitions based on isoenzyme analysis but confirm the results of former PCR-based investigations. PMID:16455904

  10. Comparison of microsatellite length polymorphism and multilocus sequence typing for DNA-Based typing of Candida albicans.

    PubMed

    Garcia-Hermoso, Dea; Cabaret, Odile; Lecellier, Gael; Desnos-Ollivier, Marie; Hoinard, Damien; Raoux, Dorothée; Costa, Jean-Marc; Dromer, Françoise; Bretagne, Stéphane

    2007-12-01

    For genotyping Candida albicans isolates, two PCR-based methods have recently emerged: multilocus sequence typing (MLST), based on the sequence of selected genes, and microsatellite length polymorphism (MLP), based on the length of PCR products containing variable numbers of short DNA repeats. To compare the two methods in their abilities to differentiate and group C. albicans isolates, we selected 50 independent isolates collected at the National Reference Center for Mycoses and Antifungals. MLST typing was performed using sequencing of seven loci as described at (http://test1.mlst.net). The MLP method consisted of a single multiplex PCR testing three different loci. Dendrograms were constructed by the unweighted pair group cluster method with Euclidean metric for both methods. The correlation between the distance matrices was performed with a Mantel test tested with 1,000 random permutations. The sensitivity and specificity of the MLP typing system were determined after allocating MLST groups for the greater number of isolates of each distinct MLP group. The discriminatory power index was >0.99, and the distances between the isolates were highly correlated with both systems. The Mantel coefficient and the Pearson product-moment correlation coefficient were 35,699 and 0.32, respectively (P < or = 1.2 x 10(-6)). Using MLP, the average specificity and sensitivity of clustering compared to MLST were 83% and 73%, respectively, when the singletons were excluded. The two methods are similarly discriminatory and can be interchangeable depending on the objectives. MLP is less expensive and faster than MLST. However, MLST is currently more accurate and additional standardization is needed for MLP.

  11. Low abundance of microsatellite repeats in the genome of the Brown-headed Cowbird (Molothrus ater)

    USGS Publications Warehouse

    Longmire, J.L.; Hahn, D.C.; Roach, J.L.

    1999-01-01

    A cosmid library made from brown-headed cowbird (Molothrus ater) DNA was examined for representation of 17 distinct microsatellite motifs including all possible mono-, di-, and trinucleotide microsatellites, and the tetranucleotide repeat (GATA)n. The overall density of microsatellites within cowbird DNA was found to be one repeat per 89 kb and the frequency of the most abundant motif, (AGC)n, was once every 382 kb. The abundance of microsatellites within the cowbird genome is estimated to be reduced approximately 15-fold compared to humans. The reduced frequency of microsatellites seen in this study is consistent with previous observations indicating reduced numbers of microsatellites and other interspersed repeats in avian DNA. In addition to providing new information concerning the abundance of microsatellites within an avian genome, these results provide useful insights for selecting cloning strategies that might be used in the development of locus-specific microsatellite markers for avian studies.

  12. Discrepancy variation of dinucleotide microsatellite repeats in eukaryotic genomes.

    PubMed

    Gao, Huan; Cai, Shengli; Yan, Binlun; Chen, Baiyao; Yu, Fei

    2009-01-01

    To address whether there are differences of variation among repeat motif types and among taxonomic groups, we present here an analysis of variation and correlation of dinucleotide microsatellite repeats in eukaryotic genomes. Ten taxonomic groups were compared, those being primates, mammalia (excluding primates and rodentia), rodentia, birds, fish, amphibians and reptiles, insects, molluscs, plants and fungi, respectively. The data used in the analysis is from the literature published in the Journal of Molecular Ecology Notes. Analysis of variation reveals that there are no significant differences between AC and AG repeat motif types. Moreover, the number of alleles correlates positively with the copy number in both AG and AC repeats. Similar conclusions can be obtained from each taxonomic group. These results strongly suggest that the increase of SSR variation is almost linear with the increase of the copy number of each repeat motif. As well, the results suggest that the variability of SSR in the genomes of low-ranking species seem to be more than that of high-ranking species, excluding primates and fungi.

  13. Efficient isolation of polymorphic microsatellites from high-throughput sequence data based on number of repeats.

    PubMed

    Cardoso, Sara D; Gonçalves, David; Robalo, Joana I; Almada, Vitor C; Canário, Adelino V M; Oliveira, Rui F

    2013-09-01

    Transcriptome data are a good resource to develop microsatellites due to their potential in targeting candidate genes. However, developing microsatellites can be a time-consuming enterprise due to the numerous primer pairs to be tested. Therefore, the use of methodologies that make it efficient to identify polymorphic microsatellites is desirable. Here we used a 62,038 contigs transcriptome assembly, obtained from pyrosequencing a peacock blenny (Salaria pavo) multi-tissue cDNA library, to mine for microsatellites and in silico evaluation of their polymorphism. A total of 4190 microsatellites were identified in 3670 unique unigenes, and from these microsatellites, in silico polymorphism was detected in 733. We selected microsatellites based either on their in silico polymorphism and annotation results or based only on their number of repeats. Using these two approaches, 28 microsatellites were successfully amplified in twenty-six individuals, and all but 2 were found to be polymorphic, being the first genetic markers for this species. Our results showed that the strategy of selection based on number of repeats is more efficient in obtaining polymorphic microsatellites than the strategy of in silico polymorphism (allelic richness was 8.2±3.85 and 4.56±2.45 respectively). This study demonstrates that combining the knowledge of number of repeats with other predictors of variability, for example in silico microsatellite polymorphism, improves the rates of polymorphism, yielding microsatellites with higher allelic richness, and decreases the number of monomorphic microsatellites obtained.

  14. Microsatellite Tandem Repeats Are Abundant in Human Promoters and Are Associated with Regulatory Elements

    PubMed Central

    Sawaya, Sterling; Bagshaw, Andrew; Buschiazzo, Emmanuel; Kumar, Pankaj; Chowdhury, Shantanu; Black, Michael A.; Gemmell, Neil

    2013-01-01

    Tandem repeats are genomic elements that are prone to changes in repeat number and are thus often polymorphic. These sequences are found at a high density at the start of human genes, in the gene’s promoter. Increasing empirical evidence suggests that length variation in these tandem repeats can affect gene regulation. One class of tandem repeats, known as microsatellites, rapidly alter in repeat number. Some of the genetic variation induced by microsatellites is known to result in phenotypic variation. Recently, our group developed a novel method for measuring the evolutionary conservation of microsatellites, and with it we discovered that human microsatellites near transcription start sites are often highly conserved. In this study, we examined the properties of microsatellites found in promoters. We found a high density of microsatellites at the start of genes. We showed that microsatellites are statistically associated with promoters using a wavelet analysis, which allowed us to test for associations on multiple scales and to control for other promoter related elements. Because promoter microsatellites tend to be G/C rich, we hypothesized that G/C rich regulatory elements may drive the association between microsatellites and promoters. Our results indicate that CpG islands, G-quadruplexes (G4) and untranslated regulatory regions have highly significant associations with microsatellites, but controlling for these elements in the analysis does not remove the association between microsatellites and promoters. Due to their intrinsic lability and their overlap with predicted functional elements, these results suggest that many promoter microsatellites have the potential to affect human phenotypes by generating mutations in regulatory elements, which may ultimately result in disease. We discuss the potential functions of human promoter microsatellites in this context. PMID:23405090

  15. Assessment of population genetic structure in the arbovirus vector midge, Culicoides brevitarsis (Diptera: Ceratopogonidae), using multi-locus DNA microsatellites.

    PubMed

    Onyango, Maria G; Beebe, Nigel W; Gopurenko, David; Bellis, Glenn; Nicholas, Adrian; Ogugo, Moses; Djikeng, Appolinaire; Kemp, Steve; Walker, Peter J; Duchemin, Jean-Bernard

    2015-09-25

    Bluetongue virus (BTV) is a major pathogen of ruminants that is transmitted by biting midges (Culicoides spp.). Australian BTV serotypes have origins in Asia and are distributed across the continent into two distinct episystems, one in the north and another in the east. Culicoides brevitarsis is the major vector of BTV in Australia and is distributed across the entire geographic range of the virus. Here, we describe the isolation and use of DNA microsatellites and gauge their ability to determine population genetic connectivity of C. brevitarsis within Australia and with countries to the north. Eleven DNA microsatellite markers were isolated using a novel genomic enrichment method and identified as useful for genetic analyses of sampled populations in Australia, northern Papua New Guinea (PNG) and Timor-Leste. Significant (P < 0.05) population genetic subdivision was observed between all paired regions, though the highest levels of genetic sub-division involved pair-wise tests with PNG (PNG vs. Australia (FST = 0.120) and PNG vs. Timor-Leste (FST = 0.095)). Analysis of multi-locus allelic distributions using STRUCTURE identified a most probable two-cluster population model, which separated PNG specimens from a cluster containing specimens from Timor-Leste and Australia. The source of incursions of this species in Australia is more likely to be Timor-Leste than PNG. Future incursions of BTV positive C. brevitarsis into Australia may be genetically identified to their source populations using these microsatellite loci. The vector's panmictic genetic structure within Australia cannot explain the differential geographic distribution of BTV serotypes.

  16. Variogram Analysis of the Spatial Genetic Structure of Continuous Populations Using Multilocus Microsatellite Data

    PubMed Central

    Wagner, Helene H.; Holderegger, Rolf; Werth, Silke; Gugerli, Felix; Hoebee, Susan E.; Scheidegger, Christoph

    2005-01-01

    A geostatistical perspective on spatial genetic structure may explain methodological issues of quantifying spatial genetic structure and suggest new approaches to addressing them. We use a variogram approach to (i) derive a spatial partitioning of molecular variance, gene diversity, and genotypic diversity for microsatellite data under the infinite allele model (IAM) and the stepwise mutation model (SMM), (ii) develop a weighting of sampling units to reflect ploidy levels or multiple sampling of genets, and (iii) show how variograms summarize the spatial genetic structure within a population under isolation-by-distance. The methods are illustrated with data from a population of the epiphytic lichen Lobaria pulmonaria, using six microsatellite markers. Variogram-based analysis not only avoids bias due to the underestimation of population variance in the presence of spatial autocorrelation, but also provides estimates of population genetic diversity and the degree and extent of spatial genetic structure accounting for autocorrelation. PMID:15654102

  17. Multilocus Microsatellite Typing reveals intra-focal genetic diversity among strains of Leishmania tropica in Chichaoua Province, Morocco.

    PubMed

    Krayter, Lena; Alam, Mohammad Zahangir; Rhajaoui, Mohamed; Schnur, Lionel F; Schönian, Gabriele

    2014-12-01

    In Morocco, cutaneous leishmaniasis (CL) caused by Leishmania (L.) tropica is a major public health threat. Strains of this species have been shown to display considerable serological, biochemical, molecular biological and genetic heterogeneity; and Multilocus Enzyme Electrophoresis (MLEE), has shown that in many countries including Morocco heterogenic variants of L. tropica can co-exist in single geographical foci. Here, the microsatellite profiles discerned by MLMT of nine Moroccan strains of L. tropica isolated in 2000 from human cases of CL from Chichaoua Province were compared to those of nine Moroccan strains of L. tropica isolated between 1988 and 1990 from human cases of CL from Marrakech Province, and also to those of 147 strains of L. tropica isolated at different times from different worldwide geographical locations within the range of distribution of the species. Several programs, each employing a different algorithm, were used for population genetic analysis. The strains from each of the two Moroccan foci separated into two phylogenetic clusters independent of their geographical origin. Genetic diversity and heterogeneity existed in both foci, which are geographically close to each other. This intra-focal distribution of genetic variants of L. tropica is not considered owing to in situ mutation. Rather, it is proposed to be explained by the importation of pre-existing variants of L. tropica into Morocco.

  18. Evidence for an association between post-fledging dispersal and microsatellite multilocus heterozygosity in a large population of greater flamingos.

    PubMed

    Gillingham, Mark A F; Cézilly, Frank; Wattier, Rémi; Béchet, Arnaud

    2013-01-01

    Dispersal can be divided into three stages: departure, transience and settlement. Despite the fact that theoretical studies have emphasized the importance of heterozygosity on dispersal strategies, empirical evidence of its effect on different stages of dispersal is lacking. Here, using multi-event capture-mark-recapture models, we show a negative association between microsatellite multilocus heterozygosity (MLH; 10 loci; n = 1023) and post-fledging dispersal propensity for greater flamingos, Phoenicopterus roseus, born in southern France. We propose that the negative effects of inbreeding depression affects competitive ability and therefore more homozygous individuals are more likely to disperse because they are less able to compete within the highly saturated natal site. Finally, a model with the effect of MLH on propensity of post-fledgling dispersers to disperse to the long-distance sites of Africa was equivalent to the null model, suggesting that MLH had low to no effect on dispersal distance. Variations in individual genetic quality thus result in context-dependent heterogeneity in dispersal strategies at each stage of dispersal. Our results have important implications on fitness since sites visited early in life are known to influence site selection later on in life and future survival.

  19. Evidence for an Association between Post-Fledging Dispersal and Microsatellite Multilocus Heterozygosity in a Large Population of Greater Flamingos

    PubMed Central

    Gillingham, Mark A. F.; Cézilly, Frank; Wattier, Rémi; Béchet, Arnaud

    2013-01-01

    Dispersal can be divided into three stages: departure, transience and settlement. Despite the fact that theoretical studies have emphasized the importance of heterozygosity on dispersal strategies, empirical evidence of its effect on different stages of dispersal is lacking. Here, using multi-event capture-mark-recapture models, we show a negative association between microsatellite multilocus heterozygosity (MLH; 10 loci; n = 1023) and post-fledging dispersal propensity for greater flamingos, Phoenicopterus roseus, born in southern France. We propose that the negative effects of inbreeding depression affects competitive ability and therefore more homozygous individuals are more likely to disperse because they are less able to compete within the highly saturated natal site. Finally, a model with the effect of MLH on propensity of post-fledgling dispersers to disperse to the long-distance sites of Africa was equivalent to the null model, suggesting that MLH had low to no effect on dispersal distance. Variations in individual genetic quality thus result in context-dependent heterogeneity in dispersal strategies at each stage of dispersal. Our results have important implications on fitness since sites visited early in life are known to influence site selection later on in life and future survival. PMID:24278385

  20. GENETIC VARIATION IN MULTILOCUS MICROSATELLITE GENOTYPES IN TWO SPECIES OF WOODRATS (NEOTOMA MACROTIS AND N. FUSCIPES) FROM CALIFORNIA

    PubMed Central

    Haynie, Michelle L.; Fulhorst, Charles F.; Rood, Michael; Bennett, Stephen G.; Hess, Barry D.; Bradley, ROBERT D.

    2009-01-01

    Five microsatellite loci were used to develop multilocus genotypes for Neotoma macrotis (n = 128) and N. fuscipes (n = 29). Several statistical analyses were used to estimate genetic structure, levels of genetic variability, and degree of relatedness within groups of these 2 species. Samples of N. macrotis represented 2 groups and 4 population clusters throughout southern California. Samples of N. fuscipes represented 2 regions in northern and southern California. Genetic structure was detected among samples of N. macrotis and N. fuscipes at a regional level. Both species displayed moderate to high genetic diversity in terms of mean expected heterozygosity (0.939 and 0.804 for N. macrotis and N. fuscipes, respectively) and mean polymorphic information content (0.930 and 0.761 for N. macrotis and N. fuscipes, respectively). Mean relatedness values within regions and populations of N. macrotis indicated 4th-order levels of relatedness within groups (e.g., distant-cousin relationships). Mean relatedness values within regions of N. fuscipes indicated 2nd-order (e.g., half-sibling) relationships within the northern region and 3rd-order (e.g., cousin) relationships in the southern region. One locus in particular (Nma04) was determined to be diagnostic in distinguishing between these 2 species. PMID:19920871

  1. A novel class of miniature inverted repeat transposable elements (MITEs) that contain hitchhiking (GTCY)n microsatellites

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The insertion of transposable elements results in the modification of genome structure and gene expression, and also facilitates the propagation of derived internal sequences. We show that (GTCY)n microsatellite loci within genomes of Lepidoptera are mobile and multilocus due to hitchhiking within ...

  2. Efficient development of highly polymorphic microsatellite markers based on polymorphic repeats in transcriptome sequences of multiple individuals.

    PubMed

    Vukosavljev, M; Esselink, G D; van 't Westende, W P C; Cox, P; Visser, R G F; Arens, P; Smulders, M J M

    2015-01-01

    The first hurdle in developing microsatellite markers, cloning, has been overcome by next-generation sequencing. The second hurdle is testing to differentiate polymorphic from nonpolymorphic loci. The third hurdle, somewhat hidden, is that only polymorphic markers with a large effective number of alleles are sufficiently informative to be deployed in multiple studies. Both steps are laborious and still performed manually. We have developed a strategy in which we first screen reads from multiple genotypes for repeats that show the most length variants, and only these are subsequently developed into markers. We validated our strategy in tetraploid garden rose using Illumina paired-end transcriptome sequences of 11 roses. Of 48 tested two markers failed to amplify, but all others were polymorphic. Ten loci amplified more than one locus, indicating duplicated genes or gene families. Completely avoiding duplicated loci will be difficult because the range of numbers of predicted alleles of highly polymorphic single- and multilocus markers largely overlapped. Of the remainder, half were replicate markers (i.e. multiple primer pairs for one locus), indicating the difficulty of correctly filtering short reads containing repeat sequences. We subsequently refined the approach to eliminate multiple primer sets to the same loci. The remaining 18 markers were all highly polymorphic, amplifying on average 11.7 alleles per marker (range = 6-20) in 11 tetraploid roses, exceeding the 8.2 alleles per marker of the 24 most polymorphic markers genotyped previously. This strategy therefore represents a major step forward in the development of highly polymorphic microsatellite markers.

  3. A novel multilocus variable number tandem repeat analysis typing scheme for African phylotype III strains of the Ralstonia solanacearum species complex.

    PubMed

    Ravelomanantsoa, Santatra; Robène, Isabelle; Chiroleu, Frédéric; Guérin, Fabien; Poussier, Stéphane; Pruvost, Olivier; Prior, Philippe

    2016-01-01

    Background. Reliable genotyping that provides an accurate description of diversity in the context of pathogen emergence is required for the establishment of strategies to improve disease management. MultiLocus variable number tandem repeat analysis (MLVA) is a valuable genotyping method. It can be performed at small evolutionary scales where high discriminatory power is needed. Strains of the Ralstonia solanacearum species complex (RSSC) are highly genetically diverse. These destructive pathogens are the causative agent of bacterial wilt on an unusually broad range of host plants worldwide. In this study, we developed an MLVA scheme for genotyping the African RSSC phylotype III. Methods. We selected different publicly available tandem repeat (TR) loci and additional TR loci from the genome of strain CMR15 as markers. Based on these loci, a new phylotype III-MLVA scheme is presented. MLVA and multiLocus sequence typing (MLST) were compared at the global, regional, and local scales. Different populations of epidemiologically related and unrelated RSSC phylotype III strains were used. Results and Discussion. Sixteen polymorphic TR loci, which included seven microsatellites and nine minisatellites, were selected. These TR loci were distributed throughout the genome (chromosome and megaplasmid) and located in both coding and intergenic regions. The newly developed RS3-MLVA16 scheme was more discriminative than MLST. RS3-MLVA16 showed good ability in differentiating strains at global, regional, and local scales, and it especially highlighted epidemiological links between closely related strains at the local scale. RS3-MLVA16 also underlines genetic variability within the same MLST-type and clonal complex, and gives a first overview of population structure. Overall, RS3-MLVA16 is a promising genotyping method for outbreak investigation at a fine scale, and it could be used for outbreak investigation as a first-line, low-cost assay for the routine screening of RSSC

  4. A novel multilocus variable number tandem repeat analysis typing scheme for African phylotype III strains of the Ralstonia solanacearum species complex

    PubMed Central

    Ravelomanantsoa, Santatra; Robène, Isabelle; Chiroleu, Frédéric; Guérin, Fabien; Poussier, Stéphane; Pruvost, Olivier

    2016-01-01

    Background. Reliable genotyping that provides an accurate description of diversity in the context of pathogen emergence is required for the establishment of strategies to improve disease management. MultiLocus variable number tandem repeat analysis (MLVA) is a valuable genotyping method. It can be performed at small evolutionary scales where high discriminatory power is needed. Strains of the Ralstonia solanacearum species complex (RSSC) are highly genetically diverse. These destructive pathogens are the causative agent of bacterial wilt on an unusually broad range of host plants worldwide. In this study, we developed an MLVA scheme for genotyping the African RSSC phylotype III. Methods. We selected different publicly available tandem repeat (TR) loci and additional TR loci from the genome of strain CMR15 as markers. Based on these loci, a new phylotype III-MLVA scheme is presented. MLVA and multiLocus sequence typing (MLST) were compared at the global, regional, and local scales. Different populations of epidemiologically related and unrelated RSSC phylotype III strains were used. Results and Discussion. Sixteen polymorphic TR loci, which included seven microsatellites and nine minisatellites, were selected. These TR loci were distributed throughout the genome (chromosome and megaplasmid) and located in both coding and intergenic regions. The newly developed RS3-MLVA16 scheme was more discriminative than MLST. RS3-MLVA16 showed good ability in differentiating strains at global, regional, and local scales, and it especially highlighted epidemiological links between closely related strains at the local scale. RS3-MLVA16 also underlines genetic variability within the same MLST-type and clonal complex, and gives a first overview of population structure. Overall, RS3-MLVA16 is a promising genotyping method for outbreak investigation at a fine scale, and it could be used for outbreak investigation as a first-line, low-cost assay for the routine screening of RSSC

  5. Microsatellite and trinucleotide-repeat evolution: evidence for mutational bias and different rates of evolution in different lineages.

    PubMed Central

    Rubinsztein, D C; Amos, B; Cooper, G

    1999-01-01

    Microsatellites are stretches of repetitive DNA, where individual repeat units comprise one to six bases. These sequences are often highly polymorphic with respect to repeat number and include trinucleotide repeats, which are abnormally expanded in a number of diseases. It has been widely assumed that microsatellite loci are as likely to gain and lose repeats when they mutate. In this review, we present population genetic and empirical data arguing that microsatellites, including normal alleles at trinucleotide-repeat disease loci, are more likely to expand in length when they mutate. In addition, our experiments suggest that the rates of expansion of such sequences differ in related species. PMID:10434312

  6. What is a microsatellite: a computational and experimental definition based upon repeat mutational behavior at A/T and GT/AC repeats.

    PubMed

    Kelkar, Yogeshwar D; Strubczewski, Noelle; Hile, Suzanne E; Chiaromonte, Francesca; Eckert, Kristin A; Makova, Kateryna D

    2010-01-01

    Microsatellites are abundant in eukaryotic genomes and have high rates of strand slippage-induced repeat number alterations. They are popular genetic markers, and their mutations are associated with numerous neurological diseases. However, the minimal number of repeats required to constitute a microsatellite has been debated, and a definition of a microsatellite that considers its mutational behavior has been lacking. To define a microsatellite, we investigated slippage dynamics for a range of repeat sizes, utilizing two approaches. Computationally, we assessed length polymorphism at repeat loci in ten ENCODE regions resequenced in four human populations, assuming that the occurrence of polymorphism reflects strand slippage rates. Experimentally, we determined the in vitro DNA polymerase-mediated strand slippage error rates as a function of repeat number. In both approaches, we compared strand slippage rates at tandem repeats with the background slippage rates. We observed two distinct modes of mutational behavior. At small repeat numbers, slippage rates were low and indistinguishable from background measurements. A marked transition in mutability was observed as the repeat array lengthened, such that slippage rates at large repeat numbers were significantly higher than the background rates. For both mononucleotide and dinucleotide microsatellites studied, the transition length corresponded to a similar number of nucleotides (approximately 10). Thus, microsatellite threshold is determined not by the presence/absence of strand slippage at repeats but by an abrupt alteration in slippage rates relative to background. These findings have implications for understanding microsatellite mutagenesis, standardization of genome-wide microsatellite analyses, and predicting polymorphism levels of individual microsatellite loci.

  7. MICdb3.0: a comprehensive resource of microsatellite repeats from prokaryotic genomes.

    PubMed

    Mudunuri, Suresh B; Patnana, Sujan; Nagarajaram, Hampapathalu A

    2014-01-01

    The MICdb is a comprehensive relational database of perfect microsatellites extracted from completely sequenced and annotated genomes of bacteria and archaea. The current version MICdb3.0 is an updated and revised version of MICdb2.0. As compared with the previous version MICdb2.0, the current release is significantly improved in terms of much larger coverage of genomes, improved presentation of queried results, user-friendly administration module to manage Simple Sequence Repeat (SSR) data such as addition of new genomes, deletion of obsolete data, etc., and also removal of certain features deemed to be redundant. The new web-interface to the database called Microsatellite Analysis Server (MICAS) version 3.0 has been improved by the addition of powerful high-quality visualization tools to view the query results in the form of pie charts and bar graphs. All the query results and graphs can be exported in different formats so that the users can use them for further analysis. MICAS3.0 is also equipped with a unique genome comparison module using which users can do pair-wise comparison of genomes with regard to their microsatellite distribution. The advanced search module can be used to filter the repeats based on certain criteria such as filtering repeats of a particular motif/repeat size, extracting repeats of coding/non-coding regions, sort repeats, etc. The MICdb database has, therefore, been made portable to be administered by a person with the necessary administrative privileges. The MICdb3.0 database and analysis server can be accessed for free from www.cdfd.org.in/micas. Database URL: http://www.cdfd.org.in/micas.

  8. Multilocus sequence and microsatellite identification of intra-specific hybrids and ancestor-like donors among natural Ethiopian isolates of Leishmania donovani.

    PubMed

    Gelanew, Tesfaye; Hailu, Asrat; Schőnian, Gabriele; Lewis, Michael D; Miles, Michael A; Yeo, Matthew

    2014-09-01

    Protozoan parasites of the genus Leishmania (Kinetoplastida: Trypanosomatidae) cause widespread and devastating human diseases. Visceral leishmaniasis is endemic in Ethiopia where it has also been responsible for fatal epidemics. It is postulated that genetic exchange in Leishmania has implications for heterosis (hybrid vigour), spread of virulent strains, resistance to chemotherapeutics, and exploitation of different hosts and vectors. Here we analyse 11 natural Ethiopian Leishmania donovani isolates consisting of four putative hybrids, seven parent-like isolates and over 90 derived biological clones. We apply a novel combination of high resolution multilocus microsatellite typing (five loci) and multilocus sequence typing (four loci) that together distinguish parent-like and hybrid L. donovani strains. Results indicate that the four isolates (and their associated biological clones) are genetic hybrids, not the results of mixed infections, each possessing heterozygous markers consistent with inheritance of divergent alleles from genetically distinct Ethiopian L. donovani lineages. The allelic profiles of the putative hybrids may have arisen from a single hybridisation event followed by inbreeding or gene conversion, or alternatively from two or more hybridisation events. Mitochondrial sequencing showed uniparental maxicircle inheritance for all of the hybrids, each possessing a single mitochondrial genotype. Fluorescence activated cell sorting analysis of DNA content demonstrated that all hybrids and their associated clones were diploid. Together the data imply that intra-specific genetic exchange is a recurrent feature of natural L. donovani populations, with substantial implications for the phyloepidemiology of Leishmania.

  9. Development of a tandem repeat-based multilocus typing system distinguishing Babesia bovis geographic isolates

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Mini and microsatellite sequences have proven to be excellent tools for the differentiation of strains and populations in several protozoan parasites due to their high variability. In the present work we have searched the genome of the tick-transmitted bovine hemoprotozoon Babesia bovis for tandem r...

  10. Microsatellite instability at tetranucleotide repeats in sporadic colorectal cancer in Japan.

    PubMed

    Yamada, Kanae; Kanazawa, Shinsaku; Koike, Junichi; Sugiyama, Hisahiko; Xu, Can; Funahashi, Kimihiko; Boland, C Richard; Koi, Minoru; Hemmi, Hiromichi

    2010-02-01

    Most tumors of patients with Lynch syndrome and a fraction of sporadic colorectal cancers (CRCs) exhibit high levels of microsatellite instability (MSI) at mono- and dinucleotide repeat loci. A different type of instability, elevated microsatellite alterations at selected tetranucleotide repeats (EMAST) has been found in non-colonic cancers. Our previous study demonstrated that EMAST is common in sporadic CRC. Here, we focused on the relationships between EMAST and other genomic instability parameters or clinicopathological features in an unselected series of 88 sporadic CRCs. Of the tumors in the sample, 4 (4.5%) were MSI-high (MSI-H), 9 (10.2%) were MSI-low (MSI-L) and 75 (85.2%) were microsatellite stable. EMAST status was determined using 7 EMAST markers. Fifty-three (60.2%) tumors without MSI-H showed instability at >or=1 EMAST loci. All 4 MSI-H tumors showed instability at several EMAST loci. Instability profiles of MSI-H tumors at EMAST loci were more complex than those of non-MSI-H tumors. A tendency of positive association was observed between MSI-L and EMAST (P=0.023). The frequency of loss of heterozygosity (LOH) for the 14 loci in EMAST-positive tumors was significantly higher than negative tumors (P=0.048). Among the clinicopathological parameters, only tumor location at the distal colon was associated with EMAST-negative tumors (P=0.0084, one-tailed). A relatively higher frequency of well-differentiated adenocarcinomas was observed in EMAST tumors as opposed to non-EMAST tumors, though the survival rate was similar. These results suggest that overlapping mechanisms that cause MSI-L, EMAST and LOH in CRCs may exist.

  11. Conservation of human chromosome 13 polymorphic microsatellite (CA){sub n} repeats in chimpanzees

    SciTech Connect

    Deka, R.; Shriver, M.D.; Yu, L.M.

    1994-07-01

    Tandemly repeated (dC-dA){sub n} {center_dot} (dG-dT){sub n} sequences occur abundantly and are found in most eukaryotic genomes. To investigate the level of conservation of these repeat sequences in nonhuman primates, the authors have analyzed seven human chromosome 13 dinucleotide (CA){sub n} repeat loci in chimpanzees by DNA amplification using primers designed for analysis of human loci. Comparable levels of polymorphism at these loci in the two species, revealed by the number of alleles, heterozygosity, and allele sizes, suggest that the (CA){sub n} repeat arrays and their genomic locations are highly conserved. Even though the proportion of shared alleles between the two species varies enormously and the modal alleles are not the same, allelic lengths at each locus in the chimpanzees are detected within the bounds of the allele size range observed in humans. A similar observation has been noted in a limited number of gorillas and orangutans. Using a new measure of genetic distance that takes into account the size of alleles, they have compared the genetic distance between humans and chimpanzees. The genetic distance between these two species was found to be ninefold smaller than expected assuming there is no selection or mutational bias toward retention of (CA){sub n} repeat arrays. These findings suggest a functional significance for these microsatellite loci. 34 refs., 1 fig., 2 tabs.

  12. Multilocus variable-number of tandem repeat analysis (MLVA) for Clostridium tyrobutyricum strains isolated from cheese production environment.

    PubMed

    Nishihara, Masaharu; Takahashi, Hajime; Sudo, Tomoko; Kyoi, Daisuke; Kawahara, Toshio; Ikeuchi, Yoshihiro; Fujita, Takashi; Kuda, Takashi; Kimura, Bon; Yanahira, Shuichi

    2014-11-03

    Clostridium tyrobutyricum is a gram-positive spore-forming anaerobe that is considered as the main causative agent for late blowing in cheese due to butyric acid fermentation. In this study, multilocus variable-number of tandem repeat (VNTR) analysis (MLVA) for C. tyrobutyricum was developed to identify the source of contamination by C. tyrobutyricum spores in the cheese production environment. For each contig constructed from the results of a whole genome draft sequence of C. tyrobutyricum JCM11008(T) based on next-generation sequencing, VNTR loci that were effective for typing were searched using the Tandem Repeat Finder program. Five VNTR loci were amplified by polymerase chain reaction (PCR) to determine their number of repeats by sequencing, and MLVA was conducted. 25 strains of C. tyrobutyricum isolated from the environment, raw milk, and silage were classified into 18 MLVA types (DI=0.963). Of the C. tyrobutyricum strains isolated from raw milk, natural cheese, and blown processed cheese, strains with identical MLVA type were detected, which suggested that these strains might have shifted from natural cheese to blown processed cheese. MLVA could be an effective tool for monitoring contamination of natural cheese with C. tyrobutyricum in the processed cheese production environment because of its high discriminability, thereby allowing the analyst to trace the source of contamination.

  13. Multi-laboratory validation study of multilocus variable-number tandem repeat analysis (MLVA) for Salmonella enterica serovar Enteritidis, 2015.

    PubMed

    Peters, Tansy; Bertrand, Sophie; Björkman, Jonas T; Brandal, Lin T; Brown, Derek J; Erdõsi, Tímea; Heck, Max; Ibrahem, Salha; Johansson, Karin; Kornschober, Christian; Kotila, Saara M; Le Hello, Simon; Lienemann, Taru; Mattheus, Wesley; Nielsen, Eva Møller; Ragimbeau, Catherine; Rumore, Jillian; Sabol, Ashley; Torpdahl, Mia; Trees, Eija; Tuohy, Alma; de Pinna, Elizabeth

    2017-03-02

    Multilocus variable-number tandem repeat analysis (MLVA) is a rapid and reproducible typing method that is an important tool for investigation, as well as detection, of national and multinational outbreaks of a range of food-borne pathogens. Salmonella enterica serovar Enteritidis is the most common Salmonella serovar associated with human salmonellosis in the European Union/European Economic Area and North America. Fourteen laboratories from 13 countries in Europe and North America participated in a validation study for MLVA of S. Enteritidis targeting five loci. Following normalisation of fragment sizes using a set of reference strains, a blinded set of 24 strains with known allele sizes was analysed by each participant. The S. Enteritidis 5-loci MLVA protocol was shown to produce internationally comparable results as more than 90% of the participants reported less than 5% discrepant MLVA profiles. All 14 participating laboratories performed well, even those where experience with this typing method was limited. The raw fragment length data were consistent throughout, and the inter-laboratory validation helped to standardise the conversion of raw data to repeat numbers with at least two countries updating their internal procedures. However, differences in assigned MLVA profiles remain between well-established protocols and should be taken into account when exchanging data.

  14. Multi-laboratory validation study of multilocus variable-number tandem repeat analysis (MLVA) for Salmonella enterica serovar Enteritidis, 2015

    PubMed Central

    Peters, Tansy; Bertrand, Sophie; Björkman, Jonas T; Brandal, Lin T; Brown, Derek J; Erdõsi, Tímea; Heck, Max; Ibrahem, Salha; Johansson, Karin; Kornschober, Christian; Kotila, Saara M; Le Hello, Simon; Lienemann, Taru; Mattheus, Wesley; Nielsen, Eva Møller; Ragimbeau, Catherine; Rumore, Jillian; Sabol, Ashley; Torpdahl, Mia; Trees, Eija; Tuohy, Alma; de Pinna, Elizabeth

    2017-01-01

    Multilocus variable-number tandem repeat analysis (MLVA) is a rapid and reproducible typing method that is an important tool for investigation, as well as detection, of national and multinational outbreaks of a range of food-borne pathogens. Salmonella enterica serovar Enteritidis is the most common Salmonella serovar associated with human salmonellosis in the European Union/European Economic Area and North America. Fourteen laboratories from 13 countries in Europe and North America participated in a validation study for MLVA of S. Enteritidis targeting five loci. Following normalisation of fragment sizes using a set of reference strains, a blinded set of 24 strains with known allele sizes was analysed by each participant. The S. Enteritidis 5-loci MLVA protocol was shown to produce internationally comparable results as more than 90% of the participants reported less than 5% discrepant MLVA profiles. All 14 participating laboratories performed well, even those where experience with this typing method was limited. The raw fragment length data were consistent throughout, and the inter-laboratory validation helped to standardise the conversion of raw data to repeat numbers with at least two countries updating their internal procedures. However, differences in assigned MLVA profiles remain between well-established protocols and should be taken into account when exchanging data. PMID:28277220

  15. Target genes of microsatellite sequences in head and neck squamous cell carcinoma: mononucleotide repeats are not detected.

    PubMed

    Wang, Yimin; Liu, Xuejuan; Li, Yulin

    2012-09-10

    Microsatellite instability (MSI) is detected in a wide variety of tumors. It is thought that mismatch repair gene mutation or inactivation is the major cause of MSI. Microsatellite sequences are predominantly distributed in intergenic or intronic DNA. However, MSI is found in the exonic sequences of some genes, causing their inactivation. In this report, we searched GenBank for candidate genes containing potential MSI sequences in exonic regions. Twenty seven target genes were selected for MSI analysis. Instability was found in 70% of these genes (14/20) with head and neck squamous cell carcinoma (HNSCC). Interestingly, no instability was detected in mononucleotide repeats in genes or in intergenic sequences. We conclude that instability of mononucleotide repeats is a rare event in HNSCC. High MSI phenotype in young HNSCC patients is limited to noncoding regions only. MSI percentage in HNSCC tumor is closely related to the repeat type, repeat location and patient's age.

  16. Multi-locus variable number tandem repeat analysis of Vibrio cholerae isolates from 2012 to 2013 cholera outbreaks in Iran.

    PubMed

    Ranjbar, R; Sadeghy, J; Shokri Moghadam, M; Bakhshi, B

    2016-08-01

    Cholera remains to be an international threat, with high rates of illness and death. In 2012 and 2013, two cholera outbreak happened in Iran, affecting lots of people. Vibrio cholerae O1 was confirmed as the etiological agent. Source identification and controlling the spread of the cholera disease are two critical approaches in cholera outbreaks. In this study, thirty V. cholerae O1 isolates were selected and has been evaluated for antimicrobial resistant as well as molecular typing by multilocus variable-number tandem-repeat analysis (MLVA) method. Twenty-nine (97%) isolates were sero-grouped as El Tor (one isolate was classical) and 100% were related to Inaba serotype. All of the isolates were susceptible to ciprofloxacin, chloramphenicol, ampicillin and gentamicin. On the other hand, 60% of the isolates were MDR (resistant to 3 or more classes). There were three resistance patterns. The most prevalent pattern was resistance to streptomycin, erythromycin, trimethoprim-sulfamethoxazole, and tetracycline (ST-SXT-E-T) which was seen in 50% of isolates. Using MLVA method 14 MLVA types were identified. MLVA type 2 (5-7-7-16-15) accounted for 43% of isolates. Isolates with the same genotype often did not have the same antibiogram. Overall, the data indicate that the Iranian V. cholerae were MDR and clonaly related. Furthermore, the results of this study shows that MLVA can be used as useful method for V. cholerae genotyping in epidemiological investigations.

  17. High mutation rate of TPE repeats: a microsatellite in the putative transposase of the hobo element in Drosophila melanogaster.

    PubMed

    Souames, Sémi; Bonnivard, Eric; Bazin, Claude; Higuet, Dominique

    2003-11-01

    The hobo transposable element contains a polymorphic microsatellite sequence located in its coding region, the TPE repeats. Previous surveys of natural populations of Drosophila melanogaster have detected at least seven different hobo transposons. These natural populations are geographically structured with regard to TPE polymorphism, and a scenario has been proposed for the invasion process. Natural populations have recently been completely invaded by hobo elements with three TPE repeats. New elements then appeared by mutation, triggering a new stage of invasion by other elements. Since TPE polymorphism appeared over a short period of time, we focused on estimating the mutation rate of these TPE repeats. We used transgenic lines harboring three TPE and/or five TPE hobo elements that had been evolving for at least 16 generations to search for a new TPE repeat polymorphism. We detected three mutants, with four, seven, and eight TPE repeats, respectively. The estimated mutation rate of the TPE repeats is therefore higher than that of neutral microsatellites in D. melanogaster (4.2 x 10-4 versus 6.5 x 10-6). The role of the transposition mechanism and the particular structure of the TPE repeats of the hobo element in this increase in the mutation rate are discussed.

  18. Ruminant rhombencephalitis-associated Listeria monocytogenes alleles linked to a multilocus variable-number tandem-repeat analysis complex.

    PubMed

    Balandyté, Lina; Brodard, Isabelle; Frey, Joachim; Oevermann, Anna; Abril, Carlos

    2011-12-01

    Listeria monocytogenes is among the most important food-borne pathogens and is well adapted to persist in the environment. To gain insight into the genetic relatedness and potential virulence of L. monocytogenes strains causing central nervous system (CNS) infections, we used multilocus variable-number tandem-repeat analysis (MLVA) to subtype 183 L. monocytogenes isolates, most from ruminant rhombencephalitis and some from human patients, food, and the environment. Allelic-profile-based comparisons grouped L. monocytogenes strains mainly into three clonal complexes and linked single-locus variants (SLVs). Clonal complex A essentially consisted of isolates from human and ruminant brain samples. All but one rhombencephalitis isolate from cattle were located in clonal complex A. In contrast, food and environmental isolates mainly clustered into clonal complex C, and none was classified as clonal complex A. Isolates of the two main clonal complexes (A and C) obtained by MLVA were analyzed by PCR for the presence of 11 virulence-associated genes (prfA, actA, inlA, inlB, inlC, inlD, inlE, inlF, inlG, inlJ, and inlC2H). Virulence gene analysis revealed significant differences in the actA, inlF, inlG, and inlJ allelic profiles between clinical isolates (complex A) and nonclinical isolates (complex C). The association of particular alleles of actA, inlF, and newly described alleles of inlJ with isolates from CNS infections (particularly rhombencephalitis) suggests that these virulence genes participate in neurovirulence of L. monocytogenes. The overall absence of inlG in clinical complex A and its presence in complex C isolates suggests that the InlG protein is more relevant for the survival of L. monocytogenes in the environment.

  19. Characterisation of Brucella suis isolates from Southeast Europe by multi-locus variable-number tandem repeat analysis.

    PubMed

    Duvnjak, Sanja; Račić, Ivana; Špičić, Silvio; Zdelar-Tuk, Maja; Reil, Irena; Cvetnić, Željko

    2015-10-22

    Porcine brucellosis is a common bacterial zoonosis which can cause significant financial losses. Its diverse and often complicated factors have hampered efforts to control disease spread. The aim of the study was to assess the epidemiological situation of porcine brucellosis primarily in Croatia and its relationship to genotypes present in other, mostly European countries. One hundred and seven Brucella suis strains isolated from swine, hares, cattle, humans, wild hares, a wild boar and a mare originating mainly from Croatia (112), but also a few from Slovenia, Bosnia and Herzegovina, Serbia and Macedonia (15) were tested using classical microbiological testing, Bruce-ladder, RFLP, Multiplex-suis and genotyped using multi-locus variable-number tandem repeat analysis (MLVA). We determined 43 Brucella suis genotypes. Strains were grouped according to phylogenetic and geographic relationships, revealing both regional specificity and uniqueness and suggesting possible sources and modes of spread among animals. Our study also confirmed problems with Bruce19 locus that may hinder comparisons of new types with those in the international database. Forty-one novel genotypes were identified and deposited into the international database. Our study supports the idea of wild animals as a source of disease in domestic animals and also gives evidence to hypothesis of cross-border animal trafficking between former Yugoslavian countries. It also highlights the need to expand such research across more of southeast Europe, especially to countries with poorer social and economical situation in order to prevent a realistic outbreak and for better understanding of the biology of this pathogen.

  20. Parallel G-Quadruplexes Formed by Guanine-Rich Microsatellite Repeats Inhibit Human Topoisomerase I.

    PubMed

    Ogloblina, A M; Bannikova, V A; Khristich, A N; Oretskaya, T S; Yakubovskaya, M G; Dolinnaya, N G

    2015-08-01

    Using UV and CD spectroscopy, we studied the thermodynamic stability and folding topology of G-quadruplexes (G4), formed by G-rich fragments in human microsatellites that differ in the number of guanosines within the repeating unit. The oligonucleotides d(GGGT)4 and d(GGT)4 were shown to form propeller-type parallel-stranded intramolecular G-quadruplexes. The G4 melting temperature is dramatically decreased (by more than 45°C) in the transition from the tri-G-tetrad to the bi-G-tetrad structure. d(GT)n-repeats do not form perfect G-quadruplexes (one-G-tetrad); folded G4-like conformation is not stable at room temperature and is not stabilized by monovalent metal ions. The minimum concentration of K+ that promotes quadruplex folding of d(GGT)4 was found to depend on the supporting Na+ concentration. It was demonstrated for the first time that the complementary regions flanking G4-motifs (as in d(CACTGG-CC-(GGGT)4-TA-CCAGTG)) cannot form a double helix in the case of a parallel G4 due to the steric remoteness, but instead destabilize the structure. Additionally, we investigated the effect of the described oligonucleotides on the activity of topoisomerase I, one of the key cell enzymes, with a focus on the relationship between the stability of the formed quadruplexes and the inhibition degree of the enzyme. The most active inhibitor with IC50 = 0.08 µM was the oligonucleotide d(CACTGG-CC-(GGGT)4-TA-CCAGTG), whose flanking G4-motif sequences reduced the extreme stability of G-quadruplex formed by d(GGGT)4.

  1. Amplification of microsatellite repeat motifs is associated with the evolutionary differentiation and heterochromatinization of sex chromosomes in Sauropsida.

    PubMed

    Matsubara, Kazumi; O'Meally, Denis; Azad, Bhumika; Georges, Arthur; Sarre, Stephen D; Graves, Jennifer A Marshall; Matsuda, Yoichi; Ezaz, Tariq

    2016-03-01

    The sex chromosomes in Sauropsida (reptiles and birds) have evolved independently many times. They show astonishing diversity in morphology ranging from cryptic to highly differentiated sex chromosomes with male (XX/XY) and female heterogamety (ZZ/ZW). Comparing such diverse sex chromosome systems thus provides unparalleled opportunities to capture evolution of morphologically differentiated sex chromosomes in action. Here, we describe chromosomal mapping of 18 microsatellite repeat motifs in eight species of Sauropsida. More than two microsatellite repeat motifs were amplified on the sex-specific chromosome, W or Y, in five species (Bassiana duperreyi, Aprasia parapulchella, Notechis scutatus, Chelodina longicollis, and Gallus gallus) of which the sex-specific chromosomes were heteromorphic and heterochromatic. Motifs (AAGG)n and (ATCC)n were amplified on the W chromosome of Pogona vitticeps and the Y chromosome of Emydura macquarii, respectively. By contrast, no motifs were amplified on the W chromosome of Christinus marmoratus, which is not much differentiated from the Z chromosome. Taken together with previously published studies, our results suggest that the amplification of microsatellite repeats is tightly associated with the differentiation and heterochromatinization of sex-specific chromosomes in sauropsids as well as in other taxa. Although some motifs were common between the sex-specific chromosomes of multiple species, no correlation was observed between this commonality and the species phylogeny. Furthermore, comparative analysis of sex chromosome homology and chromosomal distribution of microsatellite repeats between two closely related chelid turtles, C. longicollis and E. macquarii, identified different ancestry and differentiation history. These suggest multiple evolutions of sex chromosomes in the Sauropsida.

  2. Multilocus Variable Number of Tandem Repeat Analysis Reveals Multiple Introductions in Spain of Xanthomonas arboricola pv. pruni, the Causal Agent of Bacterial Spot Disease of Stone Fruits and Almond.

    PubMed

    López-Soriano, Pablo; Boyer, Karine; Cesbron, Sophie; Morente, María Clara; Peñalver, Javier; Palacio-Bielsa, Ana; Vernière, Christian; López, María M; Pruvost, Olivier

    Xanthomonas arboricola pv. pruni is the causal agent of the bacterial spot disease of stone fruits, almond and some ornamental Prunus species. In Spain it was first detected in 2002 and since then, several outbreaks have occurred in different regions affecting mainly Japanese plum, peach and almond, both in commercial orchards and nurseries. As the origin of the introduction(s) was unknown, we have assessed the genetic diversity of 239 X. arboricola pv. pruni strains collected from 11 Spanish provinces from 2002 to 2013 and 25 reference strains from international collections. We have developed an optimized multilocus variable number of tandem repeat analysis (MLVA) scheme targeting 18 microsatellites and five minisatellites. A high discriminatory power was achieved since almost 50% of the Spanish strains were distinguishable, confirming the usefulness of this genotyping technique at small spatio-temporal scales. Spanish strains grouped in 18 genetic clusters (conservatively delineated so that each cluster contained haplotype networks linked by up to quadruple-locus variations). Furthermore, pairwise comparisons among populations from different provinces showed a strong genetic differentiation. Our results suggest multiple introductions of this pathogen in Spain and redistribution through contaminated nursery propagative plant material.

  3. Multilocus Variable Number of Tandem Repeat Analysis Reveals Multiple Introductions in Spain of Xanthomonas arboricola pv. pruni, the Causal Agent of Bacterial Spot Disease of Stone Fruits and Almond

    PubMed Central

    López-Soriano, Pablo; Boyer, Karine; Cesbron, Sophie; Morente, María Clara; Peñalver, Javier; Palacio-Bielsa, Ana; Vernière, Christian; López, María M.; Pruvost, Olivier

    2016-01-01

    Xanthomonas arboricola pv. pruni is the causal agent of the bacterial spot disease of stone fruits, almond and some ornamental Prunus species. In Spain it was first detected in 2002 and since then, several outbreaks have occurred in different regions affecting mainly Japanese plum, peach and almond, both in commercial orchards and nurseries. As the origin of the introduction(s) was unknown, we have assessed the genetic diversity of 239 X. arboricola pv. pruni strains collected from 11 Spanish provinces from 2002 to 2013 and 25 reference strains from international collections. We have developed an optimized multilocus variable number of tandem repeat analysis (MLVA) scheme targeting 18 microsatellites and five minisatellites. A high discriminatory power was achieved since almost 50% of the Spanish strains were distinguishable, confirming the usefulness of this genotyping technique at small spatio-temporal scales. Spanish strains grouped in 18 genetic clusters (conservatively delineated so that each cluster contained haplotype networks linked by up to quadruple-locus variations). Furthermore, pairwise comparisons among populations from different provinces showed a strong genetic differentiation. Our results suggest multiple introductions of this pathogen in Spain and redistribution through contaminated nursery propagative plant material. PMID:27669415

  4. Chromosomal distribution of microsatellite repeats in Amazon cichlids genome (Pisces, Cichlidae).

    PubMed

    Schneider, Carlos Henrique; Gross, Maria Claudia; Terencio, Maria Leandra; de Tavares, Édika Sabrina Girão Mitozo; Martins, Cesar; Feldberg, Eliana

    2015-01-01

    Fish of the family Cichlidae are recognized as an excellent model for evolutionary studies because of their morphological and behavioral adaptations to a wide diversity of explored ecological niches. In addition, the family has a dynamic genome with variable structure, composition and karyotype organization. Microsatellites represent the most dynamic genomic component and a better understanding of their organization may help clarify the role of repetitive DNA elements in the mechanisms of chromosomal evolution. Thus, in this study, microsatellite sequences were mapped in the chromosomes of Cichla monoculus Agassiz, 1831, Pterophyllum scalare Schultze, 1823, and Symphysodon discus Heckel, 1840. Four microsatellites demonstrated positive results in the genome of Cichla monoculus and Symphysodon discus, and five demonstrated positive results in the genome of Pterophyllum scalare. In most cases, the microsatellite was dispersed in the chromosome with conspicuous markings in the centromeric or telomeric regions, which suggests that sequences contribute to chromosome structure and may have played a role in the evolution of this fish family. The comparative genome mapping data presented here provide novel information on the structure and organization of the repetitive DNA region of the cichlid genome and contribute to a better understanding of this fish family's genome.

  5. Chromosomal distribution of microsatellite repeats in Amazon cichlids genome (Pisces, Cichlidae)

    PubMed Central

    Schneider, Carlos Henrique; Gross, Maria Claudia; Terencio, Maria Leandra; de Tavares, Édika Sabrina Girão Mitozo; Martins, Cesar; Feldberg, Eliana

    2015-01-01

    Abstract Fish of the family Cichlidae are recognized as an excellent model for evolutionary studies because of their morphological and behavioral adaptations to a wide diversity of explored ecological niches. In addition, the family has a dynamic genome with variable structure, composition and karyotype organization. Microsatellites represent the most dynamic genomic component and a better understanding of their organization may help clarify the role of repetitive DNA elements in the mechanisms of chromosomal evolution. Thus, in this study, microsatellite sequences were mapped in the chromosomes of Cichla monoculus Agassiz, 1831, Pterophyllum scalare Schultze, 1823, and Symphysodon discus Heckel, 1840. Four microsatellites demonstrated positive results in the genome of Cichla monoculus and Symphysodon discus, and five demonstrated positive results in the genome of Pterophyllum scalare. In most cases, the microsatellite was dispersed in the chromosome with conspicuous markings in the centromeric or telomeric regions, which suggests that sequences contribute to chromosome structure and may have played a role in the evolution of this fish family. The comparative genome mapping data presented here provide novel information on the structure and organization of the repetitive DNA region of the cichlid genome and contribute to a better understanding of this fish family’s genome. PMID:26753076

  6. Multi-locus variable number tandem repeat analysis for Clostridium botulinum type B isolates in Japan: comparison with other isolates and genotyping methods.

    PubMed

    Umeda, Kaoru; Wada, Takayuki; Kohda, Tomoko; Kozaki, Shunji

    2013-06-01

    Clostridium botulinum produces botulinum neurotoxin (BoNT) and causes botulism in humans and animals. Recently, 15-loci multi-locus variable number tandem repeat analysis (MLVA) for C. botulinum was developed for high-resolution and inter-lab comparative genotyping. This study examines the relation between MLVA and other genotyping methods such as pulsed-field gel electrophoresis (PFGE), multi-locus sequence typing (MLST), BoNT/B subtyping and bont/b gene location to evaluate MLVA as a method applicable to the genetic markers for C. botulinum type B. Japanese isolates were genotyped using MLVA and were compared with strains from other sources reported previously. Results show that the discriminatory power of MLVA was comparable to that of PFGE and higher than that of MLST. The topology of the minimum spanning tree (MST) constructed using MLVA data was very consistent with the phylogenetic classifications of PFGE and MLST. The MST topology also represented genetic diversity between the strains possessing bont/b gene on chromosomes and plasmids. Some Japanese isolates including those associated with infant botulism were inferred to be related to isolates of Europe origin from MLVA genotyping results. The MLVA scheme used for this study is apparently useful not only for high-resolution molecular typing, but also for phylogenetic characterization of C. botulinum type B.

  7. Multi-Locus Variable Number of Tandem Repeat Analysis for Rapid and Accurate Typing of Virulent Multidrug Resistant Escherichia coli Clones

    PubMed Central

    Naseer, Umaer; Olsson-Liljequist, Barbro E.; Woodford, Neil; Dhanji, Hiran; Cantón, Rafael; Sundsfjord, Arnfinn; Lindstedt, Bjørn-Arne

    2012-01-01

    One hundred E. coli isolates from Norway (n = 37), Sweden (n = 24), UK (n = 20) and Spain (n = 19), producing CTX-M-type - (n = 84), or SHV-12 (n = 4) extended spectrum β-lactamases, or the plasmid mediated AmpC, CMY-2 (n = 12), were typed using multi-locus sequence typing (MLST) and multi-locus variable number of tandem repeat analysis (MLVA). Isolates clustered into 33 Sequence Types (STs) and 14 Sequence Type Complexes (STCs), and 58 MLVA-Types (MTs) and 25 different MLVA-Type Complexes (MTCs). A strong agreement between the MLST profile and MLVA typing results was observed, in which all ST131-isolates (n = 39) and most of the STC-648 (n = 10), STC-38 (n = 9), STC-10 (n = 9), STC-405 (n = 8) and STC-23 (n = 6) isolates were clustered distinctly into MTC-29, -36, -20, -14, -10 and -39, respectively. MLVA is a rapid and accurate tool for genotyping isolates of globally disseminated virulent multidrug resistant E. coli lineages, including ST131. PMID:22859970

  8. Unstable microsatellite repeats facilitate rapid evolution of coding and regulatory sequences.

    PubMed

    Jansen, A; Gemayel, R; Verstrepen, K J

    2012-01-01

    Tandem repeats are intrinsically highly variable sequences since repeat units are often lost or gained during replication or following unequal recombination events. Because of their low complexity and their instability, these repeats, which are also called satellite repeats, are often considered to be useless 'junk' DNA. However, recent findings show that tandem repeats are frequently found within promoters of stress-induced genes and within the coding regions of genes encoding cell-surface and regulatory proteins. Interestingly, frequent changes in these repeats often confer phenotypic variability. Examples include variation in the microbial cell surface, rapid tuning of internal molecular clocks in flies, and enhanced morphological plasticity in mammals. This suggests that instead of being useless junk DNA, some variable tandem repeats are useful functional elements that confer 'evolvability', facilitating swift evolution and rapid adaptation to changing environments. Since changes in repeats are frequent and reversible, repeats provide a unique type of mutation that bridges the gap between rare genetic mutations, such as single nucleotide polymorphisms, and highly unstable but reversible epigenetic inheritance.

  9. Relatedness of Vibrio cholerae O1/O139 Isolates from Patients and Their Household Contacts, Determined by Multilocus Variable-Number Tandem-Repeat Analysis▿ ‡

    PubMed Central

    Kendall, Emily A.; Chowdhury, Fahima; Begum, Yasmin; Khan, Ashraful I.; Li, Shan; Thierer, James H.; Bailey, Jason; Kreisel, Kristen; Tacket, Carol O.; LaRocque, Regina C.; Harris, Jason B.; Ryan, Edward T.; Qadri, Firdausi; Calderwood, Stephen B.; Stine, O. Colin

    2010-01-01

    The genetic relatedness of Vibrio cholerae O1/O139 isolates obtained from 100 patients and 146 of their household contacts in Dhaka, Bangladesh, between 2002 and 2005 was assessed by multilocus variable-number tandem-repeat analysis. Isolate genotypes were analyzed at five loci containing tandem repeats. Across the population, as well as within households, isolates with identical genotypes were clustered in time. Isolates from individuals within the same household were more likely to have similar or identical genotypes than were isolates from different households, but even within a household, isolates from different individuals often had different genotypes. When household contacts were sampled regularly for 3 weeks after the illness of the household index patient, isolates with genotypes related to the index patient appeared in contacts, on average, ∼3 days after the index patient, while isolates with unrelated genotypes appeared in contacts ∼6 days after. Limited data revealed that multiple isolates from the same individual collected within days of each other or even from a single stool sample may have identical, similar, or unrelated genotypes as well. Our results demonstrate that genetically related V. cholerae strains cluster in local outbreaks but also suggest that multiple distinct strains of V. cholerae O1 may circulate simultaneously within a household. PMID:20585059

  10. Tissue identity testing of cancer by short tandem repeat polymorphism: pitfalls of interpretation in the presence of microsatellite instability.

    PubMed

    Much, Melissa; Buza, Natalia; Hui, Pei

    2014-03-01

    Tissue identity testing by short tandem repeat (STR) polymorphism offers discriminating power in resolving tissue mix-up or contamination. However, one caveat is the presence of microsatellite unstable tumors, in which genetic alterations may drastically change the STR wild-type polymorphism leading to unexpected allelic discordance. We examined how tissue identity testing results can be altered by the presence of microsatellite instability (MSI). Eleven cases of MSI-unstable (9 intestinal and 2 endometrial adenocarcinomas) and 10 cases of MSI-stable tumors (all colorectal adenocarcinomas) were included. All had been previously tested by polymerase chain reaction testing at 5 National Cancer Institute (NCI) recommended MSI loci and/or immunohistochemistry for DNA mismatch repair proteins (MLH1, MSH2, MSH6, and PMS2). Tissue identity testing targeting 15 STR loci was performed using AmpF/STR Identifiler Amplification. Ten of 11 MSI-unstable tumors demonstrated novel alleles at 5 to 12 STR loci per case and frequently with 3 or more allelic peaks. However, all affected loci showed identifiable germline allele(s) in MSI-high tumors. A wild-type allelic profile was seen in 7 of 10 MSI-stable tumors. In the remaining 3 cases, isolated novel alleles were present at a unique single locus in addition to germline alleles. Loss of heterozygosity was observed frequently in both MSI-stable (6/11 cases) and MSI-unstable tumors (8/10 cases). In conclusion, MSI may significantly alter the wild-type allelic polymorphism, leading to potential interpretation errors of STR genotyping. Careful examination of the STR allelic pattern, high index of suspicion, and follow-up MSI testing are crucial to avoid erroneous conclusions and subsequent clinical and legal consequences.

  11. Regional bias of intratumoral genetic heterogeneity of nucleotide repeats in colon cancers with microsatellite instability.

    PubMed

    Choi, Youn Jin; Kim, Min Sung; An, Chang Hyeok; Yoo, Nam Jin; Lee, Sug Hyung

    2014-10-01

    Intratumoral heterogeneity (ITH) may produce regional biases in genotype and phenotype evaluation in a single tumor and may impede proper cancer diagnosis. To evaluate the extent of ITH in colorectal cancer (CRC) with microsatellite instability (MSI), we obtained 4-7 biopsies from 39 CRCs followed by MSI analysis either using the Bethesda MSI evaluation system or Promega system with 5 mononucleotide markers. We found decreased prevalence of MSI (+) by the Promega system compared to the Bethesda system. The overall discordance between the two systems was 54 %. In contrast to the previous studies that had shown discordance only in low MSI (MSI-L), our results showed the discordance not only in MSI-L, but also in high MSI (MSI-H) cases. Among the MSI (+) CRCs, ITH of MSI status was identified in 41.7 % of CRC by the Bethesda system and 22.2 % by the Promega system. In terms of MSI markers, the ITH originated from dinucleotide markers in most cases (69 %), but it originated from mononucleotide markers (31 %) as well. Pooling of DNA from a regional biopsy with MSI (+) with additional biopsies from stable MSI (MSS) showed that this approach was beneficial to increase the sensitivity of MSI detection. Our results indicate that ITH of MSI phenotype by the Bethesda system is more overestimated than previously identified. However, because there was considerable ITH of MSI subtypes and markers even by the Promega system, our data suggest that analysis of MSI status in multiple regional biopsies is needed for a better evaluation of MSI status in CRC.

  12. Analysis of simple sequence repeats in the Gaeumannomyces graminis var. tritici genome and the development of microsatellite markers.

    PubMed

    Li, Wei; Feng, Yanxia; Sun, Haiyan; Deng, Yuanyu; Yu, Hanshou; Chen, Huaigu

    2014-11-01

    Understanding the genetic structure of Gaeumannomyces graminis var. tritici is essential for the establishment of efficient disease control strategies. It is becoming clear that microsatellites, or simple sequence repeats (SSRs), play an important role in genome organization and phenotypic diversity, and are a large source of genetic markers for population genetics and meiotic maps. In this study, we examined the G. graminis var. tritici genome (1) to analyze its pattern of SSRs, (2) to compare it with other plant pathogenic filamentous fungi, such as Magnaporthe oryzae and M. poae, and (3) to identify new polymorphic SSR markers for genetic diversity. The G. graminis var. tritici genome was rich in SSRs; a total 13,650 SSRs have been identified with mononucleotides being the most common motifs. In coding regions, the densities of tri- and hexanucleotides were significantly higher than in noncoding regions. The di-, tri-, tetra, penta, and hexanucleotide repeats in the G. graminis var. tritici genome were more abundant than the same repeats in M. oryzae and M. poae. From 115 devised primers, 39 SSRs are polymorphic with G. graminis var. tritici isolates, and 8 primers were randomly selected to analyze 116 isolates from China. The number of alleles varied from 2 to 7 and the expected heterozygosity (He) from 0.499 to 0.837. In conclusion, SSRs developed in this study were highly polymorphic, and our analysis indicated that G. graminis var. tritici is a species with high genetic diversity. The results provide a pioneering report for several applications, such as the assessment of population structure and genetic diversity of G. graminis var. tritici.

  13. FMR1 CGG repeat distribution and linked microsatellite-SNP haplotypes in normal Mexican Mestizo and indigenous populations.

    PubMed

    Felix-López, Xóchitl Adriana; Argüello-García, Raúl; Cerda-Flores, Ricardo M; Peñaloza-Espinoza, Rosenda I; Buentello-Malo, Leonor; Estrada-Mena, Francisco Javier; Ramos-Kuri, Manuel; Gómez, Fabio Salamanca; Arenas-Aranda, Diego Julio

    2006-10-01

    The (CGG)n repeat size distribution in the FMR1 gene was studied in healthy individuals: 80 X chromosomes of Mexican Mestizos from Mexico City and 33 X chromosomes of Mexican Amerindians from three indigenous communities (Purepechas, Nahuas, and Tzeltales), along with alleles and haplotypes defined by two microsatellite polymorphic markers (DXS548 and FRAXAC1) and two single nucleotide polymorphisms (FMRA and FMRB). Genetic frequencies of Mestizo and Amerindian subpopulations were statistically similar in almost all cases and thus were considered one population for comparisons with other populations. Sixteen (CGG)n alleles in the 17-38 size range were observed, and the most common were the 25 (38.0%), 26 (28.3%), and 24 (12.3%) repeat alleles. This pattern differs from most other populations reported, but a closer relation to Amerindian, European, and African populations was found, as expected from the historical admixture that gave rise to Mexican Mestizos. The results of the CA repeats analysis at DXS548-FRAXAC1 were restricted to nine haplotypes, of which haplotypes 7-4 (52.2%), 8-4 (23.8%), and 7-3 (11.5%) were predominant. The modal haplotype 7-4, instead of the nearly universal haplotype 7-3, had been reported exclusively in Eastern Asian populations. Likewise, only seven different FRAXAC1-FMRA-FMRB haplotypes were observed, including five novel haplotypes (3TA, 4TA, 3 - A, 4 - A, and 5 - A), compared with Caucasians. Of these, haplotypes - A (78.7%) and 3 - A (13.2%) were the most common in the Mexican population. These data suggest a singular but relatively low genetic diversity at FMR1 in the studied Mexican populations that may be related to the recent origin of Mestizos and the low admixture rate of Amerindians.

  14. Survey and analysis of simple sequence repeats in the Laccaria bicolor genome, with development of microsatellite markers

    SciTech Connect

    Labbe, Jessy L; Murat, Claude; Morin, Emmanuelle; Le Tacon, F; Martin, Francis

    2011-01-01

    It is becoming clear that simple sequence repeats (SSRs) play a significant role in fungal genome organization, and they are a large source of genetic markers for population genetics and meiotic maps. We identified SSRs in the Laccaria bicolor genome by in silico survey and analyzed their distribution in the different genomic regions. We also compared the abundance and distribution of SSRs in L. bicolor with those of the following fungal genomes: Phanerochaete chrysosporium, Coprinopsis cinerea, Ustilago maydis, Cryptococcus neoformans, Aspergillus nidulans, Magnaporthe grisea, Neurospora crassa and Saccharomyces cerevisiae. Using the MISA computer program, we detected 277,062 SSRs in the L. bicolor genome representing 8% of the assembled genomic sequence. Among the analyzed basidiomycetes, L. bicolor exhibited the highest SSR density although no correlation between relative abundance and the genome sizes was observed. In most genomes the short motifs (mono- to trinucleotides) were more abundant than the longer repeated SSRs. Generally, in each organism, the occurrence, relative abundance, and relative density of SSRs decreased as the repeat unit increased. Furthermore, each organism had its own common and longest SSRs. In the L. bicolor genome, most of the SSRs were located in intergenic regions (73.3%) and the highest SSR density was observed in transposable elements (TEs; 6,706 SSRs/Mb). However, 81% of the protein-coding genes contained SSRs in their exons, suggesting that SSR polymorphism may alter gene phenotypes. Within a L. bicolor offspring, sequence polymorphism of 78 SSRs was mainly detected in non-TE intergenic regions. Unlike previously developed microsatellite markers, these new ones are spread throughout the genome; these markers could have immediate applications in population genetics.

  15. High-resolution genetic fingerprinting of European strains of Anaplasma phagocytophilum by use of multilocus variable-number tandem-repeat analysis.

    PubMed

    Bown, Kevin J; Lambin, Xavier; Ogden, Nicholas H; Petrovec, Miroslav; Shaw, Susan E; Woldehiwet, Zerai; Birtles, Richard J

    2007-06-01

    Anaplasma phagocytophilum is a widely distributed tick-borne pathogen of humans, livestock, and companion animals. We used in silico methods to identify 10 variable-number tandem-repeat (VNTR) loci within the genome sequence of the A. phagocytophilum HZ strain and used these data to develop a multilocus VNTR-based typing scheme for the species. Having confirmed the stability of four of the loci in replicates of the A. phagocytophilum strain that had been subjected to different numbers of passages through cell cocultures in vitro, we then used this typing scheme to discriminate between 20 A. phagocytophilum strains of diverse geographical and host provenances. Extensive diversity was found at each of the four loci studied, with total allele numbers ranging from 13 to 18 and Hunter-Gaston discriminatory index values ranging from 0.93 to 0.99. Only 2 of the 20 strains examined shared alleles at all four loci. The discriminatory power of VNTR analysis was found to be greater than that of either partial msp4 or 16S rRNA gene sequence comparison. The extremely high sensitivity of this novel approach to the genetic fingerprinting of A. phagocytophilum strains should serve well in molecular epidemiological studies of infection transmission, particularly when fine-scale strain delineation is required.

  16. Multilocus microsatellite typing revealed high genetic variability of Leishmania donovani strains isolated during and after a Kala-azar epidemic in Libo Kemkem district, northwest Ethiopia.

    PubMed

    Gelanew, Tesfaye; Cruz, Israel; Kuhls, Katrin; Alvar, Jorge; Cañavate, Carmen; Hailu, Asrat; Schönian, Gabriele

    2011-06-01

    In 2004, an outbreak of kala-azar (KA) occurred for the first time in Libo Kemkem district, in the highland area of northwest Ethiopia. In order to track the possible origins of the outbreak parasites, we have investigated 19 strains of Leishmania donovani that were collected during (n = 6) and after (n = 13) the outbreak by using 14 highly polymorphic microsatellite markers. Unique microsatellite profiles were obtained for all strains from Libo Kemkem. When compared to those of L. donovani strains from different Ethiopian, Kenyan and Sudanese foci, by genetic distance and Bayesian clustering model analyses, most strains from Libo Kemkem grouped with strains from: (i) Humera and Metema in the lowlands and Belessa in the highland of Ethiopia, and (ii) Sudan, at different hierarchal levels. The strains from Libo Kemkem district were assigned at least to three genetically distinct clusters (A, B1 and B2) of which only one, cluster B2, consisted exclusively of strains from Libo Kemkem. The fact that most of the outbreak strains were found to be related to strains from well-known KA foci in northwest Ethiopia and Sudan might suggest multiple introductions of L. donovani strains from these foci into Libo Kemkem district.

  17. Chromosomal mapping of microsatellite repeats in the rock bream fish Oplegnathus fasciatus, with emphasis of their distribution in the neo-Y chromosome.

    PubMed

    Xu, Dongdong; Lou, Bao; Bertollo, Luiz Antonio Carlos; Cioffi, Marcelo de Bello

    2013-03-19

    Despite the theoretical and experimental progress, our understanding on sex chromosome differentiation is still diagrammatic. The accumulation of repetitive DNA sequences is believed to occur in early stages of such differentiation. As fish species present a wide range of sex chromosome systems they are excellent models to examine the differentiation of these chromosomes. In the present study, the chromosomal distribution of 9 mono-, di- and tri-nucleotide microsatellites were analyzed using fluorescence in situ hybrization (FISH) in rock bream fish (Oplegnathus fasciatus), which is characterized by an X1X2Y sex chromosome system. Generally, the males and females exhibited the same autosomal pattern of distribution for a specific microsatellite probe. The male specific Y chromosome displays a specific amount of distinct microsatellites repeats along both arms. However, the accumulation of these repetitive sequences was not accompanied by a huge heterochromatinization process. The present data provide new insights into the chromosomal constitution of the multiple sex chromosomes and allow further investigations on the true role of the microsatellite repeats in the differentiation process of this sex system.

  18. Determination of microsatellite repeats in the human thyroid peroxidase (TPOX) gene using an automated gene analysis system with nanoscale engineered biomagnetite.

    PubMed

    Nakagawa, Takahito; Maruyama, Kohei; Takeyama, Haruko; Matsunaga, Tadashi

    2007-04-15

    The number of repeat in the microsatellite region (AATG)(5-14) of the human thyroid peroxidase gene (TOPX) was determined using an automated DNA analysis system with nano-scale engineered biomagnetite. Thermal melting curve analysis of DNA duplexes on biomagnetite indicated that shorter repeat sequences (less than 9 repeats) were easily discriminated. However, it was difficult to determine the number of repeats at more than nine. In order to improve the selectivity of this method for the longer repeats, a "double probe hybridization assay" was performed in which an intermediate probe was used to replace a target repeat sequence having more than 9 repeats with a shorter sequence possessing less than 9 repeats. Thermal probe melting curve analyses and Tm determination confirmed that the target with 10 repeats was converted to 5 repeats, 11 repeats converted to 4 and 12 to 3, respectively. Furthermore, rapid determination of repeat numbers was possible by measuring fluorescence intensities obtained by probe dissociation at 56 and 66 degrees C, and 40, 60 and 80 degrees C for signal normalization.

  19. 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.

  20. Analysis of Clostridium botulinum Serotype E Strains by Using Multilocus Sequence Typing, Amplified Fragment Length Polymorphism, Variable-Number Tandem-Repeat Analysis, and Botulinum Neurotoxin Gene Sequencing▿

    PubMed Central

    Macdonald, Thomas E.; Helma, Charles H.; Shou, Yulin; Valdez, Yolanda E.; Ticknor, Lawrence O.; Foley, Brian T.; Davis, Stephen W.; Hannett, George E.; Kelly-Cirino, Cassandra D.; Barash, Jason R.; Arnon, Stephen S.; Lindström, Miia; Korkeala, Hannu; Smith, Leonard A.; Smith, Theresa J.; Hill, Karen K.

    2011-01-01

    A total of 41 Clostridium botulinum serotype E strains from different geographic regions, including Canada, Denmark, Finland, France, Greenland, Japan, and the United States, were compared by multilocus sequence typing (MLST), amplified fragment length polymorphism (AFLP) analysis, variable-number tandem-repeat (VNTR) analysis, and botulinum neurotoxin (bont) E gene sequencing. The strains, representing environmental, food-borne, and infant botulism samples collected from 1932 to 2007, were analyzed to compare serotype E strains from different geographic regions and types of botulism and to determine whether each of the strains contained the transposon-associated recombinase rarA, involved with bont/E insertion. MLST examination using 15 genes clustered the strains into several clades, with most members within a cluster sharing the same BoNT/E subtype (BoNT/E1, E2, E3, or E6). Sequencing of the bont/E gene identified two new variants (E7, E8) that showed regions of recombination with other E subtypes. The AFLP dendrogram clustered the 41 strains similarly to the MLST dendrogram. Strains that could not be differentiated by AFLP, MLST, or bont gene sequencing were further examined using three VNTR regions. Both intact and split rarA genes were amplified by PCR in each of the strains, and their identities were confirmed in 11 strains by amplicon sequencing. The findings suggest that (i) the C. botulinum serotype E strains result from the targeted insertion of the bont/E gene into genetically conserved bacteria and (ii) recombination events (not random mutations) within bont/E result in toxin variants or subtypes within strains. PMID:22003031

  1. Virulence Gene Profile and Multilocus Variable-Number Tandem-Repeat Analysis (MLVA) of Enteroinvasive Escherichia coli (EIEC) Isolates From Patients With Diarrhea in Kerman, Iran

    PubMed Central

    Hosseini Nave, Hossein; Mansouri, Shahla; Taati Moghadam, Majid; Moradi, Mohammad

    2016-01-01

    Background Enteroinvasive Escherichia coli (EIEC) isolates cause dysentery in humans. Several virulence factors associated with EIEC pathogenesis have been characterized. Multilocus variable-number tandem-repeat analysis (MLVA) is a PCR-based method that has been used for genotyping bacterial pathogens. Objectives The aim of this study was to investigate the distribution of virulence factor genes in EIEC isolates from patients with diarrhea in Kerman, Iran, as well as the genetic relationships between these isolates. Patients and Methods A total of 620 diarrheic stool samples were collected from patients attending two hospitals in Kerman from June 2013 to August 2014. All isolates were confirmed as EIEC by PCR for the ipaH gene. The EIEC isolates were evaluated by PCR for the presence of nine virulence genes (ial, set1A, sen, virF, invE, sat, sigA, pic, and sepA). MLVA was performed for all EIEC isolates. Results A total of 11 EIEC isolates were identified, and all were positive for the ial gene. The invE and virF genes were observed in 81.8% of the isolates, while sen, sigA, and pic were detected in 72.7%, 63.6%, and 27.3% of the isolates, respectively. None of the isolates were positive for the sat, set, and sepA genes. Using MLVA, the 11 total isolates were divided into five types. Conclusions By studying the profiles of virulence genes and MLVA, it can be concluded that EIEC isolates do not have high heterogeneity and are derived from a limited number of clones. PMID:27635212

  2. Comparison of Multilocus Variable-Number Tandem-Repeat Analysis and Whole-Genome Sequencing for Investigation of Clostridium difficile Transmission

    PubMed Central

    Fawley, W. N.; Best, E. L.; Griffiths, D.; Stoesser, N. E.; Crook, D. W.; Peto, T. E. A.; Walker, A. S.; Wilcox, M. H.

    2013-01-01

    No study to date has compared multilocus variable-number tandem-repeat analysis (MLVA) and whole-genome sequencing (WGS) in an investigation of the transmission of Clostridium difficile infection. Isolates from 61 adults with ongoing and/or recurrent C. difficile infections and 17 asymptomatic carriage episodes in children (201 samples), as well as from 61 suspected outbreaks affecting 2 to 41 patients in 31 hospitals in the United Kingdom (300 samples), underwent 7-locus MLVA and WGS in parallel. When the first and last samples from the same individual taken for a median (interquartile range [IQR]) of 63 days (43 to 105 days) apart were compared, the estimated rates of the evolution of single nucleotide variants (SNVs), summed tandem-repeat differences (STRDs), and locus variants (LVs) were 0.79 (95% confidence interval [CI], 0.00 to 1.75), 1.63 (95% CI, 0.00 to 3.59), and 1.21 (95% CI, 0.00 to 2.67)/called genome/year, respectively. Differences of >2 SNVs and >10 STRDs have been used to exclude direct case-to-case transmission. With the first serial sample per individual being used to assess discriminatory power, across all pairs of samples sharing a PCR ribotype, 192/283 (68%) differed by >10 STRDs and 217/283 (77%) by >2 SNVs. Among all pairs of cases from the same suspected outbreak, 1,190/1,488 (80%) pairs had concordant results using >2 SNVs and >10 STRDs to exclude transmission. For the discordant pairs, 229 (15%) had ≥2 SNVs but ≤10 STRDs, and 69 (5%) had ≤2 SNVs but ≥10 STRDs. Discordant pairs had higher numbers of LVs than concordant pairs, supporting the more diverse measure in each type of discordant pair. Conclusions on whether the potential outbreaks were confirmed were concordant in 58/61 (95%) investigations. Overall findings using MLVA and WGS were very similar despite the fact that they analyzed different parts of the bacterial genome. With improvements in WGS technology, it is likely that MLVA locus data will be available from WGS in the

  3. New Multilocus Variable-Number Tandem-Repeat Analysis (MLVA) Scheme for Fine-Scale Monitoring and Microevolution-Related Study of Ralstonia pseudosolanacearum Phylotype I Populations.

    PubMed

    Guinard, Jérémy; Latreille, Anne; Guérin, Fabien; Poussier, Stéphane; Wicker, Emmanuel

    2017-03-01

    Bacterial wilt caused by the Ralstonia solanacearum species complex (RSSC) is considered one of the most harmful plant diseases in the world. Special attention should be paid to R. pseudosolanacearum phylotype I due to its large host range, its worldwide distribution, and its high evolutionary potential. So far, the molecular epidemiology and population genetics of this bacterium are poorly understood. Until now, the genetic structure of the RSSC has been analyzed on the worldwide and regional scales. Emerging questions regarding evolutionary forces in RSSC adaptation to hosts now require genetic markers that are able to monitor RSSC field populations. In this study, we aimed to evaluate the multilocus variable-number tandem-repeat analysis (MLVA) approach for its ability to discriminate genetically close phylotype I strains and for population genetics studies. We developed a new MLVA scheme (MLVA-7) allowing us to genotype 580 R. pseudosolanacearum phylotype I strains extracted from susceptible and resistant hosts and from different habitats (stem, soil, and rhizosphere). Based on specificity, polymorphism, and the amplification success rate, we selected seven fast-evolving variable-number tandem-repeat (VNTR) markers. The newly developed MLVA-7 scheme showed higher discriminatory power than the previously published MLVA-13 scheme when applied to collections sampled from the same location on different dates and to collections from different locations on very small scales. Our study provides a valuable tool for fine-scale monitoring and microevolution-related study of R. pseudosolanacearum phylotype I populations.IMPORTANCE Understanding the evolutionary dynamics of adaptation of plant pathogens to new hosts or ecological niches has become a key point for the development of innovative disease management strategies, including durable resistance. Whereas the molecular mechanisms underlying virulence or pathogenicity changes have been studied thoroughly, the

  4. Molecular Typing of Mycobacterium intracellulare Using Pulsed-Field Gel Electrophoresis, Variable-Number Tandem-Repeat Analysis, Mycobacteria Interspersed Repetitive-Unit-Variable-Number Tandem Repeat Typing, and Multilocus Sequence Typing: Molecular Characterization and Comparison of Each Typing Methods

    PubMed Central

    Jeon, Semi; Lim, Nara; Kwon, Seungjik; Shim, Taesun; Park, Misun; Kim, Bum-Joon; Kim, Seonghan

    2014-01-01

    Objectives Mycobacterium intracellulare is the major causative agent of nontuberculous mycobacteria-related pulmonary infections. The strain typing of M. intracellulare is important for the treatment and control of its infections. We compared the discrimination capacity and effective value of four different molecular typing methods. Methods Antibiotic susceptibility testing, hsp65 and rpoB sequencing, pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), mycobacteria interspersed repetitive-unit-variable-number tandem-repeat analysis (MIRU-VNTR), and VNTR assay targeting 44 M. intracellulare isolates obtained from patients with pulmonary infections were performed. Results All the antibiotic susceptibility patterns had no association with the molecular and sequence types tested in this study; however, the molecular and sequence types were related with each other. PFGE gave best results for discriminatory capacity, followed by VNTR, MLST, and MIRU-VNTR. Conclusion The high discriminatory power of PFGE, VNTR, and MLST is enough for differentiating between reinfection and relapse, as well as for other molecular epidemiological usages. The MLST could be regarded as a representative classification method, because it showed the clearest relation with the sequence types. PMID:25180144

  5. Three novel polymorphic microsatellite markers for the glaucoma locus GLC1B by datamining tetranucleotide repeats on chromosome 2p12-q12

    PubMed Central

    2009-01-01

    In order to identify new markers around the glaucoma locus GLC1B as a tool to refine its critical region at 2p11.2-2q11.2, we searched the critical region sequence obtained from the UCSC database for tetranucleotide (GATA)n and (GTCT)n repeats of at least 10 units in length. Three out of four potential microsatellite loci were found to be polymorphic, heterozygosity ranging from 64.56% to 79.59%. The identified markers are useful not only for GLC1B locus but also for the study of other disease loci at 2p11.2-2q11.2, a region with scarcity of microsatellite markers. PMID:21637444

  6. Three novel polymorphic microsatellite markers for the glaucoma locus GLC1B by datamining tetranucleotide repeats on chromosome 2p12-q12.

    PubMed

    Murga-Zamalloa, Carlos; Guevara-Fujita, Maria Luisa; Estrada-Cuzcano, Alejandro; Fujita, Ricardo

    2009-10-01

    In order to identify new markers around the glaucoma locus GLC1B as a tool to refine its critical region at 2p11.2-2q11.2, we searched the critical region sequence obtained from the UCSC database for tetranucleotide (GATA)n and (GTCT)n repeats of at least 10 units in length. Three out of four potential microsatellite loci were found to be polymorphic, heterozygosity ranging from 64.56% to 79.59%. The identified markers are useful not only for GLC1B locus but also for the study of other disease loci at 2p11.2-2q11.2, a region with scarcity of microsatellite markers.

  7. Organellar genome, nuclear ribosomal DNA repeat unit, and microsatellites isolated from a small-scale of 454 GS FLX sequencing on two mosses.

    PubMed

    Liu, Yang; Forrest, Laura L; Bainard, Jillian D; Budke, Jessica M; Goffinet, Bernard

    2013-03-01

    Recent innovations in high-throughput DNA sequencing methodology (next generation sequencing technologies [NGS]) allow for the generation of large amounts of high quality data that may be particularly critical for resolving ambiguous relationships such as those resulting from rapid radiations. Application of NGS technology to bryology is limited to assembling entire nuclear or organellar genomes of selected exemplars of major lineages (e.g., classes). Here we outline how organellar genomes and the entire nuclear ribosomal DNA repeat can be obtained from minimal amounts of moss tissue via small-scale 454 GS FLX sequencing. We sampled two Funariaceae species, Funaria hygrometrica and Entosthodon obtusus, and assembled nearly complete organellar genomes and the whole nuclear ribosomal DNA repeat unit (18S-ITS1-5.8S-ITS2-26S-IGS1-5S-IGS2) for both taxa. Sequence data from these species were compared to sequences from another Funariaceae species, Physcomitrella patens, revealing low overall degrees of divergence of the organellar genomes and nrDNA genes with substitutions spread rather evenly across their length, and high divergence within the external spacers of the nrDNA repeat. Furthermore, we detected numerous microsatellites among the 454 assemblies. This study demonstrates that NGS methodology can be applied to mosses to target large genomic regions and identify microsatellites.

  8. A polymorphic microsatellite repeat within the ECE-1c promoter is involved in transcriptional start site determination, human evolution, and Alzheimer's disease.

    PubMed

    Li, Yaosi; Seidel, Kerstin; Marschall, Peter; Klein, Michael; Hope, Antonia; Schacherl, Jens; Schmitz, Jennifer; Menk, Mario; Schefe, Jan H; Reinemund, Jana; Hugel, Rebecca; Walden, Peter; Schlosser, Andreas; Volkmer, Rudolf; Schimkus, Julia; Kölsch, Heike; Maier, Wolfgang; Kornhuber, Johannes; Frölich, Lutz; Klare, Sabrina; Kirsch, Sebastian; Schmerbach, Kristin; Scheele, Sylvia; Grittner, Ulrike; Zollmann, Frank; Goldin-Lang, Petra; Peters, Oliver; Kintscher, Ulrich; Unger, Thomas; Funke-Kaiser, Heiko

    2012-11-21

    Genetic factors strongly contribute to the pathogenesis of sporadic Alzheimer's disease (AD). Nevertheless, genome-wide association studies only yielded single nucleotide polymorphism loci of moderate importance. In contrast, microsatellite repeats are functionally less characterized structures within our genomes. Previous work has shown that endothelin-converting enzyme-1 (ECE-1) is able to reduce amyloid β content. Here we demonstrate that a CpG-CA repeat within the human ECE-1c promoter is highly polymorphic, harbors transcriptional start sites, is able to recruit the transcription factors poly(ADP-ribose) polymerase-1 and splicing factor proline and glutamine-rich, and is functional regarding haplotype-specific promoter activity. Furthermore, genotyping of 403 AD patients and 444 controls for CpG-CA repeat length indicated shifted allelic frequency distributions. Sequencing of 245 haplotype clones demonstrated that the overall CpG-CA repeat composition of AD patients and controls is distinct. Finally, we show that human and chimpanzee [CpG](m)-[CA](n) ECE-1c promoter repeats are genetically and functionally distinct. Our data indicate that a short genomic repeat structure constitutes a novel core promoter element, coincides with human evolution, and contributes to the pathogenesis of AD.

  9. Genetic diversity of bovine Neospora caninum determined by microsatellite markers.

    PubMed

    Salehi, N; Gottstein, B; Haddadzadeh, H R

    2015-10-01

    Neospora caninum is one of the most significant parasitic organisms causing bovine abortion worldwide. Despite the economic impact of this infection, relatively little is known about the genetic diversity of this parasite. In this study, using Nc5 and ITS1 nested PCR, N. caninum has been detected in 12 brain samples of aborted fetuses from 298 seropositive dairy cattle collected from four different regions in Tehran, Iran. These specimen (Nc-Iran) were genotyped in multilocus using 9 different microsatellite markers previously described (MS4, MS5, MS6A, MS6B, MS7, MS8, MS10, MS12 and MS21). Microsatellite amplification was completely feasible in 2 samples, semi-completely in 8 samples, and failed in 2 samples. Within the two completely performed allelic profiles of Nc-Iran strains, unique multilocus profiles were obtained for both and novel allelic patterns were found in the MS8 and MS10 microsatellite markers. The Jaccard's similarity index showed significant difference between these two strains and from other standard isolates derived from GenBank such as Nc-Liv, Nc-SweB1, Nc-GER1, KBA1, and KBA2. All samples originating from the same area showed identical allelic numbers and a correlation between the number of repeats and geographic districts was observed.

  10. The chromosomal distribution of microsatellite repeats in the genome of the wolf fish Hoplias malabaricus, focusing on the sex chromosomes.

    PubMed

    Cioffi, M B; Kejnovsky, E; Bertollo, L A C

    2011-01-01

    Distribution of 12 mono-, di- and tri-nucleotide microsatellites on the chromosomes of 2 karyomorphs with 2 distinct sex chromosome systems (a simple XX/XY - karyomorph B and a multiple X(1)X(1)X(2)X(2)/X(1)X(2)Y - karyomorph D) in Hoplias malabaricus, commonly referred to as wolf fish, was studied using their physical mapping with fluorescence in situ hybridization (FISH). The distribution patterns of different microsatellites along the chromosomes varied considerably. Strong hybridization signals were observed at subtelomeric and heterochromatic regions of several autosomes, with a different accumulation on the sex chromosomes. A massive accumulation was found in the heterochromatic region of the X chromosome of karyomorph B, whereas microsatellites were gathered at centromeres of both X chromosomes as well as in corresponding regions of the neo-Y chromosome in karyomorph D. Our findings are likely in agreement with models that predict the accumulation of repetitive DNA sequences in regions with very low recombination. This process is however in contrast with what was observed in multiple systems, where such a reduction might be facilitated by the chromosomal rearrangements that are directly associated with the origin of these systems.

  11. Reconstruction of microsatellite mutation history reveals a strong and consistent deletion bias in invasive clonal snails, Potamopyrgus antipodarum.

    PubMed

    Weetman, David; Hauser, Lorenz; Carvalho, Gary R

    2002-10-01

    Direct observations of mutations and comparative analyses suggest that nuclear microsatellites show a tendency to expand, with reports of deletion biases limited to very long alleles or a few loci in multilocus studies. Here we investigate microsatellite evolution in clonal snails, Potamopyrgus antipodarum, since their introduction to Britain in the 19th century, using an analysis based on minimum spanning networks of multilocus microsatellite genotypes. British populations consist of a small number of highly distinct genotype groups with very few outlying genotypes, suggesting clonal lineages containing minor variation generated by mutation. Network patterns suggest that a single introduced genotype was the ancestor of all extant variation and also provide support for wholly apomictic reproduction within the most common clonal lineage (group A). Microsatellites within group A showed a strong tendency to delete repeats, with an overall bias exceeding 88%, irrespective of the exact method used to infer mutations. This highly unusual pattern of deletion bias is consistent across populations and loci and is unrelated to allele size. We suggest that for persistence of microsatellites in this clone, some change in the mutation mechanism must have occurred in relatively recent evolutionary time. Possible causes of such a change in mechanism are discussed.

  12. Cytomolecular discrimination of the A(m) chromosomes of Triticum monococcum and the A chromosomes of Triticum aestivum using microsatellite DNA repeats.

    PubMed

    Megyeri, Mária; Mikó, Péter; Farkas, András; Molnár-Láng, Márta; Molnár, István

    2017-02-01

    The cytomolecular discrimination of the A(m)- and A-genome chromosomes facilitates the selection of wheat-Triticum monococcum introgression lines. Fluorescence in situ hybridisation (FISH) with the commonly used DNA probes Afa family, 18S rDNA and pSc119.2 showed that the more complex hybridisation pattern obtained in T. monococcum relative to bread wheat made it possible to differentiate the A(m) and A chromosomes within homoeologous groups 1, 4 and 5. In order to provide additional chromosomal landmarks to discriminate the A(m) and A chromosomes, the microsatellite repeats (GAA)n, (CAG)n, (CAC)n, (AAC)n, (AGG)n and (ACT)n were tested as FISH probes. These showed that T. monococcum chromosomes have fewer, generally weaker, simple sequence repeat (SSR) signals than the A-genome chromosomes of hexaploid wheat. A differential hybridisation pattern was observed on 6A(m) and 6A chromosomes with all the SSR probes tested except for the (ACT)n probe. The 2A(m) and 2A chromosomes were differentiated by the signals given by the (GAA)n, (CAG)n and (AAC)n repeats, while only (GAA)n discriminated the chromosomes 3A(m) and 3A. Chromosomes 7A(m) and 7A could be differentiated by the lack of (GAA)n and (AGG)n signals on 7A. As potential landmarks for identifying the A(m) chromosomes, SSR repeats will facilitate the introgression of T. monococcum chromatin into wheat.

  13. Estimation of genotyping error rate from repeat genotyping, unintentional recaptures and known parent-offspring comparisons in 16 microsatellite loci for brown rockfish (Sebastes auriculatus).

    PubMed

    Hess, Maureen A; Rhydderch, James G; LeClair, Larry L; Buckley, Raymond M; Kawase, Mitsuhiro; Hauser, Lorenz

    2012-11-01

    Genotyping errors are present in almost all genetic data and can affect biological conclusions of a study, particularly for studies based on individual identification and parentage. Many statistical approaches can incorporate genotyping errors, but usually need accurate estimates of error rates. Here, we used a new microsatellite data set developed for brown rockfish (Sebastes auriculatus) to estimate genotyping error using three approaches: (i) repeat genotyping 5% of samples, (ii) comparing unintentionally recaptured individuals and (iii) Mendelian inheritance error checking for known parent-offspring pairs. In each data set, we quantified genotyping error rate per allele due to allele drop-out and false alleles. Genotyping error rate per locus revealed an average overall genotyping error rate by direct count of 0.3%, 1.5% and 1.7% (0.002, 0.007 and 0.008 per allele error rate) from replicate genotypes, known parent-offspring pairs and unintentionally recaptured individuals, respectively. By direct-count error estimates, the recapture and known parent-offspring data sets revealed an error rate four times greater than estimated using repeat genotypes. There was no evidence of correlation between error rates and locus variability for all three data sets, and errors appeared to occur randomly over loci in the repeat genotypes, but not in recaptures and parent-offspring comparisons. Furthermore, there was no correlation in locus-specific error rates between any two of the three data sets. Our data suggest that repeat genotyping may underestimate true error rates and may not estimate locus-specific error rates accurately. We therefore suggest using methods for error estimation that correspond to the overall aim of the study (e.g. known parent-offspring comparisons in parentage studies).

  14. Concerted evolution of the tandemly repeated genes encoding primate U2 small nuclear RNA (the RNU2 locus) does not prevent rapid diversification of the (CT){sub n} {center_dot} (GA){sub n} microsatellite embedded within the U2 repeat unit

    SciTech Connect

    Liao, D.; Weiner, A.M.

    1995-12-10

    The RNU2 locus encoding human U2 small nuclear RNA (snRNA) is organized as a nearly perfect tandem array containing 5 to 22 copies of a 5.8-kb repeat unit. Just downstream of the U2 snRNA gene in each 5.8-kb repeat unit lies a large (CT){sub n}{center_dot}(GA){sub n} dinucleotide repeat (n {approx} 70). This form of genomic organization, in which one repeat is embedded within another, provides an unusual opportunity to study the balance of forces maintaining the homogeneity of both kinds of repeats. Using a combination of field inversion gel electrophoresis and polymerase chain reaction, we have been able to study the CT microsatellites within individual U2 tandem arrays. We find that the CT microsatellites within an RNU2 allele exhibit significant length polymorphism, despite the remarkable homogeneity of the surrounding U2 repeat units. Length polymorphism is due primarily to loss or gain of CT dinucleotide repeats, but other types of deletions, insertions, and substitutions are also frequent. Polymorphism is greatly reduced in regions where pure (CT){sub n} tracts are interrupted by occasional G residues, suggesting that irregularities stabilize both the length and the sequence of the dinucleotide repeat. We further show that the RNU2 loci of other catarrhine primates (gorilla, chimpanzee, ogangutan, and baboon) contain orthologous CT microsatellites; these also exhibit length polymorphism, but are highly divergent from each other. Thus, although the CT microsatellite is evolving far more rapidly than the rest of the U2 repeat unit, it has persisted through multiple speciation events spanning >35 Myr. The persistence of the CT microsatellite, despite polymorphism and rapid evolution, suggests that it might play a functional role in concerted evolution of the RNU2 loci, perhaps as an initiation site for recombination and/or gene conversion. 70 refs., 5 figs.

  15. New multilocus variable-number tandem-repeat analysis tool for surveillance and local epidemiology of bacterial leaf blight and bacterial leaf streak of rice caused by Xanthomonas oryzae.

    PubMed

    Poulin, L; Grygiel, P; Magne, M; Gagnevin, L; Rodriguez-R, L M; Forero Serna, N; Zhao, S; El Rafii, M; Dao, S; Tekete, C; Wonni, I; Koita, O; Pruvost, O; Verdier, V; Vernière, C; Koebnik, R

    2015-01-01

    Multilocus variable-number tandem-repeat analysis (MLVA) is efficient for routine typing and for investigating the genetic structures of natural microbial populations. Two distinct pathovars of Xanthomonas oryzae can cause significant crop losses in tropical and temperate rice-growing countries. Bacterial leaf streak is caused by X. oryzae pv. oryzicola, and bacterial leaf blight is caused by X. oryzae pv. oryzae. For the latter, two genetic lineages have been described in the literature. We developed a universal MLVA typing tool both for the identification of the three X. oryzae genetic lineages and for epidemiological analyses. Sixteen candidate variable-number tandem-repeat (VNTR) loci were selected according to their presence and polymorphism in 10 draft or complete genome sequences of the three X. oryzae lineages and by VNTR sequencing of a subset of loci of interest in 20 strains per lineage. The MLVA-16 scheme was then applied to 338 strains of X. oryzae representing different pathovars and geographical locations. Linkage disequilibrium between MLVA loci was calculated by index association on different scales, and the 16 loci showed linear Mantel correlation with MLSA data on 56 X. oryzae strains, suggesting that they provide a good phylogenetic signal. Furthermore, analyses of sets of strains for different lineages indicated the possibility of using the scheme for deeper epidemiological investigation on small spatial scales.

  16. Close linkage of a gene for X linked deafness to three microsatellite repeats at Xq21 in radiologically normal and abnormal families.

    PubMed

    Bitner-Glindzicz, M; de Kok, Y; Summers, D; Huber, I; Cremers, F P; Ropers, H H; Reardon, W; Pembrey, M E; Malcolm, S

    1994-12-01

    We have used three highly polymorphic microsatellite repeats from Xq21 to type families in whom a gene for X linked deafness with perilymphatic gusher (DFN 3) was segregating. All three markers were tightly linked to the disease in its radiologically normal and abnormal forms, with a maximum lod score of 10.37 with DXS995 and 8.44 with DXS986 at zero recombination, and 14.03 with DXS1002 at theta = 0.01. In an isolated case of deafness of this type, DXS995 indicated either the first recombination observed between the marker and the disease gene or a new mutation in the proband. Southern blotting using a cosmid fragment from the candidate region has confirmed a de novo mutation by showing a deletion in the proband which is not present in his mother as judged by dosage analysis. We also describe a family with a paracentric inversion associated with a microdeletion and discuss how deletion mapping using these and other markers in the region has helped to define a candidate region for the gene.

  17. Oligodeoxynucleotide binding to (CTG) · (CAG) microsatellite repeats inhibits replication fork stalling, hairpin formation, and genome instability.

    PubMed

    Liu, Guoqi; Chen, Xiaomi; Leffak, Michael

    2013-02-01

    (CTG)(n) · (CAG)(n) trinucleotide repeat (TNR) expansion in the 3' untranslated region of the dystrophia myotonica protein kinase (DMPK) gene causes myotonic dystrophy type 1. However, a direct link between TNR instability, the formation of noncanonical (CTG)(n) · (CAG)(n) structures, and replication stress has not been demonstrated. In a human cell model, we found that (CTG)(45) · (CAG)(45) causes local replication fork stalling, DNA hairpin formation, and TNR instability. Oligodeoxynucleotides (ODNs) complementary to the (CTG)(45) · (CAG)(45) lagging-strand template eliminated DNA hairpin formation on leading- and lagging-strand templates and relieved fork stalling. Prolonged cell culture, emetine inhibition of lagging-strand synthesis, or slowing of DNA synthesis by low-dose aphidicolin induced (CTG)(45) · (CAG)(45) expansions and contractions. ODNs targeting the lagging-strand template blocked the time-dependent or emetine-induced instability but did not eliminate aphidicolin-induced instability. These results show directly that TNR replication stalling, replication stress, hairpin formation, and instability are mechanistically linked in vivo.

  18. GT microsatellite repeats in the heme oxygenase-1 gene promoter associated with abdominal aortic aneurysm in Croatian patients.

    PubMed

    Gregorek, Andrea Crkvenac; Gornik, Kristina Crkvenac; Polancec, Darija Stupin; Dabelic, Sanja

    2013-06-01

    Abdominal aortic aneurysm (AAA) is a complex genetic disorder caused by the interplay of genetic and environmental risk factors. The number of (GT)(n) repeats in the heme oxygenase-1 (HO-1) gene promoter modulates transcription of this enzyme, which might have anti-inflammatory, antioxidant, antiapoptotic, and antiproliferative effect. The distribution of alleles and genotypes in Croatian individuals genotyped for the (GT)(n) HO-1 polymorphism was similar to that in other European populations. Frequency of the short (S) alleles (GT < 25) was higher in AAA patients (41.9%) than in non-AAA individuals (28.2%, p = 0.0026) because there were more SL heterozygotes among the AAA patients. The SL genotype appeared to increase the risk for AAA, but the increase was not statistically significant after adjustment for age, sex, smoking, hypertension, and hyperlipidemia (OR = 1.53, 95% CI 0.90-3.09, p = 0.062). These findings contradict those of the only other study performed so far on the association of (GT)(n) HO-1 polymorphism and AAA.

  19. Survey and analysis of simple sequence repeats in the Ustilaginoidea virens genome and the development of microsatellite markers.

    PubMed

    Yu, Mina; Yu, Junjie; Li, Huanhuan; Wang, Yahui; Yin, Xiaole; Bo, Huiwen; Ding, Hui; Zhou, Yuxin; Liu, Yongfeng

    2016-07-01

    Ustilaginoidea virens is the causal agent of rice false smut, causing quantitative and qualitative losses in rice industry. However, the development and application of simple sequence repeat (SSR) markers for genetic diversity studies in U. virens were limited. This study is the first to perform large-scale development of SSR markers of this pathogen at the genome level, to (1) compare these SSR markers with those of other fungi, (2) analyze the pattern of the SSRs, and (3) obtain more informative genetic markers. U. virens is rich in SSRs, and 13,778 SSRs were identified with a relative abundance of 349.7SSRs/Mb. The most common motifs in the genome or in noncoding regions were mononucleotides, whereas trinucleotides in coding sequences. A total of 6 out of 127 primers were randomly selected to be used to analyze 115 isolates, and these 6 primers showed high polymorphism in U. virens. This study may serve as an important resource for molecular genetic studies in U. virens.

  20. The analysis of microsatellites and compound microsatellites in 56 complete genomes of Herpesvirales.

    PubMed

    Wu, Xiaolong; Zhou, Lan; Zhao, Xiangyan; Tan, Zhongyang

    2014-11-01

    Simple sequence repeats (SSRs), or microsatellites, are special DNA/RNA sequences with repeated unit of 1-6 bp. The genomes of Herpesvirales have many repeating structures, which is an excellent system to study the evolution and roles of microsatellites and compound microsatellites in viruses. Therefore, 56 genomes of Herpesvirales were selected and the occurrence, composition and complexity of different repeats were investigated in the genomes. A total of 63,939 microsatellites and 5825 compound microsatellites were extracted from 56 genomes. It found that GC content has a significant strong correlation with both the counts of microsatellites (CM) and the counts of compound microsatellites (CCM). However, genome size has a moderate correlation only with CM and almost no correlation with CCM. The compound microsatellites occurring in genic regions are obviously more than that in intergenic regions. In general, the number of compound microsatellite decreases with the increase of complexity (C) (the count of individual microsatellites being part of a compound microsatellite) and the complexity hardly exceeds C=4. The vast majority of compound microsatellites exist in intergenic regions, when C≥10. The distributions of SSRs tend to be organism-specific rather than host-specific in herpesvirus genomes. The diversity of microsatellites and compound microsatellites may be helpful for a better understanding of the viral genetic diversity, genotyping, and evolutionary biology in herpesviruses genomes.

  1. Characterization of Genetic Diversity of Bacillus anthracis in France by Using High-Resolution Melting Assays and Multilocus Variable-Number Tandem-Repeat Analysis ▿ †

    PubMed Central

    Derzelle, S.; Laroche, S.; Le Flèche, P.; Hauck, Y.; Thierry, S.; Vergnaud, G.; Madani, N.

    2011-01-01

    Using high-resolution melting (HRM) analysis, we developed a cost-effective method to genotype a set of 13 phylogenetically informative single-nucleotide polymorphisms (SNPs) within the genome of Bacillus anthracis. SNP discrimination assays were performed in monoplex or duplex and applied to 100 B. anthracis isolates collected in France from 1953 to 2009 and a few reference strains. HRM provided a reliable and cheap alternative to subtype B. anthracis into one of the 12 major sublineages or subgroups. All strains could be correctly positioned on the canonical SNP (canSNP) phylogenetic tree, except the divergent Pasteur vaccine strain ATCC 4229. We detected the cooccurrence of three canSNP subgroups in France. The dominant B.Br.CNEVA sublineage was found to be prevalent in the Alps, the Pyrenees, the Auvergne region, and the Saône-et-Loire department. Strains affiliated with the A.Br.008/009 subgroup were observed throughout most of the country. The minor A.Br.001/002 subgroup was restricted to northeastern France. Multiple-locus variable-number tandem-repeat analysis using 24 markers further resolved French strains into 60 unique profiles and identified some regional patterns. Diversity found within the A.Br.008/009 and B.Br.CNEVA subgroups suggests that these represent old, ecologically established clades in France. Phylogenetic relationships with strains from other parts of the world are discussed. PMID:21998431

  2. Aridification drove repeated episodes of diversification between Australian biomes: evidence from a multi-locus phylogeny of Australian toadlets (Uperoleia: Myobatrachidae).

    PubMed

    Catullo, Renee A; Scott Keogh, J

    2014-10-01

    Australia is a large and complex landmass that comprises diverse biomes ranging from tropical rainforests to harsh deserts. While Australian biotic diversity has evolved in response to landscape and climate changes, evidence of Miocene or later biome shifts are few. The Australo-Papuan endemic frog genus Uperoleia is widely distributed across mesic, monsoonal tropic and arid regions of Australia. Thus, it represents an ideal system to evaluate biome shifts as they relate to known landscape and climate history. We comprehensively sampled the distributional range of 25 described Uperoleia species and generated a detailed molecular phylogeny for the genus based on one mitochondrial and five nuclear loci. Our results support a single origin of monsoonal tropic taxa, followed by diversification within the region under the influence of the Australian monsoon. Molecular dating analyses suggest the major divergence between eastern mesic and monsoonal species occurred in the Miocene approximately 17million years ago, with repeated evolution of species from monsoonal biomes to arid or mesic biomes in the later Miocene, early Pliocene and at the beginning of the Pleistocene. Our detailed sampling helps to clarify the true distributions of species and contributes to on-going work to improve the taxonomy of the genus. Topological differences between nuclear and mitochondrial phylogenies within major clades suggest a history of mitochondrial introgression and capture, and reduce the ability to resolve close interspecific relationships.

  3. Current Situation of Antimicrobial Resistance and Genetic Differences in Stenotrophomonas maltophilia Complex Isolates by Multilocus Variable Number of Tandem Repeat Analysis

    PubMed Central

    Song, Jae-Hoon

    2016-01-01

    Background Stenotrophomonas maltophilia is one of several opportunistic pathogens of growing significance. Several studies on the molecular epidemiology of S. maltophilia have shown clinical isolates to be genetically diverse. Materials and Methods A total of 121 clinical isolates tentatively identified as S. malophilia from seven tertiary-care hospitals in Korea from 2007 to 2011 were included. Species and groups were identified using partial gyrB gene sequences and antimicrobial susceptibility testing was performed using a broth microdilution method. Multi locus variable number of tandem repeat analysis (MLVA) surveys are used for subtyping. Results Based on partial gyrB gene sequences, 118 isolates were identified as belonging to the S. maltophilia complex. For all S. maltophilia isolates, the resistance rates to trimethoprime-sulfamethoxazole (TMP/SMX) and levofloxacin were the highest (both, 30.5%). Resistance rate to ceftazidime was 28.0%. 11.0% and 11.9% of 118 S. maltophilia isolates displayed resistance to piperacillin/tazobactam and tigecycline, respectively. Clade 1 and Clade 2 were definitely distinguished from the data of MLVA with amplification of loci. All 118 isolates were classified into several clusters as its identification. Conclusion Because of high resistance rates to TMP/SMX and levofloxacin, the clinical laboratory department should consider providing the data about other antimicrobial agents and treatment of S. maltophilia infections with a combination of antimicrobials can be considered in the current practice. The MLVA evaluated in this study provides a fast, portable, relatively low cost genotyping method that can be employed in genotypic linkage or transmission networks comparing to analysis of the gyrB gene. PMID:28032486

  4. Sequence determinants of human microsatellite variability

    PubMed Central

    2009-01-01

    Background Microsatellite loci are frequently used in genomic studies of DNA sequence repeats and in population studies of genetic variability. To investigate the effect of sequence properties of microsatellites on their level of variability we have analyzed genotypes at 627 microsatellite loci in 1,048 worldwide individuals from the HGDP-CEPH cell line panel together with the DNA sequences of these microsatellites in the human RefSeq database. Results Calibrating PCR fragment lengths in individual genotypes by using the RefSeq sequence enabled us to infer repeat number in the HGDP-CEPH dataset and to calculate the mean number of repeats (as opposed to the mean PCR fragment length), under the assumption that differences in PCR fragment length reflect differences in the numbers of repeats in the embedded repeat sequences. We find the mean and maximum numbers of repeats across individuals to be positively correlated with heterozygosity. The size and composition of the repeat unit of a microsatellite are also important factors in predicting heterozygosity, with tetra-nucleotide repeat units high in G/C content leading to higher heterozygosity. Finally, we find that microsatellites containing more separate sets of repeated motifs generally have higher heterozygosity. Conclusions These results suggest that sequence properties of microsatellites have a significant impact in determining the features of human microsatellite variability. PMID:20015383

  5. Temperature-Dependent Growth Modeling of Environmental and Clinical Legionella pneumophila Multilocus Variable-Number Tandem-Repeat Analysis (MLVA) Genotypes.

    PubMed

    Sharaby, Yehonatan; Rodríguez-Martínez, Sarah; Oks, Olga; Pecellin, Marina; Mizrahi, Hila; Peretz, Avi; Brettar, Ingrid; Höfle, Manfred G; Halpern, Malka

    2017-04-15

    Legionella pneumophila causes waterborne infections resulting in severe pneumonia. High-resolution genotyping of L. pneumophila isolates can be achieved by multiple-locus variable-number tandem-repeat analysis (MLVA). Recently, we found that different MLVA genotypes of L. pneumophila dominated different sites in a small drinking-water network, with a genotype-related temperature and abundance regime. The present study focuses on understanding the temperature-dependent growth kinetics of the genotypes that dominated the water network. Our aim was to model mathematically the influence of temperature on the growth kinetics of different environmental and clinical L. pneumophila genotypes and to compare it with the influence of their ecological niches. Environmental strains showed a distinct temperature preference, with significant differences among the growth kinetics of the three studied genotypes (Gt4, Gt6, and Gt15). Gt4 strains exhibited superior growth at lower temperatures (25 and 30°C), while Gt15 strains appeared to be best adapted to relatively higher temperatures (42 and 45°C). The temperature-dependent growth traits of the environmental genotypes were consistent with their distribution and temperature preferences in the water network. Clinical isolates exhibited significantly higher growth rates and reached higher maximal cell densities at 37°C and 42°C than the environmental strains. Further research on the growth preferences of L. pneumophila clinical and environmental genotypes will result in a better understanding of their ecological niches in drinking-water systems as well as in the human body.IMPORTANCELegionella pneumophila is a waterborne pathogen that threatens humans in developed countries. The bacteria inhabit natural and man-made freshwater environments. Here we demonstrate that different environmental L. pneumophila genotypes have different temperature-dependent growth kinetics. Moreover, Legionella strains that belong to the same species

  6. Origin of the Outbreak in France of Pseudomonas syringae pv. actinidiae Biovar 3, the Causal Agent of Bacterial Canker of Kiwifruit, Revealed by a Multilocus Variable-Number Tandem-Repeat Analysis

    PubMed Central

    Cunty, A.; Cesbron, S.; Poliakoff, F.; Jacques, M.-A.

    2015-01-01

    The first outbreaks of bacterial canker of kiwifruit caused by Pseudomonas syringae pv. actinidiae biovar 3 were detected in France in 2010. P. syringae pv. actinidiae causes leaf spots, dieback, and canker that sometimes lead to the death of the vine. P. syringae pv. actinidifoliorum, which is pathogenic on kiwi as well, causes only leaf spots. In order to conduct an epidemiological study to track the spread of the epidemics of these two pathogens in France, we developed a multilocus variable-number tandem-repeat (VNTR) analysis (MLVA). MLVA was conducted on 340 strains of P. syringae pv. actinidiae biovar 3 isolated in Chile, China, France, Italy, and New Zealand and on 39 strains of P. syringae pv. actinidifoliorum isolated in Australia, France, and New Zealand. Eleven polymorphic VNTR loci were identified in the genomes of P. syringae pv. actinidiae biovar 3 ICMP 18744 and of P. syringae pv. actinidifoliorum ICMP 18807. MLVA enabled the structuring of P. syringae pv. actinidiae biovar 3 and P. syringae pv. actinidifoliorum strains in 55 and 16 haplotypes, respectively. MLVA and discriminant analysis of principal components revealed that strains isolated in Chile, China, and New Zealand are genetically distinct from P. syringae pv. actinidiae strains isolated in France and in Italy, which appear to be closely related at the genetic level. In contrast, no structuring was observed for P. syringae pv. actinidifoliorum. We developed an MLVA scheme to explore the diversity within P. syringae pv. actinidiae biovar 3 and to trace the dispersal routes of epidemic P. syringae pv. actinidiae biovar 3 in Europe. We suggest using this MLVA scheme to trace the dispersal routes of P. syringae pv. actinidiae at a global level. PMID:26209667

  7. Origin of the Outbreak in France of Pseudomonas syringae pv. actinidiae Biovar 3, the Causal Agent of Bacterial Canker of Kiwifruit, Revealed by a Multilocus Variable-Number Tandem-Repeat Analysis.

    PubMed

    Cunty, A; Cesbron, S; Poliakoff, F; Jacques, M-A; Manceau, C

    2015-10-01

    The first outbreaks of bacterial canker of kiwifruit caused by Pseudomonas syringae pv. actinidiae biovar 3 were detected in France in 2010. P. syringae pv. actinidiae causes leaf spots, dieback, and canker that sometimes lead to the death of the vine. P. syringae pv. actinidifoliorum, which is pathogenic on kiwi as well, causes only leaf spots. In order to conduct an epidemiological study to track the spread of the epidemics of these two pathogens in France, we developed a multilocus variable-number tandem-repeat (VNTR) analysis (MLVA). MLVA was conducted on 340 strains of P. syringae pv. actinidiae biovar 3 isolated in Chile, China, France, Italy, and New Zealand and on 39 strains of P. syringae pv. actinidifoliorum isolated in Australia, France, and New Zealand. Eleven polymorphic VNTR loci were identified in the genomes of P. syringae pv. actinidiae biovar 3 ICMP 18744 and of P. syringae pv. actinidifoliorum ICMP 18807. MLVA enabled the structuring of P. syringae pv. actinidiae biovar 3 and P. syringae pv. actinidifoliorum strains in 55 and 16 haplotypes, respectively. MLVA and discriminant analysis of principal components revealed that strains isolated in Chile, China, and New Zealand are genetically distinct from P. syringae pv. actinidiae strains isolated in France and in Italy, which appear to be closely related at the genetic level. In contrast, no structuring was observed for P. syringae pv. actinidifoliorum. We developed an MLVA scheme to explore the diversity within P. syringae pv. actinidiae biovar 3 and to trace the dispersal routes of epidemic P. syringae pv. actinidiae biovar 3 in Europe. We suggest using this MLVA scheme to trace the dispersal routes of P. syringae pv. actinidiae at a global level.

  8. Survey of compound microsatellites in multiple Lactobacillus genomes.

    PubMed

    Basharat, Zarrin; Yasmin, Azra

    2015-12-01

    Distinct simple sequence repeats with 2 or more individual microsatellites joined together or lying adjacent to each other are identified as compound microsatellites. Investigation of such composite microsatellites in the genomes of genus Lactobacillus was the aim of this study. In silico inspection of microsatellite clustering in genomes of 14 Lactobacillus species revealed a wealth of compound microsatellites. All of the mined compound microsatellites were imperfect, were composed of variant motifs, and increased in all genomes, with maximum distance (dMAX) increments of 10 to 50. The majority of these repeats were present in the coding regions. A correlation of microsatellite to compound microsatellite density was detected. The difference established in compound microsatellite division among eukaryotes, Escherichia coli, and lactobacilli is suggestive of diverse genomic features and elementary distinction between creation and fixation methods of compound microsatellites among these organisms.

  9. Microsatellite DNA fingerprinting, differentiation, and genetic relationships of clones, cultivars, and varieties of six poplar species from three sections of the genus Populus.

    PubMed

    Rahman, Muhammad H; Rajora, Om P

    2002-12-01

    Accurate identification of Populus clones and cultivars is essential for effective selection, breeding, and genetic resource management programs. The unit of cultivation and breeding in poplars is a clone, and individual cultivars are normally represented by a single clone. Microsatellite DNA markers of 10 simple sequence repeat loci were used for genetic fingerprinting and differentiation of 96 clones/cultivars and varieties belonging to six Populus species (P. deltoides, P. nigra, P. balsamifera, P. trichocarpa, P. grandidentata, and P maximowiczii) from three sections of the genus. All 96 clones/cultivars could be uniquely fingerprinted based on their single- or multilocus microsatellite genotypes. The five P. grandidentata clones could be differentiated based on their single-locus genotypes, while six clones of P. trichocarpa and 11 clones of P. maximowiczii could be identified by their two-locus genotypes. Twenty clones of P. deltoides and 25 clones of P. nigra could be differentiated by their multilocus genotypes employing three loci, and 29 clones of P. balsamifera required the use of multilocus genotypes at five loci for their genetic fingerprinting and differentiation. The loci PTR3, PTR5, and PTR7 were found to be the most informative for genetic fingerprinting and differentiation of the clones. The mean number of alleles per locus ranged from 2.9 in P. trichocarpa or P. grandidentata to 6.0 in P. balsamifera and 11.2 in 96 clones of the six species. The mean number of observed genotypes per locus ranged from 2.4 in P. grandidentata to 7.4 in P. balsamifera and 19.6 in 96 clones of the six species. The mean number of unique genotypes per locus ranged from 1.3 in P. grandidentata to 3.9 in P. deltoides and 8.8 in 96 clones of the six species. The power of discrimination of the microsatellite DNA markers in the 96 clones ranged from 0.726 for PTR4 to 0.939 for PTR7, with a mean of 0.832 over the 10 simple sequence repeat loci. Clones/cultivars from the same

  10. Detection of Sequence Polymorphism in Rubus Occidentalis L. Monomorphic Microsatellite Markers by High Resolution Melting

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Microsatellite, or simple sequence repeat (SSR) markers, are valuable as co-dominant genetic markers with a variety of applications such as DNA fingerprinting, linkage mapping, and population structure analysis. Development of microsatellite primers through the identification of appropriate repeate...

  11. Isolation and characterization of microsatellite markers in the Serra Spanish mackerel, Scomberomorus brasiliensis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Thirteen nuclear-encoded microsatellites from a genomic DNA library of Serra Spanish mackerel, Scomberomorus brasiliensis, were isolated and characterized. The microsatellites include 10 perfect repeats (8 tetranucleotide and 2 dinucleotide) and 3 imperfect repeats (2 tetranucleotide and 1 dinucleo...

  12. Microsatellite primers for vulnerable and thriving Acacia (Fabaceae) species from Australia’s arid zone1

    PubMed Central

    Forrest, Cairo N.; Roberts, David G.; Denham, Andrew J.; Ayre, David J.

    2015-01-01

    Premise of the study: Microsatellite markers were developed for the common arid Australian shrub Acacia ligulata (Fabaceae) and the threatened overstory trees A. melvillei and A. pendula. Methods and Results: DNA sequence data generated by 454 sequencing were used to identify microsatellite nucleotide repeat motifs. Including previously developed primer sets, we report on the development of 10 polymorphic microsatellite loci for each species. Six of these were novel for A. melvillei and A. ligulata, and five were novel for A. pendula, while five more each were transferred from primers developed for related species (A. carneorum and A. loderi). We found three to 17 alleles per locus for each species, with high multilocus genotypic diversity within each of two A. ligulata and A. pendula stands, and one A. melvillei population. A second A. melvillei stand appeared to be monoclonal. Conclusions: These markers will allow assessment of population genetics, mating systems, and connectedness of populations of these and possibly other arid-zone acacias. PMID:25909043

  13. Transpecific microsatellites for hard pines.

    PubMed

    Shepherd, M.; Cross, M.; Maguire, L.; Dieters, J.; Williams, G.; Henry, J.

    2002-04-01

    Microsatellites are difficult to recover from large plant genomes so cross-specific utilisation is an important source of markers. Fifty microsatellites were tested for cross-specific amplification and polymorphism to two New World hard pine species, slash pine ( Pinus elliottii var. elliottii) and Caribbean pine ( P. caribaea var. hondurensis). Twenty-nine (58%) markers amplified in both hard pine species, and 23 of these 29 were polymorphic. Soft pine (subgenus Strobus) microsatellite markers did amplify, but none were polymorphic. Pinus elliottii var. elliottii and P. caribaea var. hondurensis showed mutational changes in the flanking regions and the repeat motif that were informative for Pinus spp. phylogenetic relationships. Most allele length variation could be attributed to variability in repeat unit number. There was no evidence for ascertainment bias.

  14. Blueberry Microsatellite Markers Identify Cranberries

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Forty-six blueberry simple sequence repeat (SSR) markers or microsatellites were tested for the ability to amplify a polymorphic marker in eight American cranberry accessions. Sixteen SSRs resulted in informative and polymorphic SSR primer pairs and were used to fingerprint 16 economically important...

  15. Targeted stock identification using multilocus genotype 'familyprinting'

    USGS Publications Warehouse

    Letcher, B.H.; King, T.L.

    1999-01-01

    We present an approach to stock identification of small, targeted populations that uses multilocus microsatellite genotypes of individual mating adults to uniquely identify first- and second-generation offspring in a mixture. We call the approach 'familyprinting'; unlike DNA fingerprinting where tissue samples of individuals are matched, offspring from various families are assigned to pairs of parents or sets of four grandparents with known genotypes. The basic unit of identification is the family, but families can be nested within a variety of stock units ranging from naturally reproducing groups of fish in a small tributary or pond from which mating adults can be sampled to large or small collections of families produced in hatcheries and stocked in specific locations. We show that, with as few as seven alleles per locus using four loci without error, first-generation offspring can be uniquely assigned to the correct family. For second-generation applications in a hatchery more alleles per locus (10) and loci (10) are required for correct assignment of all offspring to the correct set of grandparents. Using microsatellite DNA variation from an Atlantic salmon (Salmo solar) restoration river (Connecticut River, USA), we also show that this population contains sufficient genetic diversity in sea-run returns for 100% correct first, generation assignment and 97% correct second-generation assignment using 14 loci. We are currently using first- and second-generation familyprinting in this population with the ultimate goal of identifying stocking tributary. In addition to within-river familyprinting, there also appears to be sufficient genetic diversity within and between Atlantic salmon populations for identification of 'familyprinted' fish in a mixture of multiple populations. We also suggest that second-generation familyprinting with multiple populations may also provide a tool for examining stock structure. Familyprinting with microsatellite DNA markers is a viable

  16. Microsatellite (GT)(n) repeats and SNPs in the von Willebrand factor gene promoter do not influence circulating von Willebrand factor levels under normal conditions.

    PubMed

    Daidone, Viviana; Cattini, Maria Grazia; Pontara, Elena; Sartorello, Francesca; Gallinaro, Lisa; Marotti, Alberto; Scaroni, Carla; Pagnan, Antonio; Casonato, Alessandra

    2009-02-01

    Von Willebrand factor (VWF) levels vary considerably in normal individuals, influenced by inherited and acquired modulators. ABO blood group is the major inherited determinant of VWF levels, but a role has also been attributed to the VWF gene promoter, haplotype 1 (-3268G/-2709C/-2661A/-2527G) being associated with higher VWF levels than haplotype 2 (-3268C/-2709T/-2661G/-2527A), and the polymorphic locus (GT)(n) modulating the shear stress-induced activation of the VWF promoter. We characterized the (GT)(n) of the VWF promoter in 394 healthy individuals and assessed whether its variable length influenced VWF levels in normal conditions. (GT)(n) proved highly polymorphic, with alleles from 15 to 24 repeats long. (GT)(21) and (GT)(19) were the most common variants (37.4% and 34.4%, respectively). Short GT repeats (15-19) segregated mainly with haplotype 1, long GT repeats (20-24) with haplotype 2 (p < 0.0001). The number of GT repeats did not correlate with VWF levels, nor did such levels correlate with haplotypes 1 and 2, considered alone or in association with the (GT)(n) locus. We conclude that (GT)(n) and -3268/-2709/-2661/-2527 loci are in strong linkage disequilibrium. This polymorphic region of the VWF promoter does not affect VWF levels under normal conditions, though it might represent an environmentally activable VWF regulation site.

  17. Cost-effective development of highly polymorphic microsatellite in Japanese quail facilitated by next-generation sequencing.

    PubMed

    Tadano, R; Nunome, M; Mizutani, M; Kawahara-Miki, R; Fujiwara, A; Takahashi, S; Kawashima, T; Nirasawa, K; Ono, T; Kono, T; Matsuda, Y

    2014-12-01

    Next-generation sequencing technologies permit rapid and cost-effective identification of numerous putative microsatellite loci. Here, from the genome sequences of Japanese quail, we developed microsatellite markers containing dinucleotide repeats and employed these for characterisation of genetic diversity and population structure. A total of 385 individuals from 12 experimental and one wild-derived Japanese quail lines were genotyped with newly developed autosomal markers. The maximum number of alleles, expected heterozygosity and polymorphic information content (PIC) per locus were 10, 0.80 and 0.77 respectively. Approximately half of the markers were highly informative (PIC ≥ 0.50). The mean number of alleles per locus and observed heterozygosity within a line were in the range of 1.3-4.1 and 0.11-0.53 respectively. Compared with the wild-derived line, genetic diversity levels were low in the experimental lines. Genetic differentiation (FST ) between all pairs of the lines ranged from 0.13 to 0.83. Genetic clustering analyses based on multilocus genotypes of individuals showed that most individuals formed clearly defined clusters corresponding to the origins of the lines. These results suggest that Japanese quail experimental lines are highly structured. Microsatellite markers developed in this study may be effective for future genetic studies of Japanese quail.

  18. Short (GT)n microsatellite repeats in the heme oxygenase-1 gene promoter are associated with antioxidant and anti-inflammatory status in Mexican pediatric patients with sepsis.

    PubMed

    Vázquez-Armenta, Gabriela; González-Leal, Natalia; J Vázquez-de la Torre, Mayra; Muñoz-Valle, José Francisco; Ramos-Márquez, Martha E; Hernández-Cañaveral, Iván; Plascencia-Hernández, Arturo; Siller-López, Fernando

    2013-01-01

    An adequate immune and antioxidant response is a key to the resolution of sepsis. Heme oxygenase-1 (HMOX1) is a stress protein with a polymorphic (GT)n repeat in its gene promoter that regulates its expression in response to oxidative injury, such as that present in sepsis. HMOX1 is the rate-limiting enzyme of heme degradation, and the heme breakdown products, CO, Fe, and bilirubin, are considered to be biologically active metabolites with direct or indirect antioxidant and anti-inflammatory properties. In this study, we investigated the inflammatory and antioxidant response and the relationship with the HMOX1 levels and HMOX1 polymorphism in Mexican septic pediatric patients. In a case-control pilot study, we enrolled 64 septic patients and 72 hospitalized control patients without a diagnosis of sepsis. DNA extracted from buffy coat was genotyped for HMOX1 (GT)n polymorphism by PCR and markers of antioxidant and inflammatory status were quantified in plasma by analysis of the oxygen radical absorbance capacity (ORAC), protein carbonyl (PC), interleukin (IL) 6, IL10, and HMOX1 levels. In septic children, oxidative and inflammatory markers were elevated, and HMOX1 levels were positively correlated with IL10 levels. Genotypic and allelic distribution of HMOX1 polymorphism showed no difference between groups. HMOX1 short-allele septic carriers (< 25 GT repeats) presented favorable ORAC, PC and IL10 levels. This study confirms that an active response against pediatric sepsis involves the expression of HMOX1 and IL10, suggesting that the high antioxidant status associated with HMOX1 short-allele septic carriers might provide a beneficial environment for sepsis resolution.

  19. Comparative analysis of microsatellites and compound microsatellites in T4-like viruses.

    PubMed

    Zhou, Lan; Deng, Liang; Fu, Yongzhuo; Wu, Xiaolong; Zhao, Xiangyan; Chen, Yubao; Li, Mingfu; Tan, Zhongyang

    2016-01-10

    Microsatellites or simple sequence repeats (SSRs) are known to present ubiquitously in genomes of eukaryotes and prokaryotes, as well as viruses. A comprehensive analysis of microsatellites and compound microsatellites (CM) was performed for 67 T4-like bacteriophage genomes. We found that the number of repeats was generally proportional to the size of the genome. CM were more abundant in genic regions, while their relative abundance was higher in intergenic regions. Meanwhile, the number of CM rapidly decreased with the increase of complexity but gradually increased with higher dMAX (maximum distance between any two adjacent microsatellites). (A)n/(T)n, (AT)n/(TA)n and (AAG)n were the most abundant repeats of mono-, di- and trinucleotide microsatellites, respectively. The number of microsatellites in reference sequences was significantly lower than that in corresponding random sequences. This result was mainly attributed to mono- and dinucleotide repeats which hardly exceeded 6bp in T4-like viruses. These observations may be helpful to understand the distribution of microsatellites and viral genetic diversity in T4-like viruses.

  20. Development of microsatellites for genetic analyses and population assignment of the cat flea (Siphonaptera: Pulicidae).

    PubMed

    Husseneder, Claudia; Garner, Susan P; Foil, Lane D; Macaluso, Kevin R

    2010-11-01

    Cat fleas, Ctenocephalidesfelis (Bouché) (Siphonaptera: Pulicidae), are common ectoparasites of companion animals that negatively impact their hosts directly by causing dermatitis and blood loss during feeding and indirectly through the potential transmission of disease causing agents. We isolated and characterized seven novel microsatellite loci from a partial genomic library of the cat flea enriched for di-, tri-, and tetranucleotide repeats. We screened these loci in cat fleas from two laboratory colonies and one wild-caught population collected at a temporary animal shelter (Parker coliseum) in Baton Rouge, LA. Six loci were polymorphic, with two to 15 alleles per locus and an average observed heterozygosity of 0.21 across populations. Although the two laboratory cat flea colonies were isolated from each other for many years, they did not significantly differ in their genotypic composition. The cat flea population from Parker coliseum was genetically different from the laboratory colonies, but also showed high degrees of inbreeding. Multilocus genotypes of the polymorphic loci were sufficient to assign over 85% of cat fleas to their population of origin. Genetic markers for flea population identity will allow further studies to examine the origins and movement of cat fleas with important genetic traits such as insecticide resistance or pathogen susceptibility. The use of microsatellites also could determine if there are host-specific strains of cat fleas and add insight into the development of the different subspecies of C. felis.

  1. Survey and Analysis of Microsatellites in the Silkworm, Bombyx mori

    PubMed Central

    Prasad, M. Dharma; Muthulakshmi, M.; Madhu, M.; Archak, Sunil; Mita, K.; Nagaraju, J.

    2005-01-01

    We studied microsatellite frequency and distribution in 21.76-Mb random genomic sequences, 0.67-Mb BAC sequences from the Z chromosome, and 6.3-Mb EST sequences of Bombyx mori. We mined microsatellites of ≥15 bases of mononucleotide repeats and ≥5 repeat units of other classes of repeats. We estimated that microsatellites account for 0.31% of the genome of B. mori. Microsatellite tracts of A, AT, and ATT were the most abundant whereas their number drastically decreased as the length of the repeat motif increased. In general, tri- and hexanucleotide repeats were overrepresented in the transcribed sequences except TAA, GTA, and TGA, which were in excess in genomic sequences. The Z chromosome sequences contained shorter repeat types than the rest of the chromosomes in addition to a higher abundance of AT-rich repeats. Our results showed that base composition of the flanking sequence has an influence on the origin and evolution of microsatellites. Transitions/transversions were high in microsatellites of ESTs, whereas the genomic sequence had an equal number of substitutions and indels. The average heterozygosity value for 23 polymorphic microsatellite loci surveyed in 13 diverse silkmoth strains having 2–14 alleles was 0.54. Only 36 (18.2%) of 198 microsatellite loci were polymorphic between the two divergent silkworm populations and 10 (5%) loci revealed null alleles. The microsatellite map generated using these polymorphic markers resulted in 8 linkage groups. B. mori microsatellite loci were the most conserved in its immediate ancestor, B. mandarina, followed by the wild saturniid silkmoth, Antheraea assama. PMID:15371363

  2. Characterization of microsatellites in wild and sweet cherry (Prunus avium L.)--markers for individual identification and reproductive processes.

    PubMed

    Schueler, Silvio; Tusch, Alexandra; Schuster, Mirko; Ziegenhagen, Birgit

    2003-02-01

    Nuclear microsatellites were characterized in Prunus avium and validated as markers for individual and cultivar identification, as well as for studies of pollen- and seed-mediated gene flow. We used 20 primer pairs from a simple sequence repeat (SSR) library of Prunus persica and identified 7 loci harboring polymorphic microsatellite sequences in P. avium. In a natural population of 75 wild cherry trees, the number of alleles per locus ranged from 4 to 9 and expected heterozygosity from 0.39 to 0.77. The variability of the SSR markers allowed an unambiguous identification of individual trees and potential root suckers. Additionally, we analyzed 13 sweet cherry cultivars and differentiated 12 of them. An exclusion probability of 0.984 was calculated, which indicates that the seven loci are suitable markers for paternity analysis. The woody endocarp was successfully used for resolution of all microsatellite loci and exhibited the same multilocus genotype as the mother tree, as shown in a single seed progeny. Hence, SSR fingerprinting of the purely maternal endocarp was also successful in this Prunus species, allowing the identification of the mother tree of the dispersed seeds. The linkage of microsatellite loci with PCR-amplified alleles of the self-incompatibility locus was tested in two full-sib families of sweet cherry cultivars. From low recombination frequencies, we inferred that two loci are linked with the S locus. The present study provides markers that will significantly facilitate studies of spatial genetic variation and gene flow in wild cherry, as well as breeding programs in sweet cherry.

  3. Microsatellite DNA capture from enriched libraries.

    PubMed

    Gonzalez, Elena G; Zardoya, Rafael

    2013-01-01

    Microsatellites are DNA sequences of tandem repeats of one to six nucleotides, which are highly polymorphic, and thus the molecular markers of choice in many kinship, population genetic, and conservation studies. There have been significant technical improvements since the early methods for microsatellite isolation were developed, and today the most common procedures take advantage of the hybrid capture methods of enriched-targeted microsatellite DNA. Furthermore, recent advents in sequencing technologies (i.e., next-generation sequencing, NGS) have fostered the mining of microsatellite markers in non-model organisms, affording a cost-effective way of obtaining a large amount of sequence data potentially useful for loci characterization. The rapid improvements of NGS platforms together with the increase in available microsatellite information open new avenues to the understanding of the evolutionary forces that shape genetic structuring in wild populations. Here, we provide detailed methodological procedures for microsatellite isolation based on the screening of GT microsatellite-enriched libraries, either by cloning and Sanger sequencing of positive clones or by direct NGS. Guides for designing new species-specific primers and basic genotyping are also given.

  4. A New Electrophoresis Technique to Seperate Microsatellite Alleles

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Traditional agarose and polyacrylamide gel electrophoresis have been used commonly for microsatellite (simple sequence repeats, SSRs) analysis, but they are labor- intensive and not always able to provide accurate sizes for different alleles. Capillary sequencers provide automated analysis and accur...

  5. Rapid microsatellite marker development using next generation pyrosequencing to inform invasive Burmese python -- Python molurus bivittatus -- management

    USGS Publications Warehouse

    Hunter, Margaret E.; Hart, Kristen M.

    2013-01-01

    Invasive species represent an increasing threat to native ecosystems, harming indigenous taxa through predation, habitat modification, cross-species hybridization and alteration of ecosystem processes. Additionally, high economic costs are associated with environmental damage, restoration and control measures. The Burmese python, Python molurus bivittatus, is one of the most notable invasive species in the US, due to the threat it poses to imperiled species and the Greater Everglades ecosystem. To address population structure and relatedness, next generation sequencing was used to rapidly produce species-specific microsatellite loci. The Roche 454 GS-FLX Titanium platform provided 6616 di-, tri- and tetra-nucleotide repeats in 117,516 sequences. Using stringent criteria, 24 of 26 selected tri- and tetra-nucleotide loci were polymerase chain reaction (PCR) amplified and 18 were polymorphic. An additional six cross-species loci were amplified, and the resulting 24 loci were incorporated into eight PCR multiplexes. Multi-locus genotypes yielded an average of 61% (39%–77%) heterozygosity and 3.7 (2–6) alleles per locus. Population-level studies using the developed microsatellites will track the invasion front and monitor population-suppression dynamics. Additionally, cross-species amplification was detected in the invasive Ball, P. regius, and Northern African python, P. sebae. These markers can be used to address the hybridization potential of Burmese pythons and the larger, more aggressive P. sebae.

  6. Rapid Microsatellite Marker Development Using Next Generation Pyrosequencing to Inform Invasive Burmese Python—Python molurus bivittatus—Management

    PubMed Central

    Hunter, Margaret E.; Hart, Kristen M.

    2013-01-01

    Invasive species represent an increasing threat to native ecosystems, harming indigenous taxa through predation, habitat modification, cross-species hybridization and alteration of ecosystem processes. Additionally, high economic costs are associated with environmental damage, restoration and control measures. The Burmese python, Python molurus bivittatus, is one of the most notable invasive species in the US, due to the threat it poses to imperiled species and the Greater Everglades ecosystem. To address population structure and relatedness, next generation sequencing was used to rapidly produce species-specific microsatellite loci. The Roche 454 GS-FLX Titanium platform provided 6616 di-, tri- and tetra-nucleotide repeats in 117,516 sequences. Using stringent criteria, 24 of 26 selected tri- and tetra-nucleotide loci were polymerase chain reaction (PCR) amplified and 18 were polymorphic. An additional six cross-species loci were amplified, and the resulting 24 loci were incorporated into eight PCR multiplexes. Multi-locus genotypes yielded an average of 61% (39%–77%) heterozygosity and 3.7 (2–6) alleles per locus. Population-level studies using the developed microsatellites will track the invasion front and monitor population-suppression dynamics. Additionally, cross-species amplification was detected in the invasive Ball, P. regius, and Northern African python, P. sebae. These markers can be used to address the hybridization potential of Burmese pythons and the larger, more aggressive P. sebae. PMID:23449030

  7. Rapid Microsatellite Marker Development Using Next Generation Pyrosequencing to Inform Invasive Burmese Python-Python molurus bivittatus-Management.

    PubMed

    Hunter, Margaret E; Hart, Kristen M

    2013-02-28

    Invasive species represent an increasing threat to native ecosystems, harming indigenous taxa through predation, habitat modification, cross-species hybridization and alteration of ecosystem processes. Additionally, high economic costs are associated with environmental damage, restoration and control measures. The Burmese python, Python molurus bivittatus, is one of the most notable invasive species in the US, due to the threat it poses to imperiled species and the Greater Everglades ecosystem. To address population structure and relatedness, next generation sequencing was used to rapidly produce species-specific microsatellite loci. The Roche 454 GS-FLX Titanium platform provided 6616 di-, tri- and tetra-nucleotide repeats in 117,516 sequences. Using stringent criteria, 24 of 26 selected tri- and tetra-nucleotide loci were polymerase chain reaction (PCR) amplified and 18 were polymorphic. An additional six cross-species loci were amplified, and the resulting 24 loci were incorporated into eight PCR multiplexes. Multi-locus genotypes yielded an average of 61% (39%-77%) heterozygosity and 3.7 (2-6) alleles per locus. Population-level studies using the developed microsatellites will track the invasion front and monitor population-suppression dynamics. Additionally, cross-species amplification was detected in the invasive Ball, P. regius, and Northern African python, P. sebae. These markers can be used to address the hybridization potential of Burmese pythons and the larger, more aggressive P. sebae.

  8. New softwares for automated microsatellite marker development

    PubMed Central

    Martins, Wellington; de Sousa, Daniel; Proite, Karina; Guimarães, Patrícia; Moretzsohn, Marcio; Bertioli, David

    2006-01-01

    Microsatellites are repeated small sequence motifs that are highly polymorphic and abundant in the genomes of eukaryotes. Often they are the molecular markers of choice. To aid the development of microsatellite markers we have developed a module that integrates a program for the detection of microsatellites (TROLL), with the sequence assembly and analysis software, the Staden Package. The module has easily adjustable parameters for microsatellite lengths and base pair quality control. Starting with large datasets of unassembled sequence data in the form of chromatograms and/or text data, it enables the creation of a compact database consisting of the processed and assembled microsatellite containing sequences. For the final phase of primer design, we developed a program that accepts the multi-sequence ‘experiment file’ format as input and produces a list of primer pairs for amplification of microsatellite markers. The program can take into account the quality values of consensus bases, improving success rate of primer pairs in PCR. The software is freely available and simple to install in both Windows and Unix-based operating systems. Here we demonstrate the software by developing primer pairs for 427 new candidate markers for peanut. PMID:16493138

  9. New softwares for automated microsatellite marker development.

    PubMed

    Martins, Wellington; de Sousa, Daniel; Proite, Karina; Guimarães, Patrícia; Moretzsohn, Marcio; Bertioli, David

    2006-02-21

    Microsatellites are repeated small sequence motifs that are highly polymorphic and abundant in the genomes of eukaryotes. Often they are the molecular markers of choice. To aid the development of microsatellite markers we have developed a module that integrates a program for the detection of microsatellites (TROLL), with the sequence assembly and analysis software, the Staden Package. The module has easily adjustable parameters for microsatellite lengths and base pair quality control. Starting with large datasets of unassembled sequence data in the form of chromatograms and/or text data, it enables the creation of a compact database consisting of the processed and assembled microsatellite containing sequences. For the final phase of primer design, we developed a program that accepts the multi-sequence 'experiment file' format as input and produces a list of primer pairs for amplification of microsatellite markers. The program can take into account the quality values of consensus bases, improving success rate of primer pairs in PCR. The software is freely available and simple to install in both Windows and Unix-based operating systems. Here we demonstrate the software by developing primer pairs for 427 new candidate markers for peanut.

  10. Wide variation in microsatellite sequences within each Pfcrt mutant haplotype.

    PubMed

    Vinayak, Sumiti; Mittra, Pooja; Sharma, Yagya D

    2006-05-01

    Flanking microsatellites for each of the Pfcrt mutant haplotype of Plasmodium falciparum remain conserved among geographical isolates. We describe here heterogeneity in the intragenic microsatellites among each of the Pfcrt haplotype. There were fourteen different alleles of AT repeats of intron 2 and eight alleles of TA repeats of intron 4 of the pfcrt gene among Indian isolates. This resulted in 33 different two-locus (intron 2 plus intron 4) microsatellite genotypes among 224 isolates. There were 15 different two-locus microsatellite genotypes within the South American Pfcrt haplotype (S72V73M74N75T76S220) and 11 genotypes in the southeast Asian haplotype (C72V73I74E75T76S220) in these isolates. Indian isolates with Pfcrt haplotype C72V73I74E75T76S220 shared one of its two-locus microsatellite genotype with southeast Asian P. falciparum parasite lines from Thailand (K1) and Indochina (Dd2 and W2). Conversely, Indian isolates containing S72V73M74N75T76S220 Pfcrt haplotype did not share any of their two-locus microsatellite genotype with South American parasite line 7G8 from Brazil. Significantly, large number of newer two-locus microsatellite genotypes were detected in a 2-year time period (P<0.05). Microsatellite variation was more prominent in the areas of high malaria transmission. It is concluded that the genetic recombination in the intragenic microsatellites continues in the parasite population even after microsatellites flanking the pfcrt gene had already been fixed. Presence of various Pfcrt haplotypes and a variety of intragenic microsatellites indicates that there is a wide spectrum of chloroquine resistant parasite population in India. This information should be useful for malaria control programs of the country.

  11. Analysis of new microsatellite markers developed from reported sequences of Japanese flounder Paralichthys olivaceus

    NASA Astrophysics Data System (ADS)

    Yu, Haiyang; Jiang, Liming; Chen, Wei; Wang, Xubo; Wang, Zhigang; Zhang, Quanqi

    2010-12-01

    The expressed sequence tags (ESTs) of Japanese flounder, Paralichthys olivaceus, were selected from GenBank to identify simple sequence repeats (SSRs) or microsatellites. A bioinformatic analysis of 11111 ESTs identified 751 SSR-containing ESTs, including 440 dinucleotide, 254 trinucleotide, 53 tetranucleotide, 95 pentanucleotide and 40 hexanucleotide microsatellites respectively. The CA/TG and GA/TC repeats were the most abundant microsatellites. AT-rich types were predominant among trinucleotide and tetranucleotide microsatellites. PCR primers were designed to amplify 10 identified microsatellites loci. The PCR results from eight pairs of primers showed polymorphisms in wild populations. In 30 wild individuals, the mean observed and expected heterozygosities of these 8 polymorphic SSRs were 0.71 and 0.83 respectively and the average PIC value was 0.8. These microsatellite markers should prove to be a useful addition to the microsatellite markers that are now available for this species.

  12. Microsatellite primer resource for Populus developed from

    SciTech Connect

    Yin, Tongming; Yang, Xiaohan; Gunter, Lee E; Tuskan, Gerald A; Wullschleger, Stan D; Huang, Prof. Minren; Li, Shuxian; Zhang, Xinye

    2008-01-01

    In this study, 148 428 simple sequence repeat (SSR) primer pairs were designed from the unambiguously mapped sequence scaffolds of the Nisqually-1 genome. The physical position of the priming sites were identified along each of the 19 Populus chromosomes, and it was specified whether the priming sequences belong to intronic, intergenic, exonic or UTR regions. A subset of 150 SSR loci were amplified and a high amplification success rate (72%) was obtained in P. tremuloides, which belongs to a divergent subgenus of Populus relative to Nisqually-1. PCR reactions showed that the amplification success rate of exonic primer pairs was much higher than that of the intronic/intergenic primer pairs. Applying ANOVA and regression analyses to the flanking sequences of microsatellites, the repeat lengths, the GC contents of the repeats, the repeat motif numbers, the repeat motif length and the base composition of the repeat motif, it was determined that only the base composition of the repeat motif and the repeat motif length significantly affect the microsatellite variability in P. tremuloides samples. The SSR primer resource developed in this study provides a database for selecting highly transferable SSR markers with known physical position in the Populus genome and provides a comprehensive genetic tool to extend the genome sequence of Nisqually-1 to genetic studies in different Populus species.

  13. Microsatellite variation in red-winged blackbirds (Agelaius phoeniceus).

    PubMed

    Williams, C Lenney; Homan, H J; Johnston, J J; Linz, G M

    2004-02-01

    Territorial male red-winged blackbirds from five locations in the United States and Canada were genotyped using a suite of six microsatellite loci. Each population possessed unique alleles, but numbers of alleles per locus (range = 7.3-8.8) and expected multilocus heterozygosities (range = 0.76-0.80) were similar in all populations. Significant overall allele frequency differences were detected between some population pairs, and some pairwise Fst values were significant (but small). However, Fst among populations, although significant, was also small (0.009). Despite revealing low levels of population structure, the high multilocus polymorphism indicates these loci will be valuable in the genetic analysis of behavior and reproductive strategies in this species.

  14. Simple sequence repeat markers useful for sorghum downy mildew (Peronosclerospora sorghi) and related species

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Among the 55 primers pairs designed from clones from pathotype 3 of P. sorghi, 36 flanked microsatellite loci containing simple repeats, including 28 (55%) with dinucleotide repeats and 6 (11%) with trinucleotide repeats. A total of 22 microsatellites with CA/AC or GT/TG repeats were the most abund...

  15. The microsatellites and minisatellites in the genome of Fenneropenaeus chinensis.

    PubMed

    Gao, Huan; Kong, Jie

    2005-12-01

    Through two-time sequencing randomly in Fenneropenaeus chinensis, 2,597,000 bp cumulative length random genomic sequences about occupying 1.23 per thousand of the entire genome are obtained, in which the length of the first time sequencing is 884,000 bp, by cutting the genome DNA with Sau3AI enzyme, and the second is 1,713,000 bp by breaking the genome DNA with the physical method, ultrasonic. Using tandem repeat finder (TRF) soft to analyze the sequences, 4,588 tandem repeats are found, in which the number of microsatellites (1-6 bp) is 3,888, and 700 for minisatellites ( >or= 7 bp). The cumulative length of repeats is 305,555 bp, accounting for 11.72% of total cumulative sequence length, in which the cumulative length of microsatellites is 232,979 bp, accounting for 8.97% of total sequence length, and greater than those of other organisms, such as human and mosquito, etc. The dinucleotide repeat type is dominant in which the dominant repeat class is AT. The second abundant repeat type is trinucleotide, of which the dominant repeat class is AAT. Interestingly, of all of repeat types, the repeat numbers and repeat classes of primer number repeat types, such as pentanucleotide, heptanucleotide, elevennucleotide, etc. are less than those of repeat types beside them. The phenomena may involve the genesis and the evolution of microsatellites and minisatellites.

  16. Lessons learned from microsatellite development for nonmodel organisms using 454 pyrosequencing.

    PubMed

    Schoebel, C N; Brodbeck, S; Buehler, D; Cornejo, C; Gajurel, J; Hartikainen, H; Keller, D; Leys, M; Ríčanová, S; Segelbacher, G; Werth, S; Csencsics, D

    2013-03-01

    Microsatellites, also known as simple sequence repeats (SSRs), are among the most commonly used marker types in evolutionary and ecological studies. Next Generation Sequencing techniques such as 454 pyrosequencing allow the rapid development of microsatellite markers in nonmodel organisms. 454 pyrosequencing is a straightforward approach to develop a high number of microsatellite markers. Therefore, developing microsatellites using 454 pyrosequencing has become the method of choice for marker development. Here, we describe a user friendly way of microsatellite development from 454 pyrosequencing data and analyse data sets of 17 nonmodel species (plants, fungi, invertebrates, birds and a mammal) for microsatellite repeats and flanking regions suitable for primer development. We then compare the numbers of successfully lab-tested microsatellite markers for the various species and furthermore describe diverse challenges that might arise in different study species, for example, large genome size or nonpure extraction of genomic DNA. Successful primer identification was feasible for all species. We found that in species for which large repeat numbers are uncommon, such as fungi, polymorphic markers can nevertheless be developed from 454 pyrosequencing reads containing small repeat numbers (five to six repeats). Furthermore, the development of microsatellite markers for species with large genomes was also with Next Generation Sequencing techniques more cost and time-consuming than for species with smaller genomes. In this study, we showed that depending on the species, a different amount of 454 pyrosequencing data might be required for successful identification of a sufficient number of microsatellite markers for ecological genetic studies.

  17. Multiplexed microsatellite recovery using massively parallel sequencing.

    PubMed

    Jennings, T N; Knaus, B J; Mullins, T D; Haig, S M; Cronn, R C

    2011-11-01

    Conservation and management of natural populations requires accurate and inexpensive genotyping methods. Traditional microsatellite, or simple sequence repeat (SSR), marker analysis remains a popular genotyping method because of the comparatively low cost of marker development, ease of analysis and high power of genotype discrimination. With the availability of massively parallel sequencing (MPS), it is now possible to sequence microsatellite-enriched genomic libraries in multiplex pools. To test this approach, we prepared seven microsatellite-enriched, barcoded genomic libraries from diverse taxa (two conifer trees, five birds) and sequenced these on one lane of the Illumina Genome Analyzer using paired-end 80-bp reads. In this experiment, we screened 6.1 million sequences and identified 356,958 unique microreads that contained di- or trinucleotide microsatellites. Examination of four species shows that our conversion rate from raw sequences to polymorphic markers compares favourably to Sanger- and 454-based methods. The advantage of multiplexed MPS is that the staggering capacity of modern microread sequencing is spread across many libraries; this reduces sample preparation and sequencing costs to less than $400 (USD) per species. This price is sufficiently low that microsatellite libraries could be prepared and sequenced for all 1373 organisms listed as 'threatened' and 'endangered' in the United States for under $0.5 M (USD).

  18. Multiplexed microsatellite recovery using massively parallel sequencing

    USGS Publications Warehouse

    Jennings, T.N.; Knaus, B.J.; Mullins, T.D.; Haig, S.M.; Cronn, R.C.

    2011-01-01

    Conservation and management of natural populations requires accurate and inexpensive genotyping methods. Traditional microsatellite, or simple sequence repeat (SSR), marker analysis remains a popular genotyping method because of the comparatively low cost of marker development, ease of analysis and high power of genotype discrimination. With the availability of massively parallel sequencing (MPS), it is now possible to sequence microsatellite-enriched genomic libraries in multiplex pools. To test this approach, we prepared seven microsatellite-enriched, barcoded genomic libraries from diverse taxa (two conifer trees, five birds) and sequenced these on one lane of the Illumina Genome Analyzer using paired-end 80-bp reads. In this experiment, we screened 6.1 million sequences and identified 356958 unique microreads that contained di- or trinucleotide microsatellites. Examination of four species shows that our conversion rate from raw sequences to polymorphic markers compares favourably to Sanger- and 454-based methods. The advantage of multiplexed MPS is that the staggering capacity of modern microread sequencing is spread across many libraries; this reduces sample preparation and sequencing costs to less than $400 (USD) per species. This price is sufficiently low that microsatellite libraries could be prepared and sequenced for all 1373 organisms listed as 'threatened' and 'endangered' in the United States for under $0.5M (USD).

  19. Microsatellite instability in early sporadic breast cancer.

    PubMed Central

    Shaw, J. A.; Walsh, T.; Chappell, S. A.; Carey, N.; Johnson, K.; Walker, R. A.

    1996-01-01

    We have studied the incidence of microsatellite instability at three trinucleotide repeats and seven dinucleotide repeats from five chromosomal regions, in a group of 30 mammographically detected 'early' invasive breast cancers and correlated its occurrence with clinicopathological parameters. The myotonic dystrophy (DM-1) trinucleotide repeat was analysed in 48 additional cases. In 4 out of 78 (5%) paired tumour-normal DNA samples we found evidence of somatic microsatellite instability at DM-1: a novel allele of a different size was seen in the tumour DNA which was not present in the normal DNA sample. All four tumours that showed evidence of instability were from the core group of 30 cases (13%) and were well or moderately differentiated, oestrogen receptor-positive, infiltrating ductal carcinomas. Two of these tumours were unstable at nine of ten loci studied, both trinucleotide and dinucleotide repeats. DNA prepared from different normal tissues showed no evidence of instability, for all four instability cases. These data indicate that microsatellite instability is specific to the tumour DNA and is an early event in the genesis of some sporadic breast cancers. Images Figure 1 PMID:8645585

  20. Application of Microsatellite Loci for Molecular Identification of Elite Genotypes, Analysis of Clonality, and Genetic Diversity in Aspen Populus tremula L. (Salicaceae)

    PubMed Central

    Politov, Dmitry V.; Belokon, Maryana M.; Belokon, Yuri S.; Polyakova, Tatyana A.; Shatokhina, Anna V.; Mudrik, Elena A.; Azarova, Anna B.; Filippov, Mikhail V.; Shestibratov, Konstantin A.

    2015-01-01

    Testing systems for molecular identification of micropropagated elite aspen (Populus tremula L.) genotypes were developed on the base on microsatellite (SSR) loci. Out of 33 tested microsatellite loci, 14 were selected due to sustainable PCR amplification and substantial variability in elite clones of aspen aimed for establishment of fast-rotated forest plantations. All eight tested clones had different multilocus genotypes. Among 114 trees from three reference native stands located near the established plantations, 80 haplotypes were identified while some repeated genotypes were attributed to natural clones which appeared as a result of sprouting. The selected set of SSR markers showed reliable individual identification with low probability of appearance of identical aspen genotypes (a minimum of 4.8 · 10−10 and 1 × 10−4 for unrelated and related individuals, resp.). Case studies demonstrating practical applications of the test system are described including analysis of clonal structure and levels of genetic diversity in three natural aspen stands growing in the regions where plantations made of elite clones were established. PMID:26823661

  1. Analysis of microsatellites derived from bee Ests.

    PubMed

    Li, Bin; Xia, Qing-You; Lu, Cheng; Zhou, Ze-Yang

    2004-10-01

    To accelerate the molecular analysis of genetics,evolution and behavior, etc. in the honey bee (Apis), the frequency and density of simple sequence repeats (SSRs) have been analyzed from the bee EST (expressed sequence tag) database comprising 15 869 sequences amounting to 7.9 Mb. Results showed that the frequency of SSRs was 1/0.52 kb in bee ESTs, and hexanucleotide repeats (45.0%) motifs appeared to be the most abundant type in bee,the dinucleotide, mononucleotide, trinucleotide, tetranucleotide and pentanucleotide repeats are 17.9%, 14.1%, 11.6%, 9.2% and 2.2%, respectively. Meanwhile, the A-rich repeats are predominant in each type of SSRs, such as A, AT, AG, AC, AAT, AAG, AAC, AAAT, AAAG, AAAAG, AAAAT, AATAT, AAAAAG and AAAAAT repeats, whereas G-rich repeats are rare in the coding regions. The further analysis suggests that, apart from minor deviations, there is no significant difference in the distribution and density of microsatellites in the redundant and non-redundant set of bee ESTs. Furthermore, the availability of microsatellite markers can be expected to enhance the power and resolution of genome analysis in bee.

  2. Development of Seven Microsatellite Markers Using Next Generation Sequencing for the Conservation on the Korean Population of Dorcus hopei (E. Saunders, 1854) (Coleoptera, Lucanidae)

    PubMed Central

    Kang, Tae Hwa; Han, Sang Hoon; Park, Sun Jae

    2015-01-01

    We developed microsatellite markers for genetic structural analyses of Dorcus hopei, a stag beetle species, using next generation sequencing and polymerase chain reaction (PCR)-based genotyping for regional populations. A total of 407,070,351 base pairs of genomic DNA containing >4000 microsatellite loci except AT repeats were sequenced. From 76 loci selected for primer design, 27 were polymorphic. Of these 27 markers, 10 were tested on three regional populations: two Chinese (Shichuan and Guangxi) and one Korean (Wanju). Three markers were excluded due to inconsistent amplification, genotyping errors, and Hardy-Weinberg equilibrium (HWE). By multi-locus genotyping, the allele number, observed heterozygosity and polymorphism information content of seven microsatellite loci were ranged 2‒10, 0.1333‒1.0000, and 0.1228‒0.8509, respectively. In an analysis on the genetic differentiation among regional populations including one Japanese population and one cross-breeding population, the individual colored bar-plots showed that both Chinese populations were closer to each other than to the Far East Asian populations. In Far East Asian populations, Wanju and Nirasaki populations could not be distinguished from each other because the frequency of genetic contents was very similar in some individuals of two populations. Moreover, the cross-breeding population contained all patterns of genetic contents shown in Chinese, Korean, and Japanese populations, compared with the genetic content frequency of each regional population. As a result, we examined whether the cross-breeding population might be a hybrid population, and might contain a possibility of interbreeding with Chinese populations in parental generations. Therefore, these markers will be useful for analyses of genetic diversity in populations, genetic relationships between regional populations, genetic structure analyses, and origin tests. PMID:26370965

  3. Development of Seven Microsatellite Markers Using Next Generation Sequencing for the Conservation on the Korean Population of Dorcus hopei (E. Saunders, 1854) (Coleoptera, Lucanidae).

    PubMed

    Kang, Tae Hwa; Han, Sang Hoon; Park, Sun Jae

    2015-09-07

    We developed microsatellite markers for genetic structural analyses of Dorcus hopei, a stag beetle species, using next generation sequencing and polymerase chain reaction (PCR)-based genotyping for regional populations. A total of 407,070,351 base pairs of genomic DNA containing >4000 microsatellite loci except AT repeats were sequenced. From 76 loci selected for primer design, 27 were polymorphic. Of these 27 markers, 10 were tested on three regional populations: two Chinese (Shichuan and Guangxi) and one Korean (Wanju). Three markers were excluded due to inconsistent amplification, genotyping errors, and Hardy-Weinberg equilibrium (HWE). By multi-locus genotyping, the allele number, observed heterozygosity and polymorphism information content of seven microsatellite loci were ranged 2-10, 0.1333-1.0000, and 0.1228-0.8509, respectively. In an analysis on the genetic differentiation among regional populations including one Japanese population and one cross-breeding population, the individual colored bar-plots showed that both Chinese populations were closer to each other than to the Far East Asian populations. In Far East Asian populations, Wanju and Nirasaki populations could not be distinguished from each other because the frequency of genetic contents was very similar in some individuals of two populations. Moreover, the cross-breeding population contained all patterns of genetic contents shown in Chinese, Korean, and Japanese populations, compared with the genetic content frequency of each regional population. As a result, we examined whether the cross-breeding population might be a hybrid population, and might contain a possibility of interbreeding with Chinese populations in parental generations. Therefore, these markers will be useful for analyses of genetic diversity in populations, genetic relationships between regional populations, genetic structure analyses, and origin tests.

  4. An analysis of microsatellite loci in Arabidopsis thaliana: mutational dynamics and application.

    PubMed Central

    Symonds, V Vaughan; Lloyd, Alan M

    2003-01-01

    Microsatellite loci are among the most commonly used molecular markers. These loci typically exhibit variation for allele frequency distribution within a species. However, the factors contributing to this variation are not well understood. To expand on the current knowledge of microsatellite evolution, 20 microsatellite loci were examined for 126 accessions of the flowering plant, Arabidopsis thaliana. Substantial variability in mutation pattern among loci was found, most of which cannot be explained by the assumptions of the traditional stepwise mutation model or infinite alleles model. Here it is shown that the degree of locus diversity is strongly correlated with the number of contiguous repeats, more so than with the total number of repeats. These findings support a strong role for repeat disruptions in stabilizing microsatellite loci by reducing the substrate for polymerase slippage and recombination. Results of cluster analyses are also presented, demonstrating the potential of microsatellite loci for resolving relationships among accessions of A. thaliana. PMID:14668396

  5. Mature microsatellites: mechanisms underlying dinucleotide microsatellite mutational biases in human cells.

    PubMed

    Baptiste, Beverly A; Ananda, Guruprasad; Strubczewski, Noelle; Lutzkanin, Andrew; Khoo, Su Jen; Srikanth, Abhinaya; Kim, Nari; Makova, Kateryna D; Krasilnikova, Maria M; Eckert, Kristin A

    2013-03-01

    Dinucleotide microsatellites are dynamic DNA sequences that affect genome stability. Here, we focused on mature microsatellites, defined as pure repeats of lengths above the threshold and unlikely to mutate below it in a single mutational event. We investigated the prevalence and mutational behavior of these sequences by using human genome sequence data, human cells in culture, and purified DNA polymerases. Mature dinucleotides (≥10 units) are present within exonic sequences of >350 genes, resulting in vulnerability to cellular genetic integrity. Mature dinucleotide mutagenesis was examined experimentally using ex vivo and in vitro approaches. We observe an expansion bias for dinucleotide microsatellites up to 20 units in length in somatic human cells, in agreement with previous computational analyses of germ-line biases. Using purified DNA polymerases and human cell lines deficient for mismatch repair (MMR), we show that the expansion bias is caused by functional MMR and is not due to DNA polymerase error biases. Specifically, we observe that the MutSα and MutLα complexes protect against expansion mutations. Our data support a model wherein different MMR complexes shift the balance of mutations toward deletion or expansion. Finally, we show that replication fork progression is stalled within long dinucleotides, suggesting that mutational mechanisms within long repeats may be distinct from shorter lengths, depending on the biochemistry of fork resolution. Our work combines computational and experimental approaches to explain the complex mutational behavior of dinucleotide microsatellites in humans.

  6. Microsatellites in the Genome of the Edible Mushroom, Volvariella volvacea

    PubMed Central

    Chen, Mingjie; Wang, Hong; Bao, Dapeng

    2014-01-01

    Using bioinformatics software and database, we have characterized the microsatellite pattern in the V. volvacea genome and compared it with microsatellite patterns found in the genomes of four other edible fungi: Coprinopsis cinerea, Schizophyllum commune, Agaricus bisporus, and Pleurotus ostreatus. A total of 1346 microsatellites have been identified, with mono-nucleotides being the most frequent motif. The relative abundance of microsatellites was lower in coding regions with 21 No./Mb. However, the microsatellites in the V. volvacea gene models showed a greater tendency to be located in the CDS regions. There was also a higher preponderance of trinucleotide repeats, especially in the kinase genes, which implied a possible role in phenotypic variation. Among the five fungal genomes, microsatellite abundance appeared to be unrelated to genome size. Furthermore, the short motifs (mono- to tri-nucleotides) outnumbered other categories although these differed in proportion. Data analysis indicated a possible relationship between the most frequent microsatellite types and the genetic distance between the five fungal genomes. PMID:24575404

  7. Complex evolution of a highly conserved microsatellite locus in several fish species.

    PubMed

    Liu, J-X; Ely, B

    2009-08-01

    The evolutionary dynamics of a highly conserved microsatellite locus (Dla 11) were studied in several fish species. The data indicated that multiple types of compound microsatellites arose through point mutations that were sometimes followed by expansion of the derived motif. Furthermore, extensive length variation was detected among species in the regions immediately flanking the repeat region.

  8. Microsatellites in Pursuit of Microbial Genome Evolution

    PubMed Central

    Saeed, Abdullah F.; Wang, Rongzhi; Wang, Shihua

    2016-01-01

    Microsatellites or short sequence repeats are widespread genetic markers which are hypermutable 1–6 bp long short nucleotide motifs. Significantly, their applications in genetics are extensive due to their ceaseless mutational degree, widespread length variations and hypermutability skills. These features make them useful in determining the driving forces of evolution by using powerful molecular techniques. Consequently, revealing important questions, for example, what is the significance of these abundant sequences in DNA, what are their roles in genomic evolution? The answers of these important questions are hidden in the ways these short motifs contributed in altering the microbial genomes since the origin of life. Even though their size ranges from 1 –to- 6 bases, these repeats are becoming one of the most popular genetic probes in determining their associations and phylogenetic relationships in closely related genomes. Currently, they have been widely used in molecular genetics, biotechnology and evolutionary biology. However, due to limited knowledge; there is a significant gap in research and lack of information concerning hypermutational mechanisms. These mechanisms play a key role in microsatellite loci point mutations and phase variations. This review will extend the understandings of impacts and contributions of microsatellite in genomic evolution and their universal applications in microbiology. PMID:26779133

  9. Isolation of novel microsatellites using FIASCO by dual probe enrichment from Jatropha curcas L. and study on genetic equilibrium and diversity of Indian population revealed by isolated microsatellites.

    PubMed

    Sudheer, Pamidimarri D V N; Rahman, Hifzur; Mastan, Shaik G; Reddy, Muppala P

    2010-12-01

    Jatropha curcas L. belongs to family Euphorbiaceae, native to South America attained significant importance for its seed oil which can be converted to biodiesel, a renewable energy source alternative to conventional petrodiesel. Very few attempts were made to isolate novel microsatellite markers and assessment of the extent of genetic equilibrium and diversity that exists in J. curcas. Therefore, the present investigation was undertaken to isolate the novel microsatellites and access genetic equilibrium, diversity that exists among 44 diverse germplasm collected from distinct geographical areas in India using isolated microsatellites. The overall efficiency of the enrichment of microsatellite by dual probe in the present study found to be 54% and among the sequences obtained the percentage of sequences having suitable flanking regions for the primer designing was found to be 89.58%. The mean co-efficient of genetic similarity (CGS) was found to be 0.97. The overall diversity obtained by microsatellites was found to be low in comparison with the diversity reported by multilocus markers systems observed in earlier studies; however, the good allele polymorphism was observed. The overall dendrogram of microsatellite analysis resulted in random clustering of germplasm and not in accordance to geographical area of collection. The present study, diversity analysis using microsatellite markers concludes the low genetic diversity and genetic disequlibrium of J. curcas in India and will provide pavement for further intra-population studies on narrow geographical areas to understand the population genetic structure, phylogeography and molecular ecological studies. The germplasm characterized, and the microsatellite markers isolated and characterized in the present study can be employed efficiently in breeding programs for genetic improvement of the species through marker assisted selection and QTL analysis, for further genetic resource management and help in making the J

  10. Microsatellite-Based Fingerprinting of Western Blackberries from Plants, IQF Berries and Puree

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The blackberry industry needs a reliable method to ensure trueness-to-type of blackberry products. Microsatellite markers or simple sequence repeats (SSRs) are ideal for cultivar fingerprinting, paternity testing and identity certification. Fingerprinting is valuable for variety identification, qual...

  11. Multilocus enzyme analysis of Legionella dumoffii.

    PubMed Central

    Woods, T C; McKinney, R M; Plikaytis, B D; Steigerwalt, A G; Bibb, W F; Brenner, D J

    1988-01-01

    Variability among 29 clinical and environmental strains of Legionella dumoffii was investigated by multilocus enzyme analysis by use of starch gel electrophoresis. Based on results of analysis at 20 enzyme loci, the strains were separated into five closely related electrophoretic types (ETs), which were clearly distinguished from 53 strains representing 53 ETs of L. pneumophila. DNA hybridization results (hydroxyapatite method, 60 and 75 degrees C) for representative strains confirmed that all L. dumoffii ETs were a single genetic species. Although multilocus enzyme analysis indicated that L. dumoffii was genetically a quite uniform species, sufficient variability existed to warrant electromorph fingerprinting for epidemiologic studies. PMID:3384905

  12. Analysis of microsatellite polymorphism in inbred knockout mice.

    PubMed

    Zuo, Baofen; Du, Xiaoyan; Zhao, Jing; Yang, Huixin; Wang, Chao; Wu, Yanhua; Lu, Jing; Wang, Ying; Chen, Zhenwen

    2012-01-01

    Previously, we found that the genotype of 42 out of 198 mouse microsatellite loci, which are distributed among all chromosomes except the Y chromosome, changed from monomorphism to polymorphism (CMP) in a genetically modified inbred mouse strain. In this study, we further examined whether CMP also relates to the homologous recombination in gene knockout (KO) mouse strains. The same 42 microsatellite loci were analyzed by polymerase chain reaction (PCR) in 29 KO inbred mouse strains via short tandem sequence repeat (STR) scanning and direct sequence cloning to justify microsatellite polymorphisms. The C57BL/6J and 129 mouse strains, from which these 29 KO mice were derived, were chosen as the background controls. The results indicated that 10 out of 42 (23.8%) loci showed CMP in some of these mouse strains. Except for the trinucleotide repeat locus of D3Mit22, which had microsatellite CMP in strain number 9, the core sequences of the remaining 41 loci were dinucleotide repeats, and 9 out of 41 (21.95%) showed CMPs among detected mouse strains. However, 11 out of 29 (37.9%) KO mice strains were recognized as having CMPs. The popular dinucleotide motifs in CMP were (TG)(n) (50%, 2/4), followed by (GT)(n) (27.27%, 3/11) and (CA)(n) (23.08%, 3/13). The microsatellite CMP in (CT)(n) and (AG)(n) repeats were 20% (1/5). According to cloning sequencing results, 6 KO mouse strains showed insertions of nucleotides whereas 1 showed a deletion. Furthermore, 2 loci (D13Mit3 and D14Mit102) revealed CMP in 2 strains, and mouse strain number 9 showed CMPs in two loci (D3Mit22 and D13Mit3) simultaneously. Collectively, these results indicated that microsatellite polymorphisms were present in the examined inbred KO mice.

  13. Analysis of Microsatellite Polymorphism in Inbred Knockout Mice

    PubMed Central

    Zhao, Jing; Yang, Huixin; Wang, Chao; Wu, Yanhua; Lu, Jing; Wang, Ying; Chen, Zhenwen

    2012-01-01

    Previously, we found that the genotype of 42 out of 198 mouse microsatellite loci, which are distributed among all chromosomes except the Y chromosome, changed from monomorphism to polymorphism (CMP) in a genetically modified inbred mouse strain. In this study, we further examined whether CMP also relates to the homologous recombination in gene knockout (KO) mouse strains. The same 42 microsatellite loci were analyzed by polymerase chain reaction (PCR) in 29 KO inbred mouse strains via short tandem sequence repeat (STR) scanning and direct sequence cloning to justify microsatellite polymorphisms. The C57BL/6J and 129 mouse strains, from which these 29 KO mice were derived, were chosen as the background controls. The results indicated that 10 out of 42 (23.8%) loci showed CMP in some of these mouse strains. Except for the trinucleotide repeat locus of D3Mit22, which had microsatellite CMP in strain number 9, the core sequences of the remaining 41 loci were dinucleotide repeats, and 9 out of 41 (21.95%) showed CMPs among detected mouse strains. However, 11 out of 29 (37.9%) KO mice strains were recognized as having CMPs. The popular dinucleotide motifs in CMP were (TG)n (50%, 2/4), followed by (GT)n (27.27%, 3/11) and (CA)n (23.08%, 3/13). The microsatellite CMP in (CT)n and (AG)n repeats were 20% (1/5). According to cloning sequencing results, 6 KO mouse strains showed insertions of nucleotides whereas 1 showed a deletion. Furthermore, 2 loci (D13Mit3 and D14Mit102) revealed CMP in 2 strains, and mouse strain number 9 showed CMPs in two loci (D3Mit22 and D13Mit3) simultaneously. Collectively, these results indicated that microsatellite polymorphisms were present in the examined inbred KO mice. PMID:22509320

  14. Comparison of microsatellite distribution in genomes of Centruroides exilicauda and Mesobuthus martensii.

    PubMed

    Wang, Chen; Kubiak, Lukasz Jakub; Du, Lian-Ming; Li, Wu-Jiao; Jian, Zuo-Yi; Tang, Ce; Fan, Zhen-Xin; Zhang, Xiu-Yue; Yue, Bi-Song

    2016-12-05

    In this study, we characterized the distribution of microsatellites in the genomes and genes of Centruroides exilicauda and Mesobuthus martensii, carried out Gene Ontology (GO) analysis and GO enrichment analysis of coding sequences (CDSs) with microsatellite (SSR). In addition, over-represented GO functions related to environmental interactions, development process and methylation were identified to develop functional markers and facilitate further analysis of microsatellite function in the genes of scorpions. Location analysis indicated that microsatellites were predominantly concentrated at both ends of genes. Most genes containing microsatellite had the SSR present at only one locus, from which we infer that the number of SSRs per gene is limited even though intragenic tandem repeats can generate functional variability. Lastly, we identified 75 SSRs in 64 genes of 54 expanded gene families and 1 SSR in the toxin gene of Mesobuthus martensii, allowing future studies on the effect of microsatellites on gene function.

  15. Microsatellite instability confounds engraftment analysis of hematopoietic stem-cell transplantation.

    PubMed

    Tseng, Li-Hui; Tang, Jih-Luh; Haley, Lisa; Beierl, Katie; Gocke, Christopher D; Eshleman, James R; Lin, Ming-Tseh

    2014-07-01

    Polymorphic short tandem-repeat, or microsatellite, loci have been widely used to analyze chimerism status after allogeneic hematopoietic stem-cell transplantation. In molecular diagnostic laboratories, it is recommended to calculate mixed chimerism for at least 2 informative loci and to avoid microsatellite loci on chromosomes with copy number changes. In this report, we show that microsatellite instability observed in 2 patients with acute leukemia may confound chimerism analysis. Interpretation errors may occur even if 2 to 3 loci are analyzed because of length variation in multiple microsatellite loci. Although microsatellite loci with length variation should not be selected for chimerism analysis, the presence of microsatellite instability, like copy number alteration because of aberrant chromosomes, provides evidence of recurrent or residual cancer cells after hematopoietic stem-cell transplantation.

  16. Optimizing selection of microsatellite loci from 454 pyrosequencing via post-sequencing bioinformatic analyses.

    PubMed

    Fernandez-Silva, Iria; Toonen, Robert J

    2013-01-01

    The comparatively low cost of massive parallel sequencing technology, also known as next-generation sequencing (NGS), has transformed the isolation of microsatellite loci. The most common NGS approach consists of obtaining large amounts of sequence data from genomic DNA or enriched microsatellite libraries, which is then mined for the discovery of microsatellite repeats using bioinformatics analyses. Here, we describe a bioinformatics approach to isolate microsatellite loci, starting from the raw sequence data through a subset of microsatellite primer pairs. The primary difference to previously published approaches includes analyses to select the most accurate sequence data and to eliminate repetitive elements prior to the design of primers. These analyses aim to minimize the testing of primer pairs by identifying the most promising microsatellite loci.

  17. Using Next Generation RAD Sequencing to Isolate Multispecies Microsatellites for Pilosocereus (Cactaceae).

    PubMed

    Bonatelli, Isabel A S; Carstens, Bryan C; Moraes, Evandro M

    2015-01-01

    Microsatellite markers (also known as SSRs, Simple Sequence Repeats) are widely used in plant science and are among the most informative molecular markers for population genetic investigations, but the development of such markers presents substantial challenges. In this report, we discuss how next generation sequencing can replace the cloning, Sanger sequencing, identification of polymorphic loci, and testing cross-amplification that were previously required to develop microsatellites. We report the development of a large set of microsatellite markers for five species of the Neotropical cactus genus Pilosocereus using a restriction-site-associated DNA sequencing (RAD-seq) on a Roche 454 platform. We identified an average of 165 microsatellites per individual, with the absolute numbers across individuals proportional to the sequence reads obtained per individual. Frequency distribution of the repeat units was similar in the five species, with shorter motifs such as di- and trinucleotide being the most abundant repeats. In addition, we provide 72 microsatellites that could be potentially amplified in the sampled species and 22 polymorphic microsatellites validated in two populations of the species Pilosocereus machrisii. Although low coverage sequencing among individuals was observed for most of the loci, which we suggest to be more related to the nature of the microsatellite markers and the possible bias inserted by the restriction enzymes than to the genome size, our work demonstrates that an NGS approach is an efficient method to isolate multispecies microsatellites even in non-model organisms.

  18. Characterization of genome-wide microsatellites of Saccharina japonica based on a preliminary assembly of Illumina sequencing reads

    NASA Astrophysics Data System (ADS)

    Zhang, Linan; Peng, Jie; Li, Xiaojie; Cui, Cuiju; Sun, Juan; Yang, Guanpin

    2016-06-01

    Microsatellites or simple sequence repeats (SSR) function widely and locate dependently in genome. However, their characteristics are often ignored due to the lack of genomic sequences of most species. Kelp ( Saccharina japonica), a brown macroalga, is extensively cultured in China. In this study, the genome of S. japonica was surveyed using an Illumina sequencing platform, and its microsatellites were characterized. The preliminarily assembled genome was 469.4 Mb in size, with a scaffold N50 of 20529 bp. Among the 128370 identified microsatellites, 90671, 25726 and 11973 were found in intergenic regions, introns and exons, averaging 339.3, 178.8 and 205.4 microsatellites per Mb, respectively. These microsatellites distributed unevenly in S. japonica genome. Mononucleotide motifs were the most abundant in the genome, while trinucleotide ones were the most prevalent in exons. The microsatellite abundance decreased significantly with the increase of motif repeat numbers, and the microsatellites with a small number of repeats accounted for a higher proportion of the exons than those of the intergenic regions and introns. C/G-rich motifs were more common in exons than in intergenic regions and introns. These characteristics of microsatellites in S. japonica genome may associate with their functions, and ultimately their adaptation and evolution. Among the 120140 pairs of designed microsatellite primers, approximately 75% were predicted to be able to amplify S. japonica DNA. These microsatellite markers will be extremely useful for the genetic breeding and population evolution studies of kelp.

  19. Evolutionary dynamics of microsatellite distribution in plants: insight from the comparison of sequenced brassica, Arabidopsis and other angiosperm species.

    PubMed

    Shi, Jiaqin; Huang, Shunmou; Fu, Donghui; Yu, Jinyin; Wang, Xinfa; Hua, Wei; Liu, Shengyi; Liu, Guihua; Wang, Hanzhong

    2013-01-01

    Despite their ubiquity and functional importance, microsatellites have been largely ignored in comparative genomics, mostly due to the lack of genomic information. In the current study, microsatellite distribution was characterized and compared in the whole genomes and both the coding and non-coding DNA sequences of the sequenced Brassica, Arabidopsis and other angiosperm species to investigate their evolutionary dynamics in plants. The variation in the microsatellite frequencies of these angiosperm species was much smaller than those for their microsatellite numbers and genome sizes, suggesting that microsatellite frequency may be relatively stable in plants. The microsatellite frequencies of these angiosperm species were significantly negatively correlated with both their genome sizes and transposable elements contents. The pattern of microsatellite distribution may differ according to the different genomic regions (such as coding and non-coding sequences). The observed differences in many important microsatellite characteristics (especially the distribution with respect to motif length, type and repeat number) of these angiosperm species were generally accordant with their phylogenetic distance, which suggested that the evolutionary dynamics of microsatellite distribution may be generally consistent with plant divergence/evolution. Importantly, by comparing these microsatellite characteristics (especially the distribution with respect to motif type) the angiosperm species (aside from a few species) all clustered into two obviously different groups that were largely represented by monocots and dicots, suggesting a complex and generally dichotomous evolutionary pattern of microsatellite distribution in angiosperms. Polyploidy may lead to a slight increase in microsatellite frequency in the coding sequences and a significant decrease in microsatellite frequency in the whole genome/non-coding sequences, but have little effect on the microsatellite distribution with

  20. A Multiplexed Microsatellite Fingerprinting Set for Hazelnut Cultivar Identification

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The objective of this study was to develop a robust and cost-effective fingerprinting set for hazelnuts using microsatellite (SSR) markers. Twenty SSRs containing repeat motifs of = three nucleotides distributed throughout the hazelnut genome were screened on eight genetically diverse cultivars to a...

  1. Transferability of Rubus Microsatellite Markers for use in Black Raspberry

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Microsatellites or simple sequence repeats (SSRs) are valuable as co-dominant genetic markers with a variety of applications such as DNA fingerprinting, linkage mapping, and population structure analysis. To date, SSR marker development in Rubus has focused on red raspberry (Rubus idaeus L., subgenu...

  2. The art of attrition: development of robust oat microsatellites

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Microsatellite or simple sequence repeat (SSR) markers are important tools for genetic analyses, especially those targeting diversity, based on the fact that multiple alleles can occur at a given locus. Currently, only 160 genomic-based SSR markers are publicly available for oat, most of which have...

  3. Minisatellite and microsatellite length variation at a complex bovine VNTR locus.

    PubMed

    Nave, A; Kashi, Y; Soller, M

    1997-02-01

    Length variation at the BTGL1 variable number of tandem repeat (VNTR) locus, which includes both minisatellite and microsatellite motifs, was examined in a wide sample of cattle. A total of 22 alleles were uncovered. The distribution of allele size variation implicated mechanisms involving both minisatellite and microsatellite sequences. This was confirmed by direct sequences of two alleles. Differences between the two alleles involved repeat number variation of microsatellite motifs and a complex event involving the minisatellite motif, but point mutations were not observed.

  4. megasat: automated inference of microsatellite genotypes from sequence data.

    PubMed

    Zhan, Luyao; Paterson, Ian G; Fraser, Bonnie A; Watson, Beth; Bradbury, Ian R; Nadukkalam Ravindran, Praveen; Reznick, David; Beiko, Robert G; Bentzen, Paul

    2017-03-01

    megasat is software that enables genotyping of microsatellite loci using next-generation sequencing data. Microsatellites are amplified in large multiplexes, and then sequenced in pooled amplicons. megasat reads sequence files and automatically scores microsatellite genotypes. It uses fuzzy matches to allow for sequencing errors and applies decision rules to account for amplification artefacts, including nontarget amplification products, replication slippage during PCR (amplification stutter) and differential amplification of alleles. An important feature of megasat is the generation of histograms of the length-frequency distributions of amplification products for each locus and each individual. These histograms, analogous to electropherograms traditionally used to score microsatellite genotypes, enable rapid evaluation and editing of automatically scored genotypes. megasat is written in Perl, runs on Windows, Mac OS X and Linux systems, and includes a simple graphical user interface. We demonstrate megasat using data from guppy, Poecilia reticulata. We genotype 1024 guppies at 43 microsatellites per run on an Illumina MiSeq sequencer. We evaluated the accuracy of automatically called genotypes using two methods, based on pedigree and repeat genotyping data, and obtained estimates of mean genotyping error rates of 0.021 and 0.012. In both estimates, three loci accounted for a disproportionate fraction of genotyping errors; conversely, 26 loci were scored with 0-1 detected error (error rate ≤0.007). Our results show that with appropriate selection of loci, automated genotyping of microsatellite loci can be achieved with very high throughput, low genotyping error and very low genotyping costs.

  5. Highly Informative Simple Sequence Repeat (SSR) Markers for Fingerprinting Hazelnut

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Simple sequence repeat (SSR) or microsatellite markers have many applications in breeding and genetic studies of plants, including fingerprinting of cultivars and investigations of genetic diversity, and therefore provide information for better management of germplasm collections. They are repeatab...

  6. Polymorphic microsatellite markers in Euryale ferox Salisb. (Nymphaeaceae).

    PubMed

    Quan, Zhiwu; Pan, Lei; Ke, Weidong; Ding, Yi

    2009-01-01

    Eleven polymorphic microsatellite markers were isolated and identified in the aquatic plant Euryale ferox Salisb. (Nymphaeaceae). This species, which belongs to basal Magnoliophyta, reproduces sexually. All of these 11 microsatellite markers yielded 25 alleles in a survey of a wild population of 34 individuals. Two or three alleles per locus were detected, with expected heterozygosity ranging from 0.056 to 0.634 and observed heterozygosity from 0.000 to 0.088. These simple sequence repeat markers will be useful for evaluating the genetic structure of the E. ferox population in the future.

  7. Characterization of microsatellite loci isolated in trumpeter swan (Cygnus buccinator)

    USGS Publications Warehouse

    John, J. St; Ransler, F.A.; Quinn, T.W.; Oyler-McCance, S.J.

    2006-01-01

    Primers for 16 microsatellite loci were developed for the trumpeter swan (Cygnus buccinator), a species recovering from a recent population bottleneck. In a screen of 158 individuals, the 16 loci were found to have levels of variability ranging from two to seven alleles. No loci were found to be linked, although two loci repeatedly revealed significant departures from Hardy-Weinberg equilibrium. Amplification in the closely related tundra swan (Cygnus columbianus) was successful for all except one locus. These microsatellite loci will be applicable for population genetic analyses and ultimately aid in management efforts. ?? 2006 The Authors.

  8. Microsatellite analyses across three diverse vertebrate transcriptomes (Acipenser fulvescens, Ambystoma tigrinum, and Dipodomys spectabilis).

    PubMed

    Doyle, Jacqueline M; Siegmund, Gregor; Ruhl, Joseph D; Eo, Soo Hyung; Hale, Matthew C; Marra, Nicholas J; Waser, Peter M; Dewoody, J Andrew

    2013-07-01

    Historically, many population genetics studies have utilized microsatellite markers sampled at random from the genome and presumed to be selectively neutral. Recent studies, however, have shown that microsatellites can occur in transcribed regions, where they are more likely to be under selection. In this study, we mined microsatellites from transcriptomes generated by 454-pyrosequencing for three vertebrate species: lake sturgeon (Acipenser fulvescens), tiger salamander (Ambystoma tigrinum), and kangaroo rat (Dipodomys spectabilis). We evaluated (i) the occurrence of microsatellites across species; (ii) whether particular gene ontology terms were over-represented in genes that contained microsatellites; (iii) whether repeat motifs were located in untranslated regions or coding sequences of genes; and (iv) in silico polymorphism. Microsatellites were less common in tiger salamanders than in either lake sturgeon or kangaroo rats. Across libraries, trinucleotides were found more frequently than any other motif type, presumably because they do not cause frameshift mutations. By evaluating variation across reads assembled to a given contig, we were able to identify repeat motifs likely to be polymorphic. Our study represents one of the first comparative data sets on the distribution of vertebrate microsatellites within expressed genes. Our results reinforce the idea that microsatellites do not always occur in noncoding DNA, but commonly occur in expressed genes.

  9. Repeats in transforming acidic coiled-coil (TACC) genes.

    PubMed

    Trivedi, Seema

    2013-06-01

    Transforming acidic coiled-coil proteins (TACC1, 2, and 3) are essential proteins associated with the assembly of spindle microtubules and maintenance of bipolarity. Dysregulation of TACCs is associated with tumorigenesis, but studies of microsatellite instability in TACC genes have not been extensive. Microsatellite or simple sequence repeat instability is known to cause many types of cancer. The present in silico analysis of SSRs in human TACC gene sequences shows the presence of mono- to hexa-nucleotide repeats, with the highest densities found for mono- and di-nucleotide repeats. Density of repeats is higher in introns than in exons. Some of the repeats are present in regulatory regions and retained introns. Human TACC genes show conservation of many repeat classes. Microsatellites in TACC genes could be valuable markers for monitoring numerical chromosomal aberrations and or cancer.

  10. Cross-amplification and sequence variation of microsatellite loci in Eurasian hard pines.

    PubMed

    González-Martínez, S C; Robledo-Arnuncio, J J; Collada, C; Díaz, A; Williams, C G; Alía, R; Cervera, M T

    2004-06-01

    Microsatellite transfer across coniferous species is a valued methodology because de novo development for each species is costly and there are many species with only a limited commodity value. Cross-species amplification of orthologous microsatellite regions provides valuable information on mutational and evolutionary processes affecting these loci. We tested 19 nuclear microsatellite markers from Pinus taeda L. (subsection Australes) and three from P. sylvestris L. (subsection Pinus) on seven Eurasian hard pine species ( P. uncinata Ram., P. sylvestris L., P. nigra Arn., P. pinaster Ait., P. halepensis Mill., P. pinea L. and P. canariensis Sm.). Transfer rates to species in subsection Pinus (36-59%) were slightly higher than those to subsections Pineae and Pinaster (32-45%). Half of the trans-specific microsatellites were found to be polymorphic over evolutionary times of approximately 100 million years (ten million generations). Sequencing of three trans-specific microsatellites showed conserved repeat and flanking regions. Both a decrease in the number of perfect repeats in the non-focal species and a polarity for mutation, the latter defined as a higher substitution rate in the flanking sequence regions close to the repeat motifs, were observed in the trans-specific microsatellites. The transfer of microsatellites among hard pine species proved to be useful for obtaining highly polymorphic markers in a wide range of species, thereby providing new tools for population and quantitative genetic studies.

  11. Student's Microsatellite Project

    NASA Astrophysics Data System (ADS)

    Zelentsov, Victor; Kopik, Anatoliy; Karpenko, Stanislav; Mayorova, Victoria

    2002-01-01

    Nowadays BMSTU Youth space center carries on development of the microsatellite project. The project is based on principles of students direct involvement on all stages of development and maintenance of the satellite. The group of students was organized within the university with purpose of coordination of work at the program. Project current condition The work on creation of an experimental model of the micro satellite is performed. The aim is to define the structure and parameters of on-board devices (mass-overall dimensions characteristics, energy consumption and so on). developed. According to the simplified model an active stabilization system (three orthogonal electro-magnetic coils) and orientation characterization system (sunlight detector and magnitometer) are included in OCS structure. most suitable battery storage, power-supply controlling system. Student micro-satellite program goals 1.Scientific Information gaining in the field of Earth study- using perspective research methods. Studying of new devices behavior in space conditions. 2. Educative a. Students derive real experience of projecting, building of a spacecraft from the point of view of an experimenter, a constructor and a researcher. b. Organization of student's cooperation with key men of aerospace industry and other branches. c. Brainpower and material base preparation for micro-satellite systems' development. d. Attraction of youth interest to the topic, by: - Students' and pupils' groups attraction and involvement in experiments conduction and results processing. - Seminars and lections devoted to Earth study from the space organization - Specific scientific data distribution over World Wide Web. 3. International With purpose of program expansion, the developers' group looks to start of an international project. Within the project new experiments conduction and scientific information exchange are expected. 4. Status Bauman Moscow State Technical University's status improvement in the field

  12. Development of eighteen microsatellite loci in walleye (Sander vitreus)

    USGS Publications Warehouse

    Coykendall, Dolly K.; Morrison, Cheryl L.; Stott, Wendylee; Springmann, Marcus J.

    2014-01-01

    A suite of tri- and tetra-nucleotide microsatellite loci were developed for walleye (Sander vitreus) from 454 pyrosequencing data. Eighteen of the 50 primer sets tested amplified consistently in 35 walleye from two lakes on Isle Royale, Lake Superior: Chickenbone Lake and Whittlesey Lake. The loci displayed moderate levels of allelic diversity (average 5.5 alleles/locus) and heterozygosity (average 35.8 %). Levels of genetic diversity were sufficient to produce unique multi-locus genotypes and detect phylogeographic structuring as individuals assigned back to their population of origin. Cross-species amplification within S. canadensis(sauger) was successful for 15 loci, and 11 loci were diagnostic to species. The loci characterized here will be useful for detecting fine-scale spatial structuring, resolving the taxonomic status of Sander species and sub-species, and detecting walleye/sauger hybrids.

  13. Distribution, function and evolution characterization of microsatellite in Sargassum thunbergii (Fucales, Phaeophyta) transcriptome and their application in marker development.

    PubMed

    Liu, Fuli; Hu, Zimin; Liu, Wenhui; Li, Jingjing; Wang, Wenjun; Liang, Zhourui; Wang, Feijiu; Sun, Xiutao

    2016-01-06

    Using transcriptome data to mine microsatellite and develop markers has growingly become prevalent. However, characterizing the possible function of microsatellite is relatively rare. In this study, we explored microsatellites in the transcriptome of the brown alga Sargassum thunbergii and characterized the frequencies, distribution, function and evolution, and developed primers to validate these microsatellites. Our results showed that Tri-nucleotide is the most abundant, followed by di- and mono-nucleotide. The length of microsatellite was significantly affected by the repeat motif size. The density of microsatellite in the CDS region is significantly lower than that in the UTR region. The annotation of the transcripts containing microsatellite showed that 573 transcripts have GO terms and can be categorized into 42 groups. Pathways enrichment showed that microsatellites were significantly overrepresented in the genes involved in pathways such as Ubiquitin mediated proteolysis, RNA degradation, Spliceosome, etc. Primers flanking 961 microsatellite loci were designed, and among the 30 pairs of primer selected randomly for availability test, 23 were proved to be efficient. These findings provided new insight into the function and evolution of microsatellite in transcriptome, and the identified microsatellite loci within the annotated gene will be useful for developing functional markers in S. thunbergii.

  14. Distribution, function and evolution characterization of microsatellite in Sargassum thunbergii (Fucales, Phaeophyta) transcriptome and their application in marker development

    PubMed Central

    Liu, Fuli; Hu, Zimin; Liu, Wenhui; Li, Jingjing; Wang, Wenjun; Liang, Zhourui; Wang, Feijiu; Sun, Xiutao

    2016-01-01

    Using transcriptome data to mine microsatellite and develop markers has growingly become prevalent. However, characterizing the possible function of microsatellite is relatively rare. In this study, we explored microsatellites in the transcriptome of the brown alga Sargassum thunbergii and characterized the frequencies, distribution, function and evolution, and developed primers to validate these microsatellites. Our results showed that Tri-nucleotide is the most abundant, followed by di- and mono-nucleotide. The length of microsatellite was significantly affected by the repeat motif size. The density of microsatellite in the CDS region is significantly lower than that in the UTR region. The annotation of the transcripts containing microsatellite showed that 573 transcripts have GO terms and can be categorized into 42 groups. Pathways enrichment showed that microsatellites were significantly overrepresented in the genes involved in pathways such as Ubiquitin mediated proteolysis, RNA degradation, Spliceosome, etc. Primers flanking 961 microsatellite loci were designed, and among the 30 pairs of primer selected randomly for availability test, 23 were proved to be efficient. These findings provided new insight into the function and evolution of microsatellite in transcriptome, and the identified microsatellite loci within the annotated gene will be useful for developing functional markers in S. thunbergii. PMID:26732855

  15. Microsatellite markers for Senna spectabilis var. excelsa (Caesalpinioideae, Fabaceae)1

    PubMed Central

    López-Roberts, M. Cristina; Barbosa, Ariane R.; Paganucci de Queiroz, Luciano; van den Berg, Cássio

    2016-01-01

    Premise of the study: Senna spectabilis var. excelsa (Fabaceae) is a South and Central American tree of great ecological importance and one of the most common species in several sites of seasonally dry forests. Our goal was to develop microsatellite markers to assess the genetic diversity and structure of this species. Methods and Results: We designed and assessed 53 loci obtained from a microsatellite-enriched library and an intersimple sequence repeat library. Fourteen loci were polymorphic, and they presented a total of 39 alleles in a sample of 61 individuals from six populations. The mean values of observed and expected heterozygosities were 0.355 and 0.479, respectively. Polymorphism information content was 0.390 and the Shannon index was 0.778. Conclusions: Polymorphism information content and Shannon index indicate that at least nine of the 14 microsatellite loci developed are moderate to highly informative, and potentially useful for population genetic studies in this species. PMID:26819856

  16. High tandem repeat content in the genome of the short-lived annual fish Nothobranchius furzeri: a new vertebrate model for aging research

    PubMed Central

    Reichwald, Kathrin; Lauber, Chris; Nanda, Indrajit; Kirschner, Jeanette; Hartmann, Nils; Schories, Susanne; Gausmann, Ulrike; Taudien, Stefan; Schilhabel, Markus B; Szafranski, Karol; Glöckner, Gernot; Schmid, Michael; Cellerino, Alessandro; Schartl, Manfred; Englert, Christoph; Platzer, Matthias

    2009-01-01

    Background The annual fish Nothobranchius furzeri is the vertebrate with the shortest known life span in captivity. Fish of the GRZ strain live only three to four months under optimal laboratory conditions, show explosive growth, early sexual maturation and age-dependent physiological and behavioral decline, and express aging related biomarkers. Treatment with resveratrol and low temperature significantly extends the maximum life span. These features make N. furzeri a promising new vertebrate model for age research. Results To contribute to establishing N. furzeri as a new model organism, we provide a first insight into its genome and a comparison to medaka, stickleback, tetraodon and zebrafish. The N. furzeri genome contains 19 chromosomes (2n = 38). Its genome of between 1.6 and 1.9 Gb is the largest among the analyzed fish species and has, at 45%, the highest repeat content. Remarkably, tandem repeats comprise 21%, which is 4-12 times more than in the other four fish species. In addition, G+C-rich tandem repeats preferentially localize to centromeric regions. Phylogenetic analysis based on coding sequences identifies medaka as the closest relative. Genotyping of an initial set of 27 markers and multi-locus fingerprinting of one microsatellite provides the first molecular evidence that the GRZ strain is highly inbred. Conclusions Our work presents a first basis for systematic genomic and genetic analyses aimed at understanding the mechanisms of life span determination in N. furzeri. PMID:19210790

  17. Repeated praziquantel treatments remodel the genetic and spatial landscape of schistosomiasis risk and transmission.

    PubMed

    Barbosa, Lúcio M; Reis, Eliana A; Dos Santos, Cláudio R A; Costa, Jackson M; Carmo, Theomira M; Aminu, Peace T; Pitanga, Thassila N; Ponce-Terashima, Rafael; Blank, Walter A; Silva, Luciano K; Reis, Mitermayer G; Blanton, Ronald E

    2016-05-01

    Repeated treatments with praziquantel reduce schistosomiasis prevalence and morbidity, but transmission persists and populations often recover within a few years. To identify factors associated with persistence, we surveyed and treated all identified Schistosoma mansoni infections in two rural Brazilian communities (Jenipapo and Volta do Rio) in 2009, 2012 and 2013. Eggs were collected from all infected individuals and genotyped with 11 microsatellite markers to evaluate parasite differentiation and diversity. After successive rounds of community-wide treatment, prevalence decreased from 45% to 24% then 16%. Intensity of infection decreased by 57% over this period, and the number of eggs transmitted to the environment decreased by 92%. During all time periods the majority of eggs were excreted by those >15years of age. The incidence was 23% in 2012 and 15% in 2013, consistent with a decrease in transmission. There was little immigration or gene flow over a distance of 6km. On reinfection, infrapopulations were moderately differentiated indicating that pretreatment multilocus genotypes were not fully reacquired. The effective population size responded to census population decline more rapidly than differentiation. Reinfection was concentrated in the downstream portion of Jenipapo, consistent with the observed increased human fecal contamination. At this scale and in this area S. mansoni infections exist on a fragmented landscape with a highly focal pattern of transmission that may facilitate future elimination.

  18. Genome-wide survey and analysis of microsatellites in the Pacific oyster genome: abundance, distribution, and potential for marker development

    NASA Astrophysics Data System (ADS)

    Wang, Jiafeng; Qi, Haigang; Li, Li; Zhang, Guofan

    2014-01-01

    Microsatellites are a ubiquitous component of the eukaryote genome and constitute one of the most popular sources of molecular markers for genetic studies. However, no data are currently available regarding microsatellites across the entire genome in oysters, despite their importance to the aquaculture industry. We present the first genome-wide investigation of microsatellites in the Pacific oyster Crassostrea gigas by analysis of the complete genome, resequencing, and expression data. The Pacific oyster genome is rich in microsatellites. A total of 604 653 repeats were identified, in average of one locus per 815 base pairs (bp). A total of 12 836 genes had coding repeats, and 7 332 were expressed normally, including genes with a wide range of molecular functions. Compared with 20 different species of animals, microsatellites in the oyster genome typically exhibited 1) an intermediate overall frequency; 2) relatively uniform contents of (A)n and (C)n repeats and abundant long (C)n repeats (≥24 bp); 3) large average length of (AG)n repeats; and 4) scarcity of trinucleotide repeats. The microsatellite-flanking regions exhibited a high degree of polymorphism with a heterozygosity rate of around 2.0%, but there was no correlation between heterozygosity and microsatellite abundance. A total of 19 462 polymorphic microsatellites were discovered, and dinucleotide repeats were the most active, with over 26% of loci found to harbor allelic variations. In all, 7 451 loci with high potential for marker development were identified. Better knowledge of the microsatellites in the oyster genome will provide information for the future design of a wide range of molecular markers and contribute to further advancements in the field of oyster genetics, particularly for molecular-based selection and breeding.

  19. A Novel Approach for Characterizing Microsatellite Instability in Cancer Cells

    PubMed Central

    Lu, Yuheng; Soong, T. David; Elemento, Olivier

    2013-01-01

    Microsatellite instability (MSI) is characterized by the expansion or contraction of DNA repeat tracts as a consequence of DNA mismatch repair deficiency (MMRD). Accurate detection of MSI in cancer cells is important since MSI is associated with several cancer subtypes and can help inform therapeutic decisions. Although experimental assays have been developed to detect MSI, they typically depend on a small number of known microsatellite loci or mismatch repair genes and have limited reliability. Here, we report a novel genome-wide approach for MSI detection based on the global detection of insertions and deletions (indels) in microsatellites found in expressed genes. Our large-scale analyses of 20 cancer cell lines and 123 normal individuals revealed striking indel features associated with MSI: there is a significant increase of short microsatellite deletions in MSI samples compared to microsatellite stable (MSS) ones, suggesting a mechanistic bias of repair efficiency between insertions and deletions in normal human cells. By incorporating this observation into our MSI scoring metric, we show that our approach can correctly distinguish between MSI and MSS cancer cell lines. Moreover, when we applied this approach to primal tumor samples, our metric is also well consistent with diagnosed MSI status. Thus, our study offers new insight into DNA mismatch repair system, and also provides a novel MSI diagnosis method for clinical oncology with better reliability. PMID:23671654

  20. Genomic Microsatellites as Evolutionary Chronometers: A Test in Wild Cats

    PubMed Central

    Driscoll, Carlos A.; Menotti-Raymond, Marilyn; Nelson, George; Goldstein, David; O'Brien, Stephen J.

    2002-01-01

    Nuclear microsatellite loci (2- to 5-bp tandem repeats) would seem to be ideal markers for population genetic monitoring because of their abundant polymorphism, wide dispersal in vertebrate genomes, near selective neutrality, and ease of assessment; however, questions about their mode of generation, mutation rates and ascertainment bias have limited interpretation considerably. We have assessed the patterns of genomic diversity for ninety feline microsatellite loci among previously characterized populations of cheetahs, lions and pumas in recapitulating demographic history. The results imply that the microsatellite diversity measures (heterozygosity, allele reconstitution and microsatellite allele variance) offer proportionate indicators, albeit with large variance, of historic population bottlenecks and founder effects. The observed rate of reconstruction of new alleles plus the growth in the breadth of microsatellite allele size (variance) was used here to develop genomic estimates of time intervals following historic founder events in cheetahs (12,000 yr ago), in North American pumas (10,000–17,000 yr ago), and in Asiatic lions of the Gir Forest (1000–4000 yr ago). [Supplemental material available online at http://rex.nci.nih.gov/lgd/front_page.htm and at http://www.genome.org.] PMID:11875029

  1. Microsatellite and Chromosome Evolution of Parthenogenetic Sitobion Aphids in Australia

    PubMed Central

    Sunnucks, P.; England, P. R.; Taylor, A. C.; Hales, D. F.

    1996-01-01

    Single-locus microsatellite variation correlated perfectly with chromosome number in Sitobion miscanthi aphids. The microsatellites were highly heterozygous, with up to 10 alleles per locus in this species. Despite this considerable allelic variation, only seven different S. miscanthi genotypes were discovered in 555 individuals collected from a wide range of locations, hosts and sampling periods. Relatedness between genotypes suggests only two successful colonizations of Australia. There was no evidence for genetic recombination in 555 S. miscanthi so the occurrence of recent sexual reproduction must be near zero. Thus diversification is by mutation and chromosomal rearrangement alone. Since the aphids showed no sexual recombination, microsatellites can mutate without meiosis. Five of seven microsatellite differences were a single repeat unit, and one larger jump is likely. The minimum numbers of changes between karyotypes corresponded roughly one-to-one with microsatellite allele changes, which suggests very rapid chromosomal evolution. A chromosomal fission occurred in a cultured line, and a previously unknown chromosomal race was detected. All 121 diverse S. near fragariae were heterozygous but revealed only one genotype. This species too must have a low rate of sexual reproduction and few colonizations of Australia. PMID:8889535

  2. Genomic microsatellites as evolutionary chronometers: a test in wild cats.

    PubMed

    Driscoll, Carlos A; Menotti-Raymond, Marilyn; Nelson, George; Goldstein, David; O'Brien, Stephen J

    2002-03-01

    Nuclear microsatellite loci (2- to 5-bp tandem repeats) would seem to be ideal markers for population genetic monitoring because of their abundant polymorphism, wide dispersal in vertebrate genomes, near selective neutrality, and ease of assessment; however, questions about their mode of generation, mutation rates and ascertainment bias have limited interpretation considerably. We have assessed the patterns of genomic diversity for ninety feline microsatellite loci among previously characterized populations of cheetahs, lions and pumas in recapitulating demographic history. The results imply that the microsatellite diversity measures (heterozygosity, allele reconstitution and microsatellite allele variance) offer proportionate indicators, albeit with large variance, of historic population bottlenecks and founder effects. The observed rate of reconstruction of new alleles plus the growth in the breadth of microsatellite allele size (variance) was used here to develop genomic estimates of time intervals following historic founder events in cheetahs (12,000 yr ago), in North American pumas (10,000-17,000 yr ago), and in Asiatic lions of the Gir Forest (1000-4000 yr ago).

  3. Characterization of polymorphic microsatellite markers for Primula sikkimensis (Primulaceae) using a 454 sequencing approach1

    PubMed Central

    Li, Chang-Han; Liu, Yun-Jiao; Zhang, Cai-Yun; Yan, Hai-Fei; Ge, Xue-Jun; Hao, Gang

    2016-01-01

    Premise of the study: Microsatellite markers from Primula sikkimensis (Primulaceae) were developed for testing deep lineage divergence and speciation events. Methods and Results: A total of 3112 microsatellites were identified from 61,755 unique reads though 454 pyrosequencing technology. Twenty-nine microsatellite loci were selected for PCR amplification and polymorphic analyses. Among the 29 tested markers, 17 microsatellite loci were further used for genotyping in three wild P. sikkimensis populations. The number of alleles varied from one to eight, and the observed heterozygosity ranged from 0.111 to 1.000. Ten simple sequence repeat loci could be successfully cross-amplified in two Primula species. The transferability values were 76.5% in P. florindae and 58.8% in P. alpicola, respectively. Conclusions: These microsatellite markers will be valuable for testing the hypothesis of lineage divergence, genetic introgression, and cryptic speciation events between P. sikkimensis and its closely related taxa. PMID:27437171

  4. Multilocus microsatellite analysis of ‘Candidatus Liberibacter asiaticus’ associated with citrus Huanglongbing worldwide

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Huanglongbing (HLB) is one of the most destructive citrus diseases worldwide. In the United States (US), HLB is typically associated with the presence of a fastidious phloem-limited bacterium named ‘Candidatus Liberibacter asiaticus’, though other Liberibacter species also have been associated with ...

  5. Functional interaction between SNPs and microsatellite in the transcriptional regulation of insulin-like growth factor 1.

    PubMed

    Chen, Holly Y; Huang, Wei; Leung, Vincent H K; Fung, Simon L M; Ma, Suk Ling; Jiang, Hongling; Tang, Nelson L S

    2013-09-01

    A CA-repeat microsatellite in insulin-like growth factor 1 (IGF1) promoter was associated with interindividual variation of circulating IGF1 level. Previously, we reported that such association was due to variation of haplotype unit in a linkage disequilibrium block composed of microsatellite and single-nucleotide polymorphisms (SNPs), suggesting the presence of an interaction between them. In this study, reporter assays were performed to investigate the regulatory effect and interaction of genetic variants on gene expression. We used an in vitro system to compare the transcriptional activities of haplotypes (rs35767:T>C, the CA-repeat microsatellite, rs5742612:T>C, and rs2288377:T>A) in evolutionarily conserved region of IGF1 promoter. In haplotype C-T-T, a longer microsatellite had a lower transcriptional activity (17.6 ± 2.4-fold for 17 repeats and 8.3 ± 1.1-fold for 21 repeats), whereas in haplotype T-C-A, such trend could not be observed, as the microsatellite with 21 repeats had the highest transcriptional activity (17.5 ± 2.3-fold). Because the microsatellite and SNPs affected the transcriptional activity of each other, there may be an interaction between them in the regulation of IGF1 expression. For the first time, we demonstrated that a noncoding microsatellite polymorphism could act as a functional unit and interact with SNPs in the regulation of transcription in human genome.

  6. MICAS: a fully automated web server for microsatellite extraction and analysis from prokaryote and viral genomic sequences.

    PubMed

    Sreenu, Vattipally B; Ranjitkumar, Gundu; Swaminathan, Sugavanam; Priya, Sasidharan; Bose, Buddhaditta; Pavan, Mogili N; Thanu, Geeta; Nagaraju, Javaregowda; Nagarajaram, Hampapathalu A

    2003-01-01

    MICAS is a web server for extracting microsatellite information from completely sequenced prokaryote and viral genomes, or user-submitted sequences. This server provides an integrated platform for MICdb (database of prokaryote and viral microsatellites), W-SSRF (simple sequence repeat finding program) and Autoprimer (primer design software). MICAS, through dynamic HTML page generation, helps in the systematic extraction of microsatellite information from selected genomes hosted on MICdb or from user-submitted sequences. Further, it assists in the design of primers with the help of Autoprimer, for sequences containing selected microsatellite tracts.

  7. Effect of microsatellite selection on individual and population genetic inferences: an empirical study using cross-specific and species-specific amplifications.

    PubMed

    Queirós, J; Godinho, R; Lopes, S; Gortazar, C; de la Fuente, J; Alves, P C

    2015-07-01

    Although whole-genome sequencing is becoming more accessible and feasible for nonmodel organisms, microsatellites have remained the markers of choice for various population and conservation genetic studies. However, the criteria for choosing microsatellites are still controversial due to ascertainment bias that may be introduced into the genetic inference. An empirical study of red deer (Cervus elaphus) populations, in which cross-specific and species-specific microsatellites developed through pyrosequencing of enriched libraries, was performed for this study. Two different strategies were used to select the species-specific panels: randomly vs. highly polymorphic markers. The results suggest that reliable and accurate estimations of genetic diversity can be obtained using random microsatellites distributed throughout the genome. In addition, the results reinforce previous evidence that selecting the most polymorphic markers leads to an ascertainment bias in estimates of genetic diversity, when compared with randomly selected microsatellites. Analyses of population differentiation and clustering seem less influenced by the approach of microsatellite selection, whereas assigning individuals to populations might be affected by a random selection of a small number of microsatellites. Individual multilocus heterozygosity measures produced various discordant results, which in turn had impacts on the heterozygosity-fitness correlation test. Finally, we argue that picking the appropriate microsatellite set should primarily take into account the ecological and evolutionary questions studied. Selecting the most polymorphic markers will generally overestimate genetic diversity parameters, leading to misinterpretations of the real genetic diversity, which is particularly important in managed and threatened populations.

  8. Microsatellite instability is rare in sporadic ovarian cancer

    SciTech Connect

    Chen, S.S.; Han, H.; Schwartz, P.E.

    1994-09-01

    Microsatellite instability was first demonstrated to be a common underlying mechanism in hereditary nonpolyposis colorectal cancer (HNPCC) and has recently been implicated in the development of several other human cancers. Although numerous genetic changes have been documented in ovarian cancer, their molecular bases are poorly understood. In investigating the molecular genetics of ovarian cancer, we analyzed twelve short tandem repeats that were amplified by PCR from DNA of 48 tumors and their corresponding lymphocyte samples. All of the 48 cases studied have no noticeable family history and, of them, 42 are epithelial (benign/borderline, 5; grade I, 4; GII, 4; GIII, 29) and 6 are nonepithelial. A microsatellite instability has been shown to be inversely correlated with the occurrence of allelic losses, half of those cases chosen have a fractional allele loss of {le}15 (median = .18 of 50 tumors tested for 86 loci from every chromosomal arm). The loci examined included eight dinucleotide repeats (D2S123, D9S104, D10S197, D11S904, D16S408, D16S421, D17S250, and D17S579), two trinucleotide repeats (DM and AR) and two tetranucleotide repeats (DXS981 and VWF). Despite the fact that HNPCC phenotype includes ovarian cancer and that microsatellite instability has been shown in one ovarian cancer from an HNPCC family, the allele sizes of 12 loci were found to be identical in all paired tumor and normal samples we studied except for one tumor at a single locus. The band shift displayed on polyacrylamide gel representing an additional allele of VWF was only observed in one grade III tumor. Our results are thus a strong indication that the alteration of microsatellite repeats may not play a major role in the development of sporadic ovarian cancer.

  9. Microsatellite markers characterized in the mosquito Aedes taeniorhynchus (Diptera, Culicidae), a disease vector and major pest on the American coast and the Galápagos Islands.

    PubMed

    Bataille, Arnaud; Horsburgh, Gavin J; Dawson, Deborah A; Cunningham, Andrew A; Goodman, Simon J

    2009-09-01

    The black salt-marsh mosquito, Aedes taeniorhynchus, plays an important role in the transmission of arboviruses such as West Nile virus and other pathogens of concern for human and animal health in North and Latin America. This mosquito is notably the only widely distributed mosquito species found in the Galápagos Islands, where its impact as disease vector has not yet been studied. The use of microsatellite markers can significantly improve our understanding of the population structure and dynamics of A. taeniorhynchus and its role in the transmission of diseases. Here we report the isolation of 12 unique microsatellite loci using an enrichment protocol. We also identified other multi-locus microsatellites linked to transposable elements. The presence of such elements may explain why the isolation of useful scorable microsatellite markers in the Aedes genus is often difficult. Four of the markers isolated amplified polymorphic products in Aedes aegypti, Aedes albopictus and/or Aedes japonicus.

  10. Microsatellite spreading in the human genome: evolutionary mechanisms and structural implications.

    PubMed

    Nadir, E; Margalit, H; Gallily, T; Ben-Sasson, S A

    1996-06-25

    Microsatellites are tandem repeat sequences abundant in the genomes of higher eukaryotes and hitherto considered as "junk DNA." Analysis of a human genome representative data base (2.84 Mb) reveals a distinct juxtaposition of A-rich microsatellites and retroposons and suggests their coevolution. The analysis implies that most microsatellites were generated by a 3'-extension of retrotranscripts, similar to mRNA polyadenylylation, and that they serve in turn as "retroposition navigators," directing the retroposons via homology-driven integration into defined sites. Thus, they became instrumental in the preservation and extension of primordial genomic patterns. A role is assigned to these reiterating A-rich loci in the higher-order organization of the chromatin. The disease-associated triplet repeats are mostly found in coding regions and do not show an association with retroposons, constituting a unique set within the family of microsatellite sequences.

  11. Positive fragile X microsatellite associations point to a common mechanism of dynamic mutation evolution

    SciTech Connect

    Brown, W.T.; Zhong, N.; Dobkin, C.

    1996-03-01

    We recently reported that the size of fragile X gene (FMR1) triplet repeats and two nearby microsatellites show positive allele-size associations. The larger alleles of microsatellite DXS548, located {approximately}150 kb proximal to the FMR1 CGG repeat, and of FRAXAC1 (AC1), located 7 kb proximal to the FMR1 CGG repeat, tend to occur together, and smaller alleles also tend to occur together. Also, fragile X chromosomes are more commonly found on the larger combined microsatellite-allele haplotypes. We now have extended these observations to include two other nearby repeats, FRAXAC2 (AC2), a complex three-part polymorphism located 12 kb distal, and the FRAXE triplet repeat, located 600 kb distal. We divided the chromosomes into controls with FMR1 repeats of <60 and fragile X chromosomes with repeats {>=}60, since FMR1 alleles with repeats {>=}60 show high intergenerational instability. In the 133 controls, previously analyzed for AGG interspersions, and in 119 fragile X chromosomes, we found that these repeats show nonrandom size associations. To describe this numerically, we calculated correlation coefficients for the repeat lengths. These repeats showed significantly positive correlations with each other. Although FRAXE alleles showed no correlation with the control repeats, they did have positive correlations with fragile X chromosome microsatellites (AC1 and AC2 but not DXS548), which may reflect the larger recombinational distances involved and the possibly more recent origin of the fragile X mutations. The correlations tended to be higher for the number of 3{prime} pure CGGs than for total FMR1 repeats in controls. These findings strengthen our hypothesis that there may be a common underlying mutational mechanism that simultaneously affects these repeat loci. 13 refs., 1 tab.

  12. Microsatellite primers in the lichen symbiotic alga Trebouxia decolorans (Trebouxiophyceae)1

    PubMed Central

    Dal Grande, Francesco; Beck, Andreas; Singh, Garima; Schmitt, Imke

    2013-01-01

    • Premise of the study: Polymorphic microsatellite markers were developed for the symbiotic green alga Trebouxia decolorans to study fine-scale population structure and clonal diversity. • Methods and Results: Using Illumina pyrosequencing, 20 microsatellite primer sets were developed for T. decolorans. The primer sets were tested on 43 individuals sampled from four subpopulations in Germany. The primers amplified di-, tri-, and tetranucleotide repeats with three to 15 alleles per locus, and the unbiased haploid diversity per locus ranged from 0.636 to 0.821. • Conclusions: The identified microsatellite markers will be useful to study the genetic diversity, dispersal, and reproductive mode of this common lichen photobiont. PMID:25202529

  13. Microsatellites in the Eukaryotic DNA Mismatch Repair Genes as Modulators of Evolutionary Mutation Rate

    NASA Technical Reports Server (NTRS)

    Chang, Dong Kyung; Metzgar, David; Wills, Christopher; Boland, C. Richard

    2003-01-01

    All "minor" components of the human DNA mismatch repair (MMR) system-MSH3, MSH6, PMS2, and the recently discovered MLH3-contain mononucleotide microsatellites in their coding sequences. This intriguing finding contrasts with the situation found in the major components of the DNA MMR system-MSH2 and MLH1-and, in fact, most human genes. Although eukaryotic genomes are rich in microsatellites, non-triplet microsatellites are rare in coding regions. The recurring presence of exonal mononucleotide repeat sequences within a single family of human genes would therefore be considered exceptional.

  14. Comparative analysis of microsatellites in chloroplast genomes of lower and higher plants.

    PubMed

    George, Biju; Bhatt, Bhavin S; Awasthi, Mayur; George, Binu; Singh, Achuit K

    2015-11-01

    Microsatellites, or simple sequence repeats (SSRs), contain repetitive DNA sequence where tandem repeats of one to six base pairs are present number of times. Chloroplast genome sequences have been  shown to possess extensive variations in the length, number and distribution of SSRs. However, a comparative analysis of chloroplast microsatellites is not available. Considering their potential importance in generating genomic diversity, we have systematically analysed the abundance and distribution of simple and compound microsatellites in 164 sequenced chloroplast genomes from wide range of plants. The key findings of these studies are (1) a large number of mononucleotide repeats as compared to SSR(2-6)(di-, tri-, tetra-, penta-, hexanucleotide repeats) are present in all chloroplast genomes investigated, (2) lower plants such as algae show wide variation in relative abundance, density and distribution of microsatellite repeats as compared to flowering plants, (3) longer SSRs are excluded from coding regions of most chloroplast genomes, (4) GC content has a weak influence on number, relative abundance and relative density of mononucleotide as well as SSR(2-6). However, GC content strongly showed negative correlation with relative density (R (2) = 0.5, P < 0.05) and relative abundance (R (2) = 0.6, P < 0.05) of cSSRs. In summary, our comparative studies of chloroplast genomes illustrate the variable distribution of microsatellites and revealed that chloroplast genome of smaller plants possesses relatively more genomic diversity compared to higher plants.

  15. Joint inference of microsatellite mutation models, population history and genealogies using transdimensional Markov Chain Monte Carlo.

    PubMed

    Wu, Chieh-Hsi; Drummond, Alexei J

    2011-05-01

    We provide a framework for Bayesian coalescent inference from microsatellite data that enables inference of population history parameters averaged over microsatellite mutation models. To achieve this we first implemented a rich family of microsatellite mutation models and related components in the software package BEAST. BEAST is a powerful tool that performs Bayesian MCMC analysis on molecular data to make coalescent and evolutionary inferences. Our implementation permits the application of existing nonparametric methods to microsatellite data. The implemented microsatellite models are based on the replication slippage mechanism and focus on three properties of microsatellite mutation: length dependency of mutation rate, mutational bias toward expansion or contraction, and number of repeat units changed in a single mutation event. We develop a new model that facilitates microsatellite model averaging and Bayesian model selection by transdimensional MCMC. With Bayesian model averaging, the posterior distributions of population history parameters are integrated across a set of microsatellite models and thus account for model uncertainty. Simulated data are used to evaluate our method in terms of accuracy and precision of estimation and also identification of the true mutation model. Finally we apply our method to a red colobus monkey data set as an example.

  16. Multilocus homozygosity mapping and autozygosity patterns

    SciTech Connect

    Thompson, E.A.; Wijsman, E.M.

    1994-09-01

    Exact computation of likelihoods is often not possible for multipoint linkage analyses on complex pedigrees with missing data, such as arise in homozygosity mapping of rare recessive diseases. Markov chain Monte Carlo (MCMC) can provide an estimate of the likelihood function and lod score. Usually, the genotypes of individuals are taken as the latent variables to be sampled in MCMC analyses of computationally intractable genetic problems, but in the case of a very few sampled individuals in an extended pedigree the space of latent variables can be reduced. Our latent variables are the indicators of grandparental origins of genes for each locus and each segregation. This greatly improves the efficiency of MCMC estimates of multilocus lod scores. For homozygosity mapping, in addition to providing estimates of lod scores, MCMC provides posterior probabilities of multilocus patterns of gene identity by descent. Although in the region of the disease locus, the posterior probability of autozygosity may be high for each pedigree, the probability is low that all of a large set of inbred affected individuals exhibit autozygosity. For each segregation, in the absence of interference, the process of grandparental origin is Markov along the chromosome, but the autozygosity of a descendant individual results from grouping segregation patterns. Hence, the multilocus autozygosity of inbred individuals exhibits interesting patterns, with a clumping of autozygous loci, interspersed by small regions of non-autozygosity, these clumps being more widely separated than predicted by a simple Markov process along the chromosome. These patterns also can be analyzed by Monte Carlo, and also have practical implications in homozygosity mapping, where sometimes a heretozygous marker intervenes in a region of homozygosity. These ideas are illustrated by application to some Werner`s Syndrome pedigrees, a very rare recessive disease of premature aging.

  17. Microsatellite analysis of malaria parasites.

    PubMed

    Orjuela-Sánchez, Pamela; Brandi, Michelle C; Ferreira, Marcelo U

    2013-01-01

    Microsatellites have been increasingly used to investigate the population structure of malaria parasites, to map genetic loci contributing to phenotypes such as drug resistance and virulence in laboratory crosses and genome-wide association studies and to distinguish between treatment failures and new infections in clinical trials. Here, we provide optimized protocols for genotyping highly polymorphic microsatellites sampled from across the genomes of the human malaria parasites Plasmodium falciparum and P. vivax that have been extensively used in research laboratories worldwide.

  18. Microsatellite Length Scoring by Single Molecule Real Time Sequencing – Effects of Sequence Structure and PCR Regime

    PubMed Central

    Liljegren, Mikkel Meyn; de Muinck, Eric Jacques; Trosvik, Pål

    2016-01-01

    Microsatellites are DNA sequences consisting of repeated, short (1–6 bp) sequence motifs that are highly mutable by enzymatic slippage during replication. Due to their high intrinsic variability, microsatellites have important applications in population genetics, forensics, genome mapping, as well as cancer diagnostics and prognosis. The current analytical standard for microsatellites is based on length scoring by high precision electrophoresis, but due to increasing efficiency next-generation sequencing techniques may provide a viable alternative. Here, we evaluated single molecule real time (SMRT) sequencing, implemented in the PacBio series of sequencing apparatuses, as a means of microsatellite length scoring. To this end we carried out multiplexed SMRT sequencing of plasmid-carried artificial microsatellites of varying structure under different pre-sequencing PCR regimes. For each repeat structure, reads corresponding to the target length dominated. We found that pre-sequencing amplification had large effects on scoring accuracy and error distribution relative to controls, but that the effects of the number of amplification cycles were generally weak. In line with expectations enzymatic slippage decreased proportionally with microsatellite repeat unit length and increased with repetition number. Finally, we determined directional mutation trends, showing that PCR and SMRT sequencing introduced consistent but opposing error patterns in contraction and expansion of the microsatellites on the repeat motif and single nucleotide level. PMID:27414800

  19. Multilocus Detection of Wolf x Dog Hybridization in Italy, and Guidelines for Marker Selection

    PubMed Central

    Randi, Ettore; Hulva, Pavel; Fabbri, Elena; Galaverni, Marco; Galov, Ana; Kusak, Josip; Bigi, Daniele; Bolfíková, Barbora Černá; Smetanová, Milena; Caniglia, Romolo

    2014-01-01

    Hybridization and introgression can impact the evolution of natural populations. Several wild canid species hybridize in nature, sometimes originating new taxa. However, hybridization with free-ranging dogs is threatening the genetic integrity of grey wolf populations (Canis lupus), or even the survival of endangered species (e.g., the Ethiopian wolf C. simensis). Efficient molecular tools to assess hybridization rates are essential in wolf conservation strategies. We evaluated the power of biparental and uniparental markers (39 autosomal and 4 Y-linked microsatellites, a melanistic deletion at the β-defensin CBD103 gene, the hypervariable domain of the mtDNA control-region) to identify the multilocus admixture patterns in wolf x dog hybrids. We used empirical data from 2 hybrid groups with different histories: 30 presumptive natural hybrids from Italy and 73 Czechoslovakian wolfdogs of known hybrid origin, as well as simulated data. We assessed the efficiency of various marker combinations and reference samples in admixture analyses using 69 dogs of different breeds and 99 wolves from Italy, Balkans and Carpathian Mountains. Results confirmed the occurrence of hybrids in Italy, some of them showing anomalous phenotypic traits and exogenous mtDNA or Y-chromosome introgression. Hybridization was mostly attributable to village dogs and not strictly patrilineal. The melanistic β-defensin deletion was found only in Italian dogs and in putative hybrids. The 24 most divergent microsatellites (largest wolf-dog FST values) were equally or more informative than the entire panel of 39 loci. A smaller panel of 12 microsatellites increased risks to identify false admixed individuals. The frequency of F1 and F2 was lower than backcrosses or introgressed individuals, suggesting hybridization already occurred some generations in the past, during early phases of wolf expansion from their historical core areas. Empirical and simulated data indicated the identification of the past

  20. Multilocus detection of wolf x dog hybridization in italy, and guidelines for marker selection.

    PubMed

    Randi, Ettore; Hulva, Pavel; Fabbri, Elena; Galaverni, Marco; Galov, Ana; Kusak, Josip; Bigi, Daniele; Bolfíková, Barbora Černá; Smetanová, Milena; Caniglia, Romolo

    2014-01-01

    Hybridization and introgression can impact the evolution of natural populations. Several wild canid species hybridize in nature, sometimes originating new taxa. However, hybridization with free-ranging dogs is threatening the genetic integrity of grey wolf populations (Canis lupus), or even the survival of endangered species (e.g., the Ethiopian wolf C. simensis). Efficient molecular tools to assess hybridization rates are essential in wolf conservation strategies. We evaluated the power of biparental and uniparental markers (39 autosomal and 4 Y-linked microsatellites, a melanistic deletion at the β-defensin CBD103 gene, the hypervariable domain of the mtDNA control-region) to identify the multilocus admixture patterns in wolf x dog hybrids. We used empirical data from 2 hybrid groups with different histories: 30 presumptive natural hybrids from Italy and 73 Czechoslovakian wolfdogs of known hybrid origin, as well as simulated data. We assessed the efficiency of various marker combinations and reference samples in admixture analyses using 69 dogs of different breeds and 99 wolves from Italy, Balkans and Carpathian Mountains. Results confirmed the occurrence of hybrids in Italy, some of them showing anomalous phenotypic traits and exogenous mtDNA or Y-chromosome introgression. Hybridization was mostly attributable to village dogs and not strictly patrilineal. The melanistic β-defensin deletion was found only in Italian dogs and in putative hybrids. The 24 most divergent microsatellites (largest wolf-dog FST values) were equally or more informative than the entire panel of 39 loci. A smaller panel of 12 microsatellites increased risks to identify false admixed individuals. The frequency of F1 and F2 was lower than backcrosses or introgressed individuals, suggesting hybridization already occurred some generations in the past, during early phases of wolf expansion from their historical core areas. Empirical and simulated data indicated the identification of the past

  1. Comparative Genomics and Molecular Dynamics of DNA Repeats in Eukaryotes

    PubMed Central

    Richard, Guy-Franck; Kerrest, Alix; Dujon, Bernard

    2008-01-01

    Summary: Repeated elements can be widely abundant in eukaryotic genomes, composing more than 50% of the human genome, for example. It is possible to classify repeated sequences into two large families, “tandem repeats” and “dispersed repeats.” Each of these two families can be itself divided into subfamilies. Dispersed repeats contain transposons, tRNA genes, and gene paralogues, whereas tandem repeats contain gene tandems, ribosomal DNA repeat arrays, and satellite DNA, itself subdivided into satellites, minisatellites, and microsatellites. Remarkably, the molecular mechanisms that create and propagate dispersed and tandem repeats are specific to each class and usually do not overlap. In the present review, we have chosen in the first section to describe the nature and distribution of dispersed and tandem repeats in eukaryotic genomes in the light of complete (or nearly complete) available genome sequences. In the second part, we focus on the molecular mechanisms responsible for the fast evolution of two specific classes of tandem repeats: minisatellites and microsatellites. Given that a growing number of human neurological disorders involve the expansion of a particular class of microsatellites, called trinucleotide repeats, a large part of the recent experimental work on microsatellites has focused on these particular repeats, and thus we also review the current knowledge in this area. Finally, we propose a unified definition for mini- and microsatellites that takes into account their biological properties and try to point out new directions that should be explored in a near future on our road to understanding the genetics of repeated sequences. PMID:19052325

  2. Microsatellite variation in the Australian dingo.

    PubMed

    Wilton, A N; Steward, D J; Zafiris, K

    1999-01-01

    The dingo is thought to have arrived in Australia from Asia about 5,000 years ago. It is currently in danger because of interbreeding with domestic dogs. Several morphological, behavioral, and reproductive characteristics distinguish dingoes from domestic dog. Skull morphometrics are currently used to try to classify wild canids as pure dingo, dog, or hybrid. Molecular techniques based on diagnostic DNA differences between dogs and dingoes would make a much more reliable and practical test. A small number of markers (about 10) would allow detection of animals with domestic dog in their ancestry several generations back. We have typed 16 dingoes and 16 dogs of mixed breed for 14 microsatellites. The amount of variation in the Australian dingo is much less than in domestic dogs. The size distributions of microsatellites in the two groups usually overlap. The number of alleles in the dingo is much smaller in all cases. One dinucleotide repeat locus shows a size difference of 1 bp in allele classes between dog and dingo. This locus may be diagnostic for dog or dingo ancestry. The differences in distributions of alleles at other loci can also be used to classify animals using a likelihood method.

  3. Profile of candidate microsatellite markers in Sebastiscus marmoratus using 454 pyrosequencing

    NASA Astrophysics Data System (ADS)

    Song, Na; Chen, Muyan; Gao, Tianxiang; Yanagimoto, Takashi

    2017-01-01

    Sebastiscus marmoratus is an important sedentary ovoviparous fish distributed in near-shore coastal waters from the coast of China to Japan. Candidate S. marmoratus microsatellite markers were developed in the present study using 454 pyrosequencing, and the marker profile was analyzed. A total of 2 000 000 raw sequence reads were assembled to reduce redundancy. Among them, 1 043 dinucleotide, 925 trinucleotide, 692 tetranucleotide, and 315 pentanucleotide repeats were detected. AC repeats were the most frequent motifs among the dinucleotide repeats, and AAT was the most abundant among the trinucleotide repeats. AAAT, ATAG, and ATCC were the three most common tetranucleotide motifs, and AAGAT and AATAT were the most dominant pentanucleotide motifs. The greatest numbers of loci and potentially amplifiable loci were found in dinucleotide repeats, whereas trinucleotide repeats had the fewest. In summary, a wide range of candidate microsatellite markers were identified in the present study using a rapid and efficient 454 pyrosequencing approach.

  4. Polymorphism in Multilocus Host–Parasite Coevolutionary Interactions

    PubMed Central

    Tellier, Aurélien; Brown, James K. M.

    2007-01-01

    Numerous loci in host organisms are involved in parasite recognition, such as major histocompatibility complex (MHC) genes in vertebrates or genes involved in gene-for-gene (GFG) relationships in plants. Diversity is commonly observed at such loci and at corresponding loci encoding antigenic molecules in parasites. Multilocus theoretical models of host–parasite coevolution predict that polymorphism is more likely than in single-locus interactions because recurrent coevolutionary cycles are sustained by indirect frequency-dependent selection as rare genotypes have a selective advantage. These cycles are stabilized by direct frequency-dependent selection, resulting from repeated reinfection of the same host by a parasite, a feature of most diseases. Here, it is shown that for realistically small costs of resistance and virulence, polycyclic disease and high autoinfection rates, stable polymorphism of all possible genotypes is obtained in parasite populations. Two types of epistatic interactions between loci tend to increase the parameter space in which stable polymorphism can occur with all possible host and parasite genotypes. In the parasite, the marginal cost of each additional virulence allele should increase, while in the host, the marginal cost of each additional resistance allele should decrease. It is therefore predicted that GFG polymorphism will be stable (and hence detectable) when there is partial complementation of avirulence genes in the parasite and of resistance genes in the host. PMID:17947440

  5. Distribution and localization of microsatellites in the Perigord black truffle genome and identification of new molecular markers.

    PubMed

    Murat, C; Riccioni, C; Belfiori, B; Cichocki, N; Labbé, J; Morin, E; Tisserant, E; Paolocci, F; Rubini, A; Martin, F

    2011-06-01

    The level of genetic diversity and genetic structure in the Perigord black truffle (Tuber melanosporum Vittad.) has been debated for several years, mainly due to the lack of appropriate genetic markers. Microsatellites or simple sequence repeats (SSRs) are important for the genome organisation, phenotypic diversity and are one of the most popular molecular markers. In this study, we surveyed the T. melanosporum genome (1) to characterise its SSR pattern; (2) to compare it with SSR patterns found in 48 other fungal and three oomycetes genomes and (3) to identify new polymorphic SSR markers for population genetics. The T. melanosporum genome is rich in SSRs with 22,425 SSRs with mono-nucleotides being the most frequent motifs. SSRs were found in all genomic regions although they are more frequent in non-coding regions (introns and intergenic regions). Sixty out of 135 PCR-amplified mono-, di-, tri-, tetra, penta, and hexa-nucleotides were polymorphic (44%) within black truffle populations and 27 were randomly selected and analysed on 139 T. melanosporum isolates from France, Italy and Spain. The number of alleles varied from 2 to 18 and the expected heterozygosity from 0.124 to 0.815. One hundred and thirty-two different multilocus genotypes out of the 139 T. melanosporum isolates were identified and the genotypic diversity was high (0.999). Polymorphic SSRs were found in UTR regulatory regions of fruiting bodies and ectomycorrhiza regulated genes, suggesting that they may play a role in phenotypic variation. In conclusion, SSRs developed in this study were highly polymorphic and our results showed that T. melanosporum is a species with an important genetic diversity, which is in agreement with its recently uncovered heterothallic mating system.

  6. High frequency of RPL22 mutations in microsatellite-unstable colorectal and endometrial tumors.

    PubMed

    Ferreira, Ana M; Tuominen, Iina; van Dijk-Bos, Krista; Sanjabi, Bahram; van der Sluis, Tineke; van der Zee, Ate G; Hollema, Harry; Zazula, Monika; Sijmons, Rolf H; Aaltonen, Lauri A; Westers, Helga; Hofstra, Robert M W

    2014-12-01

    Ribosomal Protein L22 (RPL22) encodes a protein that is a component of the 60S subunit of the ribosome. Variants in this gene have recently been linked to cancer development. Mutations in an A8 repeat in exon 2 were found in a recent study in 52% of microsatellite-unstable endometrial tumors. These tumors are particularly prone to mutations in repeats due to mismatch repair deficiency. We screened this coding repeat in our collection of microsatellite-unstable endometrial tumors (EC) and colorectal tumors (CRC). We found 50% mutation frequency for EC and 77% mutation frequency for CRC. These results confirm the previous study on the involvement of RPL22 in EC and, more importantly, reports for the first time such high mutation frequency in this gene in colorectal cancer. Furthermore, considering the high mutation frequency found, our data point toward an important role for RPL22 in microsatellite instability carcinogenesis.

  7. Microsatellite analysis in the genome of Acanthaceae: An in silico approach

    PubMed Central

    Kaliswamy, Priyadharsini; Vellingiri, Srividhya; Nathan, Bharathi; Selvaraj, Saravanakumar

    2015-01-01

    Background: Acanthaceae is one of the advanced and specialized families with conventionally used medicinal plants. Simple sequence repeats (SSRs) play a major role as molecular markers for genome analysis and plant breeding. The microsatellites existing in the complete genome sequences would help to attain a direct role in the genome organization, recombination, gene regulation, quantitative genetic variation, and evolution of genes. Objective: The current study reports the frequency of microsatellites and appropriate markers for the Acanthaceae family genome sequences. Materials and Methods: The whole nucleotide sequences of Acanthaceae species were obtained from National Center for Biotechnology Information database and screened for the presence of SSRs. SSR Locator tool was used to predict the microsatellites and inbuilt Primer3 module was used for primer designing. Results: Totally 110 repeats from 108 sequences of Acanthaceae family plant genomes were identified, and the occurrence of dinucleotide repeats was found to be abundant in the genome sequences. The essential amino acid isoleucine was found rich in all the sequences. We also designed the SSR-based primers/markers for 59 sequences of this family that contains microsatellite repeats in their genome. Conclusion: The identified microsatellites and primers might be useful for breeding and genetic studies of plants that belong to Acanthaceae family in the future. PMID:25709226

  8. Genotyping the clonal structure of a gorgonian coral, Junceella juncea (Anthozoa: Octocorallia), using microsatellite loci

    NASA Astrophysics Data System (ADS)

    Liu, Shang-Yin Vanson; Yu, Hon-Tsen; Fan, Tung-Yung; Dai, Chang-Feng

    2005-11-01

    The identification of different clones is fundamental to the study of population structure among organisms with mixed reproductive modes such as cnidarians. However, due to the low genetic variation of coral mtDNA and contamination by zooxanthellate DNA, very few molecular markers are available for studying the clonal structure of cnidarians. Herein we used four polymorphic loci of microsatellite DNA isolated from a zooxanthellae-free octocoral, Junceella juncea, to study its clonal structure in seven populations collected from three localities in Taiwan. In total, 40 multilocus genotypes were found among 152 colonies, and the number of genotypes (clones) identified in the seven populations ranged from 2 to 16. Each of the 40 multilocus genotypes was restricted to a single population, even where adjacent populations were only 100 m distant. The ratio of observed to expected genotypic diversity (Go:Ge) ranged from 0.217 to 0.650, and Go showed a significant departure from Ge ( p<0.05) at each site indicating that asexual fragmentation may play a major role in the maintenance of established populations. Mean relatedness ( R) values showed that genotypes within reefs were more closely related than those between regions. The results indicate that microsatellites are useful for discerning the clonal structures among and within populations at different spatial scales.

  9. Analysis of clinical and environmental Candida parapsilosis isolates by microsatellite genotyping--a tool for hospital infection surveillance.

    PubMed

    Sabino, R; Sampaio, P; Rosado, L; Videira, Z; Grenouillet, F; Pais, C

    2015-10-01

    Candida parapsilosis emerged as an important opportunistic pathogen, causing candidaemia worldwide. Nosocomial outbreaks triggered by this species have been frequently described, particularly in cancer patients. For a better understanding of its epidemiology, several typing methods are used and microsatellite analysis has been reported as highly discriminant. The main objective of this work was to study C. parapsilosis isolates by application of microsatellite genotyping to distinguish epidemiologically related strains, compare clinical and environmental isolates and determine possible routes of dispersion of the isolates in the hospital setting. A total of 129 C. parapsilosis isolates from different origins, including hospital environment and hands of healthcare workers, were genotyped using four microsatellite markers. The isolates were recovered from different health institutions. Analysis of C. parapsilosis isolates from hospital environment showed great genotypic diversity; however, the same or very similar genotypes were also found. The same multilocus genotype was shared by isolates recovered from the hand of a healthcare worker, from the hospital environment and from patients of the same healthcare institution, suggesting that these could be possible routes of transmission and that infections due to C. parapsilosis may be mainly related with exogenous transmission to the patient. Examination of sequential isolates from the same patients showed that colonizing and bloodstream isolates had the same multilocus genotype in the majority of cases. We demonstrate that this typing method is able to distinguish clonal clusters from genetically unrelated genotypes and can be a valuable tool to support epidemiologic investigations in the hospital setting.

  10. Repeated nightmares

    MedlinePlus

    ... different from night terrors . Alternative Names Nightmares - repeated; Dream anxiety disorder References American Academy of Family Physicians. Information from your family doctor. Nightmares and night terrors in children. ...

  11. Development of novel microsatellite markers for strain-specific identification of Chlorella vulgaris.

    PubMed

    Jo, Beom-Ho; Lee, Chang Soo; Song, Hae-Ryong; Lee, Hyung-Gwan; Oh, Hee-Mock

    2014-09-01

    A strain-specific identification method is required to secure Chlorella strains with useful genetic traits, such as a fast growth rate or high lipid productivity, for application in biofuels, functional foods, and pharmaceuticals. Microsatellite markers based on simple sequence repeats can be a useful tool for this purpose. Therefore, this study developed five novel microsatellite markers (mChl-001, mChl-002, mChl-005, mChl-011, and mChl-012) using specific loci along the chloroplast genome of Chlorella vulgaris. The microsatellite markers were characterized based on their allelic diversities among nine strains of C. vulgaris with the same 18S rRNA sequence similarity. Each microsatellite marker exhibited 2~5 polymorphic allele types, and their combinations allowed discrimination between seven of the C. vulgaris strains. The two remaining strains were distinguished using one specific interspace region between the mChl-001 and mChl-005 loci, which was composed of about 27 single nucleotide polymorphisms, 13~15 specific sequence sites, and (T)n repeat sites. Thus, the polymorphic combination of the five microsatellite markers and one specific locus facilitated a clear distinction of C. vulgaris at the strain level, suggesting that the proposed microsatellite marker system can be useful for the accurate identification and classification of C. vulgaris.

  12. Development of highly transferable microsatellites for Panax ginseng (Araliaceae) using whole-genome data1

    PubMed Central

    Jiang, Peng; Shi, Feng-Xue; Li, Ya-Ling; Liu, Bao; Li, Lin-Feng

    2016-01-01

    Premise of the study: Highly transferable expressed sequence tag (EST) microsatellites were developed for Panax ginseng (Araliaceae), one of the most celebrated traditional Chinese medicines and an endangered species in East Asia, using whole-genome data. Methods and Results: Twenty-one EST microsatellites were characterized from next-generation sequencing and were composed of di- and trinucleotide repeats. Polymorphisms and genetic diversity were evaluated for 45 accessions of three ginseng landraces. The number of alleles for each locus ranged from one to five among the landraces, and the polymorphism information content varied from 0.0000 to 0.6450. These microsatellites were also tested for congeneric amplification with P. notoginseng, P. stipuleanatus, P. quinquefolius, P. bipinnatifidus, and the closely related species Aralia elata. Conclusions: These novel EST-derived microsatellite markers will facilitate further population genetic studies of the genera Panax and Aralia. PMID:27843725

  13. Development of highly variable microsatellite markers for the tetraploid Silene stellata (Caryophyllaceae)1

    PubMed Central

    Zhou, Juannan; Dudash, Michele R.; Fenster, Charles B.; Zimmer, Elizabeth A.

    2016-01-01

    Premise of the study: We designed and tested microsatellite markers for the North American native species Silene stellata (Caryophyllaceae) to investigate its population genetic structure and identify selection on floral design through male reproductive success. Methods and Results: A total of 153 candidate microsatellite loci were isolated based on next-generation sequencing. We identified 18 polymorphic microsatellite loci in three populations of S. stellata, with di- or trinucleotide repeats. Genotyping results showed the number of alleles per locus ranged from six to 45 and expected heterozygosity ranged from 0.511 to 0.951. Five of these loci were successfully amplified in S. virginica and S. caroliniana and were also polymorphic. Conclusions: The microsatellite markers reported here provide a valuable tool for paternity analysis in S. stellata. They will also be useful for investigating the population genetic structures of S. stellata and related species. PMID:28101439

  14. WebSat ‐ A web software for microsatellite marker development

    PubMed Central

    Martins, Wellington Santos; Soares Lucas, Divino César; de Souza Neves, Kelligton Fabricio; Bertioli, David John

    2009-01-01

    Simple sequence repeats (SSR), also known as microsatellites, have been extensively used as molecular markers due to their abundance and high degree of polymorphism. We have developed a simple to use web software, called WebSat, for microsatellite molecular marker prediction and development. WebSat is accessible through the Internet, requiring no program installation. Although a web solution, it makes use of Ajax techniques, providing a rich, responsive user interface. WebSat allows the submission of sequences, visualization of microsatellites and the design of primers suitable for their amplification. The program allows full control of parameters and the easy export of the resulting data, thus facilitating the development of microsatellite markers. Availability The web tool may be accessed at http://purl.oclc.org/NET/websat/ PMID:19255650

  15. An empirical review: Characteristics of plant microsatellite markers that confer higher levels of genetic variation1

    PubMed Central

    Merritt, Benjamin J.; Culley, Theresa M.; Avanesyan, Alina; Stokes, Richard; Brzyski, Jessica

    2015-01-01

    During microsatellite marker development, researchers must choose from a pool of possible primer pairs to further test in their species of interest. In many cases, the goal is maximizing detectable levels of genetic variation. To guide researchers and determine which markers are associated with higher levels of genetic variation, we conducted a literature review based on 6782 genomic microsatellite markers published from 1997–2012. We examined relationships between heterozygosity (He or Ho) or allele number (A) with the following marker characteristics: repeat type, motif length, motif region, repeat frequency, and microsatellite size. Variation across taxonomic groups was also analyzed. There were significant differences between imperfect and perfect repeat types in A and He. Dinucleotide motifs exhibited significantly higher A, He, and Ho than most other motifs. Repeat frequency and motif region were positively correlated with A, He, and Ho, but correlations with microsatellite size were minimal. Higher taxonomic groups were disproportionately represented in the literature and showed little consistency. In conclusion, researchers should carefully consider marker characteristics so they can be tailored to the desired application. If researchers aim to target high genetic variation, dinucleotide motif lengths with large repeat frequencies may be best. PMID:26312192

  16. Characterization and transferability of microsatellite markers of the cultivated peanut (Arachis hypogaea)

    PubMed Central

    Gimenes, Marcos A; Hoshino, Andrea A; Barbosa, Andrea VG; Palmieri, Dario A; Lopes, Catalina R

    2007-01-01

    Background The genus Arachis includes Arachis hypogaea (cultivated peanut) and wild species that are used in peanut breeding or as forage. Molecular markers have been employed in several studies of this genus, but microsatellite markers have only been used in few investigations. Microsatellites are very informative and are useful to assess genetic variability, analyze mating systems and in genetic mapping. The objectives of this study were to develop A. hypogaea microsatellite loci and to evaluate the transferability of these markers to other Arachis species. Results Thirteen loci were isolated and characterized using 16 accessions of A. hypogaea. The level of variation found in A. hypogaea using microsatellites was higher than with other markers. Cross-transferability of the markers was also high. Sequencing of the fragments amplified using the primer pair Ah11 from 17 wild Arachis species showed that almost all wild species had similar repeated sequence to the one observed in A. hypogaea. Sequence data suggested that there is no correlation between taxonomic relationship of a wild species to A. hypogaea and the number of repeats found in its microsatellite loci. Conclusion These results show that microsatellite primer pairs from A. hypogaea have multiple uses. A higher level of variation among A. hypogaea accessions can be detected using microsatellite markers in comparison to other markers, such as RFLP, RAPD and AFLP. The microsatellite primers of A. hypogaea showed a very high rate of transferability to other species of the genus. These primer pairs provide important tools to evaluate the genetic variability and to assess the mating system in Arachis species. PMID:17326826

  17. Microsatellites for Oenothera gayleana and O. hartwegii subsp. filifolia (Onagraceae), and their utility in section Calylophus1

    PubMed Central

    Lewis, Emily M.; Fant, Jeremie B.; Moore, Michael J.; Hastings, Amy P.; Larson, Erica L.; Agrawal, Anurag A.; Skogen, Krissa A.

    2016-01-01

    Premise of the study: Eleven nuclear and four plastid microsatellite markers were screened for two gypsum endemic species, Oenothera gayleana and O. hartwegii subsp. filifolia, and tested for cross-amplification in the remaining 11 taxa within Oenothera sect. Calylophus (Onagraceae). Methods and Results: Microsatellite markers were tested in two to three populations spanning the ranges of both O. gayleana and O. hartwegii subsp. filifolia. The nuclear microsatellite loci consisted of both di- and trinucleotide repeats with one to 17 alleles per population. Several loci showed significant deviation from Hardy–Weinberg equilibrium, which may be evidence of chromosomal rings. The plastid microsatellite markers identified one to seven haplotypes per population. The transferability of these markers was confirmed in all 11 taxa within Oenothera sect. Calylophus. Conclusions: The microsatellite loci characterized here are the first developed and tested in Oenothera sect. Calylophus. These markers will be used to assess whether pollinator foraging distance influences population genetic parameters in predictable ways. PMID:26949578

  18. Polymorphic microsatellite loci for the razor clam, Sinonovacula constricta.

    PubMed

    Ma, H-T; Jiang, H-B; Liu, X-Q; Wu, X-P; Wei, X-M

    2015-01-15

    The razor clam, Sinonovacula constricta, is an important commercial bivalve and a popular mollusca food in China. Twelve polymorphic microsatellite markers were isolated from the razor clam using a partial genomic library enriched for tandem repeat sequences of (CA)16, (GA)16. Polymorphisms of these loci were evaluated in a wild population of 30 individuals. The allele number of these polymorphic markers ranged from 5-15 per locus with an average of 9.333. Observed and expected heterozygosity values ranged from 0.192-1.000 and 0.219-0.906. Polymorphism information content ranged from 0.209-0.892 with an average of 0.704. Three loci significantly deviated from Hardy-Weinberg equilibrium after Bonferroni correction. No significant linkage disequilibrium was detected between these loci. This set of microsatellite loci are useful for genetic studies in S. constricta.

  19. Microsatellite data support subpopulation structuring among Basques.

    PubMed

    Pérez-Miranda, Ana M; Alfonso-Sánchez, Miguel A; Kalantar, Arif; García-Obregón, Susana; de Pancorbo, Marian M; Peña, José A; Herrera, Rene J

    2005-01-01

    Genomic diversity based on 13 short tandem repeat (STR) loci (D3S1358, vWA, FGA, D8S1179, D21S11, D18S51, D5S818, D13S317, D7S820, D16S539, TH01, TPOX, and CSF1PO) is reported for the first time in Basques from the provinces of Guipúzcoa and Navarre (Spain). STR data from previous studies on Basques from Alava and Vizcaya provinces were also examined using hierarchal analysis of molecular variance (AMOVA) and genetic admixture estimations to ascertain whether the Basques are genetically heterogeneous. To assess the genetic position of Basques in a broader geographic context, we conducted phylogenetic analyses based on F(ST) genetic distances [neighbor-joining trees and multidimensional scaling (MDS)] using data compiled in previous publications. The genetic profile of the Basque groups revealed distinctive regional partitioning of short tandem repeat (STR) diversity. Consistent with the above, native Basques clearly segregated from other populations from Europe (including Spain), North Africa, and the Middle East. The main line of genetic discontinuity inferred from the spatial variability of the microsatellite diversity in Basques significantly overlapped the geographic distribution of the Basque language. The genetic heterogeneity among native Basque groups correlates with the peculiar geography of peopling and marital structure in rural Basque zones and with language boundaries resulting from the uneven impact of Romance languages in the different Basque territories.

  20. Genetic Diversity and Geographic Population Structure of Bovine Neospora caninum Determined by Microsatellite Genotyping Analysis

    PubMed Central

    Regidor-Cerrillo, Javier; Díez-Fuertes, Francisco; García-Culebras, Alicia; Moore, Dadín P.; González-Warleta, Marta; Cuevas, Carmen; Schares, Gereon; Katzer, Frank; Pedraza-Díaz, Susana; Mezo, Mercedes; Ortega-Mora, Luis M.

    2013-01-01

    The cyst-forming protozoan parasite Neosporacaninum is one of the main causes of bovine abortion worldwide and is of great economic importance in the cattle industry. Recent studies have revealed extensive genetic variation among N. caninum isolates based on microsatellite sequences (MSs). MSs may be suitable molecular markers for inferring the diversity of parasite populations, molecular epidemiology and the basis for phenotypic variations in N. caninum, which have been poorly defined. In this study, we evaluated nine MS markers using a panel of 11 N. caninum-derived reference isolates from around the world and 96 N. caninum bovine clinical samples and one ovine clinical sample collected from four countries on two continents, including Spain, Argentina, Germany and Scotland, over a 10-year period. These markers were used as molecular tools to investigate the genetic diversity, geographic distribution and population structure of N. caninum. Multilocus microsatellite genotyping based on 7 loci demonstrated high levels of genetic diversity in the samples from all of the different countries, with 96 microsatellite multilocus genotypes (MLGs) identified from 108 N. caninum samples. Geographic sub-structuring was present in the country populations according to pairwise FST. Principal component analysis (PCA) and Neighbor Joining tree topologies also suggested MLG segregation partially associated with geographical origin. An analysis of the MLG relationships, using eBURST, confirmed that the close genetic relationship observed between the Spanish and Argentinean populations may be the result of parasite migration (i.e., the introduction of novel MLGs from Spain to South America) due to cattle movement. The eBURST relationships also revealed genetically different clusters associated with the abortion. The presence of linkage disequilibrium, the co-existence of specific MLGs to individual farms and eBURST MLG relationships suggest a predominant clonal propagation for

  1. Heterozygosity increases microsatellite mutation rate

    PubMed Central

    Amos, William

    2016-01-01

    Whole genome sequencing of families of Arabidopsis has recently lent strong support to the heterozygote instability (HI) hypothesis that heterozygosity locally increases mutation rate. However, there is an important theoretical difference between the impact on base substitutions, where mutation rate increases in regions surrounding a heterozygous site, and the impact of HI on sequences such as microsatellites, where mutations are likely to occur at the heterozygous site itself. At microsatellite loci, HI should create a positive feedback loop, with heterozygosity and mutation rate mutually increasing each other. Direct support for HI acting on microsatellites is limited and contradictory. I therefore analysed AC microsatellites in 1163 genome sequences from the 1000 genomes project. I used the presence of rare alleles, which are likely to be very recent in origin, as a surrogate measure of mutation rate. I show that rare alleles are more likely to occur at locus-population combinations with higher heterozygosity even when all populations carry exactly the same number of alleles. PMID:26740567

  2. Survey and analysis of simple sequence repeats (SSRs) in three genomes of Candida species.

    PubMed

    Jia, Dongmei

    2016-06-15

    Simple sequence repeats (SSRs) or microsatellites, which composed of tandem repeated short units of 1-6 bp, have been paying attention continuously. Here, the distribution, composition and polymorphism of microsatellites and compound microsatellites were analyzed in three available genomes of Candida species (Candida dubliniensis, Candida glabrata and Candida orthopsilosis). The results show that there were 118,047, 66,259 and 61,119 microsatellites in genomes of C. dubliniensis, C. glabrata and C. orthopsilosis, respectively. The SSRs covered more than 1/3 length of genomes in the three species. The microsatellites, which just consist of bases A and (or) T, such as (A)n, (T)n, (AT)n, (TA)n, (AAT)n, (TAA)n, (TTA)n, (ATA)n, (ATT)n and (TAT)n, were predominant in the three genomes. The length of microsatellites was focused on 6 bp and 9 bp either in the three genomes or in its coding sequences. What's more, the relative abundance (19.89/kbp) and relative density (167.87 bp/kbp) of SSRs in sequence of mitochondrion of C. glabrata were significantly great than that in any one of genomes or chromosomes of the three species. In addition, the distance between any two adjacent microsatellites was an important factor to influence the formation of compound microsatellites. The analysis may be helpful for further studying the roles of microsatellites in genomes' origination, organization and evolution of Candida species.

  3. Microsatellite measures of inbreeding: a meta-analysis.

    PubMed

    Coltman, D W; Slate, J

    2003-05-01

    Meta-analyses of published and unpublished correlations between phenotypic variation and two measures of genetic variation at microsatellite loci, multilocus heterozygosity (MLH) and mean d2, revealed that the strength of these associations are generally weak (mean r < 0.10). Effects on life-history trait variation were significantly greater than zero for both measures over all reported effect sizes (r = 0.0856 and 0.0479 for MLH and mean d2, respectively), whereas effects on morphometric traits were not (r = 0.0052 and r = 0.0038), which is consistent with the prediction that life-history traits exhibit greater inbreeding depression than morphometric traits. Effect sizes reported using mean d2 were smaller and more variable than those reported using MLH, suggesting that MLH may be a better metric for capturing inbreeding depression most of the time. However, analyses of paired effect sizes reported using both measures from the same data did not differ significantly. Several lines of evidence suggest that published effects sizes are upwardly biased. First, effect sizes from published studies were significantly higher than those reported in unpublished studies. Second, fail-safe numbers for reported effect sizes were generally quite low, with the exception of correlations between MLH and life-history traits. Finally, the slope of the regression of effect size on sample size was negative for most sets of traits. Taken together, these results suggest that studies designed to detect inbreeding depression on a life-history trait using microsatellites will need to sample in excess of 600 individuals to detect an average effect size (r = 0.10) with reasonable statistical power (0.80). Very few published studies have used samples sizes approaching this value.

  4. Historical and contemporary multilocus population structure of Ascochyta rabiei (teleomorph: Didymella rabiei) in the Pacific Northwest of the United States.

    PubMed

    Peever, T L; Salimath, S S; Su, G; Kaiser, W J; Muehlbauer, F J

    2004-02-01

    The historical and contemporary population genetic structure of the chickpea Ascochyta blight pathogen, Ascochyta rabiei (teleomorph: Didymella rabiei), was determined in the US Pacific Northwest (PNW) using 17 putative AFLP loci, four genetically characterized, sequence-tagged microsatellite loci (STMS) and the mating type locus (MAT). A single multilocus genotype of A. rabiei (MAT1-1) was detected in 1983, which represented the first recorded appearance of Ascochyta blight of chickpea in the PNW. During the following year many additional alleles, including the other mating type allele (MAT1-2), were detected. By 1987, all alleles currently found in the PNW had been introduced. Highly significant genetic differentiation was detected among contemporary subpopulations from different hosts and geographical locations indicating restricted gene flow and/or genetic drift occurring within and among subpopulations and possible selection by host cultivar. Two distinct populations were inferred with high posterior probability which correlated to host of origin and date of sample using Bayesian model-based population structure analyses of multilocus genotypes. Allele frequencies, genotype distributions and population assignment probabilities were significantly different between the historical and contemporary samples of isolates and between isolates sampled from a resistance screening nursery and those sampled from commercial chickpea fields. A random mating model could not be rejected in any subpopulation, indicating the importance of the sexual stage of the fungus both as a source of primary inoculum for Ascochyta blight epidemics and potentially adaptive genotypic diversity.

  5. Multilocus Sequence Typing of Clostridium difficile▿

    PubMed Central

    Griffiths, David; Fawley, Warren; Kachrimanidou, Melina; Bowden, Rory; Crook, Derrick W.; Fung, Rowena; Golubchik, Tanya; Harding, Rosalind M.; Jeffery, Katie J. M.; Jolley, Keith A.; Kirton, Richard; Peto, Tim E.; Rees, Gareth; Stoesser, Nicole; Vaughan, Alison; Walker, A. Sarah; Young, Bernadette C.; Wilcox, Mark; Dingle, Kate E.

    2010-01-01

    A robust high-throughput multilocus sequence typing (MLST) scheme for Clostridium difficile was developed and validated using a diverse collection of 50 reference isolates representing 45 different PCR ribotypes and 102 isolates from recent clinical samples. A total of 49 PCR ribotypes were represented overall. All isolates were typed by MLST and yielded 40 sequence types (STs). A web-accessible database was set up (http://pubmlst.org/cdifficile/) to facilitate the dissemination and comparison of C. difficile MLST genotyping data among laboratories. MLST and PCR ribotyping were similar in discriminatory abilities, having indices of discrimination of 0.90 and 0.92, respectively. Some STs corresponded to a single PCR ribotype (32/40), other STs corresponded to multiple PCR ribotypes (8/40), and, conversely, the PCR ribotype was not always predictive of the ST. The total number of variable nucleotide sites in the concatenated MLST sequences was 103/3,501 (2.9%). Concatenated MLST sequences were used to construct a neighbor-joining tree which identified four phylogenetic groups of STs and one outlier (ST-11; PCR ribotype 078). These groups apparently correlate with clades identified previously by comparative genomics. The MLST scheme was sufficiently robust to allow direct genotyping of C. difficile in total stool DNA extracts without isolate culture. The direct (nonculture) MLST approach may prove useful as a rapid genotyping method, potentially benefiting individual patients and informing hospital infection control. PMID:20042623

  6. Multilocus sequence analysis (MLSA) in prokaryotic taxonomy.

    PubMed

    Glaeser, Stefanie P; Kämpfer, Peter

    2015-06-01

    To obtain a higher resolution of the phylogenetic relationships of species within a genus or genera within a family, multilocus sequence analysis (MLSA) is currently a widely used method. In MLSA studies, partial sequences of genes coding for proteins with conserved functions ('housekeeping genes') are used to generate phylogenetic trees and subsequently deduce phylogenies. However, MLSA is not only suggested as a phylogenetic tool to support and clarify the resolution of bacterial species with a higher resolution, as in 16S rRNA gene-based studies, but has also been discussed as a replacement for DNA-DNA hybridization (DDH) in species delineation. Nevertheless, despite the fact that MLSA has become an accepted and widely used method in prokaryotic taxonomy, no common generally accepted recommendations have been devised to date for either the whole area of microbial taxonomy or for taxa-specific applications of individual MLSA schemes. The different ways MLSA is performed can vary greatly for the selection of genes, their number, and the calculation method used when comparing the sequences obtained. Here, we provide an overview of the historical development of MLSA and critically review its current application in prokaryotic taxonomy by highlighting the advantages and disadvantages of the method's numerous variations. This provides a perspective for its future use in forthcoming genome-based genotypic taxonomic analyses.

  7. Multilocus sequence typing of Propionibacterium freudenreichii.

    PubMed

    Dalmasso, Marion; Nicolas, Pierre; Falentin, Hélène; Valence, Florence; Tanskanen, Jarna; Jatila, Hanna; Salusjärvi, Tuomas; Thierry, Anne

    2011-01-31

    Propionibacterium freudenreichii is used as a ripening culture in Swiss cheese manufacture. This study investigates the molecular diversity and the population structure of this bacterium via multilocus sequence typing (MLST). Internal fragments of seven genes sequenced for 113 strains of different subspecies and origins allowed the resolution of 46 sequence types (STs) with occurrence frequencies ranging from 1 to 11. The core genome of the species harbours a low level of nucleotide polymorphism. In our data, single nucleotide polymorphisms account for only 2.28% of the concatenated sequences, and the average polymorphism rate in pairwise comparisons is 0.46%. The analyses reveal quantitatively comparable contributions of recombination and mutation in nucleotide changes at core genome loci along cell lineages. Remarkably, the STs exhibit little if any dairy biotope specialization. Phenotypic characterisation of the strains, based on their aptitude to use lactose and nitrate, shows that the two previously identified subspecies (freudenreichii and shermani) do not reflect the ancestral relationships in the P. freudenreichii population. The considerable phenotypic heterogeneity, found even at the ST level, suggests instead a history of recurrent switches between phenotypes.

  8. Development of microsatellite markers for buffalograss (Buchloë dactyloides; Poaceae), a drought-tolerant turfgrass alternative1

    PubMed Central

    Hadle, Jacob J.; Konrade, Lauren A.; Beasley, Rochelle R.; Lance, Stacey L.; Jones, Kenneth L.; Beck, James B.

    2016-01-01

    Premise of the study: Buchloë dactyloides (Poaceae) is an important component of Great Plains prairies and a popular drought-tolerant turfgrass alternative in North America. This species comprises an autopolyploid series, and microsatellite primers were developed to understand the distribution of genetic variation among cytotypes and across its large geographic range. Methods and Results: Fifteen microsatellite loci were designed and successfully amplified in six B. dactyloides populations. Within-population genetic diversity was comparatively high, consistent with B. dactyloides’ life history. Allelic variation at 13 loci was consistent with the cytotype established in chromosome-counted samples. Conclusions: This variable, interpretable set of loci allows for the determination of multilocus genotype in B. dactyloides individuals of varying cytotype. Data such as these from a range-wide sample set can provide important insights for germplasm conservation and crop improvement in this ecologically and economically important species. PMID:27610277

  9. Characterization of 14 microsatellite markers for genetic analysis and cultivar identification of walnut

    Technology Transfer Automated Retrieval System (TEKTRAN)

    One hundred and forty-seven primer pairs originally designed to amplify microsatellites, also known as simple sequence repeats (SSR), in black walnut (Juglans nigra L.) were screened for utility in persian walnut (J. regia L.). Based on scorability and number of informative polymorphisms, the best 1...

  10. What phylogeny and gene genealogy analyses reveal about homoplasy in citrus microsatellite alleles

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Sixty-five microsatellite alleles from three Simple Sequence Repeat (SSR) loci (cAGG9, CCT01 and GT03) of various Citrus, Fortunella or Poncirus accessions were cloned and sequenced to determine their mode of evolution. This data was used to assess sequence variation by calculating the average numb...

  11. Discrimination of American cranberry cultivars and assessment of clonal heterogeneity using microsatellite markers

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cranberries (Vaccinium macrocarpon Ait.) are an economically important fruit crop derived from a North American native species. We report the application of 12 simple sequence repeats (SSR) or microsatellite markers to assess the genetic diversity of cranberry cultivars. We studied 164 samples of 21...

  12. Development of microsatellite markers for Fusicladium effusum, the causal agent of pecan scab

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Pecan scab (caused by F. effusum) is the most important diseases of pecan in the southeastern U.S. Microsatellite (simple sequence repeat, SSR) motifs were mined from the genome of Fusicladium effusum assembled from 454 pyrosequencing and Illumina Miseq reads. A total of 278 SSR primers were designe...

  13. Development and characterization of microsatellite markers (SSR) in Sesamum (Sesamum indicum L.) species.

    PubMed

    Spandana, B; Reddy, V Prathap; Prasanna, G John; Anuradha, G; Sivaramakrishnan, S

    2012-11-01

    Microsatellites, also known as simple sequence repeats (SSRs), are the class of repetitive DNA sequences present throughout the genome of many plant and animal species. Recent advances in molecular genetics had been the introduction of microsatellite markers to investigate the genetic structuring of natural plant populations. We have employed an enrichment strategy for microsatellite isolation by using multi-enzymes digestion, microsatellite oligoprobes, and streptavidin magnetic beads in Sesamum (Sesamum indicum L.). More than 200 SSR motifs were detected (SSR motifs ≥2 repeat units or 6 bp); 80 % of the clones contained SSR motifs. When regarding SSRs with four or more repeat units and a minimum length of 10 bp, 132 of them showed repeats. Eighteen SSR markers were initially characterized for optimum annealing temperature using a gradient PCR technique. Among the 18 SSR markers characterized, five were found to be polymorphic and used to analyze 60 Sesamum germplasm accessions. The maximum number of alleles detected was four with a single primer and the least number of two alleles with three primers with an average PIC value of 0.77. SSRs are a valuable tool for estimating genetic diversity and analyzing the evolutionary and historical development of cultivars at the genomic level in sesame breeding programs.

  14. High resolution melting detects sequence polymorphism in rubus occidentalis L. monomorphic microsatellite markers

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Microsatellite, or simple sequence repeat (SSR) markers, are valuable as co-dominant genetic markers with a variety of applications such as DNA fingerprinting, linkage mapping, and population structure analysis. However, primer pairs designed from the regions that flank SSRs often generate fragment...

  15. Development and transferability of black and red raspberry microsatellite markers from short-read sequences

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The advent of next-generation sequencing technologies has been a boon to the cost-effective development of molecular markers, particularly in non-model species. Here, we demonstrate the efficiency of microsatellite or simple sequence repeat (SSR) marker development from short-read sequences using th...

  16. Multilocus typing and population structure of Cryptosporidium from children in Zaragoza, Spain.

    PubMed

    Ramo, Ana; Quílez, Joaquín; Vergara-Castiblanco, Claudia; Monteagudo, Luis; Del Cacho, Emilio; Clavel, Antonio

    2015-04-01

    A multilocus typing approach with eight variable-number tandem-repeat (VNTR) loci and the GP60 gene was used to analyze the inter- and intra-species variation of 44 Cryptosporidium isolates from pediatric patients in Zaragoza city (NE, Spain). Restriction and sequence analyses of the SSU rRNA gene revealed that Cryptosporidium transmission is mostly anthroponotic in this area, with the predominance of Cryptosporidium hominis (n: 41) over Cryptosporidium parvum (n: 3). GP60 subtyping showed limited genetic diversity and four subtypes were identified, including IbA10G2 (n: 35), IaA24R3 (n: 6), IIaA15G1R1 (n: 1) and IIaA15G2R1 (n: 2). Five out of eight VNTR loci showed a discriminatory power higher than the GP60 gene, although each locus had a predominant allele exhibited by more than 50% of isolates. All but four alleles were associated to either C. hominis or C. parvum and linked alleles at different loci were found. Multilocus typing substantially increased the discriminatory power (Hunter-Gaston index: 0.807, 95% CI, 0.683-0.926) and revealed that genetic diversity is much higher than that reported by GP60 sequencing, since 17 multilocus subtypes (MLTs) were identified. Nearly half of the specimens were allocated to a single major MLT. However, no more than three specimens were allocated to each of the remaining MLTs. Both phylogenetic and population analyses revealed a population clustering of C. hominis according to the GP60 subtype, which indicates the robustness of this marker to differentiate genetic subpopulations. Subpopulations had an overall clonal genetic structure, although traces of genetic flow between them were also observed.

  17. Multilocus Sequence Typing of Lactobacillus casei Reveals a Clonal Population Structure with Low Levels of Homologous Recombination▿ †

    PubMed Central

    Diancourt, Laure; Passet, Virginie; Chervaux, Christian; Garault, Peggy; Smokvina, Tamara; Brisse, Sylvain

    2007-01-01

    Robust genotyping methods for Lactobacillus casei are needed for strain tracking and collection management, as well as for population biology research. A collection of 52 strains initially labeled L. casei or Lactobacillus paracasei was first subjected to rplB gene sequencing together with reference strains of Lactobacillus zeae, Lactobacillus rhamnosus, and other species. Phylogenetic analysis showed that all 52 strains belonged to a single compact L. casei-L. paracasei sequence cluster, together with strain CIP107868 (= ATCC 334) but clearly distinct from L. rhamnosus and from a cluster with L. zeae and CIP103137T (= ATCC 393T). The strains were genotyped using amplified fragment length polymorphism, multilocus sequence typing based on internal portions of the seven housekeeping genes fusA, ileS, lepA, leuS, pyrG, recA, and recG, and tandem repeat variation (multilocus variable-number tandem repeats analysis [MLVA] using nine loci). Very high concordance was found between the three methods. Although amounts of nucleotide variation were low for the seven genes (π ranging from 0.0038 to 0.0109), 3 to 12 alleles were distinguished, resulting in 31 sequence types. One sequence type (ST1) was frequent (17 strains), but most others were represented by a single strain. Attempts to subtype ST1 strains by MLVA, ribotyping, clustered regularly interspaced short palindromic repeat characterization, and single nucleotide repeat variation were unsuccessful. We found clear evidence for homologous recombination during the diversification of L. casei clones, including a putative intragenic import of DNA into one strain. Nucleotides were estimated to change four times more frequently by recombination than by mutation. However, statistical congruence between individual gene trees was retained, indicating that recombination is not frequent enough to disrupt the phylogenetic signal. The developed multilocus sequence typing scheme should be useful for future studies of L. casei

  18. Multilocus DNA fingerprints in gallinaceous birds: general approach and problems.

    PubMed

    Hanotte, O; Bruford, M W; Burke, T

    1992-06-01

    Multilocus profiles were investigated in five different species of Galliformes (ring-necked pheasant Phasianus colchicus, Indian peafowl Pavo cristatus, Japanese quail Coturnix coturnix japonica, domestic chicken Gallus gallus, and red grouse Lagopus lagopus scoticus) using two human multilocus probes (33.6 and 33.15) in combination with each of four restriction enzymes (AluI, DdeI, HaeIII or HinfI). All the species show a DNA fingerprint-like pattern using at least one restriction enzyme in combination with each multilocus probe. The number of bands detected and the value of the index of similarity for each species differ significantly between the profiles obtained with each multilocus probe. Some enzyme/probe combinations reveal strong cross-hybridization of the multilocus probes with satellite or satellite-like DNA sequences in pheasant, peacock, quail and chicken, which partially or completely prevented scoring of the profile. The choice of restriction enzyme was found to influence the number of bands, the value of the index of similarity and the probability of obtaining an identical fingerprint between unrelated individuals. The Mendelian inheritance and independent segregation of the fragments detected using AluI was investigated in three species (ring-necked pheasant, Indian peafowl and red grouse). Some bands were shown to be tightly linked. An extreme case was encountered in the red grouse, where 12 of the 15 bands scored in one parent represented only two, apparently allelic, haplotypes and so derived from a single locus. However, fingerprint patterns will often be adequate for use in paternity analyses, such as in behavioural studies, despite the occurrence of haplotypic sets of bands. Identical DNA multilocus profiles were sometimes observed between captive-bred siblings in one species. These results emphasize the desirability of determining, in each new species, the optimal experimental conditions as a preliminary to any behavioural or population

  19. Tracing the origin of HLA-DRB1 alleles by microsatellite polymorphism.

    PubMed Central

    Bergström, T F; Engkvist, H; Erlandsson, R; Josefsson, A; Mack, S J; Erlich, H A; Gyllensten, U

    1999-01-01

    We analyzed the origin of allelic diversity at the class II HLA-DRB1 locus, using a complex microsatellite located in intron 2, close to the polymorphic second exon. A phylogenetic analysis of human, gorilla, and chimpanzee DRB1 sequences indicated that the structure of the microsatellite has evolved, primarily by point mutations, from a putative ancestral (GT)x(GA)y-complex-dinucleotide repeat. In all contemporary DRB1 allelic lineages, with the exception of the human *04 and the gorilla *08 lineages, the (GA)y repeat is interrupted, often by a G-->C substitution. In general, the length of the 3' (GA)y repeat correlates with the allelic lineage and thus evolves more slowly than a middle (GA)z repeat, whose length correlates with specific alleles within the lineage. Comparison of the microsatellite sequence from 30 human DRB1 alleles showed the longer 5' (GT)x to be more variable than the shorter middle (GA)z and 3' (GA)y repeats. Analysis of multiple samples with the same exon sequence, derived from different continents, showed that the 5' (GT)x repeat evolves more rapidly than the middle (GA)z and the 3' (GA)y repeats, which is consistent with findings of a higher mutation rate for longer tracts. The microsatellite-repeat-length variation was used to trace the origin of new DRB1 alleles, such as the new *08 alleles found in the Cayapa people of Ecuador and the Ticuna people of Brazil. PMID:10330359

  20. Genome-wide microsatellite characterization and marker development in the sequenced Brassica crop species.

    PubMed

    Shi, Jiaqin; Huang, Shunmou; Zhan, Jiepeng; Yu, Jingyin; Wang, Xinfa; Hua, Wei; Liu, Shengyi; Liu, Guihua; Wang, Hanzhong

    2014-02-01

    Although much research has been conducted, the pattern of microsatellite distribution has remained ambiguous, and the development/utilization of microsatellite markers has still been limited/inefficient in Brassica, due to the lack of genome sequences. In view of this, we conducted genome-wide microsatellite characterization and marker development in three recently sequenced Brassica crops: Brassica rapa, Brassica oleracea and Brassica napus. The analysed microsatellite characteristics of these Brassica species were highly similar or almost identical, which suggests that the pattern of microsatellite distribution is likely conservative in Brassica. The genomic distribution of microsatellites was highly non-uniform and positively or negatively correlated with genes or transposable elements, respectively. Of the total of 115 869, 185 662 and 356 522 simple sequence repeat (SSR) markers developed with high frequencies (408.2, 343.8 and 356.2 per Mb or one every 2.45, 2.91 and 2.81 kb, respectively), most represented new SSR markers, the majority had determined physical positions, and a large number were genic or putative single-locus SSR markers. We also constructed a comprehensive database for the newly developed SSR markers, which was integrated with public Brassica SSR markers and annotated genome components. The genome-wide SSR markers developed in this study provide a useful tool to extend the annotated genome resources of sequenced Brassica species to genetic study/breeding in different Brassica species.

  1. Partners in crime: bidirectional transcription in unstable microsatellite disease.

    PubMed

    Batra, Ranjan; Charizanis, Konstantinos; Swanson, Maurice S

    2010-04-15

    Nearly two decades have passed since the discovery that the expansion of microsatellite trinucleotide repeats is responsible for a prominent class of neurological disorders, including Huntington disease and fragile X syndrome. These hereditary diseases are characterized by genetic anticipation or the intergenerational increase in disease severity accompanied by a decrease in age-of-onset. The revelation that the variable expansion of simple sequence repeats accounted for anticipation spawned a number of pathogenesis models and a flurry of studies designed to reveal the molecular events affected by these expansions. This work led to our current understanding that expansions in protein-coding regions result in extended homopolymeric amino acid tracts, often polyglutamine or polyQ, and deleterious protein gain-of-function effects. In contrast, expansions in noncoding regions cause RNA-mediated toxicity. However, the realization that the transcriptome is considerably more complex than previously imagined, as well as the emerging regulatory importance of antisense RNAs, has blurred this distinction. In this review, we summarize evidence for bidirectional transcription of microsatellite disease genes and discuss recent suggestions that some repeat expansions produce variable levels of both toxic RNAs and proteins that influence cell viability, disease penetrance and pathological severity.

  2. Isolation and characterization of novel microsatellites for Abies koreana and A. nephrolepis (Pinaceae).

    PubMed

    Hong, J K; Lim, J; Lee, B Y; Kwak, M

    2016-04-28

    Abies koreana is an endemic and rare species from Korea and is classified as endangered by the International Union for Conservation of Nature. Although the genetic diversity assessment for current population of A. koreana needs to be performed urgently, no microsatellite markers have been developed for this species. In the present study, we developed 22 novel polymorphic microsatellite loci and the characteristics of these loci were determined in A. koreana as well as in Abies nephrolepis, the most closely related species, and these loci were compared with previously reported microsatellite markers developed for the Abies genus. Genomic sequence (161 Mbp; 325,776 reads) was obtained from one individual of A. koreana using Roche 454 GS-FLX Titanium sequencing and 19,258 repeat motifs were identified from it. A total of 288 primer pairs with high copy numbers of di-repeat motifs were evaluated for amplification in A. koreana and A. nephrolepis. A total of 71 primer pairs successfully amplified fragments, of which 22 showed polymorphisms in A. koreana and A. nephrolepis. The average expected diversity was 0.767 and 0.717 in A. koreana and A. nephrolepis, respectively; these heterozygosity levels were moderate compared to the previously reported microsatellite loci from Abies species. This is the first set of microsatellite markers developed for A. koreana as well as A. nephrolepis and further population genetic studies of both species and genetic delimitation can be carried out for the species conservation and management.

  3. Novel microsatellite markers acquired from Rubus coreanus Miq. and cross-amplification in other Rubus species.

    PubMed

    Lee, Gi-An; Song, Jae Young; Choi, Heh-Ran; Chung, Jong-Wook; Jeon, Young-Ah; Lee, Jung-Ro; Ma, Kyung-Ho; Lee, Myung-Chul

    2015-04-10

    The Rubus genus consists of more than 600 species that are distributed globally. Only a few Rubus species, including raspberries and blueberries, have been domesticated. Genetic diversity within and between Rubus species is an important resource for breeding programs. We developed genomic microsatellite markers using an SSR-enriched R. coreanus library to study the diversity of the Rubus species. Microsatellite motifs were discovered in 546 of 646 unique clones, and a dinucleotide repeat was the most frequent (75.3%) type of repeat. From 97 microsatellite loci with reproducible amplicons, we acquired 29 polymorphic microsatellite markers in the Rubus coreanus collection. The transferability values ranged from 59.8% to 84% across six Rubus species, and Rubus parvifolius had the highest transferability value (84%). The average number of alleles and the polymorphism information content were 5.7 and 0.541, respectively, in the R. coreanus collection. The diversity index of R. coreanus was similar to the values reported for other Rubus species. A phylogenetic dendrogram based on SSR profiles revealed that seven Rubus species could be allocated to three groups, and that R. coreanus was genetically close to Rubus crataegifolius (mountain berry). These new microsatellite markers might prove useful in studies of the genetic diversity, population structure, and evolutionary relationships among Rubus species.

  4. A novel microsatellite control system

    SciTech Connect

    Moore, K.R.; Frigo, J.R.; Tilden, M.W.

    1998-02-01

    The authors are researching extremely simple yet quite capable analog pulse-coded neural networks for ``smaller-faster-cheaper`` spacecraft attitude and control systems. The will demonstrate a prototype microsatellite that uses their novel control method to autonomously stabilize itself in the ambient magnetic field and point itself at the brightest available light source. Though still in design infancy, the ``Nervous Net`` controllers described could allow for space missions not currently possible given conventional satellite hardware. Result, prospects and details are presented.

  5. Contrasting microsatellite diversity in the evolutionary lineages of Phytophthora lateralis.

    PubMed

    Vettraino, AnnaMaria; Brasier, Clive M; Webber, Joan F; Hansen, Everett M; Green, Sarah; Robin, Cecile; Tomassini, Alessia; Bruni, Natalia; Vannini, Andrea

    2017-02-01

    Following recent discovery of Phytophthora lateralis on native Chamaecyparis obtusa in Taiwan, four phenotypically distinct lineages were discriminated: the Taiwan J (TWJ) and Taiwan K (TWK) in Taiwan, the Pacific Northwest (PNW) in North America and Europe and the UK in west Scotland. Across the four lineages, we analysed 88 isolates from multiple sites for microsatellite diversity. Twenty-one multilocus genotypes (MLGs) were resolved with high levels of diversity of the TWK and PNW lineages. No alleles were shared between the PNW and the Taiwanese lineages. TWK was heterozygous at three loci, whereas TWJ isolates were homozygous apart from one isolate, which exhibited a unique allele also present in the TWK lineage. PNW lineage was heterozygous at three loci. The evidence suggests its origin may be a yet unknown Asian source. North American and European PNW isolates shared all their alleles and also a dominant MLG, consistent with a previous proposal that this lineage is a recent introduction into Europe from North America. The UK lineage was monomorphic and homozygous at all loci. It shared its alleles with the PNW and the TWJ and TWK lineages, hence a possible origin in a recent hybridisation event between a Taiwan lineage and PNW cannot be ruled out.

  6. Multi-locus Analyses Reveal Four Giraffe Species Instead of One.

    PubMed

    Fennessy, Julian; Bidon, Tobias; Reuss, Friederike; Kumar, Vikas; Elkan, Paul; Nilsson, Maria A; Vamberger, Melita; Fritz, Uwe; Janke, Axel

    2016-09-26

    Traditionally, one giraffe species and up to eleven subspecies have been recognized [1]; however, nine subspecies are commonly accepted [2]. Even after a century of research, the distinctness of each giraffe subspecies remains unclear, and the genetic variation across their distribution range has been incompletely explored. Recent genetic studies on mtDNA have shown reciprocal monophyly of the matrilines among seven of the nine assumed subspecies [3, 4]. Moreover, until now, genetic analyses have not been applied to biparentally inherited sequence data and did not include data from all nine giraffe subspecies. We sampled natural giraffe populations from across their range in Africa, and for the first time individuals from the nominate subspecies, the Nubian giraffe, Giraffa camelopardalis camelopardalis Linnaeus 1758 [5], were included in a genetic analysis. Coalescence-based multi-locus and population genetic analyses identify at least four separate and monophyletic clades, which should be recognized as four distinct giraffe species under the genetic isolation criterion. Analyses of 190 individuals from maternal and biparental markers support these findings and further suggest subsuming Rothschild's giraffe into the Nubian giraffe, as well as Thornicroft's giraffe into the Masai giraffe [6]. A giraffe survey genome produced valuable data from microsatellites, mobile genetic elements, and accurate divergence time estimates. Our findings provide the most inclusive analysis of giraffe relationships to date and show that their genetic complexity has been underestimated, highlighting the need for greater conservation efforts for the world's tallest mammal.

  7. New methods employing multilocus genotypes to select or exclude populations as origins of individuals.

    PubMed Central

    Cornuet, J M; Piry, S; Luikart, G; Estoup, A; Solignac, M

    1999-01-01

    A new method for assigning individuals of unknown origin to populations, based on the genetic distance between individuals and populations, was compared to two existing methods based on the likelihood of multilocus genotypes. The distribution of the assignment criterion (genetic distance or genotype likelihood) for individuals of a given population was used to define the probability that an individual belongs to the population. Using this definition, it becomes possible to exclude a population as the origin of an individual, a useful extension of the currently available assignment methods. Using simulated data based on the coalescent process, the different methods were evaluated, varying the time of divergence of populations, the mutation model, the sample size, and the number of loci. Likelihood-based methods (especially the Bayesian method) always performed better than distance methods. Other things being equal, genetic markers were always more efficient when evolving under the infinite allele model than under the stepwise mutation model, even for equal values of the differentiation parameter F(st). Using the Bayesian method, a 100% correct assignment rate can be achieved by scoring ca. 10 microsatellite loci (H approximately 0.6) on 30-50 individuals from each of 10 populations when the F(st) is near 0.1. PMID:10581301

  8. Design and Implementation of Degenerate Microsatellite Primers for the Mammalian Clade

    PubMed Central

    Buschiazzo, Emmanuel; Beck, Josephine S.; Gemmell, Neil J.

    2011-01-01

    Microsatellites are popular genetic markers in molecular ecology, genetic mapping and forensics. Unfortunately, despite recent advances, the isolation of de novo polymorphic microsatellite loci often requires expensive and intensive groundwork. Primers developed for a focal species are commonly tested in a related, non-focal species of interest for the amplification of orthologous polymorphic loci; when successful, this approach significantly reduces cost and time of microsatellite development. However, transferability of polymorphic microsatellite loci decreases rapidly with increasing evolutionary distance, and this approach has shown its limits. Whole genome sequences represent an under-exploited resource to develop cross-species primers for microsatellites. Here we describe a three-step method that combines a novel in silico pipeline that we use to (1) identify conserved microsatellite loci from a multiple genome alignments, (2) design degenerate primer pairs, with (3) a simple PCR protocol used to implement these primers across species. Using this approach we developed a set of primers for the mammalian clade. We found 126,306 human microsatellites conserved in mammalian aligned sequences, and isolated 5,596 loci using criteria based on wide conservation. From a random subset of ∼1000 dinucleotide repeats, we designed degenerate primer pairs for 19 loci, of which five produced polymorphic fragments in up to 18 mammalian species, including the distinctly related marsupials and monotremes, groups that diverged from other mammals 120–160 million years ago. Using our method, many more cross-clade microsatellite loci can be harvested from the currently available genomic data, and this ability is set to improve exponentially as further genomes are sequenced. PMID:22216321

  9. Paternity identification in sugarcane polycrosses by using microsatellite markers.

    PubMed

    Xavier, M A; Pinto, L R; Fávero, T M; Perecin, D; Carlini-Garcia, L A; Landell, M G A

    2014-03-31

    Although polycrosses have been used to test the potential of cross-combination of a large number of sugarcane parents, the male parent of the half-sib progenies produced is unknown. The present study aimed to integrate the molecular marker technology to the sugarcane polycross approach by the application of microsatellite markers to identify the male parent of 41 elite clones derived from polycross families. Ten microsatellite [single sequence repeats (SSRs)] primer pairs were used to identify the most likely male parent considering markers present in the selected clone but absent in the female parent. The number of alleles generated by the 10 microsatellite primer pairs ranged from 102 (cross-pollination lantern 4) to 120 (cross-pollination lantern 2) with an average of 113.25 alleles per SSR. The average genetic similarity among the involved parents in the polycrosses was 45.9%. The results of the analysis of the SSR markers absent in the female parent and present only in the selected clone as well as the genetic similarity values allowed the identification of the most likely male parent in 73% of the total clones evaluated and also to detect probable contaminations. The obtained results highlight the importance of using molecular marker technology in the identification and confirmation of the male parent of high-performance clones derived from polycrosses in the sugarcane breeding programs.

  10. Microsatellite and mitochondrial DNA analyses of Atlantic bluefin tuna (Thunnus thynnus thynnus) population structure in the Mediterranean Sea.

    PubMed

    Carlsson, Jens; McDowell, Jan R; Díaz-Jaimes, Píndaro; Carlsson, Jeanette E L; Boles, Sandra B; Gold, John R; Graves, John E

    2004-11-01

    Genetic variation was surveyed at nine microsatellite loci and the mitochondrial control region (868 bp) to test for the presence of genetic stock structure in young-of-the-year Atlantic bluefin tuna (Thunnus thynnus thynnus) from the Mediterranean Sea. Bluefin tuna were sampled over a period of 5 years from the Balearic and Tyrrhenian seas in the western basin of the Mediterranean Sea, and from the southern Ionian Sea in the eastern basin of the Mediterranean Sea. Analyses of multilocus microsatellite genotypes and mitochondrial control region sequences revealed no significant heterogeneity among collections taken from the same location in different years; however, significant spatial genetic heterogeneity was observed across all samples for both microsatellite markers and mitochondrial control region sequences (FST=0.0023, P=0.038 and PhiST=0.0233, P=0.000, respectively). Significant genetic differentiation between the Tyrrhenian and Ionian collections was found for both microsatellite and mitochondrial markers (FST=0.0087, P=0.015 and PhiST=0.0367, P=0.030, respectively). These results suggest the possibility of a genetically discrete population in the eastern basin of the Mediterranean Sea.

  11. Distribution and localization of microsatellites in the Perigord black truffle genome and identification of new molecular markers (2010) Fungal Genetics and Biology

    SciTech Connect

    Murat, Claude; Riccioni, C; Belfiori, B; Cichocki, N; Labbe, Jessy L; Morin, Emmanuelle; Tisserant, Emilie; Paolocci, F; Rubini, A; Martin, Francis

    2011-01-01

    The level of genetic diversity and genetic structure in the Perigord black truffle (Tuber melanosporum Vittad.) has been debated for several years, mainly due to the lack of appropriate genetic markers. Microsatellites or simple sequence repeats (SSRs) are important for the genome organisation, phenotypic diversity and are one of the most popular molecular markers. In this study, we surveyed the T. melanosporum genome (1) to characterise its SSR pattern; (2) to compare it with SSR patterns found in 48 other fungal and three oomycetes genomes and (3) to identify new polymorphic SSR markers for population genetics. The T. melanosporum genome is rich in SSRs with 22,425 SSRs with mono-nucleotides being the most frequent motifs. SSRs were found in all genomic regions although they are more frequent in non-coding regions (introns and intergenic regions). Sixty out of 135 PCR-amplified mono-, di-, tri-, tetra, penta, and hexanucleotides were polymorphic (44%) within black truffle populations and 27 were randomly selected and analysed on 139 T. melanosporum isolates from France, Italy and Spain. The number of alleles varied from 2 to 18 and the expected heterozygosity from 0.124 to 0.815. One hundred and thirty-two different multilocus genotypes out of the 139 T. melanosporum isolates were identified and the genotypic diversity was high (0.999). Polymorphic SSRs were found in UTR regulatory regions of fruiting bodies and ectomycorrhiza regulated genes, suggesting that they may play a role in phenotypic variation. In conclusion, SSRs developed in this study were highly polymorphic and our results showed that T. melanosporum is a species with an important genetic diversity, which is in agreement with its recently uncovered heterothallic mating system.

  12. Development of an affordable typing method for Meyerozyma guilliermondii using microsatellite markers.

    PubMed

    Wrent, Petra; Rivas, Eva-María; Peinado, José M; de Silóniz, María-Isabel

    2016-01-18

    Despite previously published methods, there is still a lack of rapid and affordable methods for genotyping the Meyerozyma guilliermondii yeast species. The development of microsatellite markers is a useful genotyping method in several yeast species. Using the Tandem Repeat Finder Software, a total of 19 microsatellite motifs (di-, tri-, and tetra- repetition) were found in silico in seven of the nine scaffolds published so far. Primer pairs were designed for all of them, although only four were used in this work. All microsatellite amplifications showed size polymorphism, and the results were identical when repeated. The combination of three microsatellite markers (sc15F/R, sc32 F/R and sc72 F/R) produced a different pattern for each of the Type Culture Collection strains of M. guilliermondii used to optimize the method. The three primer pairs can be used in the same PCR reaction, which reduces costs, in tandem with the fluorescent labeling of only the forward primer in each primer pair. Microsatellite typing was applied on 40 more M. guilliermondii strains. The results showed that no pattern is repeated between the different environmental niches. Four M. guilliermondii strains were only amplified with primer pair sc32 F/R, and subsequently identified as Meyerozyma caribbica by Taq I-RFLP of the 5.8S ITS rDNA. Most out-group species gave negative results even for physiologically similarly species such as Debaryomyces hansenii. The microsatellite markers used in this work were stable over time, which enables their use as a traceability tool.

  13. Comparative analysis of the within-population genetic structure in wild cherry (Prunus avium L.) at the self-incompatibility locus and nuclear microsatellites.

    PubMed

    Schueler, Silvio; Tusch, Alexandra; Scholz, Florian

    2006-10-01

    Gametophytic self-incompatibility (SI) systems in plants exhibit high polymorphism at the SI controlling S-locus because individuals with rare alleles have a higher probability to successfully pollinate other plants than individuals with more frequent alleles. This process, referred to as frequency-dependent selection, is expected to shape number, frequency distribution, and spatial distribution of self-incompatibility alleles in natural populations. We investigated the genetic diversity and the spatial genetic structure within a Prunus avium population at two contrasting gene loci: nuclear microsatellites and the S-locus. The S-locus revealed a higher diversity (15 alleles) than the eight microsatellites (4-12 alleles). Although the frequency distribution of S-alleles differed significantly from the expected equal distribution, the S-locus showed a higher evenness than the microsatellites (Shannon's evenness index for the S-locus: E = 0.91; for the microsatellites: E = 0.48-0.83). Also, highly significant deviations from neutrality were found for the S-locus whereas only minor deviations were found for two of eight microsatellites. A comparison of the frequency distribution of S-alleles in three age-cohorts revealed no significant differences, suggesting that different levels of selection acting on the S-locus or on S-linked sites might also affect the distribution and dynamics of S-alleles. Autocorrelation analysis revealed a weak but significant spatial genetic structure for the multilocus average of the microsatellites and for the S-locus, but could not ascertain differences in the extent of spatial genetic structure between these locus types. An indirect estimate of gene dispersal, which was obtained to explain this spatial genetic pattern, indicated high levels of gene dispersal within our population (sigma(g) = 106 m). This high gene dispersal, which may be partly due to the self-incompatibility system itself, aids the effective gene flow of the

  14. Development, characterization and cross species amplification of polymorphic microsatellite markers from expressed sequence tags of turmeric (Curcuma longa L.).

    PubMed

    Siju, S; Dhanya, K; Syamkumar, S; Sasikumar, B; Sheeja, T E; Bhat, A I; Parthasarathy, V A

    2010-02-01

    Expressed sequence tags (ESTs) from turmeric (Curcuma longa L.) were used for the screening of type and frequency of Class I (hypervariable) simple sequence repeats (SSRs). A total of 231 microsatellite repeats were detected from 12,593 EST sequences of turmeric after redundancy elimination. The average density of Class I SSRs accounts to one SSR per 17.96 kb of EST. Mononucleotides were the most abundant class of microsatellite repeat in turmeric ESTs followed by trinucleotides. A robust set of 17 polymorphic EST-SSRs were developed and used for evaluating 20 turmeric accessions. The number of alleles detected ranged from 3 to 8 per loci. The developed markers were also evaluated in 13 related species of C. longa confirming high rate (100%) of cross species transferability. The polymorphic microsatellite markers generated from this study could be used for genetic diversity analysis and resolving the taxonomic confusion prevailing in the genus.

  15. Population structure of hyperinvasive serotype 12F, clonal complex 218 Streptococcus pneumoniae revealed by multilocus boxB sequence typing

    PubMed Central

    Rakov, Alexey V.; Ubukata, Kimiko; Robinson, D. Ashley

    2011-01-01

    At least four outbreaks of invasive disease caused by serotype 12F, clonal complex 218 Streptococcus pneumoniae have occurred in the United States over the past two decades. We studied the population structure of this clonal complex using a sample of 203 outbreak and surveillance isolates that were collected over 22 years from 34 US states and eight other countries. Conventional multilocus sequence typing identified five types and distinguished a single outbreak from the others. To improve typing resolution, multilocus boxB sequence typing (MLBT) was developed from 10 variable boxB minisatellite loci. MLBT identified 86 types and distinguished between each of the four outbreaks. Diversity across boxB loci tended to be positively correlated with repeat array size and, overall, best fit the infinite alleles mutation model. Multilocus linkage disequilibrium was strong, but pairwise disequilibrium decreased with the physical distance between loci and was strongest in one large region of the chromosome, indicating recent recombinations. Two major clusters were identified in the sample, and they were differentiated geographically, as western and more easterly US clusters, and temporally, as clusters that predominated before and after the licensure of pneumococcal conjugate vaccines. The diversity and linkage disequilibrium within these two clusters also differed, suggesting different population dynamics. MLBT revealed hidden aspects of the population structure of these hyperinvasive pneumococci, and it may provide a useful adjunct tool for outbreak investigations, surveillance, and population genetics studies of other pneumococcal clonal complexes. PMID:21888992

  16. Characterization of microsatellites in the coding regions

    SciTech Connect

    Tuskan, Gerald A; Li, Shuxian; Yin, Tongming; Wang, Prof. Mingxiu

    2009-01-01

    With the development of high-throughput sequencing techniques, transcriptome sequencing projects which provide valuable resources for designing simple sequence repeat (SSR) primers have been carried out for many plants. However, the utility of SSRs for molecular breeding depends on genomewide distribution and coverage, as well as moderately high allelic variability, in the available SSR library. In this study, we characterized the exonic SSRs developed from the publicly available Populus genome as a case study to determine their value for molecular breeding. As expected, our results confirmed that microsatellites occurred approximately three times less often in coding regions than in non-coding regions. Mutability test also showed that exonic SSRs contained less allelic variability than intronic SSRs. More importantly, exonic SSRs were unevenly distributed both among and within chromosomes. Large exonic SSRs deserts were observed on several chromosomes. Differential selection between paralogous chromosomes, at the gene level, appears to be responsible for these SSR deserts, though the mechanisms that cause chromosome-specific SSR deserts are not known. This work provides ample evidence that the candidate gene approach based on unigenes identified from transcribed sequences may not be the best strategy to identify highly polymorphic SSRs.

  17. RNA-binding proteins in microsatellite expansion disorders: mediators of RNA toxicity.

    PubMed

    Echeverria, Gloria V; Cooper, Thomas A

    2012-06-26

    Although protein-mediated toxicity in neurological disease has been extensively characterized, RNA-mediated toxicity is an emerging mechanism of pathogenesis. In microsatellite expansion disorders, expansion of repeated sequences in noncoding regions gives rise to RNA that produces a toxic gain of function, while expansions in coding regions can disrupt protein function as well as produce toxic RNA. The toxic RNA typically aggregates into nuclear foci and contributes to disease pathogenesis. In many cases, toxicity of the RNA is caused by the disrupted functions of RNA-binding proteins. We will discuss evidence for RNA-mediated toxicity in microsatellite expansion disorders. Different microsatellite expansion disorders are linked with alterations in the same as well as disease-specific RNA-binding proteins. Recent studies have shown that microsatellite expansions can encode multiple repeat-containing toxic RNAs through bidirectional transcription and protein species through repeat-associated non-ATG translation. We will discuss approaches that have characterized the toxic contributions of these various factors.

  18. Construction of multilocus genetic linkage maps in humans.

    PubMed Central

    Lander, E S; Green, P

    1987-01-01

    Human genetic linkage maps are most accurately constructed by using information from many loci simultaneously. Traditional methods for such multilocus linkage analysis are computationally prohibitive in general, even with supercomputers. The problem has acquired practical importance because of the current international collaboration aimed at constructing a complete human linkage map of DNA markers through the study of three-generation pedigrees. We describe here several alternative algorithms for constructing human linkage maps given a specified gene order. One method allows maximum-likelihood multilocus linkage maps for dozens of DNA markers in such three-generation pedigrees to be constructed in minutes. PMID:3470801

  19. Identification of geographically distributed sub-populations of Leishmania (Leishmania) major by microsatellite analysis

    PubMed Central

    2008-01-01

    Background Leishmania (Leishmania) major, one of the agents causing cutaneous leishmaniasis (CL) in humans, is widely distributed in the Old World where different species of wild rodent and phlebotomine sand fly serve as animal reservoir hosts and vectors, respectively. Despite this, strains of L. (L.) major isolated from many different sources over many years have proved to be relatively uniform. To investigate the population structure of the species highly polymorphic microsatellite markers were employed for greater discrimination among it's otherwise closely related strains, an approach applied successfully to other species of Leishmania. Results Multilocus Microsatellite Typing (MLMT) based on 10 different microsatellite markers was applied to 106 strains of L. (L.) major from different regions where it is endemic. On applying a Bayesian model-based approach, three main populations were identified, corresponding to three separate geographical regions: Central Asia (CA); the Middle East (ME); and Africa (AF). This was congruent with phylogenetic reconstructions based on genetic distances. Re-analysis separated each of the populations into two sub-populations. The two African sub-populations did not correlate well with strains' geographical origin. Strains falling into the sub-populations CA and ME did mostly group according to their place of isolation although some anomalies were seen, probably, owing to human migration. Conclusion The model- and distance-based analyses of the microsatellite data exposed three main populations of L. (L.) major, Central Asia, the Middle East and Africa, each of which separated into two sub-populations. This probably correlates with the different species of rodent host. PMID:18577226

  20. Campylobacter jejuni Multilocus Sequence Types in Humans, Northwest England, 2003–2004

    PubMed Central

    Birtles, Andrew; Matthews, Margaret; Fox, Andrew; Gee, Steven; Painter, Michael; Regan, Martyn; Syed, Qutub; Bolton, Eric

    2006-01-01

    Detailed understanding of the epidemiology of Campylobacter is increasingly facilitated through use of universal and reproducible techniques for accurate strain differentiation and subtyping. Multilocus sequence typing (MLST) enables discriminatory subtyping and grouping of isolate types into genetically related clonal complexes; it also has the advantage of ease of application and repeatability. Recent studies suggest that a measure of host association may be distinguishable with this system. We describe the first continuous population-based survey to investigate the potential of MLST to resolve questions of campylobacteriosis epidemiology. We demonstrate the ability of MLST to identify variations in the epidemiology of campylobacteriosis between distinct populations and describe the distribution of key subtypes of interest. PMID:17176563

  1. Genome Wide Characterization of Simple Sequence Repeats in Cucumber

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The whole genome sequence of the cucumber cultivar Gy14 was recently sequenced at 15× coverage with the Roche 454 Titanium technology. The microsatellite DNA sequences (simple sequence repeats, SSRs) in the assembled scaffolds were computationally explored and characterized. A total of 112,073 SSRs ...

  2. Developing genome-wide microsatellite markers of bamboo and their applications on molecular marker assisted taxonomy for accessions in the genus Phyllostachys

    PubMed Central

    Zhao, Hansheng; Yang, Li; Peng, Zhenhua; Sun, Huayu; Yue, Xianghua; Lou, Yongfeng; Dong, Lili; Wang, Lili; Gao, Zhimin

    2015-01-01

    Morphology-based taxonomy via exiguously reproductive organ has severely limitation on bamboo taxonomy, mainly owing to infrequent and unpredictable flowering events of bamboo. Here, we present the first genome-wide analysis and application of microsatellites based on the genome of moso bamboo (Phyllostachys edulis) to assist bamboo taxonomy. Of identified 127,593 microsatellite repeat-motifs, the primers of 1,451 microsatellites were designed and 1,098 markers were physically mapped on the genome of moso bamboo. A total of 917 markers were successfully validated in 9 accessions with ~39.8% polymorphic potential. Retrieved from validated microsatellite markers, 23 markers were selected for polymorphic analysis among 78 accessions and 64 alleles were detected with an average of 2.78 alleles per primers. The cluster result indicated the majority of the accessions were consistent with their current taxonomic classification, confirming the suitability and effectiveness of the developed microsatellite markers. The variations of microsatellite marker in different species were confirmed by sequencing and in silico comparative genome mapping were investigated. Lastly, a bamboo microsatellites database (http://www.bamboogdb.org/ssr) was implemented to browse and search large information of bamboo microsatellites. Consequently, our results of microsatellite marker development are valuable for assisting bamboo taxonomy and investigating genomic studies in bamboo and related grass species. PMID:25620112

  3. Developing genome-wide microsatellite markers of bamboo and their applications on molecular marker assisted taxonomy for accessions in the genus Phyllostachys.

    PubMed

    Zhao, Hansheng; Yang, Li; Peng, Zhenhua; Sun, Huayu; Yue, Xianghua; Lou, Yongfeng; Dong, Lili; Wang, Lili; Gao, Zhimin

    2015-01-26

    Morphology-based taxonomy via exiguously reproductive organ has severely limitation on bamboo taxonomy, mainly owing to infrequent and unpredictable flowering events of bamboo. Here, we present the first genome-wide analysis and application of microsatellites based on the genome of moso bamboo (Phyllostachys edulis) to assist bamboo taxonomy. Of identified 127,593 microsatellite repeat-motifs, the primers of 1,451 microsatellites were designed and 1,098 markers were physically mapped on the genome of moso bamboo. A total of 917 markers were successfully validated in 9 accessions with ~39.8% polymorphic potential. Retrieved from validated microsatellite markers, 23 markers were selected for polymorphic analysis among 78 accessions and 64 alleles were detected with an average of 2.78 alleles per primers. The cluster result indicated the majority of the accessions were consistent with their current taxonomic classification, confirming the suitability and effectiveness of the developed microsatellite markers. The variations of microsatellite marker in different species were confirmed by sequencing and in silico comparative genome mapping were investigated. Lastly, a bamboo microsatellites database (http://www.bamboogdb.org/ssr) was implemented to browse and search large information of bamboo microsatellites. Consequently, our results of microsatellite marker development are valuable for assisting bamboo taxonomy and investigating genomic studies in bamboo and related grass species.

  4. Isolation and characterization of novel microsatellite markers from the sika deer (Cervus nippon) genome.

    PubMed

    Li, Y M; Bai, C Y; Niu, W P; Yu, H; Yang, R J; Yan, S Q; Zhang, J Y; Zhang, M J; Zhao, Z H

    2015-09-28

    Microsatellite markers are widely and evenly distributed, and are highly polymorphic. Rapid and convenient detection through automated analysis means that microsatellite markers are widely used in the construction of plant and animal genetic maps, in quantitative trait loci localization, marker-assisted selection, identification of genetic relationships, and genetic diversity and phylogenetic tree construction. However, few microsatellite markers remain to be isolated. We used streptavidin magnetic beads to affinity-capture and construct a (CA)n microsatellite DNA-enriched library from sika deer. We selected sequences containing more than six repeats to design primers. Clear bands were selected, which were amplified using non-specific primers following PCR amplification to screen polymorphisms in a group of 65 unrelated sika deer. The positive clone rate reached 82.9% by constructing the enriched library, and we then selected positive clones for sequencing. There were 395 sequences with CA repeats, and the CA repeat number was 4-105. We selected sequences containing more than six repeats to design primers, of which 297 pairs were designed. We next selected clear bands and used non-specific primers to amplify following PCR amplification. In total, 245 pairs of primers were screened. We then selected 50 pairs of primers to randomly screen for polymorphisms. We detected 47 polymorphic and 3 monomorphic loci in 65 unrelated sika deer. These newly isolated and characterized microsatellite loci can be used to construct genetic maps and for lineage testing in deer. In addition, they can be used for comparative genomics between Cervidae species.

  5. Application of Microsatellite Markers in Conservation Genetics and Fisheries Management: Recent Advances in Population Structure Analysis and Conservation Strategies

    PubMed Central

    Abdul-Muneer, P. M.

    2014-01-01

    Microsatellites are the most popular and versatile genetic marker with myriads of applications in population genetics, conservation biology, and evolutionary biology. These are the arrays of DNA sequences, consisting of tandemly repeating mono-, di-, tri-, and tetranucleotide units, which are distributed throughout the genomes of most eukaryotic species. Microsatellites are codominant in nature, highly polymorphic, easily typed, and Mendelian inherited, all properties which make them very suitable for the study of population structure and pedigree analysis and capable of detecting differences among closely related species. PCR for microsatellites can be automated for identifying simple sequence repeat polymorphism. Small amount of blood samples or alcohol preserved tissue is adequate for analyzing them. Most of the microsatellites are noncoding, and therefore variations are independent of natural selection. These properties make microsatellites ideal genetic markers for conservation genetics and fisheries management. This review addresses the applications of microsatellite markers in conservation genetics and recent advances in population structure analysis in the context of fisheries management. PMID:24808959

  6. Population size estimation in Yellowstone wolves with error-prone noninvasive microsatellite genotypes.

    PubMed

    Creel, Scott; Spong, Goran; Sands, Jennifer L; Rotella, Jay; Zeigle, Janet; Joe, Lawrence; Murphy, Kerry M; Smith, Douglas

    2003-07-01

    Determining population sizes can be difficult, but is essential for conservation. By counting distinct microsatellite genotypes, DNA from noninvasive samples (hair, faeces) allows estimation of population size. Problems arise because genotypes from noninvasive samples are error-prone, but genotyping errors can be reduced by multiple polymerase chain reaction (PCR). For faecal genotypes from wolves in Yellowstone National Park, error rates varied substantially among samples, often above the 'worst-case threshold' suggested by simulation. Consequently, a substantial proportion of multilocus genotypes held one or more errors, despite multiple PCR. These genotyping errors created several genotypes per individual and caused overestimation (up to 5.5-fold) of population size. We propose a 'matching approach' to eliminate this overestimation bias.

  7. DNA fingerprints of farm animals generated by microsatellite and minisatellite DNA probes.

    PubMed

    Haberfeld, A; Cahaner, A; Yoffe, O; Plotsky, Y; Hillel, J

    1991-01-01

    A multi-locus DNA probe, R18.1, derived from a bovine genomic library, detected DNA fingerprints of highly polymorphic loci in hybridization to genomic DNA from poultry and sheep, and of moderate polymorphic loci in cattle and human DNA. The average numbers of detected bands in chickens and sheep were 27.8 and 21.4, and the average band sharing levels were 0.25 and 0.33, respectively. In hybridization to cattle and human DNA, the results were less polymorphic; nevertheless, individual identification is feasible using probe R18.1. The results obtained by R18.1 were compared to results obtained by Jeffreys minisatellite probe 33.6 and two microsatellite oligonucleotides, (GT)12 and (GTG)5. The total number of detected loci using probes R18.1 and 33.6 were estimated in chickens through family analysis of broilers and the maximal number of detectable loci was calculated.

  8. Development and characterization of microsatellite markers for the Brazil nut tree Bertholletia excelsa Humb. & Bonpl. (Lecythidaceae).

    PubMed

    Reis, Alessandra M M; Braga, Aline C; Lemes, Maristerra R; Gribel, Rogério; Collevatti, Rosane G

    2009-05-01

    Twelve polymorphic microsatellite markers were developed for the Brazil nut (Bertholletia excelsa), one of the most valuable non-timber forest products from the Amazon, based on enrichment protocol. Six to 18 (mean 10.4) alleles per locus were identified and the expected heterozygosity ranged from 0.663 to 0.923 based on a screen of 40 individuals from one population of B. excelsa. The combined probabilities of genetic identity (8.39 × 10(-17) ) and paternity exclusion (0.999999) indicated that multilocus genotypes are likely to be unique allowing precise analyses of genetic structure, gene flow, and mating system of this economically important species.

  9. Microsatellite DNA markers for delineating population structure and kinship among the endangered Kirtland's warbler (Dendroica kirtlandii)

    USGS Publications Warehouse

    King, T.L.; Eackles, M.S.; Henderson, A.P.; Bocetti, C.I.; Currie, D.; Wunderle, J.M.

    2005-01-01

    We document the isolation and characterization of 23 microsatellite DNA markers for the endangered Kirtland's warbler (Dendroica kirtlandii), a Nearctic/Neotropical migrant passerine. This suite of markers revealed moderate to high levels of allelic diversity (averaging 7.7 alleles per locus) and heterozygosity (averaging 72%). Genotypic frequencies at 22 of 23 (95%) markers conformed to Hardy-Weinberg equilibrium expectations, and no linkage disequilibrium was observed in blood samples taken from 14 warblers found on the wintering grounds in the Bahamas archipelago. Multilocus genotypes resulting from this suite of markers should reduce the amount of resources required for initiating new genetic studies assessing breeding structure, parentage, demographics, and individual-level ecological interactions for D. kirtlandii. ?? 2005 Blackwell Publishing Ltd.

  10. Development of twelve microsatellite loci in the red tree corals Primnoa resedaeformis and Primnoa pacifica

    USGS Publications Warehouse

    Morrison, Cheryl; Springmann, Marcus J.; Shroades, Kelsey; Stone, Robert P.

    2015-01-01

    A suite of tetra-, penta-, and hexa-nucleotide microsatellite loci were developed from Roche 454 pyrosequencing data for the cold-water octocorals Primnoa resedaeformis and P. pacifica. Twelve of 98 primer sets tested consistently amplified in 30 P. resedaeformis samples from Baltimore Canyon (western North Atlantic Ocean) and in 24 P. pacifica samples (Shutter Ridge, eastern Gulf of Alaska). The loci displayed moderate levels of allelic diversity (average 7.5 alleles/locus) and heterozygosity (average 47 %). Levels of genetic diversity were sufficient to produce unique multi-locus genotypes and to distinguish species. These common species are long-lived (hundreds of years) and provide essential fish habitat (P. pacifica), yet populations are provided little protection from human activities. These loci will be used to determine regional patterns of population connectivity to inform effective marine spatial planning and ecosystem-based fisheries management.

  11. Isolation and characterization of microsatellite loci in Alasmidonta heterodon (Bivalvia: Unionidae)

    USGS Publications Warehouse

    Shaw, K.M.; King, T.L.; Lellis, W.A.; Eackles, M.S.

    2006-01-01

    We developed 13 species-specific microsatellite markers for the federally endangered Atlantic slope unionid Alasmidonta heterodon. Four to 18 alleles per locus were observed among 30 individuals. Observed heterozygosity throughout the loci ranged from 26.9 to 86.2% and averaged 63.6%. Estimates of individual pairwise genetic distances indicated that levels of genetic diversity among loci were sufficient to produce unique multilocus genotypes for all animals surveyed. Randomization tests showed that genotypes for this collection were consistent with Hardy-Weinberg expectations, and no significant linkage disequilibrium was observed between loci. These loci therefore appear suitable for population surveys, kinship assessment and other such applications. ?? 2006 Blackwell Publishing Ltd.

  12. Rapid Microsatellite Isolation from a Butterfly by De Novo Transcriptome Sequencing: Performance and a Comparison with AFLP-Derived Distances

    PubMed Central

    Mikheyev, Alexander S.; Vo, Tanya; Wee, Brian; Singer, Michael C.; Parmesan, Camille

    2010-01-01

    Background The isolation of microsatellite markers remains laborious and expensive. For some taxa, such as Lepidoptera, development of microsatellite markers has been particularly difficult, as many markers appear to be located in repetitive DNA and have nearly identical flanking regions. We attempted to circumvent this problem by bioinformatic mining of microsatellite sequences from a de novo-sequenced transcriptome of a butterfly (Euphydryas editha). Principal Findings By searching the assembled sequence data for perfect microsatellite repeats we found 10 polymorphic loci. Although, like many expressed sequence tag-derived microsatellites, our markers show strong deviations from Hardy-Weinberg equilibrium in many populations, and, in some cases, a high incidence of null alleles, we show that they nonetheless provide measures of population differentiation consistent with those obtained by amplified fragment length polymorphism analysis. Estimates of pairwise population differentiation between 23 populations were concordant between microsatellite-derived data and AFLP analysis of the same samples (r = 0.71, p<0.00001, 425 individuals from 23 populations). Significance De novo transcriptional sequencing appears to be a rapid and cost-effective tool for developing microsatellite markers for difficult genomes. PMID:20585453

  13. Microsatellite markers for the yam bean Pachyrhizus (Fabaceae)1

    PubMed Central

    Delêtre, Marc; Soengas, Beatriz; Utge, José; Lambourdière, Josie; Sørensen, Marten

    2013-01-01

    • Premise of the study: Microsatellite loci were developed for the understudied root crop yam bean (Pachyrhizus spp.) to investigate intraspecific diversity and interspecific relationships within the genus Pachyrhizus. • Methods and Results: Seventeen nuclear simple sequence repeat (SSR) markers with perfect di- and trinucleotide repeats were developed from 454 pyrosequencing of SSR-enriched genomic libraries. Loci were characterized in P. ahipa and wild and cultivated populations of four closely related species. All loci successfully cross-amplified and showed high levels of polymorphism, with number of alleles ranging from three to 12 and expected heterozygosity ranging from 0.095 to 0.831 across the genus. • Conclusions: By enabling rapid assessment of genetic diversity in three native neotropical crops, P. ahipa, P. erosus, and P. tuberosus, and two wild relatives, P. ferrugineus and P. panamensis, these markers will allow exploration of the genetic diversity and evolutionary history of the genus Pachyrhizus. PMID:25202568

  14. RNA toxicity and foci formation in microsatellite expansion diseases.

    PubMed

    Zhang, Nan; Ashizawa, Tetsuo

    2017-02-13

    More than 30 incurable neurological and neuromuscular diseases are caused by simple microsatellite expansions consisted of 3-6 nucleotides. These repeats can occur in non-coding regions and often result in a dominantly inherited disease phenotype that is characteristic of a toxic RNA gain-of-function. The expanded RNA adopts unusual secondary structures, sequesters various RNA binding proteins to form insoluble nuclear foci, and causes cellular defects at a multisystem level. Nuclear foci are dynamic in size, shape and colocalization of RNA binding proteins in different expansion diseases and tissue types. This review sets to provide new insights into the disease mechanisms of RNA toxicity and foci modulation, in light of recent advancement on bi-directional transcription, antisense RNA, repeat-associated non-ATG translation and beyond.

  15. Genome Variability and Gene Content in Chordopoxviruses: Dependence on Microsatellites

    PubMed Central

    Hatcher, Eneida L.; Wang, Chunlin; Lefkowitz, Elliot J.

    2015-01-01

    To investigate gene loss in poxviruses belonging to the Chordopoxvirinae subfamily, we assessed the gene content of representative members of the subfamily, and determined whether individual genes present in each genome were intact, truncated, or fragmented. When nonintact genes were identified, the early stop mutations (ESMs) leading to gene truncation or fragmentation were analyzed. Of all the ESMs present in these poxvirus genomes, over 65% co-localized with microsatellites—simple sequence nucleotide repeats. On average, microsatellites comprise 24% of the nucleotide sequence of these poxvirus genomes. These simple repeats have been shown to exhibit high rates of variation, and represent a target for poxvirus protein variation, gene truncation, and reductive evolution. PMID:25912716

  16. Microsatellite instability in thyroid tumours and tumour-like lesions

    PubMed Central

    Lazzereschi, D; Palmirotta, R; Ranieri, A; Ottini, L; Verì, M C; Cama, A; Cetta, F; Nardi, F; Colletta, G; Mariani-Costantini, R

    1999-01-01

    Fifty-one thyroid tumours and tumour-like lesions were analysed for instability at ten dinucleotide microsatellite loci and at two coding mononucleotide repeats within the transforming growth factor β (TGF-β) type II receptor (TβRII) and insulin-like growth factor II (IGF-II) receptor (IGFIIR) genes respectively. Microsatellite instability (MI) was detected in 11 out of 51 cases (21.5%), including six (11.7%) with MI at one or two loci and five (9.8%) with Ml at three or more loci (RER+ phenotype). No mutations in the TβRII and IGFIIR repeats were observed. The overall frequency of MI did not significantly vary in relation to age, gender, benign versus malignant status and tumour size. However, widespread MI was significantly more frequent in follicular adenomas and carcinomas than in papillary and Hürthle cell tumours: three out of nine tumours of follicular type (33.3%) resulted in replication error positive (RER+), versus 1 out of 29 papillary carcinomas (3.4%, P = 0.01), and zero out of eight Hürthle cell neoplasms. Regional lymph node metastases were present in five MI-negative primary cancers and resulted in MI-positive in two cases. © 1999 Cancer Research Campaign PMID:9888478

  17. Microsatellite primers for red drum (Sciaenops ocellatus)

    Technology Transfer Automated Retrieval System (TEKTRAN)

    In this note, we document polymerase-chain-reaction (PCR) primer pairs for 101, nuclear-encoded microsatellites designed and developed from a red drum (Sciaenops ocellatus) genomic library. The 101 microsatellites (Genbank Accession Numbers EU015882-EU015982) were amplified successfully and used to...

  18. Genetic variation at microsatellite loci in the tropical herb Aphelandra aurantiaca (Acanthaceae)1

    PubMed Central

    Suárez-Montes, Pilar; Tapia-López, Rosalinda; Núñez-Farfán, Juan

    2015-01-01

    Premise of the study: To assess the effect of forest fragmentation on genetic variation and population structure of Aphelandra aurantiaca (Acanthaceae), a tropical and ornamental herbaceous perennial plant, we developed the first microsatellite primers for the species. Methods and Results: Fourteen microsatellite markers were isolated and characterized from A. aurantiaca genomic libraries enriched for di-, tri-, and tetranucleotide repeat motifs. Polymorphism was evaluated in 107 individuals from four natural populations. Twelve out of 14 genetic markers were polymorphic. The number of alleles per locus ranged from two to 12, and the observed and expected heterozygosities ranged from 0.22 to 0.96 and from 0.20 to 0.87, respectively. Fixation indices ranged from −0.41 to 0.44. Conclusions: These newly developed microsatellite markers for A. aurantiaca will be useful for future population genetic studies, specifically to detect the possible loss of genetic diversity due to habitat fragmentation. PMID:26649265

  19. Fourteen polymorphic microsatellite markers for the threatened Arnica montana (Asteraceae)1

    PubMed Central

    Duwe, Virginia K.; Ismail, Sascha A.; Buser, Andres; Sossai, Esther; Borsch, Thomas; Muller, Ludo A. H.

    2015-01-01

    • Premise of the study: Microsatellite markers were developed to investigate population genetic structure in the threatened species Arnica montana. • Methods and Results: Fourteen microsatellite markers with di-, tetra-, and hexanucleotide repeat motifs were developed for A. montana using 454 pyrosequencing without and with library-enrichment methods, resulting in 56,545 sequence reads and 14,467 sequence reads, respectively. All loci showed a high level of polymorphism, with allele numbers ranging from four to 11 in five individuals from five populations (25 samples) and an expected heterozygosity ranging from 0.192 to 0.648 across the loci. • Conclusions: This set of microsatellite markers is the first one described for A. montana and will facilitate conservation genetic applications as well as the understanding of phylogeographic patterns in this species. PMID:25606354

  20. Isolation and characterization of 12 microsatellite loci in soapbark, Quillaja saponaria (Quillajaceae)1

    PubMed Central

    Letelier, Luis; Harvey, Nick; Valderrama, Aly; Stoll, Alexandra; González-Rodríguez, Antonio

    2015-01-01

    Premise of the study: Microsatellite primers were developed for the endemic Chilean tree Quillaja saponaria (Quillajaceae), a common member of the sclerophyllous Mediterranean forest, to investigate intraspecific patterns of genetic diversity and structure. Methods and Results: Using an enriched library, 12 polymorphic microsatellite loci were developed in Q. saponaria. All loci consisted of dinucleotide repeats. The average number of alleles per locus was 5.3 (2–13), with a total of 64 alleles recorded in 39 individuals from three populations. Conclusions: The microsatellite markers described here are the first characterized for Q. saponaria. The polymorphic loci will be useful in studies of genetic diversity and genetic population differentiation in natural populations of this species. PMID:25995980

  1. Isolation and Characterization of Sixteen Polymorphic Microsatellite Loci in the Golden Apple Snail Pomacea canaliculata

    PubMed Central

    Chen, Lian; Xu, Haigen; Li, Hong; Wu, Jun; Ding, Hui; Liu, Yan

    2011-01-01

    We report the characterization of 16 polymorphic microsatellite markers in the golden apple snail, Pomacea canaliculata, a pest registered in the list of “100 of the world’s worst invasive alien species”. The fast isolation by AFLP (Amplified Fragment Length Polymorphism) of sequences containing repeats (FIASCO) method was used to isolate microsatellite loci, and polymorphism was explored with 29 individuals collected in an invasive region from China. These primers showed a number of alleles per locus ranging from three to 13. The ranges of observed and expected heterozygosity were 0.310–0.966 and 0.523–0.898, respectively. These microsatellite markers described here will be useful for population genetic studies of P. canaliculata. PMID:22016640

  2. Characterization of nuclear microsatellite markers for Rumex bucephalophorus (Polygonaceae) using 454 sequencing1

    PubMed Central

    Viruel, Juan; Ortiz, Pedro L.; Arista, Montserrat; Talavera, María

    2015-01-01

    Premise of the study: Nuclear microsatellite markers were developed in Rumex bucephalophorus subsp. canariensis (Polygonaceae) to investigate its genetic diversity and structure. Methods and Results: Sixteen polymorphic microsatellite markers were obtained using 454 next-generation sequencing with di-, tri-, and tetranucleotide repeats. The average number of alleles was 5.688 and 3.813 for R. bucephalophorus subsp. canariensis var. canariensis and var. fruticescens, respectively. Slightly higher levels of mean genetic diversity were found in var. canariensis (expected heterozygosity = 0.600) than in var. fruticescens (expected heterozygosity = 0.514). Cross-amplifications in related taxa within R. bucephalophorus showed good amplification and polymorphic patterns. Conclusions: These 16 novel nuclear microsatellite markers are the first in the genus Rumex and may serve as valuable tools to carry out studies on genetic diversity and structure as well as progeny studies. PMID:26697279

  3. Isolation and characterization of sixteen polymorphic microsatellite loci in the golden apple snail Pomacea canaliculata.

    PubMed

    Chen, Lian; Xu, Haigen; Li, Hong; Wu, Jun; Ding, Hui; Liu, Yan

    2011-01-01

    We report the characterization of 16 polymorphic microsatellite markers in the golden apple snail, Pomacea canaliculata, a pest registered in the list of "100 of the world's worst invasive alien species". The fast isolation by AFLP (Amplified Fragment Length Polymorphism) of sequences containing repeats (FIASCO) method was used to isolate microsatellite loci, and polymorphism was explored with 29 individuals collected in an invasive region from China. These primers showed a number of alleles per locus ranging from three to 13. The ranges of observed and expected heterozygosity were 0.310-0.966 and 0.523-0.898, respectively. These microsatellite markers described here will be useful for population genetic studies of P. canaliculata.

  4. Development and characterization of nine microsatellites for an endangered tree, Pinus wangii (Pinaceae)1

    PubMed Central

    Dou, Jing-Jing; Zhou, Ren-Chao; Tang, An-Jun; Ge, Xue-Jun; Wu, Wei

    2013-01-01

    • Premise of the study: Pinus wangii is an endemic and endangered species in southwestern China, and microsatellite primers were developed to characterize its genetic diversity and population structure. • Methods and Results: Using the Fast Isolation by AFLP of Sequences COntaining repeats (FIASCO) protocol, nine sets of microsatellite primers were developed in P. wangii. One population with 26 individuals of P. wangii, as well as 11 individuals each for two congeneric species, P. taiwanensis and P. squamata, were used to test their polymorphism and transferability. The number of alleles per locus ranged from one to seven with an average of 3.7, and the observed heterozygosity and expected heterozygosity ranged from 0 to 0.91 and 0 to 0.75, respectively. • Conclusions: We developed nine sets of polymorphic microsatellite loci that are suitable for investigating genetic diversity and population structure of P. wangii, and these markers may be useful for other Pinus species. PMID:25202511

  5. Infrequent microsatellite instability in oesophageal cancers.

    PubMed Central

    Muzeau, F.; Fléjou, J. F.; Belghiti, J.; Thomas, G.; Hamelin, R.

    1997-01-01

    Alterations of microsatellites have been found at relatively high frequency in hereditary and sporadic colorectal cancer and gastric and pancreatic cancers and at lower frequency in some other cancers. We determined the frequency of instability at 39 poly-CA microsatellite loci in 20 squamous cell carcinomas and 26 Barrett's adenocarcinomas of the oesophagus. None of the tumours presented instability for a high percentage of the tested loci. Four squamous cell carcinomas and six Barrett's adenocarcinomas showed microsatellite instability at one locus, and three Barrett's adenocarcinomas showed microsatellite instability at two loci. The presence of few loci showing microsatellite instability could be due to an instability background. We conclude that genetic defects in the DNA mismatch repair system do not play an important role in oesophageal cancers. Images Figure 1 PMID:9155055

  6. Isolation and characterization of microsatellite loci in the intertidal sponge Halichondria panicea

    USGS Publications Warehouse

    Knowlton, A.L.; Pierson, Barbara J.; Talbot, S.L.; Highsmith, R.C.

    2003-01-01

    GA- and CA-enriched genomic libraries were constructed for the intertidal sponge Halichondria panicea. Unique repeat motifs identified varied from the expected simple dinucleotide repeats to more complex repeat units. All sequences tended to be highly repetitive but did not necessarily contain the targeted motifs. Seven microsatellite loci were evaluated on sponges from the clone source population. All seven were polymorphic with 5.43??0.92 mean number of alleles. Six of the seven loci that could be resolved had mean heterozygosities of 0.14-0.68. The loci identified here will be useful for population studies.

  7. The number of alleles at a microsatellite defines the allele frequency spectrum and facilitates fast accurate estimation of theta.

    PubMed

    Haasl, Ryan J; Payseur, Bret A

    2010-12-01

    Theoretical work focused on microsatellite variation has produced a number of important results, including the expected distribution of repeat sizes and the expected squared difference in repeat size between two randomly selected samples. However, closed-form expressions for the sampling distribution and frequency spectrum of microsatellite variation have not been identified. Here, we use coalescent simulations of the stepwise mutation model to develop gamma and exponential approximations of the microsatellite allele frequency spectrum, a distribution central to the description of microsatellite variation across the genome. For both approximations, the parameter of biological relevance is the number of alleles at a locus, which we express as a function of θ, the population-scaled mutation rate, based on simulated data. Discovered relationships between θ, the number of alleles, and the frequency spectrum support the development of three new estimators of microsatellite θ. The three estimators exhibit roughly similar mean squared errors (MSEs) and all are biased. However, across a broad range of sample sizes and θ values, the MSEs of these estimators are frequently lower than all other estimators tested. The new estimators are also reasonably robust to mutation that includes step sizes greater than one. Finally, our approximation to the microsatellite allele frequency spectrum provides a null distribution of microsatellite variation. In this context, a preliminary analysis of the effects of demographic change on the frequency spectrum is performed. We suggest that simulations of the microsatellite frequency spectrum under evolutionary scenarios of interest may guide investigators to the use of relevant and sometimes novel summary statistics.

  8. High-throughput microsatellite isolation through 454 GS-FLX Titanium pyrosequencing of enriched DNA libraries.

    PubMed

    Malausa, Thibaut; Gilles, André; Meglécz, Emese; Blanquart, Hélène; Duthoy, Stéphanie; Costedoat, Caroline; Dubut, Vincent; Pech, Nicolas; Castagnone-Sereno, Philippe; Délye, Christophe; Feau, Nicolas; Frey, Pascal; Gauthier, Philippe; Guillemaud, Thomas; Hazard, Laurent; Le Corre, Valérie; Lung-Escarmant, Brigitte; Malé, Pierre-Jean G; Ferreira, Stéphanie; Martin, Jean-François

    2011-07-01

    Microsatellites (or SSRs: simple sequence repeats) are among the most frequently used DNA markers in many areas of research. The use of microsatellite markers is limited by the difficulties involved in their de novo isolation from species for which no genomic resources are available. We describe here a high-throughput method for isolating microsatellite markers based on coupling multiplex microsatellite enrichment and next-generation sequencing on 454 GS-FLX Titanium platforms. The procedure was calibrated on a model species (Apis mellifera) and validated on 13 other species from various taxonomic groups (animals, plants and fungi), including taxa for which severe difficulties were previously encountered using traditional methods. We obtained from 11,497 to 34,483 sequences depending on the species and the number of detected microsatellite loci ranged from 199 to 5791. We thus demonstrated that this procedure can be readily and successfully applied to a large variety of taxonomic groups, at much lower cost than would have been possible with traditional protocols. This method is expected to speed up the acquisition of high-quality genetic markers for nonmodel organisms.

  9. Morphological and microsatellite diversity associated with ecological factors in natural populations of Medicago laciniata Mill. (Fabaceae).

    PubMed

    Badri, Mounawer; Zitoun, Adel; Ilahi, Houcine; Huguet, Thierry; Aouani, Mohamed Elarbi

    2008-12-01

    Genetic variability in 10 natural Tunisian populations of Medicago laciniata were analysed using 19 quantitative traits and 12 polymorphic microsatellite loci. A large degree of genetic variability within-populations and among-populations was detected for both quantitative characters and molecular markers. High genetic differentiation among populations for quantitative traits was seen, with Q(ST) = 0.47, and F(ST) = 0.47 for microsatellite markers. Several quantitative traits displayed no statistical difference in the levels of Q(ST) and F(ST). Further, significant correlations between quantitative traits and eco-geographical factors suggest that divergence in the traits among populations may track environmental differences. There was no significant correlation between genetic variability at quantitative traits and microsatellite markers within populations. The site-of-origin of eco-geographical factors explain between 18.13% and 23.40% of genetic variance among populations at quantitative traits and microsatellite markers, respectively. The environmental factors that most influence variation in measured traits among populations are assimilated phosphorus (P(2)0(5)) and mean annual rainfall, followed by climate and soil texture, altitude and organic matter. Significant associations between eco-geographical factors and gene diversity, He, were established in five microsatellite loci suggesting that these simple sequence repeats (SSRs) are not necessarily biologically neutral.

  10. FANCJ is essential to maintain microsatellite structure genome-wide during replication stress

    PubMed Central

    Barthelemy, Joanna; Hanenberg, Helmut; Leffak, Michael

    2016-01-01

    Microsatellite DNAs that form non-B structures are implicated in replication fork stalling, DNA double strand breaks (DSBs) and human disease. Fanconi anemia (FA) is an inherited disorder in which mutations in at least nineteen genes are responsible for the phenotypes of genome instability and cancer predisposition. FA pathway proteins are active in the resolution of non-B DNA structures including interstrand crosslinks, G quadruplexes and DNA triplexes. In FANCJ helicase depleted cells, we show that hydroxyurea or aphidicolin treatment leads to loss of microsatellite polymerase chain reaction signals and to chromosome recombination at an ectopic hairpin forming CTG/CAG repeat in the HeLa genome. Moreover, diverse endogenous microsatellite signals were also lost upon replication stress after FANCJ depletion, and in FANCJ null patient cells. The phenotype of microsatellite signal instability is specific for FANCJ apart from the intact FA pathway, and is consistent with DSBs at microsatellites genome-wide in FANCJ depleted cells following replication stress. PMID:27179029

  11. Development of microsatellite markers in Cratylia mollis and their transferability to C. argentea (Fabaceae)1

    PubMed Central

    López-Roberts, M. Cristina; de Queiroz, Luciano Paganucci; van den Berg, Cássio

    2013-01-01

    • Premise of the study: This work aimed to develop microsatellite markers for Cratylia mollis as tools to assess its genetic diversity and structure and to evaluate their potential cross-amplification in related species. • Methods and Results: Microsatellite markers were developed using a microsatellite-enriched library and an intersimple sequence repeat library. From a set of 19 markers, 12 microsatellite loci were polymorphic and presented considerable variation in allele number (2–11), expected heterozygosity (0.226–0.883), and polymorphism information content per locus (0.212–0.870). Cross-amplification in C. argentea was successful in 16 loci, 12 of which were polymorphic (2–10 alleles). • Conclusions: The polymorphism of this set of microsatellite markers for C. mollis, as well as their successful cross-amplification in C. intermedia and C. bahiensis and their transferability to C. argentea, supports their use in future comparative studies to understand the mechanism involved in population divergence and speciation in the genus. PMID:25202484

  12. Bulk development and stringent selection of microsatellite markers in the western flower thrips Frankliniella occidentalis

    PubMed Central

    Cao, Li-Jun; Li, Ze-Min; Wang, Ze-Hua; Zhu, Liang; Gong, Ya-Jun; Chen, Min; Wei, Shu-Jun

    2016-01-01

    Recent improvements in next-generation sequencing technologies have enabled investigation of microsatellites on a genome-wide scale. Faced with a huge amount of candidates, the use of appropriate marker selection criteria is crucial. Here, we used the western flower thrips Frankliniella occidentalis for an empirical microsatellite survey and validation; 132,251 candidate microsatellites were identified, 92,102 of which were perfect. Dinucleotides were the most abundant category, while (AG)n was the most abundant motif. Sixty primer pairs were designed and validated in two natural populations, of which 30 loci were polymorphic, stable, and repeatable, but not all in Hardy–Weinberg equilibrium (HWE) and linkage equilibrium. Four marker panels were constructed to understand effect of marker selection on population genetic analyses: (i) only accept loci with single nucleotide insertions (SNI); (ii) only accept the most polymorphic loci (MP); (iii) only accept loci that did not deviate from HWE, did not show SNIs, and had unambiguous peaks (SS) and (iv) all developed markers (ALL). Although the MP panel resulted in microsatellites of highest genetic diversity followed by the SNI, the SS performed best in individual assignment. Our study proposes stringent criteria for selection of microsatellites from a large-scale number of genomic candidates for population genetic studies. PMID:27197749

  13. Radiation hybrid mapping of 304 novel microsatellites in the domestic cat genome.

    PubMed

    Menotti-Raymond, M; David, V A; Agarwala, R; Schäffer, A A; Stephens, R; O'Brien, S J; Murphy, W J

    2003-01-01

    Effective utilization of the domestic cat as an animal model for hereditary and infectious disease requires the development and implementation of high quality gene maps incorporating microsatellites and conserved coding gene markers. Previous feline linkage and radiation hybrid maps have lacked sufficient microsatellite coverage on all chromosomes to make effective use of full genome scans. Here we report the isolation and genomic mapping of 304 novel polymorphic repeat loci in the feline genome. The new loci were mapped in the domestic cat radiation hybrid panel using an automated fluorescent TAQ-Man based assay. The addition of these 304 microsatellites brings the total number of microsatellites mapped in the feline genome to 580, and the total number of loci placed onto the RH map to 1,126. Microsatellites now span every autosome with an average spacing of roughly one polymorphic STR every five centimorgans, and full genome coverage of one marker every 2.7 megabases. These loci now provide a useful tool for undertaking full-genome scans to identify genes associated with phenotypes of interest, such as those relating to hereditary disease, coat color, patterning and morphology. These resources can also be extended to the remaining 36 species of the cat family for population genetic and evolutionary genomic analyses.

  14. GGGGCC microsatellite RNA is neuritically localized, induces branching defects, and perturbs transport granule function.

    PubMed

    Burguete, Alondra Schweizer; Almeida, Sandra; Gao, Fen-Biao; Kalb, Robert; Akins, Michael R; Bonini, Nancy M

    2015-12-09

    Microsatellite expansions are the leading cause of numerous neurodegenerative disorders. Here we demonstrate that GGGGCC and CAG microsatellite repeat RNAs associated with C9orf72 in amyotrophic lateral sclerosis/frontotemporal dementia and with polyglutamine diseases, respectively, localize to neuritic granules that undergo active transport into distal neuritic segments. In cultured mammalian spinal cord neurons, the presence of neuritic GGGGCC repeat RNA correlates with neuronal branching defects, and the repeat RNA localizes to granules that label with fragile X mental retardation protein (FMRP), a transport granule component. Using a Drosophila GGGGCC expansion disease model, we characterize dendritic branching defects that are modulated by FMRP and Orb2. The human orthologs of these modifiers are misregulated in induced pluripotent stem cell-differentiated neurons (iPSNs) from GGGGCC expansion carriers. These data suggest that expanded repeat RNAs interact with the messenger RNA transport and translation machinery, causing transport granule dysfunction. This could be a novel mechanism contributing to the neuronal defects associated with C9orf72 and other microsatellite expansion diseases.

  15. GGGGCC microsatellite RNA is neuritically localized, induces branching defects, and perturbs transport granule function

    PubMed Central

    Burguete, Alondra Schweizer; Almeida, Sandra; Gao, Fen-Biao; Kalb, Robert; Akins, Michael R; Bonini, Nancy M

    2015-01-01

    Microsatellite expansions are the leading cause of numerous neurodegenerative disorders. Here we demonstrate that GGGGCC and CAG microsatellite repeat RNAs associated with C9orf72 in amyotrophic lateral sclerosis/frontotemporal dementia and with polyglutamine diseases, respectively, localize to neuritic granules that undergo active transport into distal neuritic segments. In cultured mammalian spinal cord neurons, the presence of neuritic GGGGCC repeat RNA correlates with neuronal branching defects, and the repeat RNA localizes to granules that label with fragile X mental retardation protein (FMRP), a transport granule component. Using a Drosophila GGGGCC expansion disease model, we characterize dendritic branching defects that are modulated by FMRP and Orb2. The human orthologs of these modifiers are misregulated in induced pluripotent stem cell-differentiated neurons (iPSNs) from GGGGCC expansion carriers. These data suggest that expanded repeat RNAs interact with the messenger RNA transport and translation machinery, causing transport granule dysfunction. This could be a novel mechanism contributing to the neuronal defects associated with C9orf72 and other microsatellite expansion diseases. DOI: http://dx.doi.org/10.7554/eLife.08881.001 PMID:26650351

  16. Passive cryocooler for microsatellite payload

    NASA Astrophysics Data System (ADS)

    Mullins, Mayes; Thomas, Paul J.; Harron, John W.; Duggan, Philip; Sinclair, Peter M.; Khanna, Shyam M.

    1998-11-01

    A passive cryocooler has been developed for the cooling of small payloads to temperatures as low as 145 K. Although designed for a specific electronics experiment on the STRV-1d microsatellite, the device is suitable for a wide range of applications. The cryocooler uses coated surfaces for tailored radiative cooling. Mechanical support between components is provided by fiberglass struts. The measured end temperature reached is 151 K in a liquid nitrogen dewar which extrapolates to an end temperature of lower than 145 K in space. Thermal vacuum testing and random vibration testing at levels consistent with an Ariane 5 launch have been performed as part of formal qualification for the STRV mission. In this paper, details of the design, analysis, fabrication and testing of the passive cryocooler are presented.

  17. Microsatellite instability in prostate cancer

    SciTech Connect

    Shan, A.L.; Wick, M.J.; Persons, D.L.

    1994-09-01

    Microsatellite instability (MIN) has been documented in hereditary nonpolyposis colorectal cancer (HNPCC) as well as in sporadic forms of human cancers. Two of the genes which appear to be responsible for this particular tumor phenotype, hMSH2 and hMLH1, have now been identified. To determine the potential role of these mutator genes in prostate cancer, we have examined 95 prostate adenocarcinomas (40 paraffin embedded and 55 fresh frozen) for the presence of genetic instability at four microsatellite markers. The markers are localized to chromosome arms 5q(APC-CA1), 8p(Mfd 210Z), 15q(635/636), and 17q(p53-CA). Patients from whom paraffin embedded material was obtained were divided into short term (<3 years, n=18), and long term (>3 years, n=22) survivors. Of the 95 tumors examined, only four tumors (4%) demonstrated MIN: two tumors demonstrated MIN at 3 loci (p53-CA, APC-CA1, 635/636), one tumor demonstrated MIN at 2 loci (APC-CA1 and 635/636), and one tumor demonstrated instability at 635/636 only. All tumors exhibiting MIN had Gleason scores of {ge} 4+4. A correlation between MIN and survival was not observed. Information on family history was limited. However, of the two patients demonstrating MIN at three loci, one patient was diagnosed with a second malignancy (TCC of the ureter), but otherwise had a negative family history, while the second patient had one first degree relative with esophageal cancer. The patient demonstrating MIN at two loci had a negative family history, while the remaining patient had two first degree relatives with cancer (prostate and stomach). These results suggest that hMSH2 and hMLH1 (as reflected by the small percentage of tumors displaying MIN) do not play a prominent role in the process of prostate tumorigenesis.

  18. Tandem repeat distribution of gene transcripts in three plant families

    PubMed Central

    2009-01-01

    Tandem repeats (microsatellites or SSRs) are molecular markers with great potential for plant genetic studies. Modern strategies include the transfer of these markers among widely studied and orphan species. In silico analyses allow for studying distribution patterns of microsatellites and predicting which motifs would be more amenable to interspecies transfer. Transcribed sequences (Unigene) from ten species of three plant families were surveyed for the occurrence of micro and minisatellites. Transcripts from different species displayed different rates of tandem repeat occurrence, ranging from 1.47% to 11.28%. Both similar and different patterns were found within and among plant families. The results also indicate a lack of association between genome size and tandem repeat fractions in expressed regions. The conservation of motifs among species and its implication on genome evolution and dynamics are discussed. PMID:21637460

  19. Development, inheritance, and linkage-group assignment of 60 novel microsatellite markers for the gray, short-tailed opossum Monodelphis domestica.

    PubMed

    Gouin, Nicolas; Westenberger, Scott J; Mahaney, Susan M; Lindley, Peter; VandeBerg, John L; Samollow, Paul B

    2005-12-01

    Short-tandem-repeat (SSR) or microsatellite polymorphisms are some of the most extensively employed genetic markers in contemporary linkage mapping studies. To date, only a limited number of microsatellites have been isolated in the gray, short-tailed opossum Monodelphis domestica, a South American marsupial widely used for comparative biological and biomedical research. To increase the number of potentially useful mapping markers, we screened 2 microsatellite-enriched genomic libraries containing alternatively (CA)n or (GA)n repeats. A total of 184 clones were sequenced, from which 60 polymorphic microsatellite markers were successfully optimized. The efficiency of this enrichment protocol for M. domestica microsatellite isolation is discussed, and suggestions to improve the outcome are made. All 60 loci showed high allelic diversity, with allele numbers ranging from 2 to 10 in a subset of 33 unrelated animals. Normal Mendelian inheritance was confirmed for all loci by analyzing allelic segregation in 5 two-generation families. One microsatellite appeared to be X linked, and null alleles were found in 5 others. Two-point linkage analyses were implemented using the data on the 5 families, leading to the assignment of 59 of these loci to the existing linkage groups. The 60 novel microsatellites developed in this study will contribute significantly to the M. domestica linkage map, and further QTL mapping studies.

  20. Population genetic structure of the tropical moss Acanthorrhynchium papillatum as measured with microsatellite markers.

    PubMed

    Leonardía, A A P; Tan, B C; Kumar, P P

    2013-03-01

    Mosses and other bryophytes are vital components of forests, because they sustain a tremendous diversity of invertebrates and influence significant ecological functions. There have been few studies on moss population diversity in Southeast Asia, despite the escalating deforestation in this region of rich biodiversity. The genetic diversity of the tropical moss Acanthorrhynchium papillatum (Harv.) Fleisch., collected from forested areas in Singapore and Peninsular Malaysia, was elucidated using eight microsatellite markers developed for this species. Significant levels of allelic and haplotypic diversity were observed among clumps of the moss. Differences in allelic richness and genotypic diversity among the populations were higher in less disturbed forests compared to the more disturbed areas, suggesting that genetic diversity is affected by habitat quality. Genetic diversity levels within the clumps studied were low, indicating that vegetative reproduction was more important within clumps than sexual reproduction. However, multilocus genotypes of samples within the clumps studied were not all alike, providing evidence of microsatellite mutation or of occasional sexuality. Despite the isolation of populations, A. papillatum can introduce genetic variability by mutation among vegetatively propagated individuals. This study provides baseline information on the genetic diversity of A. papillatum tropical rain forests.

  1. Colonization of Ireland: revisiting 'the pygmy shrew syndrome' using mitochondrial, Y chromosomal and microsatellite markers.

    PubMed

    McDevitt, A D; Vega, R; Rambau, R V; Yannic, G; Herman, J S; Hayden, T J; Searle, J B

    2011-12-01

    There is great uncertainty about how Ireland attained its current fauna and flora. Long-distance human-mediated colonization from southwestern Europe has been seen as a possible way that Ireland obtained many of its species; however, Britain has (surprisingly) been neglected as a source area for Ireland. The pygmy shrew has long been considered an illustrative model species, such that the uncertainty of the Irish colonization process has been dubbed 'the pygmy shrew syndrome'. Here, we used new genetic data consisting of 218 cytochrome (cyt) b sequences, 153 control region sequences, 17 Y-intron sequences and 335 microsatellite multilocus genotypes to distinguish between four possible hypotheses for the colonization of the British Isles, formulated in the context of previously published data. Cyt b sequences from western Europe were basal to those found in Ireland, but also to those found in the periphery of Britain and several offshore islands. Although the central cyt b haplotype in Ireland was found in northern Spain, we argue that it most likely occurred in Britain also, from where the pygmy shrew colonized Ireland as a human introduction during the Holocene. Y-intron and microsatellite data are consistent with this hypothesis, and the biological traits and distributional data of pygmy shrews argue against long-distance colonization from Spain. The compact starburst of the Irish cyt b expansion and the low genetic diversity across all markers strongly suggests a recent colonization. This detailed molecular study of the pygmy shrew provides a new perspective on an old colonization question.

  2. [Genetic diversity of microsatellite loci in captive Amur tigers].

    PubMed

    Zhang, Yu-Gaung; Li, Di-Qiang; Xiao, Qi-Ming; Rao, Li-Qun; Zhang, Xue-Wen

    2004-09-01

    microsatellie loci had high or medium polymorphism in these Amur tigers and had high genetic diversity. At the same time, we only found even bases variability which showed the even bases repeat sequence (CA/GT) maybe the basic unit for length variability of microsatellite in all loci. In this study, the samples were made up of 75 hair specimens, 23 blood specimens and 15 tissue specimens, we obtained the genome DNA from hairs using the non-invasive DNA technology and demonstrated that DNA derived from hair samples is as good as that obtained from blood samples for the analysis of microsatellite polymorphism. These results imply that microsatellite DNA markers and non-invasive DNA technology can help study the genetic diversity of Amur tiger. This method could be used in the captive management of other endangered species.

  3. Multilocus heterozygosity, parental relatedness and individual fitness components in a wild mountain goat, Oreamnos americanus population.

    PubMed

    Mainguy, Julien; Côté, Steeve D; Coltman, David W

    2009-05-01

    Matings between relatives lead to a decrease in offspring genetic diversity which can reduce fitness, a phenomenon known as inbreeding depression. Because alpine ungulates generally live in small structured populations and often exhibit a polygynous mating system, they are susceptible to inbreeding. Here, we used marker-based measures of pairwise genetic relatedness and inbreeding to investigate the fitness consequences of matings between relatives in a long-term study population of mountain goats (Oreamnos americanus) at Caw Ridge, Alberta, Canada. We first assessed whether individuals avoided mating with kin by comparing actual and random mating pairs according to their estimated genetic relatedness, which was derived from 25 unlinked polymorphic microsatellite markers and reflected pedigree relatedness. We then examined whether individual multilocus heterozygosity H, used as a measure of inbreeding, was predicted by parental relatedness and associated with yearling survival and the annual probability of giving birth to a kid in adult females. Breeding pairs identified by genetic parentage analyses of offspring that survived to 1 year of age were less genetically related than expected under random matings. Parental relatedness was negatively correlated with offspring H, and more heterozygous yearlings had higher survival to 2 years of age. The probability of giving birth was not affected by H in adult females. Because kids that survived to yearling age were mainly produced by less genetically related parents, our results suggest that some individuals experienced inbreeding depression in early life. Future research will be required to quantify the levels of gene flow between different herds, and evaluate their effects on population genetic diversity and dynamics.

  4. Genetic evidence that both dNTP-stabilized and strand slippage mechanisms may dictate DNA polymerase errors within mononucleotide microsatellites.

    PubMed

    Baptiste, Beverly A; Jacob, Kimberly D; Eckert, Kristin A

    2015-05-01

    Mononucleotide microsatellites are tandem repeats of a single base pair, abundant within coding exons and frequent sites of mutation in the human genome. Because the repeated unit is one base pair, multiple mechanisms of insertion/deletion (indel) mutagenesis are possible, including strand-slippage, dNTP-stabilized, and misincorportion-misalignment. Here, we examine the effects of polymerase identity (mammalian Pols α, β, κ, and η), template sequence, dNTP pool size, and reaction temperature on indel errors during in vitro synthesis of mononucleotide microsatellites. We utilized the ratio of insertion to deletion errors as a genetic indicator of mechanism. Strikingly, we observed a statistically significant bias toward deletion errors within mononucleotide repeats for the majority of the 28 DNA template and polymerase combinations examined, with notable exceptions based on sequence and polymerase identity. Using mutator forms of Pol β did not substantially alter the error specificity, suggesting that mispairing-misalignment mechanism is not a primary mechanism. Based on our results for mammalian DNA polymerases representing three structurally distinct families, we suggest that dNTP-stabilized mutagenesis may be an alternative mechanism for mononucleotide microsatellite indel mutation. The change from a predominantly dNTP-stabilized mechanism to a strand-slippage mechanism with increasing microsatellite length may account for the differential rates of tandem repeat mutation that are observed genome-wide.

  5. A multilocus sequence typing scheme for Streptococcus pneumoniae: identification of clones associated with serious invasive disease.

    PubMed

    Enright, M C; Spratt, B G

    1998-11-01

    The population biology of Streptococcus pneumoniae is poorly understood. Most of the important issues could be addressed by the molecular characterization of large, well sampled populations from carriage and from the different manifestations of pneumococcal disease. The authors have therefore developed a pneumococcal multilocus sequence typing scheme and database by sequencing approximately 450 bp fragments of seven housekeeping loci from 295 isolates. The combination of alleles at the seven loci provided an allelic profile, or sequence type (ST), and the relatedness between isolates was obtained by constructing a dendrogram from the matrix of pairwise differences between STs. The typing scheme was validated using pneumococci of known genetic relatedness and could resolve >6 billion STs. Among 274 isolates from recent cases of invasive pneumococcal disease in eight countries, 143 STs were resolved, but 12 STs contained at least five isolates (range 5-21 isolates). The repeated recovery of indistinguishable isolates from invasive disease in different countries implies that these STs define strains with an increased capacity to cause invasive disease. The relationship between STs and serotypes suggested that, in the longer term, capsular genes have been distributed horizontally within the pneumococcal population, but in the short term, expansion of clones occurs with only occasional changes of serotype. The multilocus sequence typing scheme provides a powerful new approach to the characterization of pneumococci, since it provides molecular typing data that are electronically portable between laboratories, and which can be used to probe aspects of the population and evolutionary biology of these organisms. A Web site for the molecular characterization of pneumococci by MLST is available (http ://mlst.zoo.ox.ac.uk).

  6. Characterization of 10 new nuclear microsatellite markers in Acca sellowiana (Myrtaceae)1

    PubMed Central

    Klabunde, Gustavo H. F.; Olkoski, Denise; Vilperte, Vinicius; Zucchi, Maria I.; Nodari, Rubens O.

    2014-01-01

    • Premise of the study: Microsatellite primers were identified and characterized in Acca sellowiana in order to expand the limited number of pre-existing polymorphic markers for use in population genetic studies for conservation, phylogeography, breeding, and domestication. • Methods and Results: A total of 10 polymorphic microsatellite primers were designed from clones obtained from a simple sequence repeat (SSR)–enriched genomic library. The primers amplified di- and trinucleotide repeats with four to 27 alleles per locus. In all tested populations, the observed heterozygosity ranged from 0.269 to 1.0. • Conclusions: These new polymorphic SSR markers will allow future genetic studies to be denser, either for genetic structure characterization of natural populations or for studies involving genetic breeding and domestication process in A. sellowiana. PMID:25202632

  7. Gametogenesis processes and multilocus gene identity by descent.

    PubMed Central

    Guo, S. W.

    1996-01-01

    With few exceptions, the determination of unconditional probability of genes shared identical by descent (IBD) by relatives can be very difficult, especially if the relationship is complex or if multiple loci are involved. It is particularly difficult if one needs the IBD probability in a explicit form, expressed in terms of interlocus recombination fractions. In this paper, I will further extend the concept of gametogenesis process introduced elsewhere and indicate that it completely determines the gene IBD events of interest in pedigrees. I will demonstrate that the gametogenesis process not only serves as a convenient conceptual framework in considering IBD events in pedigrees but also provides a simple yet powerful tool to solve a wide range of seemingly difficult problems. In particular, I consider the problem of multilocus IBD probability for relative pairs, k siblings, and a group of pedigree members. In addition, I consider the problem of multilocus autozygosity probability and the problem of gene preservation in close relatives. PMID:8571968

  8. Likelihoods of multilocus DNA fingerprints in extended families.

    PubMed Central

    Bockel, B; Nürnberg, P; Krawczak, M

    1992-01-01

    A concept for the application of complex pedigree analysis to multilocus DNA fingerprinting is described. By following this approach, the extent to which the DNA fingerprints of grandparents influence the phenotype likelihoods of their offspring was determined. It was demonstrated by simulation that approximately 90% of paternity disputes can be solved if mother, child, and paternal grandparents, instead of the putative father, are tested. If only phenotype information on a single paternal sib is allowed for, true paternity will be detected with reasonable persuasive power in up to 64% of cases. Exclusion of false paternity remains possible for 40% of cases. Finally, the analysis concept is modified by reducing the number of genotype variations considered in likelihood computations. This time-saving procedure is shown to yield sufficiently accurate likelihoods in the analysis of both simulation data and multilocus DNA fingerprints obtained in two large families. PMID:1496987

  9. Multilocus Sequence Typing (MLST) for Characterization of Enterobacter cloacae

    PubMed Central

    Miyoshi-Akiyama, Tohru; Hayakawa, Kayoko; Ohmagari, Norio; Shimojima, Masahiro; Kirikae, Teruo

    2013-01-01

    Enterobacter cloacae is an important emerging pathogen, which sometime causes respiratory infection, surgical site infection, urinary infection, sepsis, and outbreaks at neonatal units. We have developed a multilocus sequence typing (MLST) scheme utilizing seven housekeeping genes and evaluated the performance in 101 clinical isolates. The MLST scheme yielded 83 sequence types (ST) including 78 novel STs found in the clinical isolates. These findings supported the robustness of the MLST scheme developed in this study. PMID:23776664

  10. The Unstable Repeats - Three Evolving Faces of Neurological Disease

    PubMed Central

    Nelson, David L.; Orr, Harry T.; Warren, Stephen T.

    2013-01-01

    Disorders characterized by expansion of an unstable nucleotide repeat account for a number of inherited neurological diseases. Here, we review examples of unstable repeat disorders that nicely illustrate the three of the major pathogenic mechanisms associated with these diseases: loss-of-function typically by disrupting transcription of the mutated gene, RNA toxic gain-of-function, and protein toxic gain-of-function. In addition to providing insight into the mechanisms underlying these devastating neurological disorders, the study of these unstable microsatellite repeat disorders has provided insight into very basic aspects of neuroscience. PMID:23473314

  11. Microsatellites as targets of natural selection.

    PubMed

    Haasl, Ryan J; Payseur, Bret A

    2013-02-01

    The ability to survey polymorphism on a genomic scale has enabled genome-wide scans for the targets of natural selection. Theory that connects patterns of genetic variation to evidence of natural selection most often assumes a diallelic locus and no recurrent mutation. Although these assumptions are suitable to selection that targets single nucleotide variants, fundamentally different types of mutation generate abundant polymorphism in genomes. Moreover, recent empirical results suggest that mutationally complex, multiallelic loci including microsatellites and copy number variants are sometimes targeted by natural selection. Given their abundance, the lack of inference methods tailored to the mutational peculiarities of these types of loci represents a notable gap in our ability to interrogate genomes for signatures of natural selection. Previous theoretical investigations of mutation-selection balance at multiallelic loci include assumptions that limit their application to inference from empirical data. Focusing on microsatellites, we assess the dynamics and population-level consequences of selection targeting mutationally complex variants. We develop general models of a multiallelic fitness surface, a realistic model of microsatellite mutation, and an efficient simulation algorithm. Using these tools, we explore mutation-selection-drift equilibrium at microsatellites and investigate the mutational history and selective regime of the microsatellite that causes Friedreich's ataxia. We characterize microsatellite selective events by their duration and cost, note similarities to sweeps from standing point variation, and conclude that it is premature to label microsatellites as ubiquitous agents of efficient adaptive change. Together, our models and simulation algorithm provide a powerful framework for statistical inference, which can be used to test the neutrality of microsatellites and other multiallelic variants.

  12. Nonradioactive sequence-tagged microsatellite site analyses: a method transferable to the tropics.

    PubMed

    Lagoda, P J; Dambier, D; Grapin, A; Baurens, F C; Lanaud, C; Noyer, J L

    1998-02-01

    Utilization of existing isozyme analysis facilities to detect sequence-tagged microsatellite site (STMS) polymorphism or any simple sequence repeat (SSR) variation is described. Different parameters concerning the difficulties in transferring molecular techniques to less sophisticated laboratory infrastructures (i.e. tropical outstations) are discussed (e.g. reproducibility, efficacy, precision). Nonradioactive STMS analysis is bound to foster collaborative research between "biodiversity" and "biotechnology" centers.

  13. The report of my death was an exaggeration: A review for researchers using microsatellites in the 21st century1

    PubMed Central

    Hodel, Richard G. J.; Segovia-Salcedo, M. Claudia; Landis, Jacob B.; Crowl, Andrew A.; Sun, Miao; Liu, Xiaoxian; Gitzendanner, Matthew A.; Douglas, Norman A.; Germain-Aubrey, Charlotte C.; Chen, Shichao; Soltis, Douglas E.; Soltis, Pamela S.

    2016-01-01

    Microsatellites, or simple sequence repeats (SSRs), have long played a major role in genetic studies due to their typically high polymorphism. They have diverse applications, including genome mapping, forensics, ascertaining parentage, population and conservation genetics, identification of the parentage of polyploids, and phylogeography. We compare SSRs and newer methods, such as genotyping by sequencing (GBS) and restriction site associated DNA sequencing (RAD-Seq), and offer recommendations for researchers considering which genetic markers to use. We also review the variety of techniques currently used for identifying microsatellite loci and developing primers, with a particular focus on those that make use of next-generation sequencing (NGS). Additionally, we review software for microsatellite development and report on an experiment to assess the utility of currently available software for SSR development. Finally, we discuss the future of microsatellites and make recommendations for researchers preparing to use microsatellites. We argue that microsatellites still have an important place in the genomic age as they remain effective and cost-efficient markers. PMID:27347456

  14. Multilocus inference of species trees and DNA barcoding

    PubMed Central

    2016-01-01

    The unprecedented amount of data resulting from next-generation sequencing has opened a new era in phylogenetic estimation. Although large datasets should, in theory, increase phylogenetic resolution, massive, multilocus datasets have uncovered a great deal of phylogenetic incongruence among different genomic regions, due both to stochastic error and to the action of different evolutionary process such as incomplete lineage sorting, gene duplication and loss and horizontal gene transfer. This incongruence violates one of the fundamental assumptions of the DNA barcoding approach, which assumes that gene history and species history are identical. In this review, we explain some of the most important challenges we will have to face to reconstruct the history of species, and the advantages and disadvantages of different strategies for the phylogenetic analysis of multilocus data. In particular, we describe the evolutionary events that can generate species tree—gene tree discordance, compare the most popular methods for species tree reconstruction, highlight the challenges we need to face when using them and discuss their potential utility in barcoding. Current barcoding methods sacrifice a great amount of statistical power by only considering one locus, and a transition to multilocus barcodes would not only improve current barcoding methods, but also facilitate an eventual transition to species-tree-based barcoding strategies, which could better accommodate scenarios where the barcode gap is too small or inexistent. This article is part of the themed issue ‘From DNA barcodes to biomes’. PMID:27481787

  15. Microsatellite characterization of Cimarron Uruguayo dogs

    PubMed Central

    Gagliardi, Rosa; Llambí, Silvia; García, Cristina; Arruga, María Victoria

    2011-01-01

    Various genetic markers, including microsatellites, have been used to analyze the genetic polymorphism and heterozygosity in canine breeds. In this work, we used nine microsatellite markers to investigate the genetic variability in Cimarron Uruguayo dogs, the only officially recognized native canine breed in Uruguay. DNA from 30 Cimarron Uruguayo dogs from northeastern and southern Uruguay was analyzed. The allelic frequencies for each microsatellite, the genetic variability and the consanguinity were calculated, as were the polymorphic information content (PIC) and the probability of exclusion (PE). All of the microsatellites studied were polymorphic. FH 2361, FH 2305 and PEZ 03 were the most informative, with PIC values > 0.7, in agreement with results for other canine breeds. The PE values for the markers were within the ranges previously described and were generally greater for microsatellites with higher PIC values. The heterozygosity value (0.649) was considered high since only nine microsatellites were analyzed. Compared with data for other breeds, the results obtained here indicate that Cimarron Uruguayo dogs have high genetic diversity. PMID:21637561

  16. FullSSR: Microsatellite Finder and Primer Designer

    PubMed Central

    Metz, Sebastián; Cabrera, Juan Manuel; Rueda, Eva; Giri, Federico; Amavet, Patricia

    2016-01-01

    Microsatellites are genomic sequences comprised of tandem repeats of short nucleotide motifs widely used as molecular markers in population genetics. FullSSR is a new bioinformatic tool for microsatellite (SSR) loci detection and primer design using genomic data from NGS assay. The software was tested with 2000 sequences of Oryza sativa shotgun sequencing project from the National Center of Biotechnology Information Trace Archive and with partial genome sequencing with ROCHE 454® from Caiman latirostris, Salvator merianae, Aegla platensis, and Zilchiopsis collastinensis. FullSSR performance was compared against other similar SSR search programs. The results of the use of this kind of approach depend on the parameters set by the user. In addition, results can be affected by the analyzed sequences because of differences among the genomes. FullSSR simplifies the detection of SSRs and primer design on a big data set. The command line interface of FullSSR was intended to be used as part of genomic analysis tools pipeline; however, it can be used as a stand-alone program because the results are easily interpreted for a nonexpert user. PMID:27366148

  17. FullSSR: Microsatellite Finder and Primer Designer.

    PubMed

    Metz, Sebastián; Cabrera, Juan Manuel; Rueda, Eva; Giri, Federico; Amavet, Patricia

    2016-01-01

    Microsatellites are genomic sequences comprised of tandem repeats of short nucleotide motifs widely used as molecular markers in population genetics. FullSSR is a new bioinformatic tool for microsatellite (SSR) loci detection and primer design using genomic data from NGS assay. The software was tested with 2000 sequences of Oryza sativa shotgun sequencing project from the National Center of Biotechnology Information Trace Archive and with partial genome sequencing with ROCHE 454® from Caiman latirostris, Salvator merianae, Aegla platensis, and Zilchiopsis collastinensis. FullSSR performance was compared against other similar SSR search programs. The results of the use of this kind of approach depend on the parameters set by the user. In addition, results can be affected by the analyzed sequences because of differences among the genomes. FullSSR simplifies the detection of SSRs and primer design on a big data set. The command line interface of FullSSR was intended to be used as part of genomic analysis tools pipeline; however, it can be used as a stand-alone program because the results are easily interpreted for a nonexpert user.

  18. Toward fully automated genotyping: genotyping microsatellite markers by deconvolution.

    PubMed Central

    Perlin, M W; Lancia, G; Ng, S K

    1995-01-01

    Dense genetic linkage maps have been constructed for the human and mouse genomes, with average densities of 2.9 cM and 0.35 cM, respectively. These genetic maps are crucial for mapping both Mendelian and complex traits and are useful in clinical genetic diagnosis. Current maps are largely comprised of abundant, easily assayed, and highly polymorphic PCR-based microsatellite markers, primarily dinucleotide (CA)n repeats. One key limitation of these length polymorphisms is the PCR stutter (or slippage) artifact that introduces additional stutter bands. With two (or more) closely spaced alleles, the stutter bands overlap, and it is difficult to accurately determine the correct alleles; this stutter phenomenon has all but precluded full automation, since a human must visually inspect the allele data. We describe here novel deconvolution methods for accurate genotyping that mathematically remove PCR stutter artifact from microsatellite markers. These methods overcome the manual interpretation bottleneck and thereby enable full automation of genetic map construction and use. New functionalities, including the pooling of DNAs and the pooling of markers, are described that may greatly reduce the associated experimentation requirements. PMID:7485172

  19. (GTG)5 microsatellite regions in citrinin-producing Penicillium.

    PubMed

    Di Conza, José Alejandro; Nepote, Andrea Fabiana; González, Ana María; Lurá, María Cristina

    2007-03-01

    Morphological and cultural characteristics, as well as biochemical properties, are the main criteria used in fungal taxonomy and in the standard description of fungi species. Sometimes, however, this criterion is difficult to apply due to fungal phenotypic variations. This is particularly true in the genus Penicillium. The aims of this work were to determine (GTG)5 microsatellite sequence in potentially citrinin-producing Penicillium strains and to investigate if this sequence could be useful to characterize such fungi. Penicillium citrinum Thom and Penicillium chrysogenum Thom were isolated from different foods. The identification of the isolates at species level was carried out according to classical taxonomy. The production of citrinin was determined by thin layer chromatography. This study proved that microsatellite regions exist as short repeated sequences in all tested strains. The patterns were very similar for all P. citrinum isolates and it was possible to group them in function of the quantity of citrinin produced. Yet, not similar clusters were obtained when P. chrysogenum isolates were analyzed.

  20. Survey of simple sequence repeats in woodland strawberry (Fragaria vesca).

    PubMed

    Guan, L; Huang, J F; Feng, G Q; Wang, X W; Wang, Y; Chen, B Y; Qiao, Y S

    2013-07-30

    The use of simple sequence repeats (SSRs), or microsatellites, as genetic markers has become popular due to their abundance and variation in length among individuals. In this study, we investigated linkage groups (LGs) in the woodland strawberry (Fragaria vesca) and demonstrated variation in the abundances, densities, and relative densities of mononucleotide, dinucleotide, and trinucleotide repeats. Mononucleotide, dinucleotide, and trinucleotide repeats were more common than longer repeats in all LGs examined. Perfect SSRs were the predominant SSR type found and their abundance was extremely stable among LGs and chloroplasts. Abundances of mononucleotide, dinucleotide, and trinucleotide repeats were positively correlated with LG size, whereas those of tetranucleotide and hexanucleotide SSRs were not. Generally, in each LG, the abundance, relative abundance, relative density, and the proportion of each unique SSR all declined rapidly as the repeated unit increased. Furthermore, the lengths and frequencies of SSRs varied among different LGs.

  1. Identification and Evaluation of 21 Novel Microsatellite Markers from the Autumnal Moth (Epirrita autumnata) (Lepidoptera: Geometridae)

    PubMed Central

    Aarnes, Siv Grethe; Fløystad, Ida; Schregel, Julia; Vindstad, Ole Petter Laksforsmo; Jepsen, Jane Uhd; Eiken, Hans Geir; Ims, Rolf A.; Hagen, Snorre B.

    2015-01-01

    The autumnal moth (Epirrita autumnata) is a cyclically outbreaking forest Lepidoptera with circumpolar distribution and substantial impact on Northern ecosystems. We have isolated 21 microsatellites from the species to facilitate population genetic studies of population cycles, outbreaks, and crashes. First, PCR primers and PCR conditions were developed to amplify 19 trinucleotide loci and two tetranucleotide loci in six multiplex PCR approaches and then analyzed for species specificity, sensitivity and precision. Twelve of the loci showed simple tandem repeat array structures while nine loci showed imperfect repeat structures, and repeat numbers varied in our material between six and 15. The application in population genetics for all the 21 microsatellites were further validated in 48 autumnal moths sampled from Northern Norway, and allelic variation was detected in 19 loci. The detected numbers of alleles per locus ranged from two to 13, and the observed and expected heterozygosities varied from 0.04 to 0.69 and 0.04 to 0.79, respectively. Evidence for linkage disequilibrium was found for six loci as well as indication of one null allele. We find that these novel microsatellites and their multiplex-PCR assays are suitable for further research on fine- and large-scale population-genetic studies of Epirrita autumnata. PMID:26393576

  2. DNA Slippage Occurs at Microsatellite Loci without Minimal Threshold Length in Humans: A Comparative Genomic Approach

    PubMed Central

    Leclercq, Sébastien; Rivals, Eric; Jarne, Philippe

    2010-01-01

    The dynamics of microsatellite, or short tandem repeats (STRs), is well documented for long, polymorphic loci, but much less is known for shorter ones. For example, the issue of a minimum threshold length for DNA slippage remains contentious. Model-fitting methods have generally concluded that slippage only occurs over a threshold length of about eight nucleotides, in contradiction with some direct observations of tandem duplications at shorter repeated sites. Using a comparative analysis of the human and chimpanzee genomes, we examined the mutation patterns at microsatellite loci with lengths as short as one period plus one nucleotide. We found that the rates of tandem insertions and deletions at microsatellite loci strongly deviated from background rates in other parts of the human genome and followed an exponential increase with STR size. More importantly, we detected no lower threshold length for slippage. The rate of tandem duplications at unrepeated sites was higher than expected from random insertions, providing evidence for genome-wide action of indel slippage (an alternative mechanism generating tandem repeats). The rate of point mutations adjacent to STRs did not differ from that estimated elsewhere in the genome, except around dinucleotide loci. Our results suggest that the emergence of STR depends on DNA slippage, indel slippage, and point mutations. We also found that the dynamics of tandem insertions and deletions differed in both rates and size at which these mutations take place. We discuss these results in both evolutionary and mechanistic terms. PMID:20624737

  3. Development of microsatellite markers for the Korean Mussel, Mytilus coruscus (Mytilidae) using next-generation sequencing.

    PubMed

    An, Hye Suck; Lee, Jang Wook

    2012-01-01

    Mytilus coruscus (family Mytilidae) is one of the most important marine shellfish species in Korea. During the past few decades, this species has become endangered due to the loss of habitats and overfishing. Despite this species' importance, information on its genetic background is scarce. In this study, we developed microsatellite markers for M. coruscus using next-generation sequencing. A total of 263,900 raw reads were obtained from a quarter-plate run on the 454 GS-FLX titanium platform, and 176,327 unique sequences were generated with an average length of 381 bp; 2569 (1.45%) sequences contained a minimum of five di- to tetra-nucleotide repeat motifs. Of the 51 loci screened, 46 were amplified successfully, and 22 were polymorphic among 30 individuals, with seven of trinucleotide repeats and three of tetranucleotide repeats. All loci exhibited high genetic variability, with an average of 17.32 alleles per locus, and the mean observed and expected heterozygosities were 0.67 and 0.90, respectively. In addition, cross-amplification was tested for all 22 loci in another congener species, M. galloprovincialis. None of the primer pairs resulted in effective amplification, which might be due to their high mutation rates. Our work demonstrated the utility of next-generation 454 sequencing as a method for the rapid and cost-effective identification of microsatellites. The high degree of polymorphism exhibited by the 22 newly developed microsatellites will be useful in future conservation genetic studies of this species.

  4. Identification and Evaluation of 21 Novel Microsatellite Markers from the Autumnal Moth (Epirrita autumnata) (Lepidoptera: Geometridae).

    PubMed

    Aarnes, Siv Grethe; Fløystad, Ida; Schregel, Julia; Vindstad, Ole Petter Laksforsmo; Jepsen, Jane Uhd; Eiken, Hans Geir; Ims, Rolf A; Hagen, Snorre B

    2015-09-17

    The autumnal moth (Epirrita autumnata) is a cyclically outbreaking forest Lepidoptera with circumpolar distribution and substantial impact on Northern ecosystems. We have isolated 21 microsatellites from the species to facilitate population genetic studies of population cycles, outbreaks, and crashes. First, PCR primers and PCR conditions were developed to amplify 19 trinucleotide loci and two tetranucleotide loci in six multiplex PCR approaches and then analyzed for species specificity, sensitivity and precision. Twelve of the loci showed simple tandem repeat array structures while nine loci showed imperfect repeat structures, and repeat numbers varied in our material between six and 15. The application in population genetics for all the 21 microsatellites were further validated in 48 autumnal moths sampled from Northern Norway, and allelic variation was detected in 19 loci. The detected numbers of alleles per locus ranged from two to 13, and the observed and expected heterozygosities varied from 0.04 to 0.69 and 0.04 to 0.79, respectively. Evidence for linkage disequilibrium was found for six loci as well as indication of one null allele. We find that these novel microsatellites and their multiplex-PCR assays are suitable for further research on fine- and large-scale population-genetic studies of Epirrita autumnata.

  5. Transcriptome-Derived Tetranucleotide Microsatellites and Their Associated Genes from the Giant Panda (Ailuropoda melanoleuca).

    PubMed

    Song, Xuhao; Shen, Fujun; Huang, Jie; Huang, Yan; Du, Lianming; Wang, Chengdong; Fan, Zhenxin; Hou, Rong; Yue, Bisong; Zhang, Xiuyue

    2016-09-01

    Recently, an increasing number of microsatellites or simple sequence repeats (SSRs) have been found and characterized from transcriptomes. Such SSRs can be employed as putative functional markers to easily tag corresponding genes, which play an important role in biomedical studies and genetic analysis. However, the transcriptome-derived SSRs for giant panda (Ailuropoda melanoleuca) are not yet available. In this work, we identified and characterized 20 tetranucleotide microsatellite loci from a transcript database generated from the blood of giant panda. Furthermore, we assigned their predicted transcriptome locations: 16 loci were assigned to untranslated regions (UTRs) and 4 loci were assigned to coding regions (CDSs). Gene identities of 14 transcripts contained corresponding microsatellites were determined, which provide useful information to study the potential contribution of SSRs to gene regulation in giant panda. The polymorphic information content (PIC) values ranged from 0.293 to 0.789 with an average of 0.603 for the 16 UTRs-derived SSRs. Interestingly, 4 CDS-derived microsatellites developed in our study were also polymorphic, and the instability of these 4 CDS-derived SSRs was further validated by re-genotyping and sequencing. The genes containing these 4 CDS-derived SSRs were embedded with various types of repeat motifs. The interaction of all the length-changing SSRs might provide a way against coding region frameshift caused by microsatellite instability. We hope these newly gene-associated biomarkers will pave the way for genetic and biomedical studies for giant panda in the future. In sum, this set of transcriptome-derived markers complements the genetic resources available for giant panda.

  6. Genome-wide microsatellite identification in the fungus Anisogramma anomala using Illumina sequencing and genome assembly.

    PubMed

    Cai, Guohong; Leadbetter, Clayton W; Muehlbauer, Megan F; Molnar, Thomas J; Hillman, Bradley I

    2013-01-01

    High-throughput sequencing has been dramatically accelerating the discovery of microsatellite markers (also known as Simple Sequence Repeats). Both 454 and Illumina reads have been used directly in microsatellite discovery and primer design (the "Seq-to-SSR" approach). However, constraints of this approach include: 1) many microsatellite-containing reads do not have sufficient flanking sequences to allow primer design, and 2) difficulties in removing microsatellite loci residing in longer, repetitive regions. In the current study, we applied the novel "Seq-Assembly-SSR" approach to overcome these constraints in Anisogramma anomala. In our approach, Illumina reads were first assembled into a draft genome, and the latter was then used in microsatellite discovery. A. anomala is an obligate biotrophic ascomycete that causes eastern filbert blight disease of commercial European hazelnut. Little is known about its population structure or diversity. Approximately 26 M 146 bp Illumina reads were generated from a paired-end library of a fungal strain from Oregon. The reads were assembled into a draft genome of 333 Mb (excluding gaps), with contig N50 of 10,384 bp and scaffold N50 of 32,987 bp. A bioinformatics pipeline identified 46,677 microsatellite motifs at 44,247 loci, including 2,430 compound loci. Primers were successfully designed for 42,923 loci (97%). After removing 2,886 loci close to assembly gaps and 676 loci in repetitive regions, a genome-wide microsatellite database of 39,361 loci was generated for the fungus. In experimental screening of 236 loci using four geographically representative strains, 228 (96.6%) were successfully amplified and 214 (90.7%) produced single PCR products. Twenty-three (9.7%) were found to be perfect polymorphic loci. A small-scale population study using 11 polymorphic loci revealed considerable gene diversity. Clustering analysis grouped isolates of this fungus into two clades in accordance with their geographic origins. Thus, the

  7. Identification and Characterization of Microsatellite Markers Derived from the Whole Genome Analysis of Taenia solium

    PubMed Central

    Pajuelo, Mónica J.; Eguiluz, María; Dahlstrom, Eric; Requena, David; Guzmán, Frank; Ramirez, Manuel; Sheen, Patricia; Frace, Michael; Sammons, Scott; Cama, Vitaliano; Anzick, Sarah; Bruno, Dan; Mahanty, Siddhartha; Wilkins, Patricia; Nash, Theodore; Gonzalez, Armando; García, Héctor H.; Gilman, Robert H.; Porcella, Steve; Zimic, Mirko

    2015-01-01

    Background Infections with Taenia solium are the most common cause of adult acquired seizures worldwide, and are the leading cause of epilepsy in developing countries. A better understanding of the genetic diversity of T. solium will improve parasite diagnostics and transmission pathways in endemic areas thereby facilitating the design of future control measures and interventions. Microsatellite markers are useful genome features, which enable strain typing and identification in complex pathogen genomes. Here we describe microsatellite identification and characterization in T. solium, providing information that will assist in global efforts to control this important pathogen. Methods For genome sequencing, T. solium cysts and proglottids were collected from Huancayo and Puno in Peru, respectively. Using next generation sequencing (NGS) and de novo assembly, we assembled two draft genomes and one hybrid genome. Microsatellite sequences were identified and 36 of them were selected for further analysis. Twenty T. solium isolates were collected from Tumbes in the northern region, and twenty from Puno in the southern region of Peru. The size-polymorphism of the selected microsatellites was determined with multi-capillary electrophoresis. We analyzed the association between microsatellite polymorphism and the geographic origin of the samples. Results The predicted size of the hybrid (proglottid genome combined with cyst genome) T. solium genome was 111 MB with a GC content of 42.54%. A total of 7,979 contigs (>1,000 nt) were obtained. We identified 9,129 microsatellites in the Puno-proglottid genome and 9,936 in the Huancayo-cyst genome, with 5 or more repeats, ranging from mono- to hexa-nucleotide. Seven microsatellites were polymorphic and 29 were monomorphic within the analyzed isolates. T. solium tapeworms were classified into two genetic groups that correlated with the North/South geographic origin of the parasites. Conclusions/Significance The availability of draft

  8. Evolution of hypervariable microsatellites in apomictic polyploid lineages of Ranunculus carpaticola: directional bias at dinucleotide loci.

    PubMed

    Paun, Ovidiu; Hörandl, Elvira

    2006-09-01

    Microsatellites are widely used in genetic and evolutionary analyses, but their own evolution is far from simple. The mechanisms maintaining the mutational patterns of simple repeats and the typical stable allele-frequency distributions are still poorly understood. Asexual lineages may provide particularly informative models for the indirect study of microsatellite evolution, because their genomes act as complete linkage groups, with mutations being the only source of genetic variation. Here, we study the direction of accumulated dinucleotide microsatellite mutations in wild asexual lineages of hexaploid Ranunculus carpaticola. Whereas the overall number of contractions is not significantly different from that of expansions, the within-locus frequency of contractions, but not of expansions, significantly increases with allele length. Moreover, within-locus polymorphism is positively correlated with allele length, but this relationship is due solely to the influence of contraction mutations. Such asymmetries may explain length constraints generally observed with microsatellites and are consistent with stable, bell-shaped allele-frequency distributions. Although apomictic and allohexaploid, the R. carpaticola lineages show mutational patterns resembling the trends observed in a broad range of organisms, including sexuals and diploids, suggesting that, even if not of germline origin, the mutations in these apomicts may be the consequence of similar mechanisms.

  9. Characterization of novel microsatellite markers for Hyphantria cunea and implications for other Lepidoptera.

    PubMed

    Cao, L J; Wen, J B; Wei, S J; Liu, J; Yang, F; Chen, M

    2015-06-01

    This is the first report of microsatellite markers (simple sequence repeats, SSR) for fall webworm, Hyphantria cunea (Drury) (Lepidoptera: Arctiidae), an important quarantine pest in some European and Asian countries. Here, we developed 48 microsatellite markers for H. cunea from SSR enrichment libraries. Sequences isolated from libraries were sorted into four categories and analyzed. Our results suggest that sequences classified as Grouped should not be used for microsatellite primer design. The genetic diversity of microsatellite loci was assessed in 72 individuals from three populations. The number of alleles per locus ranged from 2 to 5 with an average of 3. The observed and expected heterozygosities of loci ranged from 0 to 0.958 and 0 to 0.773, respectively. A total of 18 out of 153 locus/population combinations deviated significantly from Hardy-Weinberg equilibrium. Moreover, significant linkage disequilibrium was detected in one pair of loci (1275 pairs in total). In the neutral test, two loci were grouped into the candidate category for positive selection and the remainder into the neutral category. In addition, a complex mutation pattern was observed for these loci, and F ST performed better than did R ST for the estimation of population differentiation in different mutation patterns. The results of the present study can be used for population genetic studies of H. cunea.

  10. Development of microsatellite markers for six Tetranychus species by transfer from Tetranychus urticae genome.

    PubMed

    Zhang, Jia; Sun, Jing-Tao; Jin, Peng-Yu; Hong, Xiao-Yue

    2016-09-01

    Microsatellite markers are frequently used to explore the population genetic structure of organisms. Spider mites (genus Tetranychus) are important agricultural pests. Several markers have been developed for T. urticae, but for other spider mites, few such markers are available, hampering studies of their population genetics. In this study, we developed and characterized microsatellite markers for six non-model spider mite species (T. truncatus, T. kanzawai, T. ludeni, T. piercei, T. phaselus and T. pueraricola) by cross-species amplification of markers in the T. urticae genome, in order to better understand the population structure of Tetranychus species. Among 228 screened loci, many were polymorphic, including 13 loci in T. urticae, 11 loci in T. truncatus, 15 loci in T. pueraricola, 23 loci in T. kanzawai, 19 loci in T. piercei, 11 loci in T. phaselus and 9 loci in T. ludeni. Sequence analysis determined that the fragment length variations of the transferred microsatellites were mainly due to the variations of the numbers of repeats. These new microsatellite markers should be useful for studying the population genetics of the seven Tetranychus species.

  11. Development of 23 polymorphic microsatellite loci in invasive silver wattle, Acacia dealbata (Fabaceae)1

    PubMed Central

    Guillemaud, Thomas; Broadhurst, Linda; Legoff, Isabelle; Henery, Martin; Blin, Aurélie; Ducatillion, Catherine; Ferrando, Nathalie; Malausa, Thibaut

    2015-01-01

    Premise of the study: Microsatellite markers were developed for silver wattle, Acacia dealbata (Fabaceae), which is both an ornamental and an invasive weed species. It is native to southeastern Australia and invasive in Europe, Africa, Asia, and the Americas. Methods and Results: The pyrosequencing of a microsatellite-enriched genomic DNA library of A. dealbata produced 33,290 sequences and allowed the isolation of 201 loci with a minimum of seven repeats of microsatellite motifs. Amplification tests led to the setup of two multiplex PCR mixes allowing the amplification of 21 loci. The polymorphism of these markers was evaluated on a sample of 32 individuals collected in southeastern Australia. The number of alleles and the expected heterozygosity varied between two and 11, and between 0.11 and 0.88, respectively. Conclusions: The level of polymorphism of this set of 23 microsatellites is large enough to provide valuable information on the genetic structure and the invasion history of A. dealbata. PMID:25995979

  12. CRISPR–Cas9-targeted fragmentation and selective sequencing enable massively parallel microsatellite analysis

    PubMed Central

    Shin, GiWon; Grimes, Susan M.; Lee, HoJoon; Lau, Billy T.; Xia, Li C.; Ji, Hanlee P.

    2017-01-01

    Microsatellites are multi-allelic and composed of short tandem repeats (STRs) with individual motifs composed of mononucleotides, dinucleotides or higher including hexamers. Next-generation sequencing approaches and other STR assays rely on a limited number of PCR amplicons, typically in the tens. Here, we demonstrate STR-Seq, a next-generation sequencing technology that analyses over 2,000 STRs in parallel, and provides the accurate genotyping of microsatellites. STR-Seq employs in vitro CRISPR–Cas9-targeted fragmentation to produce specific DNA molecules covering the complete microsatellite sequence. Amplification-free library preparation provides single molecule sequences without unique molecular barcodes. STR-selective primers enable massively parallel, targeted sequencing of large STR sets. Overall, STR-Seq has higher throughput, improved accuracy and provides a greater number of informative haplotypes compared with other microsatellite analysis approaches. With these new features, STR-Seq can identify a 0.1% minor genome fraction in a DNA mixture composed of different, unrelated samples. PMID:28169275

  13. Microsatellite primers for the Pacific Northwest conifer Callitropsis nootkatensis (Cupressaceae)1

    PubMed Central

    Jennings, Tara N.; Knaus, Brian J.; Alderman, Katherine; Hennon, Paul E.; D’Amore, David V.; Cronn, Richard

    2013-01-01

    • Premise of the study: Microsatellite primers were developed for Nootka cypress (Callitropsis nootkatensis) to provide quantitative measures for gene conservation that can assist in guiding management decisions for a species experiencing climate-induced decline. • Methods and Results: Using multiplexed massively parallel sequencing, we identified 136,785 microsatellite-containing sequences from 489,625 Illumina paired-end 80-bp reads. After stringent filtering, we selected 144 primer pairs and screened variation at these loci in five populations of C. nootkatensis. Loci show between three and 36 dinucleotide repeats per locus, with an average of 13. Screening of these markers in the Pacific Northwest relative Chamaecyparis lawsoniana demonstrated no marker transferability. This finding highlights the narrow taxonomic utility of microsatellite markers in Callitropsis. • Conclusions: These microsatellites show high polymorphism and can be used for routine screening of natural variation in Callitropsis nootkatensis, and will be particularly helpful in identifying clones and inbred relatives at the stand-level. PMID:25202587

  14. [Frequency of various mini- and micro-satellite sequences in DNA of human chromosome 13].

    PubMed

    Ryskov, A P; Kupriianova, N S; Kapanadze, B I; Nechvolodov, K K; Pozmogova, G E; Prosniak, M I; Iankovskiĭ, N K

    1993-10-01

    The frequency of specific mini- and micro-satellites known also as short tandem repeated sequences (STR) in the human 13 chromosome was estimated by hybridization of STR core oligonucleotides to recombinant cosmid clones transferred to a grid from a human 13 chromosome specific cosmid library ICRF Lawrist 4 C108 (DN L4/HS 13). Oligonucleotides: M13 and Jeffreys minisatellite core sequences and micro-satellite core sequences (TCC)5, (CAC)5, and (GACA)4 were [gamma-32P] end labeled and hybridized to membrane filters carrying good ordered cosmid clones. It was shown that great number of all these mini- and micro-satellite copies (besides of Jeffreys minisatellite) are spread independently along the 13th chromosome. It was also estimated that two or more (GACA)n blocks present in the same cosmid (i.e. on the stretch of 40-50 kb) forming similar groups of clustered micro-satellites. The interesting peculiarity has been recorded that some (GACA)n+ cosmids are also hybridizable to conservative 28SrDNA 3'-fragment that indicates that (GACA)n localization in the nucleoli area. As the result of it we began the creation of a new highly polymorphic markers collections for these chromosome.

  15. Genetic polymorphism of Hucul horse population based on 17 microsatellite loci.

    PubMed

    Fornal, Agnieszka; Radko, Anna; Piestrzyńska-Kajtoch, Agata

    2013-01-01

    Short tandem repeat (STR) loci, i.e. microsatellites are a class of genetic markers commonly used for population studies and parentage control. This study determined the usefulness of microsatellite markers recommended by International Society for Animal Genetics (ISAG) for identification and pedigree analysis in horses based on the example of Polish Hucul horse population (Equus caballus). The set of seventeen microsatellites loci was tested (AHT4, AHT5, ASB2, HMS2, HMS3, HMS6, HMS7, HTG10, HTG4, HTG6, HTG7, VHL20, ASB17, ASB23, CA425, HMS1, LEX3) for 216 individuals. All samples were genotyped and mean number of alleles per locus was estimated (7.00). Means of observed (Ho) and expected (He) heterozygosity were calculated 0.7288 and 0.7027, respectively. The observed heterozygosity was similar to the results of research on Hucul horse population in another area of Carpathians Mountains. The average polymorphism information content (PIC) for analyses of seventeen microsatellite markers indicates the usefulness of this set of markers for Hucul horse parentage testing.

  16. Development of novel tetra- and trinucleotide microsatellite markers for giant grouper Epinephelus lanceolatus using 454 pyrosequencing.

    PubMed

    Kim, Keun-Sik; Noh, Choong Hwan; Moon, Shin-Joo; Han, Seung-Hee; Bang, In-Chul

    2016-06-01

    Giant grouper (Epinephelus lanceolatus) is a commercially important species, but its wild population has recently been classified as vulnerable. This species has significant potential for use in aquaculture, though a greater understanding of population genetics is necessary for selective breeding programs to minimize kinship for genetically healthy individuals. High-throughput pyrosequencing of genomic DNA was used to identify and characterize novel tetra- and trinucleotide microsatellite markers in giant grouper from Sabah, Malaysia. In total, of 62,763 sequences containing simple sequence repeats (SSRs) were obtained, and 78 SSR loci were selected to possibly contain tetra- and trinucleotide repeats. Of these loci, 16 had tetra- and 8 had trinucleotide repeats, all of which exhibited polymorphisms within easily genotyped regions. A total of 143 alleles were identified with an average of 5.94 alleles per locus, with mean observed and expected heterozygosities of 0.648 and 0.620, respectively. Among of them, 15 microsatellite markers were identified without null alleles and with Hardy-Weinberg equilibrium. These alleles showed a combined non-exclusion probability of 0.01138. The probability of individual identification (PID) value combined with in descending order 12 microsatellite markers was 0.00008, which strongly suggests that the use of the microsatellite markers developed in this study in various combinations would result in a high resolution method for parentage analysis and individual identification. These markers could be used to establish a broodstock management program for giant grouper and to provide a foundation for genetic studies such as population structure, parentage analysis, and kinship selection.

  17. Development and diversity of Andean-derived, gene-based microsatellites for common bean (Phaseolus vulgaris L.)

    PubMed Central

    2009-01-01

    Background Gene-based (genic) microsatellites are a useful tool for plant genetics and simple sequence repeat loci can often be found in coding regions of the genome. While EST sequencing can be used to discover genic microsatellites, direct screening of cDNA libraries for repeat motifs can save on overall sequencing costs. The objective of this research was to screen a large cDNA library from and Andean common bean genotype for six di-nucleotide and tri-nucleotide repeat motifs through a filter hybridization approach and to develop microsatellite markers from positive clones. Results Robotics were used for high-throughput colony picking and to create a high-density filter of 18,432 double spotted cDNA clones which was followed by hybridization with repeat motif containing probes based on GA, CA, AAT, CAG, CAA and ACG repeats. A total of 1203 positive clones were identified by their addresses and sequenced from 5' ends and if required from 3' ends to confirm repeat motif and length. Out of 886 high quality sequences, 497 had complete microsatellite loci that were not truncated by the sequencing reaction and of these tri-nucleotide repeats were more common than di-nucleotide repeats. Different motifs were found in different frequencies in the 5' and 3' ends of the cDNAs. In a microsatellite development program, primers were designed for 248 SSR loci which were tested on a panel of 18 common bean genotypes to determine their potential as genetic markers finding higher average polymorphism information content for di-nucleotide repeat markers (0.3544) than for tri-nucleotide repeat markers (0.1536). Conclusion The present study provides a set of validated gene-based markers for common bean that are derived from G19833, an Andean landrace that is an important source of disease and abiotic stress tolerance which has been used for physical map development and as a mapping parent. Gene-based markers appear to be very efficient at separating divergent wild and cultivated

  18. Development and characterization of polymorphic microsatellite markers in taro (Colocasia esculenta).

    PubMed

    Mace, Emma S; Godwin, Ian D

    2002-10-01

    Microsatellite-containing sequences were isolated from enriched genomic libraries of taro (Colocasia esculenta (L.) Schott). The sequencing of 269 clones yielded 77 inserts containing repeat motifs. The majority of these (81.7%) were dinucleotide or trinucleotide repeats. The GT/CA repeat motif was the most common, accounting for 42% of all repeat types. From a total of 43 primer pairs designed, 41 produced markers within the expected size range. Sixteen (39%) were polymorphic when screened against a restricted set of taro genotypes from Southeast Asia and Oceania, with an average of 3.2 alleles detected on each locus. These markers represent a useful resource for taro germplasm management, genome mapping, and marker-assisted selection.

  19. [Microsatellite data verify low genetic differentiation between western and eastern subspecies of the common crane Grusgrus L. (Gruidae, Aves)].

    PubMed

    Mudrik, E A; Kashentseva, T A; Redchuk, P S; Politov, D V

    2015-01-01

    Using a set of 10 microsatellite loci (Gram-22, Gram-30, Gpa-12, Gpa-38, Gpa-39, Gj-M15, Gj-M34, Gj-4066, Gj-8077, Gj-2298) a high level of genetic variability (N(A) = 10.2, H(O) = = 0.684, H(E) = 0.728) and low genetic differentiation (F(ST) = 0.011) in the Common crane (Grus grus L.) was detected throughout its range. Genetic diversity in disputed western (G. g. grus) and eastern (G. g. lilfordi) sub- species was shown to be similar. Spatial distribution of multi-locus individual genotypes has not been revealed (R(XY) = 0.017). Despite low differentiation, subspecies and local populations of the Common crane should be considered as separate conservational units. Organization of programs for gene pool conservation and monitor- ing requires more detailed analysis based on combined analysis of various molecular markers.

  20. Diversity, genetic structure and evidence of outcrossing in British populations of the rock fern Adiantum capillus-veneris using microsatellites.

    PubMed

    Pryor, K V; Young, J E; Rumsey, F J; Edwards, K J; Bruford, M W; Rogers, H J

    2001-08-01

    Microsatellites were isolated and a marker system was developed in the fern Adiantum capillus-veneris. Polymorphic markers were then used to study the genetic diversity and structure of populations within the UK and Ireland where this species grows at the northern edge of its range, requiring a specific rock habitat and limited to a few scattered populations. Three dinucleotide loci detected a high level of diversity (23 alleles and 28 multilocus genotypes) across the UK and Ireland, with nearly all variation partitioned among rather than within populations. Of 17 populations represented by multiple samples, all except four were monomorphic. Heterozygosity was detected in three populations, all within Glamorgan, Wales (UK), showing evidence of outcrossing. We make inferences on the factors determining the observed levels and patterns of genetic variation and the possible evolutionary history of the populations.

  1. Development of ten microsatellite loci in the invasive giant African land snail, Achatina (=Lissachatina) fulica Bowdich, 1822

    USGS Publications Warehouse

    Morrison, Cheryl L.; Springmann, Marcus J.; Iwanowicz, Deborah D.; Wade, Christopher M.

    2015-01-01

    A suite of tetra-nucleotide microsatellite loci were developed for the invasive giant African land snail, Achatina (=Lissachatina) fulica Bowdich, 1822, from Ion Torrent next-generation sequencing data. Ten of the 96 primer sets tested amplified consistently in 30 snails from Miami, Florida, plus 12 individuals representative of their native East Africa, Indian and Pacific Ocean regions. The loci displayed moderate levels of allelic diversity (average 5.6 alleles/locus) and heterozygosity (average 42 %). Levels of genetic diversity were sufficient to produce unique multi-locus genotypes and detect phylogeographic structuring among regional samples. The invasive A. fulica can cause extensive damage to important food crops and natural resources, including native flora and fauna. The loci characterized here will be useful for determining the origins and tracking the spread of invasions, detecting fine-scale spatial structuring and estimating demographic parameters.

  2. Microsatellite primers resource developed from the mapped sequence scaffolds of Nisqually-1 genome. Submitted to New Phytologist

    SciTech Connect

    Yin, Tongming; ZHANG, Dr. XINYE; Gunter, Lee E; Li, Shuxian; Wullschleger, Stan D; Huang, Prof. Minren; Tuskan, Gerald A

    2009-01-01

    In this study, 148 428 simple sequence repeat (SSR) primer pairs were designed from the unambiguously mapped sequence scaffolds of the Nisqually-1 genome. The physical position of the priming sites were identified along each of the 19 Populus chromosomes, and it was specified whether the priming sequences belong to intronic, intergenic, exonic or UTR regions. A subset of 150 SSR loci were amplified and a high amplification success rate (72%) was obtained in P. tremuloides, which belongs to a divergent subgenus of Populus relative to Nisqually-1. PCR reactions showed that the amplification success rate of exonic primer pairs was much higher than that of the intronic/intergenic primer pairs. Applying ANOVA and regression analyses to the flanking sequences of microsatellites, the repeat lengths, the GC contents of the repeats, the repeat motif numbers, the repeat motif length and the base composition of the repeat motif, it was determined that only the base composition of the repeat motif and the repeat motif length significantly affect the microsatellite variability in P. tremuloides samples. The SSR primer resource developed in this study provides a database for selecting highly transferable SSR markers with known physical position in the Populus genome and provides a comprehensive genetic tool to extend the genome sequence of Nisqually-1 to genetic studies in different Populus species.

  3. Development of 18 novel microsatellite primers for Begonia fimbristipula (Begoniaceae), an endangered medicinal plant in China1

    PubMed Central

    Zhao, Bo; Du, Yun-Qian; Li, Jing-Jian; Tang, Wen-Xiu; Zhong, Shu-Hua

    2016-01-01

    Premise of the study: Begonia fimbristipula (Begoniaceae) is a medicinal herb distributed in the Chinese provinces of Fujian, Guangdong, Guangxi, Hainan, Hunan, Jiangxi, and Zhejiang, and it is on the verge of extinction due to habitat destruction and deterioration of its ecosystem. Here we developed a set of highly polymorphic microsatellite markers for population genetic and conservation studies of this endangered medicinal plant. Methods and Results: Using the Fast Isolation by AFLP of Sequences COntaining repeats (FIASCO) protocol, 18 polymorphic microsatellite markers were identified within 48 individuals from two geographic locations. The observed and expected heterozygosities ranged from 0.208 to 1.000 and from 0.291 to 0.812, respectively. These microsatellite markers were cross-amplified in five related Begonia species, and six loci were successfully amplified in all species. Conclusions: These 18 markers will be useful for better conservation and utilization of wild resources of B. fimbristipula and other Begonia species in the future. PMID:27437176

  4. Development of 14 microsatellite markers in Odontites vernus s.l. (Orobanchaceae) and cross-amplification in related taxa1

    PubMed Central

    Pinto-Carrasco, Daniel; Košnar, Jiří; López-González, Noemí; Koutecký, Petr; Těšitel, Jakub; Rico, Enrique; Martínez-Ortega, M. Montserrat

    2016-01-01

    Premise of the study: Microsatellite primers were developed for the first time in the root hemiparasite herb Odontites vernus (Orobanchaceae). These markers will be useful to investigate the role of polyploidization in the evolution of this diploid-tetraploid complex, as well as the extent of gene flow between different ploidy levels. Methods and Results: Fourteen polymorphic and reproducible loci were identified and optimized from O. vernus using a microsatellite-enriched library and 454 Junior sequencing. The set of primers amplified di- to pentanucleotide repeats and showed two to 13 alleles per locus. Transferability was tested in 30 taxa (19 belonging to Odontites and 11 from eight other genera of Orobanchaceae tribe Rhinantheae). Conclusions: The results indicate the utility of the newly developed microsatellites in O. vernus and several other species, which will be useful for taxon delimitation and conservation genetics studies. PMID:27011897

  5. Mining and characterization of sequence tagged microsatellites from the brown planthopper Nilaparvata lugens.

    PubMed

    Sun, Jing-Tao; Zhang, Yan-Kai; Ge, Cheng; Hong, Xiao-Yue

    2011-01-01

    The brown planthopper, Nilaparvata lugens (Stål) (Hemiptera: Delphacidae), is an important pest of rice. To better understand the migration pattern and population structure of the Chinese populations of N. lugens, we developed and characterized 12 polymorphic microsatellites from the expressed sequence tags database of N. lugens. The occurrence of these simple sequence repeats was assessed in three populations collected from three provinces of China. The number of alleles per locus ranged from 3 to 13 with an average of 6.5 alleles per locus. The mean observed heterozygosity of the three populations ranged from 0.051 to 0.772 and the expected heterozygosity ranged from 0.074 to 0.766. The sequences of the 12 markers were highly variable. The polymorphism information content of the 12 markers was high and ranged from 0.074 to 0.807 (mean = 0.503). Sequencing of microsatellite alleles revealed that the fragment length differences were mainly due to the variation of the repeat motif. Significant genetic differentiation was detected among the three N. lugens populations as the Fst ranged from 0.034 to 0.273. Principle coordinates analysis also revealed significant genetic differentiation between populations of different years. We conclude that these microsatellite markers will be a powerful tools to study the migration routine of the N. lugens.

  6. Conservation implications of the mating system of the Pampa Hermosa landrace of peach palm analyzed with microsatellite markers

    PubMed Central

    Picanço-Rodrigues, Doriane; Astolfi-Filho, Spartaco; Lemes, Maristerra R.; Gribel, Rogerio; Sebbenn, Alexandre M.; Clement, Charles R.

    2015-01-01

    Peach palm (Bactris gasipaes) is cultivated by many indigenous and traditional communities from Amazonia to Central America for its edible fruits, and is currently important for its heart-of-palm. The objective of this study was to investigate the mating system of peach palm, as this is important for conservation and breeding. Eight microsatellite loci were used to genotype 24 open-pollinated progenies from three populations of the Pampa Hermosa landrace maintained in a progeny trial for genetic improvement. Both the multi-locus outcrossing rates (0.95 to 0.99) and the progeny level multi-locus outcrossing rates (0.9 to 1.0) were high, indicating that peach palm is predominantly allogamous. The outcrossing rates among relatives were significantly different from zero (0.101 to 0.202), providing evidence for considerable biparental inbreeding within populations, probably due to farmers planting seeds of a small number of open-pollinated progenies in the same plot. The correlations of paternity estimates were low (0.051 to 0.112), suggesting a large number of pollen sources (9 to 20) participating in pollination of individual fruit bunches. Effective population size estimates suggest that current germplasm collections are insufficient for long-term ex situ conservation. As with most underutilized crops, on farm conservation is the most important component of an integrated conservation strategy. PMID:25983626

  7. Comparison of Leishmania killicki (syn. L. tropica) and Leishmania tropica Population Structure in Maghreb by Microsatellite Typing.

    PubMed

    Chaara, Dhekra; Bañuls, Anne-Laure; Haouas, Najoua; Talignani, Loïc; Lami, Patrick; Mezhoud, Habib; Harrat, Zoubir; Dedet, Jean-Pierre; Babba, Hamouda; Pratlong, Francine

    2015-12-01

    Leishmania (L.) killicki (syn. L. tropica), which causes cutaneous leishmaniasis in Maghreb, was recently described in this region and identified as a subpopulation of L. tropica. The present genetic analysis was conducted to explore the spatio-temporal distribution of L. killicki (syn. L. tropica) and its transmission dynamics. To better understand the evolution of this parasite, its population structure was then compared with that of L. tropica populations from Morocco. In total 198 samples including 85 L. killicki (syn. L. tropica) (from Tunisia, Algeria and Libya) and 113 L. tropica specimens (all from Morocco) were tested. Theses samples were composed of 168 Leishmania strains isolated from human skin lesions, 27 DNA samples from human skin lesion biopsies, two DNA samples from Ctenodactylus gundi bone marrow and one DNA sample from a Phlebotomus sergenti female. The sample was analyzed by using MultiLocus Enzyme Electrophoresis (MLEE) and MultiLocus Microsatellite Typing (MLMT) approaches. Analysis of the MLMT data support the hypothesis that L. killicki (syn. L. tropica) belongs to the L. tropica complex, despite its strong genetic differentiation, and that it emerged from this taxon by a founder effect. Moreover, it revealed a strong structuring in L. killicki (syn. L. tropica) between Tunisia and Algeria and within the different Tunisian regions, suggesting low dispersion of L. killicki (syn. L. tropica) in space and time. Comparison of the L. tropica (exclusively from Morocco) and L. killicki (syn. L. tropica) population structures revealed distinct genetic organizations, reflecting different epidemiological cycles.

  8. Molecular characterization of Giardia psittaci by multilocus sequence analysis.

    PubMed

    Abe, Niichiro; Makino, Ikuko; Kojima, Atsushi

    2012-12-01

    Multilocus sequence analyses targeting small subunit ribosomal DNA (SSU rDNA), elongation factor 1 alpha (ef1α), glutamate dehydrogenase (gdh), and beta giardin (β-giardin) were performed on Giardia psittaci isolates from three Budgerigars (Melopsittacus undulates) and four Barred parakeets (Bolborhynchus lineola) kept in individual households or imported from overseas. Nucleotide differences and phylogenetic analyses at four loci indicate the distinction of G. psittaci from the other known Giardia species: Giardia muris, Giardia microti, Giardia ardeae, and Giardia duodenalis assemblages. Furthermore, G. psittaci was related more closely to G. duodenalis than to the other known Giardia species, except for G. microti. Conflicting signals regarded as "double peaks" were found at the same nucleotide positions of the ef1α in all isolates. However, the sequences of the other three loci, including gdh and β-giardin, which are known to be highly variable, from all isolates were also mutually identical at every locus. They showed no double peaks. These results suggest that double peaks found in the ef1α sequences are caused not by mixed infection with genetically different G. psittaci isolates but by allelic sequence heterogeneity (ASH), which is observed in diplomonad lineages including G. duodenalis. No sequence difference was found in any G. psittaci isolates at the gdh and β-giardin, suggesting that G. psittaci is indeed not more diverse genetically than other Giardia species. This report is the first to provide evidence related to the genetic characteristics of G. psittaci obtained using multilocus sequence analysis.

  9. Multilocus phylogenetic analyses of Hispaniolan and Bahamian trunk anoles (distichus species group).

    PubMed

    Geneva, Anthony J; Hilton, Jared; Noll, Sabina; Glor, Richard E

    2015-06-01

    The distichus species group includes six species and 21 subspecies of trunk ecomorph anoles distributed across Hispaniola and its satellite islands as well as the northern Bahamas. Although this group has long served as a model system for studies of reproductive character displacement, adaptation, behavior and speciation, it has never been the subject of a comprehensive phylogenetic analysis. Our goal here is to generate a multilocus phylogenetic dataset (one mitochondrial and seven nuclear loci) and to use this dataset to infer phylogenetic relationships among the majority of the taxa assigned to the distichus species group. We use these phylogenetic trees to address three topics about the group's evolution. First, we consider longstanding taxonomic controversies about the status of several species and subspecies assigned to the distichus species group. Second, we investigate the biogeographic history of the group and specifically test the hypotheses that historical division of Hispaniola into two paleo-islands contributed to the group's diversification and that Bahamian and Hispaniolan satellite island populations are derived from colonists from the main Hispaniolan landmass. Finally, third, we use comparative phylogenetic analyses to test the hypothesis that divergence between pale yellow and darkly pigmented orange or red dewlap coloration has occurred repeatedly across the distichus species group.

  10. Comparative multi-locus phylogeography confirms multiple vicariance events in co-distributed rainforest frogs

    PubMed Central

    Bell, Rayna C.; MacKenzie, Jason B.; Hickerson, Michael J.; Chavarría, Krystle L.; Cunningham, Michael; Williams, Stephen; Moritz, Craig

    2012-01-01

    Though Pleistocene refugia are frequently cited as drivers of species diversification, comparisons of molecular divergence among sister species typically indicate a continuum of divergence times from the Late Miocene, rather than a clear pulse of speciation events at the Last Glacial Maximum. Community-scale inference methods that explicitly test for multiple vicariance events, and account for differences in ancestral effective population size and gene flow, are well suited for detecting heterogeneity of species' responses to past climate fluctuations. We apply this approach to multi-locus sequence data from five co-distributed frog species endemic to the Wet Tropics rainforests of northeast Australia. Our results demonstrate at least two episodes of vicariance owing to climate-driven forest contractions: one in the Early Pleistocene and the other considerably older. Understanding how repeated cycles of rainforest contraction and expansion differentially affected lineage divergence among co-distributed species provides a framework for identifying evolutionary processes that underlie population divergence and speciation. PMID:21900325

  11. genepopedit: a simple and flexible tool for manipulating multilocus molecular data in R.

    PubMed

    Stanley, Ryan R E; Jeffery, Nicholas W; Wringe, Brendan F; DiBacco, Claudio; Bradbury, Ian R

    2017-01-01

    Advances in genetic sequencing technologies and techniques have made large, genome-wide data sets comprised of hundreds or even thousands of individuals and loci the norm rather than the exception even for nonmodel organisms. While such data present new opportunities for evaluating population structure and demographic processes, the large size of these genomic data sets brings new computational challenges for researchers needing to parse, convert and manipulate data often into a variety of software-specific formats required of genomic analyses. We developed genepopedit as a flexible tool for the manipulation of multilocus molecular data sets. Functionality can be divided among diagnostic-, manipulation-, sampling-, simulation-, and transformation-based tools. Metadata from large genomic data sets can be efficiently extracted, without the need to view data in a text-editing program. genepopedit provides tools to manipulate loci, individual samples and populations included in genomic data sets, in addition to the ability to convert directly to a variety of software formats. Functions are compiled as an R package, which can integrate into existing analysis workflows. Importantly, genepopedit provides a simple yet robust code-based tool for repeatable genomic data manipulation, which has been proven to be stable for data sets in excess of 200 000 SNPs. The latest version of the package and associated documentation are available on Github (github.com/rystanley/genepopedit).

  12. Identification of Coxiella burnetii genotypes in Croatia using multi-locus VNTR analysis.

    PubMed

    Račić, Ivana; Spičić, Silvio; Galov, Ana; Duvnjak, Sanja; Zdelar-Tuk, Maja; Vujnović, Anja; Habrun, Boris; Cvetnić, Zeljko

    2014-10-10

    Although Q fever affects humans and animals in Croatia, we are unaware of genotyping studies of Croatian strains of the causative pathogen Coxiella burnetii, which would greatly assist monitoring and control efforts. Here 3261 human and animal samples were screened for C. burnetii DNA by conventional PCR, and 335 (10.3%) were positive. Of these positive samples, 82 were genotyped at 17 loci using the relatively new method of multi-locus variable number tandem repeat analysis (MLVA). We identified 13 C. burnetii genotypes not previously reported anywhere in the world. Two of these 13 genotypes are typical of the continental part of Croatia and share more similarity with genotypes outside Croatia than with genotypes within the country. The remaining 11 novel genotypes are typical of the coastal part of Croatia and show more similarity to one another than to genotypes outside the country. Our findings shed new light on the phylogeny of C. burnetii strains and may help establish MLVA as a standard technique for Coxiella genotyping.

  13. Inter-allelic interactions play a major role in microsatellite evolution.

    PubMed

    Amos, William; Kosanović, Danica; Eriksson, Anders

    2015-11-07

    Microsatellite mutations identified in pedigrees confirm that most changes involve the gain or loss of single repeats. However, an unexpected pattern is revealed when the resulting data are plotted on standardized scales that range from the shortest to longest allele at a locus. Both mutation rate and mutation bias reveal a strong dependency on allele length relative to other alleles at the same locus. We show that models in which alleles mutate independently cannot explain these patterns. Instead, both mutation probability and direction appear to involve interactions between homologues in heterozygous individuals. Simple models in which the longer homologue in heterozygotes is more likely to mutate and/or biased towards contraction readily capture the observed trends. The exact model remains unclear in all its details but inter-allelic interactions are a vital component, implying a link between demographic history and the mode and tempo of microsatellite evolution.

  14. Isolation and characterization of microsatellite loci in the invasive herb Solidago altissima (Asteraceae)1

    PubMed Central

    Sakata, Yuzu; Kaneko, Shingo; Hayano, Azusa; Inoue-Murayama, Miho; Ohgushi, Takayuki; Isagi, Yuji

    2013-01-01

    • Premise of the study: Microsatellite markers were developed in the perennial herb Solidago altissima from populations within its introduced range in Japan to assess its population structure and to facilitate tracking of invasion expansion. • Methods and Results: Using 454 pyrosequencing, 16 microsatellite primer sets were developed for S. altissima. The primer sets were tested on 70 individuals sampled from three populations in Japan. The primers amplified di- and trinucleotide repeats with five to 25 alleles per locus, and the expected heterozygosity ranged from 0.46 to 0.92. • Conclusions: These results indicate the utility of primers in S. altissima for future research on a wide range of applications, including tracking of invasion dynamics and investigating population genetics of the species. PMID:25202531

  15. Development and characterization of 32 microsatellite loci in Genipa americana (Rubiaceae)1

    PubMed Central

    Manoel, Ricardo O.; Freitas, Miguel L. M.; Barreto, Mariana A.; Moraes, Mário L. T.; Souza, Anete P.; Sebbenn, Alexandre M.

    2014-01-01

    • Premise of the study: Microsatellite primers were developed for the tree species Genipa americana (Rubiaceae) for further population genetic studies. • Methods and Results: We identified 144 clones containing 65 repeat motifs from a genomic library enriched for (CT)8 and (GT)8 motifs. Primer pairs were developed for 32 microsatellite loci and validated in 40 individuals of two natural G. americana populations. Seventeen loci were polymorphic, revealing from three to seven alleles per locus. The observed and expected heterozygosities ranged from 0.24 to 1.00 and from 0.22 to 0.78, respectively. • Conclusions: The 17 primers identified as polymorphic loci are suitable to study the genetic diversity and structure, mating system, and gene flow in G. americana. PMID:25202610

  16. FANCJ suppresses microsatellite instability and lymphomagenesis independent of the Fanconi anemia pathway.

    PubMed

    Matsuzaki, Kenichiro; Borel, Valerie; Adelman, Carrie A; Schindler, Detlev; Boulton, Simon J

    2015-12-15

    Microsatellites are short tandem repeat sequences that are highly prone to expansion/contraction due to their propensity to form non-B-form DNA structures, which hinder DNA polymerases and provoke template slippage. Although error correction by mismatch repair plays a key role in preventing microsatellite instability (MSI), which is a hallmark of Lynch syndrome, activities must also exist that unwind secondary structures to facilitate replication fidelity. Here, we report that Fancj helicase-deficient mice, while phenotypically resembling Fanconi anemia (FA), are also hypersensitive to replication inhibitors and predisposed to lymphoma. Whereas metabolism of G4-DNA structures is largely unaffected in Fancj(-/-) mice, high levels of spontaneous MSI occur, which is exacerbated by replication inhibition. In contrast, MSI is not observed in Fancd2(-/-) mice but is prevalent in human FA-J patients. Together, these data implicate FANCJ as a key factor required to counteract MSI, which is functionally distinct from its role in the FA pathway.

  17. Development and Characterization of Microsatellite Markers for the Cape Gooseberry Physalis peruviana

    PubMed Central

    Simbaqueba, Jaime; Sánchez, Pilar; Sanchez, Erika; Núñez Zarantes, Victor Manuel; Chacon, Maria Isabel; Barrero, Luz Stella; Mariño-Ramírez, Leonardo

    2011-01-01

    Physalis peruviana, commonly known as Cape gooseberry, is an Andean Solanaceae fruit with high nutritional value and interesting medicinal properties. In the present study we report the development and characterization of microsatellite loci from a P. peruviana commercial Colombian genotype. We identified 932 imperfect and 201 perfect Simple Sequence Repeats (SSR) loci in untranslated regions (UTRs) and 304 imperfect and 83 perfect SSR loci in coding regions from the assembled Physalis peruviana leaf transcriptome. The UTR SSR loci were used for the development of 162 primers for amplification. The efficiency of these primers was tested via PCR in a panel of seven P. peruviana accessions including Colombia, Kenya and Ecuador ecotypes and one closely related species Physalis floridana. We obtained an amplification rate of 83% and a polymorphic rate of 22%. Here we report the first P. peruviana specific microsatellite set, a valuable tool for a wide variety of applications, including functional diversity, conservation and improvement of the species. PMID:22039540

  18. Development and characterization of 15 microsatellite markers for Cephalotaxus fortunei (Cephalotaxaceae)1

    PubMed Central

    Wang, Chunbo; Guo, Zhiyou; Huang, Xilian; Huang, Lu

    2016-01-01

    Premise of the study: To survey population variation and the adaptive evolution of Cephalotaxus fortunei (Cephalotaxaceae), an endemic and endangered conifer in China, microsatellite markers were developed and characterized for this species. Methods and Results: Based on the Fast Isolation by AFLP of Sequences COntaining repeats (FIASCO) protocol, 15 microsatellite markers were developed for C. fortunei, 13 of which were polymorphic within a sample of 75 individuals representing five natural populations. The number of alleles per locus ranged from one to seven. The expected and observed heterozygosities were 0.108–0.738 and 0.000–1.000, respectively. Ten polymorphic loci were also successfully amplified in C. oliveri. Conclusions: These polymorphic loci provide a valuable tool for population genetic analysis of C. fortunei, which will contribute to its management and conservation. PMID:27213121

  19. Development and characterization of microsatellite loci in the mistletoe Psittacanthus schiedeanus (Loranthaceae)1

    PubMed Central

    González, Clementina; Harvey, Nick; Ornelas, Juan Francisco

    2015-01-01

    • Premise of the study: Microsatellite primers were developed for the parasitic Psittacanthus schiedeanus, a common mistletoe species on cloud forest–adapted tree hosts in Mesoamerica, to investigate intraspecific genetic patterns of diversity and genetic structure. • Methods and Results: Using an enriched library, 10 polymorphic microsatellite loci were developed in P. schiedeanus. All loci consisted of dinucleotide repeats. Average alleles per locus were 12 (4–17), and a total of 120 alleles were recorded across 39 individuals from four populations in Mexico. Primers were tested in 11 additional species, but only amplified successfully in P. calyculatus and P. angustifolius. • Conclusions: The polymorphic loci described will be useful in studies of genetic diversity and genetic population differentiation in natural populations of these parasitic plants, and will provide valuable information to understand the importance of host distribution. PMID:25606357

  20. Microsatellite primers in the foundation tree species Pinus edulis and P. monophylla (Pinaceae)1

    PubMed Central

    Krohn, Andrew L.; Flores-Rentería, Lluvia; Gehring, Catherine A.

    2013-01-01

    • Premise of the study: Microsatellite primers were developed in the foundational tree species Pinus edulis to investigate population differentiation of P. edulis and hybridization among closely related species. • Methods and Results: Using a hybridization protocol, primer sets for 11 microsatellite loci were developed using megagametophyte tissue from P. edulis and scored for polymorphism in three populations of P. edulis and a single P. monophylla population. The primers amplified simple and compound di-, tri-, and pentanucleotide repeats with two to 18 alleles per locus. • Conclusions: These results demonstrate the utility of the described primers for studies of population differentiation within and among P. edulis populations as well as across putative hybrid zones where P. edulis may coexist with sister species. PMID:25202571

  1. Inter-allelic interactions play a major role in microsatellite evolution

    PubMed Central

    Amos, William; Kosanović, Danica; Eriksson, Anders

    2015-01-01

    Microsatellite mutations identified in pedigrees confirm that most changes involve the gain or loss of single repeats. However, an unexpected pattern is revealed when the resulting data are plotted on standardized scales that range from the shortest to longest allele at a locus. Both mutation rate and mutation bias reveal a strong dependency on allele length relative to other alleles at the same locus. We show that models in which alleles mutate independently cannot explain these patterns. Instead, both mutation probability and direction appear to involve interactions between homologues in heterozygous individuals. Simple models in which the longer homologue in heterozygotes is more likely to mutate and/or biased towards contraction readily capture the observed trends. The exact model remains unclear in all its details but inter-allelic interactions are a vital component, implying a link between demographic history and the mode and tempo of microsatellite evolution. PMID:26511050

  2. The Repeat Expansion Diseases: the dark side of DNA repair?

    PubMed Central

    Zhao, Xiao-Nan; Usdin, Karen

    2015-01-01

    DNA repair normally protects the genome against mutations that threaten genome integrity and thus cell viability. However, growing evidence suggests that in the case of the Repeat Expansion Diseases, disorders that result from an increase in the size of a disease-specific microsatellite, the disease-causing mutation is actually the result of aberrant DNA repair. A variety of proteins from different DNA repair pathways have thus far been implicated in this process. This review will summarize recent findings from patients and from mouse models of these diseases that shed light on how these pathways may interact to cause repeat expansion. PMID:26002199

  3. Microsatellite markers for raspberry and blackberry

    Technology Transfer Automated Retrieval System (TEKTRAN)

    welve microsatellites were isolated from SSR-enriched genomic libraries of Rubus idaeus L.‘Meeker’ red raspberry (diploid) and R. loganobaccus L. H. Bailey ‘Marion’ blackberry-raspberry hybrid (hexaploid). These primer pairs, with the addition of one developed from a GenBank R. idaeus sequence, we...

  4. Microsatellite Markers for Raspberries and Blackberries

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Twelve microsatellites were isolated from SSR-enriched genomic libraries of Rubus idaeus L.‘Meeker’ red raspberry (diploid) and R. loganobaccus L. H. Bailey ‘Marion’ blackberry-raspberry hybrid (hexaploid). These primer pairs, with the addition of one developed from a GenBank R. idaeus sequence, w...

  5. Microsatellite markers in plant pathogenic fungi

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Knowing the genetic diversity of plant pathogenic fungi is essential in the management of crops and disease. The genetic variability of fungal pathogens can be evaluated using molecular markers, among which, microsatellites are a relatively inexpensive source of information. We have developed an e...

  6. Automated genotyping of dinucleotide repeat markers

    SciTech Connect

    Perlin, M.W.; Hoffman, E.P. |

    1994-09-01

    The dinucleotide repeats (i.e., microsatellites) such as CA-repeats are a highly polymorphic, highly abundant class of PCR-amplifiable markers that have greatly streamlined genetic mapping experimentation. It is expected that over 30,000 such markers (including tri- and tetranucleotide repeats) will be characterized for routine use in the next few years. Since only size determination, and not sequencing, is required to determine alleles, in principle, dinucleotide repeat genotyping is easily performed on electrophoretic gels, and can be automated using DNA sequencers. Unfortunately, PCR stuttering with these markers generates not one band for each allele, but a pattern of bands. Since closely spaced alleles must be disambiguated by human scoring, this poses a key obstacle to full automation. We have developed methods that overcome this obstacle. Our model is that the observed data is generated by arithmetic superposition (i.e., convolution) of multiple allele patterns. By quantitatively measuring the size of each component band, and exploiting the unique stutter pattern associated with each marker, closely spaced alleles can be deconvolved; this unambiguously reconstructs the {open_quotes}true{close_quotes} allele bands, with stutter artifact removed. We used this approach in a system for automated diagnosis of (X-linked) Duchenne muscular dystrophy; four multiplexed CA-repeats within the dystrophin gene were assayed on a DNA sequencer. Our method accurately detected small variations in gel migration that shifted the allele size estimate. In 167 nonmutated alleles, 89% (149/167) showed no size variation, 9% (15/167) showed 1 bp variation, and 2% (3/167) showed 2 bp variation. We are currently developing a library of dinucleotide repeat patterns; together with our deconvolution methods, this library will enable fully automated genotyping of dinucleotide repeats from sizing data.

  7. Multilocus phylogeography and population structure of common eiders breeding in North America and Scandinavia

    USGS Publications Warehouse

    Sonsthagen, S.A.; Talbot, S.L.; Scribner, K.T.; McCracken, K.G.

    2011-01-01

    Aim Glacial refugia during the Pleistocene had major impacts on the levels and spatial apportionment of genetic diversity of species in northern latitude ecosystems. We characterized patterns of population subdivision, and tested hypotheses associated with locations of potential Pleistocene refugia and the relative contribution of these refugia to the post-glacial colonization of North America and Scandinavia by common eiders (Somateria mollissima). Specifically, we evaluated localities hypothesized as ice-free areas or glacial refugia for other Arctic vertebrates, including Beringia, the High Arctic Canadian Archipelago, Newfoundland Bank, Spitsbergen Bank and north-west Norway. Location Alaska, Canada, Norway and Sweden. Methods Molecular data from 12 microsatellite loci, the mitochondrial DNA (mtDNA) control region, and two nuclear introns were collected and analysed for 15 populations of common eiders (n=716) breeding throughout North America and Scandinavia. Population genetic structure, historical population fluctuations and gene flow were inferred using F-statistics, analyses of molecular variance, and multilocus coalescent analyses. Results Significant inter-population variation in allelic and haplotypic frequencies were observed (nuclear DNA FST=0.004-0.290; mtDNA ??ST=0.051-0.927). Whereas spatial differentiation in nuclear genes was concordant with subspecific designations, geographic proximity was more predictive of inter-population variance in mitochondrial DNA haplotype frequency. Inferences of historical population demography were consistent with restriction of common eiders to four geographic areas during the Last Glacial Maximum: Belcher Islands, Newfoundland Bank, northern Alaska and Svalbard. Three of these areas coincide with previously identified glacial refugia: Newfoundland Bank, Beringia and Spitsbergen Bank. Gene-flow and clustering analyses indicated that the Beringian refugium contributed little to common eider post-glacial colonization

  8. Multilocus phylogeography and population structure of common eiders breeding in North America and Scandinavia

    USGS Publications Warehouse

    Sonsthagen, Sarah A.; Talbot, Sandra L.; Scribner, Kim T.; McCracken, Kevin G.

    2014-01-01

    Aim:  Glacial refugia during the Pleistocene had major impacts on the levels and spatial apportionment of genetic diversity of species in northern latitude ecosystems. We characterized patterns of population subdivision, and tested hypotheses associated with locations of potential Pleistocene refugia and the relative contribution of these refugia to the post-glacial colonization of North America and Scandinavia by common eiders (Somateria mollissima). Specifically, we evaluated localities hypothesized as ice-free areas or glacial refugia for other Arctic vertebrates, including Beringia, the High Arctic Canadian Archipelago, Newfoundland Bank, Spitsbergen Bank and north-west Norway. Location: Alaska, Canada, Norway and Sweden. Methods: Molecular data from 12 microsatellite loci, the mitochondrial DNA (mtDNA) control region, and two nuclear introns were collected and analysed for 15 populations of common eiders (n = 716) breeding throughout North America and Scandinavia. Population genetic structure, historical population fluctuations and gene flow were inferred using F-statistics, analyses of molecular variance, and multilocus coalescent analyses. Results: Significant inter-population variation in allelic and haplotypic frequencies were observed (nuclear DNA FST = 0.004–0.290; mtDNA ΦST = 0.051–0.927). Whereas spatial differentiation in nuclear genes was concordant with subspecific designations, geographic proximity was more predictive of inter-population variance in mitochondrial DNA haplotype frequency. Inferences of historical population demography were consistent with restriction of common eiders to four geographic areas during the Last Glacial Maximum: Belcher Islands, Newfoundland Bank, northern Alaska and Svalbard. Three of these areas coincide with previously identified glacial refugia: Newfoundland Bank, Beringia and Spitsbergen Bank. Gene-flow and clustering analyses indicated that the Beringian refugium contributed little to common eider post

  9. Rediscovering Medicinal Plants' Potential with OMICS: Microsatellite Survey in Expressed Sequence Tags of Eleven Traditional Plants with Potent Antidiabetic Properties

    PubMed Central

    Sahu, Jagajjit; Sen, Priyabrata; Choudhury, Manabendra Dutta; Dehury, Budheswar; Barooah, Madhumita; Modi, Mahendra Kumar

    2014-01-01

    Abstract Herbal medicines and traditionally used medicinal plants present an untapped potential for novel molecular target discovery using systems science and OMICS biotechnology driven strategies. Since up to 40% of the world's poor people have no access to government health services, traditional and folk medicines are often the only therapeutics available to them. In this vein, North East (NE) India is recognized for its rich bioresources. As part of the Indo-Burma hotspot, it is regarded as an epicenter of biodiversity for several plants having myriad traditional uses, including medicinal use. However, the improvement of these valuable bioresources through molecular breeding strategies, for example, using genic microsatellites or Simple Sequence Repeats (SSRs) or Expressed Sequence Tags (ESTs)-derived SSRs has not been fully utilized in large scale to date. In this study, we identified a total of 47,700 microsatellites from 109,609 ESTs of 11 medicinal plants (pineapple, papaya, noyontara, bitter orange, bermuda brass, ratalu, barbados nut, mango, mulberry, lotus, and guduchi) having proven antidiabetic properties. A total of 58,159 primer pairs were designed for the non-redundant 8060 SSR-positive ESTs and putative functions were assigned to 4483 unique contigs. Among the identified microsatellites, excluding mononucleotide repeats, di-/trinucleotides are predominant, among which repeat motifs of AG/CT and AAG/CTT were most abundant. Similarity search of SSR containing ESTs and antidiabetic gene sequences revealed 11 microsatellites linked to antidiabetic genes in five plants. GO term enrichment analysis revealed a total of 80 enriched GO terms widely distributed in 53 biological processes, 17 molecular functions, and 10 cellular components associated with the 11 markers. The present study therefore provides concrete insights into the frequency and distribution of SSRs in important medicinal resources. The microsatellite markers reported here markedly add to

  10. Characterization of perfect microsatellite based on genome-wide and chromosome level in Rhesus monkey (Macaca mulatta).

    PubMed

    Xu, Yongtao; Hu, Zongxiu; Wang, Chen; Zhang, Xiuyue; Li, Jing; Yue, Bisong

    2016-11-05

    Microsatellite studies based on chromosomes level would contribute to the biometric correlation analysis of chromosome and microsatellite applications on the specific chromosome. In this study, the total microsatellite length of 1,141,024 loci was 21.8Mb, which covered about 0.74% of the male Rhesus monkey genome. Perfect mononucleotide SSRs were the most abundant, followed by the pattern: perfect di->tetra->tri->penta->hexanucleotide SSRs. The main range of repeat times focused on 12-32 times (mono-), 7-23 times (di-), 5-10 times (tri-), 4-14 times (tetra-), 4-9 times (penta-), 4-8 times (hexa-), respectively. The largest SSRs number was found in chromosome 1 with 94,347 loci, followed by chromosome 3, 2, 7 and 5, and the smallest number was in chromosome 18. The predominant repeat types in male Rhesus monkey genome and chromosome Y were basically A, AC, AG, AAT, AAC, AAAT, AAAC, AAAG, AAACA and AAACAA. SSRs number of all chromosomes was closely positively correlated with chromosome sequence size (r=0.969, p<0.01), and significantly negatively correlated with abundance (r=-0.24, 0.01microsatellite density (r=-0.456, 0.01repeat sequences in each chromosome for primer design would facilitate the exploration of microsatellites structural function, composition mode and molecular markers development in Rhesus monkey genome.

  11. SSR_pipeline: a bioinformatic infrastructure for identifying microsatellites from paired-end Illumina high-throughput DNA sequencing data

    USGS Publications Warehouse

    Miller, Mark P.; Knaus, Brian J.; Mullins, Thomas D.; Haig, Susan M.

    2013-01-01

    SSR_pipeline is a flexible set of programs designed to efficiently identify simple sequence repeats (e.g., microsatellites) from paired-end high-throughput Illumina DNA sequencing data. The program suite contains 3 analysis modules along with a fourth control module that can automate analyses of large volumes of data. The modules are used to 1) identify the subset of paired-end sequences that pass Illumina quality standards, 2) align paired-end reads into a single composite DNA sequence, and 3) identify sequences that possess microsatellites (both simple and compound) conforming to user-specified parameters. The microsatellite search algorithm is extremely efficient, and we have used it to identify repeats with motifs from 2 to 25bp in length. Each of the 3 analysis modules can also be used independently to provide greater flexibility or to work with FASTQ or FASTA files generated from other sequencing platforms (Roche 454, Ion Torrent, etc.). We demonstrate use of the program with data from the brine fly Ephydra packardi (Diptera: Ephydridae) and provide empirical timing benchmarks to illustrate program performance on a common desktop computer environment. We further show that the Illumina platform is capable of identifying large numbers of microsatellites, even when using unenriched sample libraries and a very small percentage of the sequencing capacity from a single DNA sequencing run. All modules from SSR_pipeline are implemented in the Python programming language and can therefore be used from nearly any computer operating system (Linux, Macintosh, and Windows).

  12. SSR_pipeline: a bioinformatic infrastructure for identifying microsatellites from paired-end Illumina high-throughput DNA sequencing data.

    PubMed

    Miller, Mark P; Knaus, Brian J; Mullins, Thomas D; Haig, Susan M

    2013-01-01

    SSR_pipeline is a flexible set of programs designed to efficiently identify simple sequence repeats (e.g., microsatellites) from paired-end high-throughput Illumina DNA sequencing data. The program suite contains 3 analysis modules along with a fourth control module that can automate analyses of large volumes of data. The modules are used to 1) identify the subset of paired-end sequences that pass Illumina quality standards, 2) align paired-end reads into a single composite DNA sequence, and 3) identify sequences that possess microsatellites (both simple and compound) conforming to user-specified parameters. The microsatellite search algorithm is extremely efficient, and we have used it to identify repeats with motifs from 2 to 25 bp in length. Each of the 3 analysis modules can also be used independently to provide greater flexibility or to work with FASTQ or FASTA files generated from other sequencing platforms (Roche 454, Ion Torrent, etc.). We demonstrate use of the program with data from the brine fly Ephydra packardi (Diptera: Ephydridae) and provide empirical timing benchmarks to illustrate program performance on a common desktop computer environment. We further show that the Illumina platform is capable of identifying large numbers of microsatellites, even when using unenriched sample libraries and a very small percentage of the sequencing capacity from a single DNA sequencing run. All modules from SSR_pipeline are implemented in the Python programming language and can therefore be used from nearly any computer operating system (Linux, Macintosh, and Windows).

  13. Haplogroup-specific deviation from the stepwise mutation model at the microsatellite loci DYS388 and DYS392.

    PubMed

    Nebel, A; Filon, D; Hohoff, C; Faerman, M; Brinkmann, B; Oppenheim, A

    2001-01-01

    Deviation from the stepwise mutation model (SMM) at specific human microsatellite loci has implications for population genetic and forensic investigations. In the present study, data on six Y chromosome-specific microsatellites were pooled for 455 paternally unrelated males from six Middle Eastern populations. All chromosomes were assigned to three haplogroups defined by six binary polymorphisms. Two of the microsatellite loci tested, DYS388 and DYS392, displayed marked haplogroup-specific differences in their allele variability. A bimodal distribution of short and long alleles was observed for DYS388 in haplogroup 1 and for DYS392 in haplogroups 1 and 2. Further investigation showed that the short/long alleles segregated almost completely between genealogically distinct haplogroups defined by additional binary markers. Thus, these two loci have a discriminatory power similar to a binary polymorphism. DYS388 was characterised by an extremely low mutation rate in haplogroups 2 and 3, as was DYS392 in haplogroup 3. Sequence analysis of the repeat regions at the two loci revealed no irregularities, indicating that the triplet expansion in these loci is not controlled by sequence variation at the repeat level. A high frequency of long DYS388 alleles has, so far, been found only in populations originating in the Middle East, suggesting that this microsatellite is useful as a region-specific marker.

  14. Microsatellite variability among wild and cultivated hops (Humulus lupulus L.).

    PubMed

    Jakse, Jernej; Satovic, Zlatko; Javornik, Branka

    2004-10-01

    Hop (Humulus lupulus L.) is a dioecious perennial plant native to the northern hemisphere cultivated for its use in the brewing industry. To investigate the genetic diversity present in wild hop accessions in comparison with cultivated hops, microsatellite marker variation was assessed at four loci in 124 accessions of wild (from Europe, Asia and from North America) and cultivated (varieties and breeding lines) hops. A total of 63 alleles were identified, with an average of 15.7 alleles per locus and an average PIC of 0.64 over four loci. The average number of alleles per locus in groups of accessions ranged from 5.75 to 8.30, with the highest number detected in groups of wild hops either of European (EU) or North American (NA) origin. Accessions from NA revealed the highest number of unique alleles indicating the high diversity present in this gene pool. Cluster analysis based on the D(D) or D(sw) distance matrix divided accessions into 10 different clusters, which reflect the relationship among geographically diverse wild accessions and hop cultivars. The highest genetic differences were found between NA wild accessions, forming one distant cluster, and all the other accessions. The differentiation between European wild and cultivated accessions was revealed by PCoA based on the D(D) distance matrix and by AMOVA results. Cultivated hops differ significantly from wild ones, although most of the variability was found within groups. The molecular variances within groups of cultivated and wild hops were homogeneous, suggesting that a similar level of molecular variability is found in both groups of accessions. The analysis of allele polymorphism and of allele sequences showed that hop germplasm can be differentiated to NA and EU geographic types according to the differences of allele sizes at three loci or by the specific microsatellite repeat type at one locus. The analysis also indicates the different evolutionary dynamics and complex mutations of microsatellite

  15. Microsatellite flanking region similarities among different loci within insect species.

    PubMed

    Meglécz, E; Anderson, S J; Bourguet, D; Butcher, R; Caldas, A; Cassel-Lundhagen, A; d'Acier, A C; Dawson, D A; Faure, N; Fauvelot, C; Franck, P; Harper, G; Keyghobadi, N; Kluetsch, C; Muthulakshmi, M; Nagaraju, J; Patt, A; Péténian, F; Silvain, J-F; Wilcock, H R

    2007-04-01

    Although microsatellites are ubiquitous in eukaryota, the number of available markers varies strongly among taxa. This meta-analysis was conducted on 32 insect species. Sequences were obtained from two assembled whole genomes, whole genome shotgun (WGS) sequences from 10 species and screening partial genomic libraries for microsatellites from 23 species. We have demonstrated: (1) strong differences in the abundance of microsatellites among species; (2) that microsatellites within species are often grouped into families based on similarities in their flanking sequences; (3) that the proportion of microsatellites grouped into families varies strongly among taxa; and (4) that microsatellite families were significantly more often associated with transposable elements - or their remnants - than unique microsatellite sequences.

  16. Microsatellite Genotyping of Plasmodium vivax Isolates from Pregnant Women in Four Malaria Endemic Countries

    PubMed Central

    Menegon, Michela; Bardají, Azucena; Martínez-Espinosa, Flor; Bôtto-Menezes, Camila; Ome-Kaius, Maria; Mueller, Ivo; Betuela, Inoni; Arévalo-Herrera, Myriam; Kochar, Swati; Kochar, Sanjay K.; Jaju, Puneet; Hans, Dhiraj; Chitnis, Chetan; Padilla, Norma; Castellanos, María Eugenia; Ortiz, Lucía; Sanz, Sergi; Piqueras, Mireia; Desai, Meghna; Mayor, Alfredo; del Portillo, Hernando; Menéndez, Clara; Severini, Carlo

    2016-01-01

    Plasmodium vivax is the most widely distributed human parasite and the main cause of human malaria outside the African continent. However, the knowledge about the genetic variability of P. vivax is limited when compared to the information available for P. falciparum. We present the results of a study aimed at characterizing the genetic structure of P. vivax populations obtained from pregnant women from different malaria endemic settings. Between June 2008 and October 2011 nearly 2000 pregnant women were recruited during routine antenatal care at each site and followed up until delivery. A capillary blood sample from the study participants was collected for genotyping at different time points. Seven P. vivax microsatellite markers were used for genotypic characterization on a total of 229 P. vivax isolates obtained from Brazil, Colombia, India and Papua New Guinea. In each population, the number of alleles per locus, the expected heterozygosity and the levels of multilocus linkage disequilibrium were assessed. The extent of genetic differentiation among populations was also estimated. Six microsatellite loci on 137 P. falciparum isolates from three countries were screened for comparison. The mean value of expected heterozygosity per country ranged from 0.839 to 0.874 for P. vivax and from 0.578 to 0.758 for P. falciparum. P. vivax populations were more diverse than those of P. falciparum. In some of the studied countries, the diversity of P. vivax population was very high compared to the respective level of endemicity. The level of inter-population differentiation was moderate to high in all P. vivax and P. falciparum populations studied. PMID:27011010

  17. Isolation and characterization of microsatellite DNA loci in the threatened flat-spired three-toothed land snail Triodopsis platysayoides

    USGS Publications Warehouse

    King, Timothy L.; Eackles, Michael S.; Garner, B. A.; van Tuinen, M.; Arbogast, B. S.

    2015-01-01

    The hermaphroditic flat-spired three-tooth land snail (Triodopsis platysayoides) is endemic to a 21-km stretch of the Cheat River Gorge of northeastern West Virginia, USA. We document isolation and characterization of ten microsatellite DNA markers in this at-risk species. The markers displayed a moderate level of allelic diversity (averaging 7.1 alleles/locus) and heterozygosity (averaging 58.6 %). Allelic diversity at seven loci was sufficient to produce unique multilocus genotypes; no indication of selfing was detected in this cosexual species. Minimal deviations from Hardy–Weinberg equilibrium and no linkage disequilibrium were observed within subpopulations. All loci deviated from Hardy–Weinberg expectations when individuals from subpopulations were pooled. Microsatellite markers developed for T. platysayoides yielded sufficient genetic diversity to (1) distinguish all individuals sampled and the level of selfing; (2) be appropriate for addressing fine-scale population structuring; (3) provide novel demographic insights for the species; and (4) cross-amplify and detect allelic diversity in the congeneric T. juxtidens.

  18. STAMP: Extensions to the STADEN sequence analysis package for high throughput interactive microsatellite marker design

    PubMed Central

    Kraemer, Lars; Beszteri, Bánk; Gäbler-Schwarz, Steffi; Held, Christoph; Leese, Florian; Mayer, Christoph; Pöhlmann, Kevin; Frickenhaus, Stephan

    2009-01-01

    Background Microsatellites (MSs) are DNA markers with high analytical power, which are widely used in population genetics, genetic mapping, and forensic studies. Currently available software solutions for high-throughput MS design (i) have shortcomings in detecting and distinguishing imperfect and perfect MSs, (ii) lack often necessary interactive design steps, and (iii) do not allow for the development of primers for multiplex amplifications. We present a set of new tools implemented as extensions to the STADEN package, which provides the backbone functionality for flexible sequence analysis workflows. The possibility to assemble overlapping reads into unique contigs (provided by the base functionality of the STADEN package) is important to avoid developing redundant markers, a feature missing from most other similar tools. Results Our extensions to the STADEN package provide the following functionality to facilitate microsatellite (and also minisatellite) marker design: The new modules (i) integrate the state-of-the-art tandem repeat detection and analysis software PHOBOS into workflows, (ii) provide two separate repeat detection steps – with different search criteria – one for masking repetitive regions during assembly of sequencing reads and the other for designing repeat-flanking primers for MS candidate loci, (iii) incorporate the widely used primer design program PRIMER3 into STADEN workflows, enabling the interactive design and visualization of flanking primers for microsatellites, and (iv) provide the functionality to find optimal locus- and primer pair combinations for multiplex primer design. Furthermore, our extensions include a module for storing analysis results in an SQLite database, providing a transparent solution for data access from within as well as from outside of the STADEN Package. Conclusion The STADEN package is enhanced by our modules into a highly flexible, high-throughput, interactive tool for conventional and multiplex

  19. Major clades of Agaricales: a multilocus phylogenetic overview.

    PubMed

    Matheny, P Brandon; Curtis, Judd M; Hofstetter, Valérie; Aime, M Catherine; Moncalvo, Jean-Marc; Ge, Zai-Wei; Slot, Jason C; Ammirati, Joseph F; Baroni, Timothy J; Bougher, Neale L; Hughes, Karen W; Lodge, D Jean; Kerrigan, Richard W; Seidl, Michelle T; Aanen, Duur K; DeNitis, Matthew; Daniele, Graciela M; Desjardin, Dennis E; Kropp, Bradley R; Norvell, Lorelei L; Parker, Andrew; Vellinga, Else C; Vilgalys, Rytas; Hibbett, David S

    2006-01-01

    An overview of the phylogeny of the Agaricales is presented based on a multilocus analysis of a six-gene region supermatrix. Bayesian analyses of 5611 nucleotide characters of rpb1, rpb1-intron 2, rpb2 and 18S, 25S, and 5.8S ribosomal RNA genes recovered six major clades, which are recognized informally and labeled the Agaricoid, Tricholomatoid, Marasmioid, Pluteoid, Hygrophoroid and Plicaturopsidoid clades. Each clade is discussed in terms of key morphological and ecological traits. At least 11 origins of the ectomycorrhizal habit appear to have evolved in the Agaricales, with possibly as many as nine origins in the Agaricoid plus Tricholomatoid clade alone. A family-based phylogenetic classification is sketched for the Agaricales, in which 30 families, four unplaced tribes and two informally named clades are recognized.

  20. Characterization of twelve novel microsatellite markers of Sogatella furcifera (Horváth) (Hemiptera: Delphacidae) identified from next generation sequence data

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The white-backed planthopper, Sogatella furcifera (Horváth) (Hemiptera: Delphacidae), is a major pest of rice and has long-range migratory behavior in Asia. Microsatellite markers (simple sequence repeats, SSRs) have been widely used to determine the origins and genetic diversity of insect pests. ...

  1. Development of 304 new microsatellite markers for carrot. Analysis of their potential for linkage mapping, assessment of genetic diversity and cross-taxa utilization

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Two different approaches were used to isolate carrot SSRs: 1) Construction and analysis of a genomic DNA library enriched for SSR loci (GSSRs) and 2) Bioinformatic mining for SSR motifs in a 1.7 Mb BAC-end sequence database (BSSR). The SSR-enriched library yielded microsatellites with more repeats b...

  2. Phylogenetic inference of Indian malaria vectors from multilocus DNA sequences.

    PubMed

    Dixit, Jyotsana; Srivastava, Hemlata; Sharma, Meenu; Das, Manoj K; Singh, O P; Raghavendra, K; Nanda, Nutan; Dash, Aditya P; Saksena, D N; Das, Aparup

    2010-08-01

    Inferences on the taxonomic positions, phylogenetic interrelationships and divergence time among closely related species of medical importance is essential to understand evolutionary patterns among species, and based on which, disease control measures could be devised. To this respect, malaria is one of the important mosquito borne diseases of tropical and sub-tropical parts of the globe. Taxonomic status of malaria vectors has been so far documented based on morphological, cytological and few molecular genetic features. However, utilization of multilocus DNA sequences in phylogenetic inferences are still in dearth. India contains one of the richest resources of mosquito species diversity but little molecular taxonomic information is available in Indian malaria vectors. We herewith utilized the whole genome sequence information of An. gambiae to amplify and sequence three orthologous nuclear genetic regions in six Indian malaria vector species (An. culicifacies, An. minimus, An. sundaicus, An. fluviatilis, An. annularis and An. stephensi). Further, we utilized the previously published DNA sequence information on the COII and ITS2 genes in all the six species, making the total number of loci to five. Multilocus molecular phylogenetic study of Indian anophelines and An. gambiae was conducted at each individual genetic region using Neighbour Joining (NJ), Maximum Likelihood (ML), Maximum Parsimony (MP) and Bayesian approaches. Although tree topologies with COII, and ITS2 genes were similar, for no other three genetic regions similar tree topologies were observed. In general, the reconstructed phylogenetic status of Indian malaria vectors follows the pattern based on morphological and cytological classifications that was reconfirmed with COII and ITS2 genetic regions. Further, divergence times based on COII gene sequences were estimated among the seven Anopheles species which corroborate the earlier hypothesis on the radiation of different species of the Anopheles

  3. Genomic sequencing and microsatellite marker development for Boswellia papyrifera, an economically important but threatened tree native to dry tropical forests

    PubMed Central

    Addisalem, A. B.; Esselink, G. Danny; Bongers, F.; Smulders, M. J. M.

    2015-01-01

    Microsatellite (or simple sequence repeat, SSR) markers are highly informative DNA markers often used in conservation genetic research. Next-generation sequencing enables efficient development of large numbers of SSR markers at lower costs. Boswellia papyrifera is an economically important tree species used for frankincense production, an aromatic resinous gum exudate from bark. It grows in dry tropical forests in Africa and is threatened by a lack of rejuvenation. To help guide conservation efforts for this endangered species, we conducted an analysis of its genomic DNA sequences using Illumina paired-end sequencing. The genome size was estimated at 705 Mb per haploid genome. The reads contained one microsatellite repeat per 5.7 kb. Based on a subset of these repeats, we developed 46 polymorphic SSR markers that amplified 2–12 alleles in 10 genotypes. This set included 30 trinucleotide repeat markers, four tetranucleotide repeat markers, six pentanucleotide markers and six hexanucleotide repeat markers. Several markers were cross-transferable to Boswellia pirrotae and B. popoviana. In addition, retrotransposons were identified, the reads were assembled and several contigs were identified with similarity to genes of the terpene and terpenoid backbone synthesis pathways, which form the major constituents of the bark resin. PMID:25573702

  4. Genome-wide identification of microsatellites in white clover (Trifolium repens L.) using FIASCO and phpSSRMiner

    PubMed Central

    Zhang, Yan; He, Ji; Zhao, Patrick X; Bouton, Joseph H; Monteros, Maria J

    2008-01-01

    Background Allotetraploid white clover (Trifolium repens L.) is an important forage legume widely cultivated in most temperate regions. Only a small number of microsatellite markers are publicly available and can be utilized in white clover breeding programs. The objectives of this study were to develop an integrated approach for microsatellite development and to evaluate the approach for the development of new SSR markers for white clover. Results Genomic libraries containing simple sequence repeat (SSR) sequences were constructed using a modified Fast Isolation by AFLP of Sequences COntaining repeats (FIASCO) procedure and phpSSRMiner was used to develop the microsatellite markers. SSR motifs were isolated using two biotin-labeled probes, (CA)17 and (ATG)12. The sequences of 6,816 clones were assembled into 1,698 contigs, 32% of which represented novel sequences based on BLASTN searches. Approximately 32%, 28%, and 16% of these SSRs contained hexa-, tri-, and di-nucleotide repeats, respectively. The most frequent motifs were the CA and ATG complementary repeats and the associated compound sequences. Primer pairs were designed for 859 SSR loci based on sequences from these genomic libraries and from GenBank white clover nucleotide sequences. A total of 191 SSR primers developed from the two libraries were tested for polymorphism in individual clones from the parental genotypes GA43 ('Durana'), 'SRVR' and six F1 progeny from a mapping population. Ninety two percent produced amplicons and 66% of these were polymorphic. Conclusion The combined approach of identifying SSR-enriched fragments by FIASCO coupled with the primer design and in silico amplification using phpSSRMiner represents an efficient and low cost pipeline for the large-scale development of microsatellite markers in plants. The approach described here could be readily adapted and utilized in other non-related species with none or limited genomic resources. PMID:18631390

  5. A microsatellite marker for studying the ecology and diversity of fungal endophytes (Epichloë spp.) in grasses.

    PubMed Central

    Groppe, K; Sanders, I; Wiemken, A; Boller, T

    1995-01-01

    Randomly amplified polymorphic DNA fingerprinting, which is based on PCR with arbitrary 10-nucleotide primers, were used to analyze genetic diversity among isolates of the endophytic ascomycete Epichloë typhina, which were collected at a single field site from a population of one of its hosts, the grass Bromus erectus. One of the polymorphic randomly amplified polymorphic DNA PCR products occurred in all isolates as single bands with different but closely related sizes. Two of the size variants of this product were cloned and sequenced, and they were found to represent the same DNA sequence, except for a stretch of tandem repeats of the trinucleotide AAG.TTC, which differed in size, consisting of 8 and 18 repeats, respectively. Tandem repeats of this type are called microsatellites. Oligonucleotides were synthesized corresponding to portions of the sequence flanking the microsatellite and were used for PCR amplification of the loci from the genomic DNAs of different Epichloë isolates. A single PCR product was found for most isolates, indicating that the sequence represented a single genetic locus. Five alleles that could clearly be distinguished in size were found in a population of 91 field isolates. PCR with (AAC)8 and (AAG)8 as primers yielded a number of amplified bands from genomic DNA of Epichloë isolates, indicating that these types of microsatellites occur frequently in the genome of this fungus. A survey of all fungal DNA sequences currently deposited in the DNA sequence databases of EMBL and GenBank revealed that microsatellites of different repeating units are widespread in fungi.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:8526508

  6. New microsatellite markers developed from Urochloa humidicola (Poaceae) and cross amplification in different Urochloa species

    PubMed Central

    2011-01-01

    Background Urochloa humidicola is a forage grass that grows in tropical regions and is recognized for its tolerance to seasonal flooding. It is a polyploid and apomictic species with high phenotypic plasticity. As molecular tools are important in facilitating the development of new cultivars and in the classification of related species, the objectives of this study were to develop new polymorphic microsatellite markers from an enriched library constructed from U. humidicola and to evaluate their transferability to other Urochloa species. Findings Microsatellite sequences were identified from a previously constructed enriched library, and specific primers were designed for 40 loci. Isolated di-nucleotide repeat motifs were the most abundant followed by tetra-nucleotide repeats. Of the tested loci, 38 displayed polymorphism when screened across 34 polyploid Urochloa sp. genotypes, including 20 accessions and six hybrids of U. humidicola and two accessions each from U. brizantha, U. dictyoneura, U. decumbens and U. ruziziensis. The number of bands per Simple Sequence Repeat (SSR) locus ranged from one to 29 with a mean of 11.5 bands per locus. The mean Polymorphism Information Content (PIC) of all loci was 0.7136, and the mean Discrimination Power (DP) was 0.7873. Six loci amplified in all species tested. STRUCTURE analysis revealed six different allelic pools, and the genetic similarity values analyzed using Jaccard's coefficient ranged from 0.000 to 0.913. Conclusions This work reports new polymorphic microsatellite markers that will be useful for breeding programs for Urochloa humidicola and other Urochloa species as well as for genetic map development, germplasm characterization, evolutionary and taxonomic studies and marker-assisted trait selection. PMID:22142493

  7. Microsatellite Instability Assay — EDRN Public Portal

    Cancer.gov

    Microsatellite analysis (MSA) is a promising new technique for the surveillance of bladder cancer. The technology, which permits the separation by electrophoresis of polymerase chain reaction (PCR)-amplified sequences from non-malignant and malignant sources, has been applied to the diagnosis of solid tumors arising in colon, lung, oropharynx, kidney and bladder. MSA can detect genetic changes indicative of carcinoma from urothelial cells obtained in voided urine specimens. The genetic profile of DNA purified from urine is compared to that of DNA purified from peripheral lymphocytes that are considered normal. Once the DNA from uroepithelial cells has been obtained, PCR is performed with specific oligonucleotide primers for each chromosomal locus. The PCR products are then examined for evidence of microsatellite instability (MSI) and loss of heterozygosity (LOH), which are genetic characteristics of epithelial tumors. Preliminary work shows that MSA detects 95% of cancers.

  8. Microsatellite data analysis for population genetics.

    PubMed

    Kim, Kyung Seok; Sappington, Thomas W

    2013-01-01

    Theories and analytical tools of population genetics have been widely applied for addressing various questions in the fields of ecological genetics, conservation biology, and any context where the role of dispersal or gene flow is important. Underlying much of population genetics is the analysis of variation at selectively neutral marker loci, and microsatellites continue to be a popular choice of marker. In recent decades, software programs to estimate population genetics parameters have been developed at an increasing pace as computational science and theoretical knowledge advance. Numerous population genetics software programs are presently available to analyze microsatellite genotype data, but only a handful are commonly employed for calculating parameters such as genetic variation, genetic structure, patterns of spatial and temporal gene flow, population demography, individual population assignment, and genetic relationships within and between populations. In this chapter, we introduce statistical analyses and relevant population genetic software programs that are commonly employed in the field of population genetics and molecular ecology.

  9. Autosomal microsatellite data from Northwestern Colombia.

    PubMed

    Palacio, Oscar Darío; Triana, Omar; Gaviria, Aníbal; Ibarra, Adriana Alexandra; Ochoa, Luz Mariela; Posada, Yeny; Maya, María Clara; Lareu, María Victoria; Brión, María; Acosta, María Amparo; Carracedo, Angel

    2006-07-13

    Allele frequencies and some forensic parameters for 12 autosomal microsatellites (CSF1PO, TPOX, THO1, VWA, D16S539, D7S820, D13S317, D5S818, F13A1, FESFPS, F13B, LPL) were estimated from three departments from Northwestern Colombia. The total number of samples analysed was 1045 individuals. Comparative analysis among the three studied departments and with other published Colombian populations were also performed and discussed.

  10. Spatio-temporal Genetic Structuring of Leishmania major in Tunisia by Microsatellite Analysis

    PubMed Central

    Harrabi, Myriam; Bettaieb, Jihène; Ghawar, Wissem; Toumi, Amine; Zaâtour, Amor; Yazidi, Rihab; Chaâbane, Sana; Chalghaf, Bilel; Hide, Mallorie; Bañuls, Anne-Laure; Ben Salah, Afif

    2015-01-01

    In Tunisia, cases of zoonotic cutaneous leishmaniasis caused by Leishmania major are increasing and spreading from the south-west to new areas in the center. To improve the current knowledge on L. major evolution and population dynamics, we performed multi-locus microsatellite typing of human isolates from Tunisian governorates where the disease is endemic (Gafsa, Kairouan and Sidi Bouzid governorates) and collected during two periods: 1991–1992 and 2008–2012. Analysis (F-statistics and Bayesian model-based approach) of the genotyping results of isolates collected in Sidi Bouzid in 1991–1992 and 2008–2012 shows that, over two decades, in the same area, Leishmania parasites evolved by generating genetically differentiated populations. The genetic patterns of 2008–2012 isolates from the three governorates indicate that L. major populations did not spread gradually from the south to the center of Tunisia, according to a geographical gradient, suggesting that human activities might be the source of the disease expansion. The genotype analysis also suggests previous (Bayesian model-based approach) and current (F-statistics) flows of genotypes between governorates and districts. Human activities as well as reservoir dynamics and the effects of environmental changes could explain how the disease progresses. This study provides new insights into the evolution and spread of L. major in Tunisia that might improve our understanding of the parasite flow between geographically and temporally distinct populations. PMID:26302440

  11. Reduced microsatellite heterozygosity does not affect natal dispersal in three contrasting roe deer populations.

    PubMed

    Vanpé, Cécile; Debeffe, Lucie; Hewison, A J Mark; Quéméré, Erwan; Lemaître, Jean-François; Galan, Maxime; Amblard, Britany; Klein, François; Cargnelutti, Bruno; Capron, Gilles; Merlet, Joël; Warnant, Claude; Gaillard, Jean-Michel

    2015-03-01

    Although theoretical studies have predicted a link between individual multilocus heterozygosity and dispersal, few empirical studies have investigated the effect of individual heterozygosity on dispersal propensity or distance. We investigated this link using measures of heterozygosity at 12 putatively neutral microsatellite markers and natal dispersal behaviour in three contrasting populations of European roe deer (Capreolus capreolus), a species displaying pre-saturation condition-dependent natal dispersal. We found no effect of individual heterozygosity on either dispersal propensity or dispersal distance. Average heterozygosity was similar across the three studied populations, but dispersal propensity and distance differed markedly among them. In Aurignac, dispersal propensity and distance were positively related to individual body mass, whereas there was no detectable effect of body mass on dispersal behaviour in Chizé and Trois Fontaines. We suggest that we should expect both dispersal propensity and distance to be greater when heterozygosity is lower only in those species where dispersal behaviour is driven by density-dependent competition for resources.

  12. Genetic characterization of Mytilus coruscus and M. galloprovincialis using microsatellite markers.

    PubMed

    Kang, J H; Lee, J M; Noh, E S; Park, J Y; An, C M

    2013-11-13

    Korean (hard-shelled) mussels (Mytilus coruscus) are an economically important endemic marine bivalve mollusk of Korea; yet, the population has rapidly declined because of overharvesting and habitat competition from the invasive Mytilus galloprovincialis species. The population structures of M. coruscus and M. galloprovincialis were analyzed by next-generation sequencing using 5 microsatellite markers specifically developed for M. coruscus. M. galloprovincialis had an average of 5.4 alleles per locus (range = 2-10), with an average allelic richness of 4.9 per locus (range = 2.0-9.3). M. coruscus had an average of 5.7 alleles per locus (range = 2-13), with an average allelic richness of 5.2 per locus (range = 2.0-11.9). Excessive homozygosity was observed at 3 loci, which was assumed to be due to the presence of null alleles at these loci. Pairwise multilocus FST estimates showed that the M. coruscus and M. galloprovincialis populations were clearly separated. Six populations of M. galloprovincialis from the western, eastern, and southern coast of Korea formed 2 separate clusters, indicating that more than 2 populations of M. galloprovincialis have been introduced to the Korean Peninsula. Hybrids between M. coruscus and M. galloprovincialis were not identified, probably because of genetic differences or different habitat preferences. Further genetic information is required to perform selective breeding, population management, and restoration of M. coruscus.

  13. Estimating black bear population density and genetic diversity at Tensas River, Louisiana using microsatellite DNA markers

    USGS Publications Warehouse

    Boersen, Mark R.; Clark, Joseph D.; King, Tim L.

    2003-01-01

    The Recovery Plan for the federally threatened Louisiana black bear (Ursus americanus luteolus) mandates that remnant populations be estimated and monitored. In 1999 we obtained genetic material with barbed-wire hair traps to estimate bear population size and genetic diversity at the 329-km2 Tensas River Tract, Louisiana. We constructed and monitored 122 hair traps, which produced 1,939 hair samples. Of those, we randomly selected 116 subsamples for genetic analysis and used up to 12 microsatellite DNA markers to obtain multilocus genotypes for 58 individuals. We used Program CAPTURE to compute estimates of population size using multiple mark-recapture models. The area of study was almost entirely circumscribed by agricultural land, thus the population was geographically closed. Also, study-area boundaries were biologically discreet, enabling us to accurately estimate population density. Using model Chao Mh to account for possible effects of individual heterogeneity in capture probabilities, we estimated the population size to be 119 (SE=29.4) bears, or 0.36 bears/km2. We were forced to examine a substantial number of loci to differentiate between some individuals because of low genetic variation. Despite the probable introduction of genes from Minnesota bears in the 1960s, the isolated population at Tensas exhibited characteristics consistent with inbreeding and genetic drift. Consequently, the effective population size at Tensas may be as few as 32, which warrants continued monitoring or possibly genetic augmentation.

  14. Genetic diversity, paraphyly and incomplete lineage sorting of mtDNA, ITS2 and microsatellite flanking region in closely related Heliopora species (Octocorallia).

    PubMed

    Yasuda, Nina; Taquet, Coralie; Nagai, Satoshi; Fortes, Miguel; Fan, Tung-Yung; Harii, Saki; Yoshida, Terutoyo; Sito, Yuta; Nadaoka, Kazuo

    2015-12-01

    Examining genetic diversity and lineage sorting of different genes in closely related species provide useful information for phylogenetic analyses and ultimately for understanding the origins of biodiversity. In this study, we examined inter- and intraspecific genetic variation in internal transcribed spacer 2 (ITS2), partial mitochondrial gene (mtMutS), and nuclear microsatellite flanking region in two closely related octocoral species (Heliopora coerulea, HC-A and HC-B). These species were recently identified in a population genetic study using microsatellite markers. The two species have different reproductive timing, which ecologically promotes lineage sorting. In this study, we examined whether species boundaries could be detected by the commonly used nuclear ITS2 and mtMutS, as well as by possibly neutral microsatellite flanking sequences. Haplotype network analysis of microsatellite flanking region revealed that a possible ancestral haplotype was still shared between the two species, indicating on-going lineage sorting. Haplotype network analysis of ITS2 and microsatellite flanking region revealed shared haplotypes between the two lineages. The two species shared fewer ITS2 sequences than microsatellite flanking region sequences. The almost fixed point mutation at the tip of helix 3 of ITS2 was not associated with the secondary structure or compensatory base changes (CBCs). The phylogenetic tree of ITS2 showed paraphyly and that of the microsatellite flanking region indicated that lineage sorting for the two species may be incomplete. Much higher intra- and inter-individual variation of ITS2 was observed in HC-B than that in HC-A, highlighting the importance of examining ITS2 from multiple individuals to estimate genetic diversity. The mitochondrial mtMutS gene sequences from 39 individuals, including both species collected from Japan and Taiwan, showed no variation because of slow rates of mitochondrial nucleotide substitution. This study suggests caution

  15. A Genome-Wide Scan of Selective Sweeps and Association Mapping of Fruit Traits Using Microsatellite Markers in Watermelon

    PubMed Central

    Reddy, Umesh K.; Abburi, Lavanya; Abburi, Venkata Lakshmi; Saminathan, Thangasamy; Cantrell, Robert; Vajja, Venkata Gopinath; Reddy, Rishi; Tomason, Yan R.; Levi, Amnon; Wehner, Todd C.; Nimmakayala, Padma

    2015-01-01

    Our genetic diversity study uses microsatellites of known map position to estimate genome level population structure and linkage disequilibrium, and to identify genomic regions that have undergone selection during watermelon domestication and improvement. Thirty regions that showed evidence of selective sweep were scanned for the presence of candidate genes using the watermelon genome browser (www.icugi.org). We localized selective sweeps in intergenic regions, close to the promoters, and within the exons and introns of various genes. This study provided an evidence of convergent evolution for the presence of diverse ecotypes with special reference to American and European ecotypes. Our search for location of linked markers in the whole-genome draft sequence revealed that BVWS00358, a GA repeat microsatellite, is the GAGA type transcription factor located in the 5′ untranslated regions of a structure and insertion element that expresses a Cys2His2 Zinc finger motif, with presumed biological processes related to chitin response and transcriptional regulation. In addition, BVWS01708, an ATT repeat microsatellite, located in the promoter of a DTW domain-containing protein (Cla002761); and 2 other simple sequence repeats that association mapping link to fruit length and rind thickness. PMID:25425675

  16. Allele frequencies of microsatellite loci for genetic characterization of a Sicilian bovine population.

    PubMed

    Cosenza, M; Reale, S; Lupo, T; Vitale, F; Caracappa, S

    2015-01-30

    Short tandem repeats are used as an effective method to trace DNA markers in genotyping. Using a standardized kit, we tested 11 microsatellite markers recommended by the International Society for Animal Genetics (ISAG) in a sample of 495 Sicilian cattle. The aim of this study was to investigate the allele frequencies in the Sicilian cattle population to provide a reference database and at the same time to assess the use of the ISAG microsatellite panel for pedigree analysis. DNA samples were collected from blood and amplified in an 11-plex polymerase chain reaction (PCR); PCR products were injected in a 3130 Genetic Analyzer. All loci showed high mean polymorphism information content (0.768), and the observed mean heterozygosity was less than the expected value (0.732 vs 0.794, respectively). The exact test for Hardy-Weinberg proportions, allele number, and inbreeding coefficient were calculated. Our results indicated that equilibrium was not always maintained. The observed mean homozygote value exceeded the expected value (132.81 vs 102.14), but no evidence for allele dropout was found. These results could be explained by a non-random mating; further studies using a larger number of animals could confirm or invalidate this hypothesis. The probability of identity and exclusion of a locus were also estimated and proved to be useful in paternity testing. The ISAG microsatellite panel is useful to screen the Sicilian bovine kinship. Currently, an allele frequency database is being constructed.

  17. A web-based microsatellite database for the Magnaporthe oryzae genome

    PubMed Central

    Singh, Pankaj Kumar; Singh, Akshay; Pawar, Deepak V.; Devanna, B. N.; Singh, Jyoti; Sharma, Vinay; Sharma, Tilak R.

    2016-01-01

    Microsatellites have been widely utilized for molecular marker development. Codominant and multiallelic nature of these simple repeats have several advantages over other types of molecular markers. Their broad applicability in the area of molecular biology like gene mapping, genome characterization, genome evolution, and gene regulation has been reported in various crop plants, animals and fungi. Considering these benefits of the SSR markers, a MMDB (Magnaporthe oryzae Microsatellite Database) was developed to help in understanding about the pathogen and its diversity at strains level of a particular geographic region, which can help us to make a proper utilization of blast resistance genes in the region. This microsatellite database is based on whole genome sequence of two M. oryzae isolates, RML-29 (2665 SSRs from 43037792 bp) and RP-2421 (3169 SSRs from 45510614 bp). Although, first M. oryzae genome (70-15) was sequenced in 2005, but this sequenced isolate is not a true field isolate of M. oryzae. Therefore, MMDB has great potential in the study of diversification and characterization of M. oryzae and other related fungi. Availability: http://14.139.229.199/home.aspx PMID:28293068

  18. Somatic microsatellite variability as a predictive marker for colorectal cancer and liver cancer progression

    PubMed Central

    Vaksman, Zalman; Garner, Harold R.

    2015-01-01

    Microsatellites (MSTs) are short tandem repeated genetic motifs that comprise ~3% of the genome. MST instability (MSI), defined as acquired/lost primary alleles at a small subset of microsatellite loci (e.g. Bethesda markers), is a clinically relevant marker for colorectal cancer. However, these markers are not applicable to other types of cancers, specifically, for liver cancer which has a high mortality rate. Here we show that somatic MST variability (SMV), defined as the presence of additional, non-primary (aka minor) alleles at MST loci, is a complementary measure of MSI, and a genetic marker for colorectal and liver cancer. Re-analysis of Illumina sequenced exomes from The Cancer Genome Atlas indicates that SMV may distinguish a subpopulation of African American patients with colorectal cancer, which represents ~33% of the population in this study. Further, for liver cancer, a higher rate of SMV may be indicative of an earlier age of onset. The work presented here suggests that classical MSI should be expanded to include SMV, going beyond alterations of the primary alleles at a small number of microsatellite loci. This measure of SMV may represent a potential new diagnostic for a variety of cancers and may provide new information for colorectal cancer patients. PMID:25691061

  19. Isolation and characterization of microsatellite markers for Axonopus compressus (Sw.) Beauv. (Poaceae) using 454 sequencing technology.

    PubMed

    Wang, X-L; Li, Y; Liao, L; Bai, C-J; Wang, Z-Y

    2015-05-11

    Axonopus compressus (Sw.) Beauv. is a perennial herb widely used as a garden lawn grass. In this study, we used Roche 454 pyrosequencing, combined with the magnetic bead enrichment method FIASCO, to isolate simple sequence repeat markers from the A. compressus genome. A total of 1942 microsatellite loci were identified, with 53,193 raw sequencing reads. One hundred micro-satellite loci were selected to test the primer amplification efficiency in 24 individuals; 14 primer pairs yielded polymorphic amplification products. The number of observed alleles ranged from two to six, with an average of 3.5. Shannon's Information index values ranged from 0.169 to 0.650, with an average of 0.393. Nei's genetic diversity values ranged from 0.108 to 0.457, with an average of 0.271. This first set of microsatellite markers developed for Axonopus will assist in the development of molecular marker-assisted breeding and the assessment of genetic diversity in A. compressus.

  20. Development of polymorphic microsatellite markers issued from pyrosequencing technology for the medicinal mushroom Agaricus subrufescens.

    PubMed

    Foulongne-Oriol, Marie; Spataro, Cathy; Moinard, Magalie; Cabannes, Delphine; Callac, Philippe; Savoie, Jean-Michel

    2012-09-01

    The recently described procedure of microsatellite-enriched library pyrosequencing was used to isolate microsatellite loci in the gourmet and medicinal mushroom Agaricus subrufescens. Three hundred and five candidate loci containing at least one simple sequence repeats (SSR) locus and for which primers design was successful, were obtained. From a subset of 95 loci, 35 operational and polymorphic SSR markers were developed and characterized on a sample of 14 A. subrufescens genotypes from diverse origins. These SubSSR markers each displayed from two to 10 alleles with an average of 4.66 alleles per locus. The observed heterozygosity ranged from 0 to 0.71. Several multiplex combinations can be set up, making it possible to genotype up to six markers easily and simultaneously. Cross-amplification in some closely congeneric species was successful for a subset of loci. The 35 microsatellite markers developed here provide a highly valuable molecular tool to study genetic diversity and reproductive biology of A. subrufescens.

  1. Polymorphic microsatellite loci identified through development and cross-species amplification within shorebirds

    USGS Publications Warehouse

    Williams, I.; Guzzetti, B.M.; Gust, Judy R.; Sage, G.K.; Gill, R.E.; Tibbitts, T.L.; Sonsthagen, S.A.; Talbot, S.L.

    2012-01-01

    We developed microsatellite loci for demographic assessments of shorebirds, a group with limited markers. First, we isolated five dinucleotide repeat microsatellite loci from the Black Oystercatcher (Haematopodidae: Haematopus bachmani), and three from the Bristle-thighed Curlew (Scolopacidae: Numenius tahitiensis); both species are of conservation concern. All eight loci were polymorphic in their respective target species. Hbaμ loci were characterized by two to three alleles with observed heterozygosity ranging from 0.07 to 0.33, and two to nine alleles were detected for Nut loci with observed heterozygosity ranging from 0.08 to 0.72. No linkage disequilibrium or departures from Hardy–Weinberg equilibrium were observed. The eight loci were also tested for cross-species amplification in 12 other species within Charadriidae and Scolopacidae, and the results demonstrated transferability across several genera. We further tested all 14 species at 12 additional microsatellite markers developed for other shorebirds: Dunlin (Calidris alpina; four loci) and Ruff (Philomachus pugnax; eight loci). Two markers (Hbaμ4 and Ruff6) were polymorphic in 13 species, while two (Calp6 and Ruff9) were monomorphic. The remaining eight markers revealed polymorphism in one to nine species each. Our results provide further evidence that locus Ruff10 is sex-linked, contrary to the initial description. These markers can be used to enhance our understanding of shorebird biology by, for example, helping to determine migratory connectivity among breeding and wintering populations and detecting relatedness among individuals.

  2. A multilocus perspective on the speciation history of a North American aridland toad (Anaxyrus punctatus).

    PubMed

    Bryson, Robert W; Jaeger, Jef R; Lemos-Espinal, Julio A; Lazcano, David

    2012-09-01

    Interpretations of phylogeographic patterns can change when analyses shift from single gene-tree to multilocus coalescent analyses. Using multilocus coalescent approaches, a species tree and divergence times can be estimated from a set of gene trees while accounting for gene-tree stochasticity. We utilized the conceptual strengths of a multilocus coalescent approach coupled with complete range-wide sampling to examine the speciation history of a broadly distributed, North American warm-desert toad, Anaxyrus punctatus. Phylogenetic analyses provided strong support for three major lineages within A. punctatus. Each lineage broadly corresponded to one of three desert regions. Early speciation in A. punctatus appeared linked to late Miocene-Pliocene development of the Baja California peninsula. This event was likely followed by a Pleistocene divergence associated with the separation of the Chihuahuan and Sonoran Deserts. Our multilocus coalescent-based reconstruction provides an informative contrast to previous single gene-tree estimates of the evolutionary history of A. punctatus.

  3. Multilocus genotyping reveals high heterogeneity and strong local population structure of the Plasmodium vivax population in the Peruvian Amazon

    PubMed Central

    2010-01-01

    Background Peru is one of the Latin American countries with the highest malaria burden, mainly due to Plasmodium vivax infections. However, little is known about P. vivax transmission dynamics in the Peruvian Amazon, where most malaria cases occur. The genetic diversity and population structure of P. vivax isolates collected in different communities around Iquitos city, the capital of the Peruvian Amazon, was determined. Methods Plasmodium vivax population structure was determined by multilocus genotyping with 16 microsatellites on 159 P. vivax infected blood samples (mono-infections) collected in four sites around Iquitos city. The population characteristics were assessed only in samples with monoclonal infections (n = 94), and the genetic diversity was determined by calculating the expected heterozygosity and allelic richness. Both linkage disequilibrium and the genetic differentiation (θ) were estimated. Results The proportion of polyclonal infections varied substantially by site (11% - 70%), with the expected heterozygosity ranging between 0.44 and 0.69; no haplotypes were shared between the different populations. Linkage disequilibrium was present in all populations (IAS 0.14 - 0.61) but was higher in those with fewer polyclonal infections, suggesting inbreeding and a clonal population structure. Strong population differentiation (θ = 0.45) was found and the Bayesian inference cluster analysis identified six clusters based on distinctive allele frequencies. Conclusion The P. vivax populations circulating in the Peruvian Amazon basin are genetically diverse, strongly differentiated and they have a low effective recombination rate. These results are in line with the low and clustered pattern of malaria transmission observed in the region around Iquitos city. PMID:20525233

  4. Toward Microsatellite Based Space Situational Awareness

    NASA Astrophysics Data System (ADS)

    Scott, L.; Wallace, B.; Sale, M.; Thorsteinson, S.

    2013-09-01

    The NEOSSat microsatellite is a dual mission space telescope which will perform asteroid detection and Space Situational Awareness (SSA) observation experiments on deep space, earth orbiting objects. NEOSSat was launched on 25 February 2013 into a 800 dawn-dusk sun synchronous orbit and is currently undergoing satellite commissioning. The microsatellite consists of a small aperture optical telescope, GPS receiver, high performance attitude control system, and stray light rejection baffle designed to reject stray light from the Sun while searching for asteroids with elongations 45 degrees along the ecliptic. The SSA experimental mission, referred to as HEOSS (High Earth Orbit Space Surveillance), will focus on objects in deep space orbits. The HEOSS mission objective is to evaluate the utility of microsatellites to perform catalog maintenance observations of resident space objects in a manner consistent with the needs of the Canadian Forces. The advantages of placing a space surveillance sensor in low Earth orbit are that the observer can conduct observations without the day-night interruption cycle experienced by ground based telescopes, the telescope is insensitive to adverse weather and the system has visibility to deep space resident space objects which are not normally visible from ground based sensors. Also, from a photometric standpoint, the microsatellite is able to conduct observations on objects with a rapidly changing observer position. The possibility of spin axis estimation on geostationary satellites may be possible and an experiment characterize spin axis of distant resident space objects is being planned. Also, HEOSS offers the ability to conduct observations of satellites at high phase angles which can potentially extend the trackable portion of space in which deep space objects' orbits can be monitored. In this paper we describe the HEOSS SSA experimental data processing system and the preliminary findings of the catalog maintenance experiments

  5. Mining of expressed sequence tag libraries of cacao for microsatellite markers using five computational tools.

    PubMed

    Riju, Aikkal; Rajesh, M K; Sherin, P T P Fasila; Chandrasekar, A; Apshara, S Elain; Arunachalam, Vadivel

    2009-08-01

    Expressed sequence tags (ESTs) provide researchers with a quick and inexpensive route for discovering new genes, data on gene expression and regulation, and also provide genic markers that help in constructing genome maps. Cacao is an important perennial crop of humid tropics. Cacao EST sequences, as available in the public domain, were downloaded and made into contigs. Microsatellites were located in these ESTs and contigs using five softwares (MISA, TRA, TROLL, SSRIT and SSR primer). MISA gave maximum coverage of SSRs in cacao ESTs and contigs, although TRA was able to detect higher order (5-mer) repeats. The frequency of SSRs was one per 26.9 kb in the known set of ESTs. One-third of the repeats in EST-contigs were found to be trimeric. A few rare repeats like 21-mer repeat were also located. A/T repeats were most abundant among the mononucleotide repeats and the AG/GA/TC/CT type was the most frequent among dimerics. Flanking primers were designed using Primer3 program and verified experimentally for PCR amplification. The results of the study are made available freely online database (http://riju.byethost31.com/cocoa/). Seven primer pairs amplified genomic DNA isolated from leaves were used to screen a representative set of 12 accessions of cacao.

  6. Development and multiplexing of microsatellite markers using pyrosequencing in the clonal plant Comarum palustre (Rosaceae).

    PubMed

    Somme, L; Raabová, J; Jacquemart, A L; Raspé, O

    2012-01-01

    Microsatellites represent one of the most commonly used genetic markers for population genetic studies. Traditionally, their development is quite time consuming, requiring construction of a genomic library enriched for repeated motifs. Using pyrosequencing, a fast and cost-effective new generation sequencing technique, we produced 24,340,862 bases in 63,860 short fragment reads, including 1170 dinucleotide motifs with a minimum of six repeats and 1383 trinucleotide motifs with a minimum of four repeats for the Marsh Cinquefoil, Comarum palustre L., an endangered marsh pioneer species. We selected 58 loci with SSR (Short Sequence Repeat) segments (at least 10 repeats) for a preliminary screening. Out of them, we screened 29 loci on a capillary sequencer after ligation in a vector and PCR using T7 forward primer labelled with FAM fluorescent dye and the specific unlabeled reverse primers. This procedure allowed us to screen large number of candidate loci with the same labelled primer and unlabelled specific primers. Finally, we characterized 20 polymorphic microsatellite markers, nine dinucleotides and 11 trinucleotides. We used these markers to assess genetic diversity and clonal structure in two Belgian populations. All loci showed a maximum of two alleles per individual, suggesting that they are from a diploid genome. One genet was detected in a newly extending population while 53 different genets in a long-term ecologically managed population. The number of alleles per locus ranged from 6 to 14 in this old population with an expected heterozygosity, ranging from 0.5964 to 0.8278. These preliminary results show a genet size up to 7.2 m.

  7. Genome-wide characterization of perfect microsatellites in yak (Bos grunniens).

    PubMed

    Ma, Zhijie

    2015-08-01

    Microsatellites or simple sequence repeats (SSRs) constitute a significant portion of genomes and play an important role in gene function and genome organization. The availability of a complete genome sequence for yak (Bos grunniens) has made it possible to carry out genome-wide analysis of microsatellites in this species. We analyzed the abundance and density of perfect SSRs in the yak genome. We found a total of 723,172 SSRs with 1-6 bp nucleotide motifs, indicating that about 0.47 % of the yak whole genome sequence (2.66 Gb) comprises perfect SSRs, the average length of which was 17.34 bp/Mb. The average frequency and density of perfect SSRs was 272.18 loci/Mb and 4719.25 bp/Mb, respectively. The proportion of the six classes of perfect SSRs was not evenly distributed in the yak genome. Mononucleotide repeats (44.04 %) with a total number of 318,435 and a average length of 14.71 bp appeared to be the most abundant SSRs class, while the percentages of dinucleotide, trinucleotide, pentanucleotide, tetranucleotide and hexanucleotide repeats was 24.11 %, 15.80 %, 9.50 %, 6.40 % and 0.15 %, respectively. Different repeat classes of SSRs varied in their repeat number with the highest being 1206. Our results suggest that 15 motifs comprised the predominant categories with a frequency above 1 loci/Mb: A, AC, AT, AG, AGC, AAC, AAT, ACC, ATTT, GTTT, AATG, CTTT, ATGG, AACTG and ATCTG.

  8. Development of microsatellite loci in Artocarpus altilis (Moraceae) and cross-amplification in congeneric species

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Microsatellite loci were isolated and characterized from enriched genomic libraries of Artocarpus altilis (breadfruit) and tested in three other Artocarpus species and one hybrid. The microsatellite markers provide new tools for further studies in Artocarpus. Nineteen microsatellite primers were tes...

  9. Biogeography of Paenibacillus larvae, the causative agent of American foulbrood, using a new multilocus sequence typing scheme

    PubMed Central

    Morrissey, Barbara J; Helgason, Thorunn; Poppinga, Lena; Fünfhaus, Anne; Genersch, Elke; Budge, Giles E

    2015-01-01

    American foulbrood is the most destructive brood disease of honeybees (Apis mellifera) globally. The absence of a repeatable, universal typing scheme for the causative bacterium Paenibacillus larvae has restricted our understanding of disease epidemiology. We have created the first multilocus sequence typing scheme (MLST) for P. larvae, which largely confirms the previous enterobacterial repetitive intergenic consensus (ERIC)–polymerase chain reaction (PCR)-based typing scheme's divisions while providing added resolution and improved repeatability. We have used the new scheme to determine the distribution and biogeography of 294 samples of P. larvae from across six continents. We found that of the two most epidemiologically important ERIC types, ERIC I was more diverse than ERIC II. Analysis of the fixation index (FST) by distance suggested a significant relationship between genetic and geographic distance, suggesting that population structure exists in populations of P. larvae. Interestingly, this effect was only observed within the native range of the host and was absent in areas where international trade has moved honeybees and their disease. Correspondence analysis demonstrated similar sequence type (ST) distributions between native and non-native countries and that ERIC I and II STs mainly have differing distributions. The new typing scheme facilitates epidemiological study of this costly disease of a key pollinator. PMID:25244044

  10. Development of novel polymorphic microsatellite markers in Siganus fuscescens.

    PubMed

    Mao, X Q; Li, Z B; Ning, Y F; Shangguan, J B; Yuan, Y; Huang, Y S; Li, B B

    2016-07-29

    Rabbitfish, Siganus fuscescens, is widely distributed in the Indo-Pacific regions and eastern Mediterranean. Its dwelling place includes reef flats, coral reef regions, and seagrass meadows in tropical area and reef areas or shallow waters in locations at high latitudes. In the present study, 10 new polymorphic microsatellite markers were screened from 30 wild S. fuscescens individuals, using a method of fast isolation protocol and amplified fragment length polymorphism of sequences containing repeats. The number of polymorphic alleles per locus was 3 to 5 with a mean of 4.3, while the value of polymorphic information content ranged from 0.283 to 0.680. The values of the observed and expected heterozygosities were in the range 0.3333-0.8462 and 0.3011-0.7424, respectively. Deviation from Hardy-Weinberg equilibrium was not observed in this study. These polymorphic loci are expected to be effective in evaluating the genetic diversity, population structure, and gene flow and in determining the paternity in S. fuscescens, as well as for conservation management.

  11. Allele Frequencies at Microsatellite Loci: The Stepwise Mutation Model Revisited

    PubMed Central

    Valdes, A. M.; Slatkin, M.; Freimer, N. B.

    1993-01-01

    We summarize available data on the frequencies of alleles at microsatellite loci in human populations and compare observed distributions of allele frequencies to those generated by a simulation of the stepwise mutation model. We show that observed frequency distributions at 108 loci are consistent with the results of the model under the assumption that mutations cause an increase or decrease in repeat number by one and under the condition that the product Nu, where N is the effective population size and u is the mutation rate, is larger than one. We show that the variance of the distribution of allele sizes is a useful estimator of Nu and performs much better than previously suggested estimators for the stepwise mutation model. In the data, there is no correlation between the mean and variance in allele size at a locus or between the number of alleles and mean allele size, which suggests that the mutation rate at these loci is independent of allele size. PMID:8454213

  12. A microsatellite genetic linkage map of human chromosome 13

    SciTech Connect

    Petrukhin, K.E.; Speer, M.C.; Vayanis, E.; Fatima Bonaldo, M. de; Soares, M.B.; Fischer, S.G.; Warburton, D. ); Gilliam, C.; Ott, J. New York State Psychiatric Institute, New York, NY ); Tantravahi, U. )

    1993-01-01

    We have characterized 21 polymorphic (CA), microsatellites for the development of a genetic map of chromosome 13. Fifteen markers were isolated from a flow- sorted chromosome 13 library, four CA repeats were derived from NotI-containing cosmid clones, and two polymorphic markers were described previously (J. L. Weber, A. E. Kwitek, and P. E. May, 1990, Nucleic Acids Res. IS: 4638; L. Warnich, 1. Groenwald, L. Laubscher, and A. E. Retief, 1991, Am. J. Hum. Genet. 49(Suppl.): 372 (Abstract)). Regional localization for all of the markers was performed by amplification of DNA from five somatic cell hybrids containing different deletions of chromosome 13. Genetic markers were shown to be distributed throughout 6 of the 11 resolvable chromosomal subregions. Using data from nine families provided by the Centre d'Etude du Polymorphisme Humain (CEPH), a framework map of 12 of these 21 markers was developed. Six of the 12 markers form three pairs, with each two members of a pair being tightly linked, such that nine systems of markers can be distinguished. The average heterozygosity of these 12 markers is 0.75. The total length of the sex-averaged map is 65.4 cM (Kosambi), with an average distance of 8.2 cM between systems of markers (eight intervals). Seven remaining markers were placed provisionally into the framework map. 41 refs., 3 figs., 4 tabs.

  13. Microsatellite Interruptions Stabilize Primate Genomes and Exist as Population-Specific Single Nucleotide Polymorphisms within Individual Human Genomes

    PubMed Central

    Ananda, Guruprasad; Hile, Suzanne E.; Breski, Amanda; Wang, Yanli; Kelkar, Yogeshwar; Makova, Kateryna D.; Eckert, Kristin A.

    2014-01-01

    Interruptions of microsatellite sequences impact genome evolution and can alter disease manifestation. However, human polymorphism levels at interrupted microsatellites (iMSs) are not known at a genome-wide scale, and the pathways for gaining interruptions are poorly understood. Using the 1000 Genomes Phase-1 variant call set, we interrogated mono-, di-, tri-, and tetranucleotide repeats up to 10 units in length. We detected ∼26,000–40,000 iMSs within each of four human population groups (African, European, East Asian, and American). We identified population-specific iMSs within exonic regions, and discovered that known disease-associated iMSs contain alleles present at differing frequencies among the populations. By analyzing longer microsatellites in primate genomes, we demonstrate that single interruptions result in a genome-wide average two- to six-fold reduction in microsatellite mutability, as compared with perfect microsatellites. Centrally located interruptions lowered mutability dramatically, by two to three orders of magnitude. Using a biochemical approach, we tested directly whether the mutability of a specific iMS is lower because of decreased DNA polymerase strand slippage errors. Modeling the adenomatous polyposis coli tumor suppressor gene sequence, we observed that a single base substitution interruption reduced strand slippage error rates five- to 50-fold, relative to a perfect repeat, during synthesis by DNA polymerases α, β, or η. Computationally, we demonstrate that iMSs arise primarily by base substitution mutations within individual human genomes. Our biochemical survey of human DNA polymerase α, β, δ, κ, and η error rates within certain microsatellites suggests that interruptions are created most frequently by low fidelity polymerases. Our combined computational and biochemical results demonstrate that iMSs are abundant in human genomes and are sources of population-specific genetic variation that may affect genome stability. The

  14. Multilocus phylogeography of the European ground squirrel: cryptic interglacial refugia of continental climate in Europe.

    PubMed

    Říčanová, Štěpánka; Koshev, Yordan; Říčan, Oldřich; Ćosić, Nada; Ćirović, Duško; Sedláček, František; Bryja, Josef

    2013-08-01

    The theory of classical and cryptic Pleistocene refugia is based mainly on historical changes in temperature, and the refugia are usually defined within a latitudinal gradient. However, the gradient of oceanic-continental climate (i.e. longitudinal) was also significantly variable during glacial cycles with important biotic consequences. Range-wide phylogeography of the European ground squirrel (EGS) was used to interpret the evolutionary and palaeogeographical history of the species in Europe and to shed light on its glacial-interglacial dynamic. The EGS is a steppe-inhabiting species and the westernmost member of the genus in the Palaearctic region. We have analysed 915 specimens throughout the present natural range by employing mitochondrial DNA sequences (cytochrome b gene) and 12 nuclear microsatellite markers. The reconstructed phylogeography divides the species into two main geographical groups, with deep substructuring within both groups. Bulgaria is the centre of the ancestral area, and it also has the highest genetic diversity within the species. The northernmost group of the EGS survived in the southern part of Pannonia throughout several glacial-interglacial cycles. Animals from this population probably repeatedly colonized areas further to the north and west during the glacial periods, while in the interglacial periods, the EGS distribution contracted back to this Pannonian refugium. The EGS thus represents a species with a glacial expansion/interglacial contraction palaeogeographical dynamics, and the Pannonian and southeastern Balkanian steppes are supported as cryptic refugia of continental climate during Pleistocene interglacials.

  15. An optimized microsatellite genotyping strategy for assessing genetic identity and kinship in Azara's owl monkeys (Aotus azarai).

    PubMed

    Babb, Paul L; McIntosh, Annick M; Fernandez-Duque, Eduardo; Di Fiore, Anthony; Schurr, Theodore G

    2011-01-01

    In this study, we characterize a panel of 20 microsatellite markers that reproducibly amplify in Azara's owl monkeys (Aotus azarai) for use in genetic profiling analyses. A total of 128 individuals from our study site in Formosa, Argentina, were genotyped for 20 markers, 13 of which were found to be polymorphic. The levels of allelic variation at these loci provided paternity exclusion probabilities of 0.852 when neither parent was known, and 0.981 when one parent was known. In addition, our analysis revealed that, although genotypes can be rapidly scored using fluorescence-based fragment analysis, the presence of complex or multiple short tandem repeat (STR) motifs at a microsatellite locus could generate similar fragment patterns from alleles that have different nucleotide sequences and perhaps different evolutionary origins. Even so, this collection of microsatellite loci is suitable for parentage analyses and will allow us to test various hypotheses about the relationship between social behavior and kinship in wild owl monkey populations. Furthermore, given the limited number of platyrrhine-specific microsatellite loci available in the literature, this STR panel represents a valuable tool for population studies of other cebines and callitrichines.

  16. Characterization of 12 Novel Microsatellite Markers of Sogatella furcifera (Hemiptera: Delphacidae) Identified From Next-Generation Sequence Data

    PubMed Central

    Nam, Hwa Yeun; Coates, Brad; Kim, Kyung Seok; Park, Marana; Lee, Joon-Ho

    2015-01-01

    The white-backed planthopper, Sogatella furcifera (Horváth) (Hemiptera: Delphacidae), is a major pest of rice and has long-range migratory behavior in Asia. Microsatellite markers (simple sequence repeats) have been widely used to determine the origins and genetic diversity of insect pests. We identified novel microsatellite loci for S. furcifera samples collected from Laos, Vietnam, and three localities in Bangladesh from next-generation Roche 454 pyrosequencing data. Size polymorphism at 12 microsatellite loci was verified for 40 adult individuals collected from Shinan, South Korea. The average number of alleles per locus was 7.92. The mean values of observed (Ho) and expected heterozygosities (HE) were 0.615 and 0.757, respectively. These new microsatellite markers will be a resource for future ecological genetic studies of S. furcifera samples across more broad geographic regions in Asia and may assist in estimations of genetic differentiation and gene flow among populations for implementation of more effective management strategies to control this serious rice pest. PMID:26163593

  17. Characterization of 12 Novel Microsatellite Markers of Sogatella furcifera (Hemiptera: Delphacidae) Identified From Next-Generation Sequence Data.

    PubMed

    Nam, Hwa Yeun; Coates, Brad; Kim, Kyung Seok; Park, Marana; Lee, Joon-Ho

    2015-01-01

    The white-backed planthopper, Sogatella furcifera (Horváth) (Hemiptera: Delphacidae), is a major pest of rice and has long-range migratory behavior in Asia. Microsatellite markers (simple sequence repeats) have been widely used to determine the origins and genetic diversity of insect pests. We identified novel microsatellite loci for S. furcifera samples collected from Laos, Vietnam, and three localities in Bangladesh from next-generation Roche 454 pyrosequencing data. Size polymorphism at 12 microsatellite loci was verified for 40 adult individuals collected from Shinan, South Korea. The average number of alleles per locus was 7.92. The mean values of observed (H(o)) and expected heterozygosities (H(E)) were 0.615 and 0.757, respectively. These new microsatellite markers will be a resource for future ecological genetic studies of S. furcifera samples across more broad geographic regions in Asia and may assist in estimations of genetic differentiation and gene flow among populations for implementation of more effective management strategies to control this serious rice pest.

  18. A genome-wide view of microsatellite instability: old stories of cancer mutations revisited with new sequencing technologies

    PubMed Central

    Kim, Tae-Min; Park, Peter J

    2014-01-01

    Microsatellites are simple tandem repeats that are present at millions of loci in the human genome. Microsatellite instability (MSI) refers to DNA slippage events on microsatellites that occur frequently in cancer genomes when there is a defect in the DNA mismatch repair system. These somatic mutations can result in inactivation of tumor suppressor genes or disrupt other non-coding regulatory sequences, thereby playing a role in carcinogenesis. Here, we will discuss the ways in which high-throughput sequencing data can facilitate a genome- or exome-wide discovery and more detailed investigation of MSI events in microsatellite-unstable cancer genomes. We will address the methodological aspects of this approach and highlight insights from recent analyses of colorectal and endometrial cancer genomes from The Cancer Genome Atlas project. These include identification of novel MSI targets within and across tumor types and the relationship between the likelihood of MSI events to chromatin structure. Given the increasing popularity of exome and genome sequencing of cancer genomes, a comprehensive characterization of MSI may serve as a valuable marker of cancer evolution and aid in a search for therapeutic targets. PMID:25371413

  19. Isolation and characterization of genomic microsatellite markers for small cardamom (Elettaria cardamomum Maton) for utility in genetic diversity analysis.

    PubMed

    Cyriac, Anu; Paul, Ritto; Anupama, K; Senthil Kumar, R; Sheeja, T E; Nirmal Babu, K; Parthasarathy, V A

    2016-04-01

    Microsatellite markers in small cardamom (Elettaria cardamomum Maton) were developed using the selective hybridization enrichment method. A total of 140 microsatellite repeats were identified from 270 clones. Primers were designed for 58 microsatellites and 44 primer pairs amplified products of expected size in cardamom. These markers were used for studying the diversity of 20 important small cardamom genotypes, and six markers were found to be polymorphic. The number of alleles ranged from 2 to 7 with an average of 3.6 per locus. Polymorphic information content values ranged from 0.14 to 0.38 based on dominant scoring. The two markers ECM 47a and ECMG 28 generated specific banding patterns for the genotypes MCC7 (Pink tiller) and APG434 (MA18) respectively. Dendrogram illustrated the genetic similarity between different genotypes of Kerala and Karnataka regions. It differentiated the closely related genotypes and released varieties into separate groups. Principal coordinate analysis revealed PV1 and ICRI 1 as the most divergent genotypes. The study demonstrated that these markers are informative and can be further utilized for generating reliable molecular data for assisting the crop improvement of small cardamom. Cross generic transferability (71.4 %) of the developed primers proved that they are useful for phylogenetic studies in the family Zingiberaceae. This is the first report of de novo isolation, characterisation and utilization of microsatellite markers for the genetic diversity analysis of small cardamom.

  20. Microsatellite marker development by multiplex ion torrent PGM sequencing: a case study of the endangered Odorrana narina complex of frogs.

    PubMed

    Igawa, Takeshi; Nozawa, Masafumi; Nagaoka, Mai; Komaki, Shohei; Oumi, Shohei; Fujii, Tamotsu; Sumida, Masayuki

    2015-01-01

    The endangered Ryukyu tip-nosed frog Odorrana narina and its related species, Odorrana amamiensis, Odorrana supranarina, and Odorrana utsunomiyaorum, belong to the family Ranidae and are endemically distributed in Okinawa (O. narina), Amami and Tokunoshima (O. amamiensis), and Ishigaki and Iriomote (O. supranarina and O. utsunomiyaorum) Islands. Because of varying distribution patterns, this species complex is an intrinsic model for speciation and adaptation. For effective conservation and molecular ecological studies, further genetic information is needed. For rapid, cost-effective development of several microsatellite markers for these and 2 other species, we used next-generation sequencing technology of Ion Torrent PGM™. Distribution patterns of repeat motifs of microsatellite loci in these modern frog species (Neobatrachia) were similarly skewed. We isolated and characterized 20 new microsatellite loci of O. narina and validated cross-amplification in the three-related species. Seventeen, 16, and 13 loci were cross-amplified in O. amamiensis, O. supranarina, and O. utsunomiyaorum, respectively, reflecting close genetic relationships between them. Mean number of alleles and expected heterozygosity of newly isolated loci varied depending on the size of each inhabited island. Our findings suggested the suitability of Ion Torrent PGM™ for microsatellite marker development. The new markers developed for the O. narina complex will be applicable in conservation genetics and molecular ecological studies.

  1. Multi-locus genotypes of Enterocytozoon bieneusi in captive Asiatic black bears in southwestern China: High genetic diversity, broad host range, and zoonotic potential

    PubMed Central

    Cao, Xuefeng; Song, Yuan; Wang, Wuyou; Huang, Xiangming; Liu, Xuehan; Hu, Yanchun; Fu, Hualin; He, Min; Wang, Ya; Zhang, Yue; Wu, Kongju; Peng, Guangneng

    2017-01-01

    Enterocytozoon bieneusi is an obligate eukaryotic intracellular parasite that infects a wide variety of vertebrate and invertebrate hosts. Although considerable research has been conducted on this organism, relatively little information is available on the occurrence of E. bieneusi in captive Asiatic black bears. The present study was performed to determine the prevalence, genetic diversity, and zoonotic potential of E. bieneusi in captive Asiatic black bears in zoos in southwestern China. Fecal specimens from Asiatic black bears in four zoos, located in four different cities, were collected and analyzed for the prevalence of E. bieneusi. The average prevalence of E. bieneusi was 27.4% (29/106), with the highest prevalence in Guiyang Zoo (36.4%, 16/44). Altogether, five genotypes of E. bieneusi were identified among the 29 E. bieneusi-positive samples, including three known genotypes (CHB1, SC02, and horse2) and two novel genotypes named ABB1 and ABB2. Multi-locus sequence typing using three microsatellites (MS1, MS3, and MS7) and one minisatellite (MS4) revealed V, III, V, and IV genotypes at these four loci, respectively. Phylogenetic analysis showed that the genotypes SC02 and ABB2 were clustered into group 1 of zoonotic potential, the genotypes CHB1 and ABB1 were clustered into a new group, and the genotype horse2 was clustered into group 6 of unclear zoonotic potential. In conclusion, this study identified two novel E. bieneusi genotypes in captive Asiatic black bears, and used microsatellite and minisatellite markers to reveal E. bieneusi genetic diversity. Moreover, our findings show that genotypes SC02 (identified in humans) and ABB2 belong to group 1 with zoonotic potential, suggesting the risk of transmission of E. bieneusi from Asiatic black bears to humans and other animals. PMID:28182656

  2. Multilocus sequence typing of Blastocystis isolates in Aydin, Turkey.

    PubMed

    Ertug, Sema; Malatyali, Erdogan; Ertabaklar, Hatice; Bozdogan, Bulent

    2016-12-01

    Blastocystis, a stramenopile protozoon of the human gastrointestinal tract, has a worldwide distribution. Multilocus sequence typing (MLST) is described as a portable, universal, and definitive method for accurate strain identification of microorganisms and was recently used for detecting intra-subtype variability of Blastocystis. The present study aimed to determine MLST sequence types in Blastocystis isolates from a routine diagnostic laboratory at Adnan Menderes University, Training and Research Hospital, Turkey. Samples were inoculated into Jones's medium after native-Lugol examination. Total genomic DNA was isolated from positive cultures with a commercially available kit. A total of 11 polymerase chain reactions were performed for each isolate (five loci for subtype 3 and six loci for subtype 4) using previously published MLST primers. The amplicons were sequenced and queried against a MLST database. A total of 11 isolates were amplified with subtype 3 MLST primers and could be sequenced at all loci; however none of the isolates were amplified with subtype 4 MLST primers. The isolates in our study population contained ten new alleles and nine sequence types. The present study contributes to existing knowledge of MLST data for Blastocystis isolates and is the first MLST study from Turkey. Our study confirms the extensive genetic diversity in Blastocystis that was reported in previous studies using different methods.

  3. A multilocus timescale for the origin of extant amphibians.

    PubMed

    San Mauro, Diego

    2010-08-01

    One of the most hotly debated topics in vertebrate evolution is the origin of extant amphibians (Lissamphibia). The recent contribution of molecular data is shedding new light on this debate, but many important questions still remain unresolved. I have assembled a large and comprehensive multilocus dataset (the largest to date in terms of number and heterogeneity of sequence characters) combining mitogenomic and nuclear information from 23 genes for a sufficiently dense taxon sampling with the key major lineages of extant amphibians. This dataset has been used to infer a robust phylogenetic framework and molecular timescale for the origin of extant amphibians employing the most recent phylogenetic and dating methods, as well as several alternative calibration schemes. The monophyly of each extant amphibian order and the sister group relationship between frogs and salamanders (Batrachia hypothesis) are all strongly supported. Dating analyses (all methods and calibration schemes used) suggest that the origin of extant amphibians (divergence between caecilian and batrachians) occurred in the Late Carboniferous, around 315 Mya, and the divergence between frogs and salamanders occurred in the Early Permian, around 290 Mya. These age estimates are more consistent with the fossil record than previous older estimates, and more in line with the Temnospondyli or the Lepospondyli hypotheses of lissamphibian ancestry (although the polyphyly hypothesis cannot be completely ruled out).

  4. Multilocus Sequence Typing for Interpreting Blood Isolates of Staphylococcus epidermidis.

    PubMed

    Sharma, Prannda; Satorius, Ashley E; Raff, Marika R; Rivera, Adriana; Newton, Duane W; Younger, John G

    2014-01-01

    Staphylococcus epidermidis is an important cause of nosocomial infection and bacteremia. It is also a common contaminant of blood cultures and, as a result, there is frequently uncertainty as to its diagnostic significance when recovered in the clinical laboratory. One molecular strategy that might be of value in clarifying the interpretation of S. epidermidis identified in blood culture is multilocus sequence typing. Here, we examined 100 isolates of this species (50 blood isolates representing true bacteremia, 25 likely contaminant isolates, and 25 skin isolates) and the ability of sequence typing to differentiate them. Three machine learning algorithms (classification regression tree, support vector machine, and nearest neighbor) were employed. Genetic variability was substantial between isolates, with 44 sequence types found in 100 isolates. Sequence types 2 and 5 were most commonly identified. However, among the classification algorithms we employed, none were effective, with CART and SVM both yielding only 73% diagnostic accuracy and nearest neighbor analysis yielding only 53% accuracy. Our data mirror previous studies examining the presence or absence of pathogenic genes in that the overlap between truly significant organisms and contaminants appears to prevent the use of MLST in the clarification of blood cultures recovering S. epidermidis.

  5. Multilocus sequence typing of oenological Saccharomyces cerevisiae strains.

    PubMed

    Muñoz, Rosario; Gómez, Alicia; Robles, Virginia; Rodríguez, Patricia; Cebollero, Eduardo; Tabera, Laura; Carrascosa, Alfonso V; Gonzalez, Ramon

    2009-12-01

    This study describes the application of a multilocus sequence typing (MLST) analysis for molecular discrimination at the strain level of Spanish wine yeast strains. The discrimination power of MLST is compared to mitochondrial RFLP analysis. Fragments of the ADP1, ACC1, RPN2, GLN4, and ALA1 genes were amplified by PCR from chromosomal DNA of 18 wine Saccharomyces cerevisiae strains. Ten polymorphic sites were found in the five loci analyzed showing 13 different genotypes, with 11 of them represented by only one strain. RFLP analysis of the same 18 wine yeast strains showed seventeen different mitochondrial patterns. Phylogenetic relationships among the strains analyzed, inferred by MLST data, showed wine isolates of S. cerevisiae as a rather homogeneous group. The discrimination potential of mitochondrial RFLP analysis was superior to the MLST scheme used in this work. However, MLST analysis allowed an easy construction of reliable phylogenetic trees. MLST analysis offers the possibility of typing wine S. cerevisiae strains simultaneously to the study of the genetic relationship among them.

  6. Environmental stress induces trinucleotide repeat mutagenesis in human cells

    PubMed Central

    Chatterjee, Nimrat; Lin, Yunfu; Santillan, Beatriz A.; Yotnda, Patricia; Wilson, John H.

    2015-01-01

    The dynamic mutability of microsatellite repeats is implicated in the modification of gene function and disease phenotype. Studies of the enhanced instability of long trinucleotide repeats (TNRs)—the cause of multiple human diseases—have revealed a remarkable complexity of mutagenic mechanisms. Here, we show that cold, heat, hypoxic, and oxidative stresses induce mutagenesis of a long CAG repeat tract in human cells. We show that stress-response factors mediate the stress-induced mutagenesis (SIM) of CAG repeats. We show further that SIM of CAG repeats does not involve mismatch repair, nucleotide excision repair, or transcription, processes that are known to promote TNR mutagenesis in other pathways of instability. Instead, we find that these stresses stimulate DNA rereplication, increasing the proportion of cells with >4 C-value (C) DNA content. Knockdown of the replication origin-licensing factor CDT1 eliminates both stress-induced rereplication and CAG repeat mutagenesis. In addition, direct induction of rereplication in the absence of stress also increases the proportion of cells with >4C DNA content and promotes repeat mutagenesis. Thus, environmental stress triggers a unique pathway for TNR mutagenesis that likely is mediated by DNA rereplication. This pathway may impact normal cells as they encounter stresses in their environment or during development or abnormal cells as they evolve metastatic potential. PMID:25775519

  7. A Microsatellite Genetic Linkage Map for Xiphophorus

    PubMed Central

    Walter, R. B.; Rains, J. D.; Russell, J. E.; Guerra, T. M.; Daniels, C.; Johnston, Dennis A.; Kumar, Jay; Wheeler, A.; Kelnar, K.; Khanolkar, V. A.; Williams, E. L.; Hornecker, J. L.; Hollek, L.; Mamerow, M. M.; Pedroza, A.; Kazianis, S.

    2004-01-01

    Interspecies hybrids between distinct species of the genus Xiphophorus are often used in varied research investigations to identify genomic regions associated with the inheritance of complex traits. There are 24 described Xiphophorus species and a greater number of pedigreed strains; thus, the number of potential interspecies hybrid cross combinations is quite large. Previously, select Xiphophorus experimental crosses have been shown to exhibit differing characteristics between parental species and among the hybrid fishes derived from crossing them, such as widely differing susceptibilities to chemical or physical agents. For instance, genomic regions harboring tumor suppressor and oncogenes have been identified via linkage association of these loci with a small set of established genetic markers. The power of this experimental strategy is related to the number of genetic markers available in the Xiphophorus interspecies cross of interest. Thus, we have undertaken the task of expanding the suite of easily scored markers by characterization of Xiphophorus microsatellite sequences. Using a cross between Xiphophorus maculatus and X. andersi, we report a linkage map predominantly composed of microsatellite markers. All 24 acrocentric chromosome sets of Xiphophorus are represented in the assembled linkage map with an average intergenomic distance of 7.5 cM. Since both male and female F1 hybrids were used to produce backcross progeny, these recombination rates were compared between “male” and “female” maps. Although several genomic regions exhibit differences in map length, male- and female-derived maps are similar. Thus Xiphophorus, in contrast to zebrafish, Danio rerio, and several other vertebrate species, does not show sex-specific differences in recombination. The microsatellite markers we report can be easily adapted to any Xiphophorus interspecies and some intraspecies crosses, and thus provide a means to directly compare results derived from independent

  8. Microsatellite DNA markers for assessing phylogeographic and population structure in Preble's meadow jumping mice (Zapus hudsonius preblei) and cross-amplification among neighbouring taxa

    USGS Publications Warehouse

    King, T.L.; Eackles, M.S.; Young, C.C.

    2006-01-01

    We document the isolation and characterization of 14 tetranucleotide microsatellite DNA markers in Preble's meadow jumping mouse (Zapus hudsonius preblei). The identified markers displayed moderate levels of allelic diversity (averaging 4.9 alleles per locus) and heterozygosity (averaging 55.1%). Genotypic and allelic frequencies in a collection of 30 individuals conformed to Hardy-Weinberg equilibrium expectations and indicated no linkage disequilibrium. High levels of cross-amplification (95% overall) among neighbouring subspecies and two congeners (Zapus princeps and Zapus trinotatus) were observed. Multilocus genotypes resulting from these markers appear to provide ample genetic diversity for studies assessing individual- and population-level ecological interactions within Z. h. preblei and evolutionary relationships among neighbouring subspecies (Z. h. campestris, Z. h. intermedius, Z. h. pallidus and Z. h. luteus). ?? 2006 The Authors.

  9. New microsatellite markers for bananas (Musa spp).

    PubMed

    Amorim, E P; Silva, P H; Ferreira, C F; Amorim, V B O; Santos, V J; Vilarinhos, A D; Santos, C M R; Souza Júnior, M T; Miller, R N G

    2012-04-27

    Thirty-four microsatellite markers (SSRs) were identified in EST and BAC clones from Musa acuminata burmannicoides var. Calcutta 4 and validated in 22 Musa genotypes from the Banana Germplasm Bank of Embrapa-CNPMF, which includes wild and improved diploids. The number of alleles per locus ranged from 2 to 14. The markers were considered highly informative based on their polymorphism information content values; more than 50% were above 0.5. These SSRs will be useful for banana breeding programs, for studies of genetic diversity, germplasm characterization and selection, development of saturated genetic linkage maps, and marker assisted selection.

  10. In-silico analysis of simple and imperfect microsatellites in diverse tobamovirus genomes.

    PubMed

    Alam, Chaudhary Mashhood; Singh, Avadhesh Kumar; Sharfuddin, Choudhary; Ali, Safdar

    2013-11-10

    An in-silico analysis of simple sequence repeats (SSRs) in 30 species of tobamoviruses was done. SSRs (mono to hexa) were present with variant frequency across species. Compound microsatellites, primarily of variant motifs accounted for up to 11.43% of the SSRs. Motif duplications were observed for A, T, AT, and ACA repeats. (AG)-(TC) was the most prevalent SSR-couple. SSRs were differentially localized in the coding region with ~54% on the 128 kDa protein while 20.37% was exclusive to 186 kDa protein. Characterization of such variations is important for elucidating the origin, sequence variations, and structure of these widely used, but incompletely understood sequences.

  11. Microplate array diagonal gel electrophoresis for cohort studies of microsatellite loci.

    PubMed

    Chen, Xiao-he; O'Dell, Sandra D; Day, Ian N M

    2002-05-01

    After PCR amplification, we have achieved precise sizing of trinucleotide and tetranucleotide microsatellite alleles on 96-well open-faced polyacrylamide microplate array diagonal gel electrophoresis (MADGE) gels: two tetranucleotide repeats, HUMTHOI (five alleles 248-263 bp) and DYS390 (eight alleles 200-228 bp), and DYS392, a trinucleotide repeat (eight alleles 210-231 bp). A gel matrix of Duracryl, a high mechanical strength polyacrylamide derivative, and appropriate ionic conditions provide the 1.3%-1.5% band resolution required. No end-labeling of primers is needed, as the sensitive Vistra Green intercalating dye is used for the visualization of bands. Co-run markers bracketing the PCR fragments ensure accurate sizing without inter-lane variability. Electrophoresis of multiple gels in a thermostatically controlled tank allows up to 1000 samples to be run in 90 min. Gel images were analyzed using a Fluorlmager 595 fluorescent scanning system, and alleles were identified using Phoretix software for band migration measurement and Microsoft Excel to compute fragment sizes. Estimated sizes were interpolated precisely to achieve accurate binning. Microsatellite-MADGE represents a utilitarian methodfor high-throughput genotyping in cohort studies, using standard laboratory equipment.

  12. VNTR and microsatellite polymorphisms within the subtelomeric region of 7q

    SciTech Connect

    Helms, C.; Donis-Keller, H. ); Hing, A.V.

    1993-08-01

    The molecular basis of a highly polymorphic RFLP marker, HTY146c3 (D7S591), within the subtelomeric region of human chromosome 7q was determined by restriction-fragment and DNA sequence analysis. Two polymorphic systems were found - a simple base-substitution polymorphism and a GC-rich VNTR element with a core structure of C[sub 3]AG[sub 2]C[sub 2]. In addition, a compound-imperfect CA dinucleotide-repeat element was identified approximately 10-20 kb from the telomeric sequence repeat (T[sub 2]AG[sub 3]), demonstrating that microsatellites can extend essentially to the ends of human chromosomes. The microsatellite marker, sAVH-6 (D7S594), is highly polymorphic, with 10 alleles and an observed heterozygosity of 84% found with the CEPH (Centre d'Etude du Polymorphisme Humain) reference pedigree collection. In combination with the RFLPs, the informativeness of the markers contained within 240 kb at the telomere approaches 100%. A unique genetic and physical STS marker, sAVH-6, defines the endpoint of the long arm of human chromosome 7. 33 refs., 4 figs., 2 tabs.

  13. Microsatellite typing as a new tool for identification of Saccharomyces cerevisiae strains.

    PubMed

    Hennequin, C; Thierry, A; Richard, G F; Lecointre, G; Nguyen, H V; Gaillardin, C; Dujon, B

    2001-02-01

    Since Saccharomyces cerevisiae appears to be an emerging pathogen, there is a need for a valuable molecular marker able to distinguish among strains. In this work, we investigated the potential value of microsatellite length polymorphism with a panel of 91 isolates, including 41 clinical isolates, 14 laboratory strains, and 28 strains with industrial relevance. Testing seven polymorphic regions (five trinucleotide repeats and two dinucleotide repeats) in a subgroup of 58 unrelated strains identified a total of 69 alleles (6 to 13 per locus) giving 52 different patterns with a discriminatory power of 99.03%. We found a cluster of clinical isolates sharing their genotype with a bakery strain, suggesting a digestive colonization following ingestion of this strain with diet. With the exception of this cluster of isolates and isolates collected from the same patient or from patients treated with Saccharomyces boulardii, all clinical isolates gave different and unique patterns. The genotypes are stable, and the method is reproducible. The possibility to make the method portable is of great interest for further studies using this technique. This work shows the possibility to readily identify S. boulardii (a strain increasingly isolated from invasive infections) using a unique and specific microsatellite allele.

  14. Scientific technological module of plasma injection for microsatellite

    NASA Astrophysics Data System (ADS)

    Sorokin, A. F.; Tsyukh, A. M.; Nesteruk, V. N.; Vlasenko, V. P.; Zaluzhnyj, R. N.; Sorokin, A. A.; Zakharova, M. Y.; Arsentiev, I. N.; Tarasov, I. S.

    The possibility to accomodate aboard a microsatellite a multifunctional module consisting of a plasma injector and heterolaser is considered. It is shown that a module of this kind allows one not only to make orbit correction and microsatellite orientation but also may be used in active space experiments.

  15. A microsatellite genetic linkage map of black rockfish ( Sebastes schlegeli)

    NASA Astrophysics Data System (ADS)

    Chu, Guannan; Jiang, Liming; He, Yan; Yu, Haiyang; Wang, Zhigang; Jiang, Haibin; Zhang, Quanqi

    2014-12-01

    Ovoviviparous black rockfish ( Sebastes schlegeli) is an important marine fish species for aquaculture and fisheries in China. Genetic information of this species is scarce because of the lack of microsatellite markers. In this study, a large number of microsatellite markers of black rockfish were isolated by constructing microsatellite-enriched libraries. Female- and male-specific genetic linkage maps were constructed using 435 microsatellite markers genotyped in a full-sib family of the fish species. The female linkage map contained 140 microsatellite markers, in which 23 linkage groups had a total genetic length of 1334.1 cM and average inter-marker space of 13.3 cM. The male linkage map contained 156 microsatellite markers, in which 25 linkage groups had a total genetic length of 1359.6 cM and average inter-marker distance of 12.4 cM. The genome coverage of the female and male linkage maps was 68.6% and 69.3%, respectively. The female-to-male ratio of the recombination rate was approximately 1.07:1 in adjacent microsatellite markers. This paper presents the first genetic linkage map of microsatellites in black rockfish. The collection of polymorphic markers and sex-specific linkage maps of black rockfish could be useful for further investigations on parental assignment, population genetics, quantitative trait loci mapping, and marker-assisted selection in related breeding programs.

  16. Isolation, cloning and characterisation of motifs containing (GA/TC)n repeats isolated from vetch, Vicia bithynica.

    PubMed

    Sakowicz, Tomasz; Bowater, Richard; Parniewski, Paweł

    2004-01-01

    Microsatellites are widely distributed in plant genomes and comprise unstable regions that undergo mutational changes at rates much greater than that observed for non-repetitive sequences. They demonstrate intrinsic genetic instability, manifested as frequent length changes due to insertions or deletions of repeat units. Detailed analysis of 1600 clones containing genomic sequences of Vicia bithynica revealed the presence of microsatellite repeats in its genome. Based on the screening of a partial DNA library of plasmids, 13 clones harbouring (GA/TC)n tracts of various lengths of repeated motif were identified for further analysis of their internal sequence organization. Sequence analyses revealed the precise length, number of repeats, interruptions within tracts, as well as sequence composition flanking the repeat motifs. Representative plasmids containing different lengths of (GA/TC)n embedded in their original flanking sequence were used to investigate the genetic stability of the repeats. In the study presented herein, we employed a well characterised and tractable bacterial genetic system. Recultivations of Escherichia coli harbouring plasmids containing (GA/TC)n inserts demonstrated that the genetic instability of (GA/TC)n microsatellites depends highly on their length (number of repeats). These observations are in agreement with similar studies performed on repetitive sequences from humans and other organisms.

  17. [Molecular structure of the allelic variants of (AAT)n microsatellite locus Du47D in the parthenogenetic species Darevskia unisexualis and bisexual parental species D. valentini and D. raddei].

    PubMed

    Korchagin, V I; Tokarskaia, O N

    2010-05-01

    Microsatellite repeats are one of the most widespread elements of the eukaryotic genome, but are poorly studied in species with clonal reproduction. PCR analysis and DNA sequencing were used to study the molecular structure of the allelic variants of microsatellite locus Du47D in the parthenogenetic species Darevskia unisexualis and its evolutionary ancestors, bisexual species D. raddei and D. valentini, of the genus Darevskia (Lacerta saxicola complex). Sequencing showed that the allelic variants of the D. unisexualis Du47D locus and the alleles of its D. raddei and D. valentini orthologs have a perfect microsatellite cluster structure, differ in number of ATT monomeric units, and have certain species-specific combinations of nucleotide substitutions, deletions, and insertions in the microsatellite-flanking DNA sequences. The Du47D alleles that the parthenogenetic species inherited from D. valentini or from D. raddei were identified.

  18. Development of Nuclear Microsatellite Loci and Mitochondrial Single Nucleotide Polymorphisms for the Natterjack Toad, Bufo (Epidalea) calamita (Bufonidae), Using Next Generation Sequencing and Competitive Allele Specific PCR (KASPar).

    PubMed

    Faucher, Leslie; Godé, Cécile; Arnaud, Jean-François

    2016-01-01

    Amphibians are undergoing a major decline worldwide and the steady increase in the number of threatened species in this particular taxa highlights the need for conservation genetics studies using high-quality molecular markers. The natterjack toad, Bufo (Epidalea) calamita, is a vulnerable pioneering species confined to specialized habitats in Western Europe. To provide efficient and cost-effective genetic resources for conservation biologists, we developed and characterized 22 new nuclear microsatellite markers using next-generation sequencing. We also used sequence data acquired from Sanger sequencing to develop the first mitochondrial markers for KASPar assay genotyping. Genetic polymorphism was then analyzed for 95 toads sampled from 5 populations in France. For polymorphic microsatellite loci, number of alleles and expected heterozygosity ranged from 2 to 14 and from 0.035 to 0.720, respectively. No significant departures from panmixia were observed (mean multilocus F IS = -0.015) and population differentiation was substantial (mean multilocus F ST = 0.222, P < 0.001). From a set of 18 mitochondrial SNPs located in the 16S and D-loop region, we further developed a fast and cost-effective SNP genotyping method based on competitive allele-specific PCR amplification (KASPar). The combination of allelic states for these mitochondrial DNA SNP markers yielded 10 different haplotypes, ranging from 2 to 5 within populations. Populations were highly differentiated (G ST = 0.407, P < 0.001). These new genetic resources will facilitate future parentage, population genetics and phylogeographical studies and will be useful for both evolutionary and conservation concerns, especially for the set-up of management strategies and the definition of distinct evolutionary significant units.

  19. Differentiation among Spanish sheep breeds using microsatellites

    PubMed Central

    Arranz, Juan-José; Bayón, Yolanda; Primitivo, Fermín San

    2001-01-01

    Genetic variability at 18 microsatellites was analysed on the basis of individual genotypes in five Spanish breeds of sheep – Churra, Latxa, Castellana, Rasa-Aragonesa and Merino -, with Awassi also being studied as a reference breed. The degree of population subdivision calculated between Spanish breeds from FST diversity indices was around 7% of total variability. A high degree of reliability was obtained for individual-breed assignment from the 18 loci by using different approaches among which the Bayesian method provided to be the most efficient, with an accuracy for nine microsatellites of over 99%. Analysis of the Bayesian assignment criterion illustrated the divergence between any one breed and the others, which was highest for Awassi sheep, while no great differences were evident among the Spanish breeds. Relationships between individuals were analysed from the proportion of shared alleles. The resulting dendrogram showed a remarkable breed structure, with the highest level of clustering among members of the Spanish breeds in Latxa and the lowest in Merino sheep, the latter breed exhibiting a peculiar pattern of clustering, with animals grouped into several closely set nodes. Analysis of individual genotypes provided valuable information for understanding intra- and inter-population genetic differences and allowed for a discussion with previously reported results using populations as taxonomic units. PMID:11712973

  20. Risk management for micro-satellite design

    NASA Astrophysics Data System (ADS)

    Santoni, Fabio

    2004-02-01

    It is shown how the methods of risk analysis have been used in the University of Rome micro-satellite program. One of the driving research topics related to this program is the reduction of cost in building spacecraft. The probability risk analysis techniques seem to be a powerful tool in the field of micro-satellites design, to outline possible faults. Innovation and limited budget forcing the designer to move in a very "risky" environment and can be faced with an as rigorous as possible decision making method. In our project, cost reduction is often attained relying on commercial, not space-rated components, which of course increases risk. This is why the design process should be led by the careful analysis of the risk associated with the selection of components and construction techniques. In standard applications risk can be evaluated from reliability data obtained in previous and well known similar applications. In our case, for many components, there is a lack of reliability data, due to the obvious missing experience when dealing with not yet space qualified, or even never flown before components. This lack is overcome using numerical simulations and practical engineering considerations, but does not allow a rigorous reliability assessment. A simple qualitative analysis is used to rank priorities among subsystems and allocate economic resources and development efforts. The main risk source is space radiation effect on CMOS electronic components. Therefore, development resources are directed to radiation effect mitigation. A procedure is proposed to lower risk without using space rated components.

  1. Endometrial cancer and microsatellite instability status

    PubMed Central

    Vidugiriene, Jolanta; Valuckas, Konstantinas Povilas; Smailyte, Giedre; Uleckiene, Saule; Bacher, Jeff

    2015-01-01

    Microsatellite instability (MSI) is an important factor in the development of various cancers as an identifier of a defective DNA mismatch repair system. The objective of our study was to define the association between microsatellite instability status and traditional clinicopathologic characteristics of endometrioid type adenocarcinoma. Material and methods MSI status of endometrial cancer was examined by employing the Promega MSI Analysis System. This system uses 5 mononucleotide markers to identify MSI in tumour and normal tissue DNA (BAT-25, BAT-26, NR-21, NR-24, and MONO-27), and 2 pentanucleotide markers (Penta C and Penta D) for specimen identification. In this study, we investigated MSI status in 109 endometrial carcinomas. Results and conclusions One hundred (92%) of 109 endometrial cancers showed endometrioid type histology and only 9 (8%) non-endometrioid type. MSI-high was found in 17% (17/100) of endometrioid type adenocarcinomas, in 0% (0/9) of non-endometrioid carcinomas. Selected clinicopathologic parameters for endometrioid type adenocarcinomas were compared to the MSI status which was separated into two groups – MSI-high and MSI stable. The results showed that MSI-high status was related to clinicopathologic parameters such as deep myometrial invasion and higher histologic grade in endometrioid type adenocarcinomas.

  2. Perils of gene mapping with microsatellite markers

    SciTech Connect

    Knowles, J.A.; Gilliam, T.C. ); Vieland, V.J. )

    1992-10-01

    The discovery of microsatellite polymorphisms has revitalized the genetic mapping of the human genome and promises to have a dramatic effect on human disease gene mapping. The high polymorphicity, relative abundance, and amenability of these markers to assay by PCR amplification gives them a significant advantage over previous markers, which explains their general acceptance and widespread use (Litt and Luty 1989; Weber and May 1989). Preliminary chromosome maps have been constructed using microsatellites exclusively (Weber et al. 1991; Hazen et al. 1992; Kwiatkowski et al. 1992), and disease loci have been mapped by linkage to these markers (Wijmenga et al. 1991). The markers provide new optimism for the mapping of disease genes, particularly for the mapping of complex genetic disorders. The authors present evidence that the very qualities that render these markers so efficient for chromosome mapping in large reference pedigrees can lead to dramatic lod score bias when applied to the typical pedigrees used to study genetic disorders, particularly when the disorder under study is complex. 11 refs., 2 figs., 1 tab.

  3. Adult survival selection in relation to multilocus heterozygosity and body size in a tropical bird species, the Zenaida dove, Zenaida aurita.

    PubMed

    Cézilly, Frank; Quinard, Aurélie; Motreuil, Sébastien; Pradel, Roger

    2016-01-01

    Both phenotypic and genetic quality can influence the survival of individuals through time, although their relative influences are rarely addressed simultaneously. Here we used capture-mark-recapture modelling to assess the influence of both multilocus heterozygosity (MLH) and body size on apparent adult survival in a tropical bird species, the Zenaida dove, Zenaida aurita, using a sample of 391 individuals genotyped at 11 microsatellites, while controlling for the effects of sex. No effect of body size on either adult survival or capture rate was found. In the best model, survival was a logit linear function of MLH, whereas detection probability was a sex-dependent logit linear function of the logarithm of field effort, increasing with time and affected by a random individual effect. Using a Bayesian approach, we found that MLH explained 1.14% of the total deviance, as expected from theory and previous studies of heterozygosity-fitness correlations, with no evidence for local effects. However, results from capture-mark-recapture modelling indicated that expected longevity varied from 4.8 years in the least heterozygous individuals (MLH = 0.37) to 10.6 years in the most heterozygous ones (MLH = 1), thus suggesting that MLH had potentially a substantial effect on survival. We discuss our results in relation to current hypotheses about the origin of heterozygosity-fitness correlations.

  4. Combined use of a new SNP-based assay and multilocus SSR markers to assess genetic diversity of Xylella fastidiosa subsp. pauca infecting citrus and coffee plants.

    PubMed

    Montes-Borrego, Miguel; Lopes, Joao R S; Jiménez-Díaz, Rafael M; Landa, Blanca B

    2015-03-01

    Two haplotypes of Xylella fastidiosa subsp. pauca (Xfp) that correlated with their host of origin were identified in a collection of 90 isolates infecting citrus and coffee plants in Brazil, based on a single-nucleotide polymorphism in the gyrB sequence. A new single-nucleotide primer extension (SNuPE) protocol was designed for rapid identification of Xfp according to the host source. The protocol proved to be robust for the prediction of the Xfp host source in blind tests using DNA from cultures of the bacterium, infected plants, and insect vectors allowed to feed on Xfp-infected citrus plants. AMOVA and STRUCTURE analyses of microsatellite data separated most Xfp populations on the basis of their host source, indicating that they were genetically distinct. The combined use of the SNaPshot protocol and three previously developed multilocus SSR markers showed that two haplotypes and distinct isolates of Xfp infect citrus and coffee in Brazil and that multiple, genetically different isolates can be present in a single orchard or infect a single tree. This combined approach will be very useful in studies of the epidemiology of Xfp-induced diseases, host specificity of bacterial genotypes, the occurrence of Xfp host jumping, vector feeding habits, etc., in economically important cultivated plants or weed host reservoirs of Xfp in Brazil and elsewhere.

  5. Epidemiological tracing of bovine tuberculosis in Switzerland, multilocus variable number of tandem repeat analysis of Mycobacterium bovis and Mycobacterium caprae

    PubMed Central

    Scherrer, Simone; Friedel, Ute; Frei, Daniel; Suter, Dominique; Perler, Lukas; Wittenbrink, Max M.

    2017-01-01

    Background After 15 years of absence, in 2013 bovine tuberculosis (bTB), caused by Mycobacterium (M.) bovis and M. caprae, reemerged in the Swiss dairy cattle population. In order to identify the sources of infection as well as the spread of the agents, molecular-epidemiologic tracing by MIRU-VNTR analysis in combination with spoligotyping was performed. A total of 17 M. bovis and 7 M. caprae isolates were cultured from tuberculous bovine lymph nodes and analyzed with a set of 49 genetic markers by using automated capillary electrophoresis. Results The outbreak in the western part of Switzerland was caused by M. bovis spoligotype SB0120. With the exception of four single-locus variations observed in MIRU 20, the MIRU-VNTR profiles of the 17 M. bovis isolates were identical, indicating a single source of infection. M. bovis detected in one archival bovine specimen from the outbreak region showed an identical MIRU-VNTR profile, suggesting persistence of the agent in a dairy herd for nearly fifteen years. The outbreak in the eastern part of Switzerland was caused by M. caprae spoligotype SB0418. All Swiss M. caprae isolates showed the Lechtal-type MIRU-VNTR profile, described as endemic in wild ruminants and in dairy cattle in Austrian bordering regions. This suggests the agent was most likely introduced by Swiss dairy cattle summering on Austrian pastures. Conclusions The present study is the first MIRU-VNTR analysis of Swiss bTB mycobacterial isolates. The genotyping assay was found to be highly discriminating and suitable for the epidemiological tracing of further outbreaks. These findings will contribute to the development of an international MIRU-VNTR database aiming to improve bTB surveillance. PMID:28222182

  6. Genome wide characterization of simple sequence repeats in watermelon genome and their application in comparative mapping and genetic diversity analysis

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Simple sequence repeats (SSR) or microsatellite markers are one of the most informative and versatile DNA-based markers. The use of next-generation sequencing technologies allow whole genome sequencing and make it possible to develop large numbers of SSRs through bioinformatic analysis of genome da...

  7. A high-density simple sequence repeat and single nucleotide polymorphism genetic map of the tetraploid cotton genome

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cotton genome complexity was investigated with a saturated molecular genetic map that combined several sets of microsatellites or simple sequence repeats (SSR) and the first major public set of single nucleotide polymorphism (SNP) markers in cotton genomes (Gossypium spp.), and that was constructed ...

  8. Genome-wide In Silico Analysis, Characterization and Identification of Microsatellites in Spodoptera littoralis Multiple nucleopolyhedrovirus (SpliMNPV)

    PubMed Central

    Atia, Mohamed A. M.; Osman, Gamal H.; Elmenofy, Wael H.

    2016-01-01

    In this study, we undertook a survey to analyze the distribution and frequency of microsatellites or Simple Sequence Repeats (SSRs) in Spodoptera littoralis multiple nucleopolyhedrovirus (SpliMNPV) genome (isolate AN–1956). Out of the 55 microsatellite motifs, identified in the SpliMNPV-AN1956 genome using in silico analysis (inclusive of mono-, di-, tri- and hexa-nucleotide repeats), 39 were found to be distributed within coding regions (cSSRs), whereas 16 were observed to lie within intergenic or noncoding regions. Among the 39 motifs located in coding regions, 21 were located in annotated functional genes whilst 18 were identified in unknown functional genes (hypothetical proteins). Among the identified motifs, trinucleotide (80%) repeats were found to be the most abundant followed by dinucleotide (13%), mononucleotide (5%) and hexanucleotide (2%) repeats. The 39 motifs located within coding regions were further validated in vitro by using PCR analysis, while the 21 motifs located within known functional genes (15 genes) were characterized using nucleotide sequencing. A comparison of the sequence analysis data of the 21 sequenced cSSRs with the published sequences is presented. Finally, the developed SSR markers of the 39 motifs were further mapped/localized onto the SpliMNPV-AN1956 genome. In conclusion, the SSR markers specific to SpliMNPV, developed in this study, could be a useful tool for the identification of isolates and analysis of genetic diversity and viral evolutionary status. PMID:27650818

  9. Single-molecule PCR analysis of an unstable microsatellite for detecting mutations in sperm of mice exposed to chemical mutagens.

    PubMed

    Beal, Marc A; Rowan-Carroll, Andrea; Campbell, Caleigh; Williams, Andrew; Somers, Christopher M; Marchetti, Francesco; Yauk, Carole L

    2015-05-01

    Single-molecule PCR (SM-PCR) analysis of long and repetitive DNA sequences, known as expanded simple tandem repeats (ESTRs), has been the most efficient method for studying germline mutation induction in endogenous sequences to date. However, the long length of these sequences makes mutation detection imprecise and laborious, and they have been characterized only in mice. Here, we explore the use of unstable microsatellite sequences that can be typed with high precision by capillary electrophoresis as alternative loci for detecting germline mutations. We screened 24 microsatellite loci across inbred mouse strains and identified Mm2.2.1 as the most polymorphic microsatellite locus. We then optimized SM-PCR of Mm2.2.1 to detect mutations in sperm. SM-PCR analysis of sperm from untreated B6C3F1 and Muta(™)Mouse samples revealed mutation frequencies that are consistent with rates derived from family pedigree analysis (∼ 5 × 10(-3)). To determine whether this locus can be used to detect chemically induced germline mutations, Muta(™)Mouse males were exposed by oral gavage to a single dose of 100mg/kg of N-ethyl-N-nitrosourea (ENU) or to 100mg/kg of benzo(a)pyrene (BaP) for 28 days alongside vehicle treated controls. Sperm were collected 10 weeks post-ENU exposure to sample sperm exposed as spermatogonial stem cells and 6 weeks post-BaP exposure to sample sperm that were dividing spermatogonia when the exposure was terminated. Both treatments resulted in a significant (approximately 2-fold) increase in mutation frequency in sperm compared to the control animals. The work establishes the utility of this microsatellite for studying mutation induction in the germ cells of mice. Because microsatellites are found in virtually every species, this approach holds promise for other organisms, including humans.

  10. Characterization of 35 novel microsatellite DNA markers from the duck (Anas platyrhynchos) genome and cross-amplification in other birds

    PubMed Central

    Huang, Yinhua; Tu, Jianfeng; Cheng, Xuebo; Tang, Bo; Hu, Xiaoxiang; Liu, Zhaoliang; Feng, Jidong; Lou, Yankun; Lin, Li; Xu, Ke; Zhao, Yulong; Li, Ning

    2005-01-01

    In order to study duck microsatellites, we constructed a library enriched for (CA)n, (CAG)n, (GCC)n and (TTTC)n. A total of 35 pairs of primers from these microsatellites were developed and used to detect polymorphisms in 31 unrelated Peking ducks. Twenty-eight loci were polymorphic and seven loci were monomorphic. A total of 117 alleles were observed from these polymorphic microsatellite markers, which ranged from 2 to 14 with an average of 4.18 per locus. The frequencies of the 117 alleles ranged from 0.02 to 0.98. The highest heterozygosity (0.97) was observed at the CAUD019 microsatellite locus and the lowest heterozygosity (0.04) at the CAUD008 locus, and 11 loci had heterozygosities greater than 0.50 (46.43%). The polymorphism information content (PIC) of 28 loci ranged from 0.04 to 0.88 with an average of 0.42. All the above markers were used to screen the polymorphism in other bird species. Two markers produced specific monomorphic products with the chicken DNA. Fourteen markers generated specific fragments with the goose DNA: 5 were polymorphic and 9 were monomorphic. But no specific product was detected with the peacock DNA. Based on sequence comparisons of the flanking sequence and repeat, we conclude that 2 chicken loci and 14 goose loci were true homologous loci of the duck loci. The microsatellite markers identified and characterized in the present study will contribute to the genetic map, quantitative traits mapping, and phylogenetic analysis in the duck and goose. PMID:15943922

  11. Selecting representative microsatellite loci for genetic monitoring and analyzing genetic structure of an outbred population of orange tabby cats in China.

    PubMed

    Du, X Y; Yi, S; Huo, X Y; Wang, C; Liu, D F; Ren, W Z; Chen, Z W

    2015-03-13

    We optimized a panel of microsatellite markers from cat and tiger genetic data for efficient genetic monitoring and used it to analyze the genetic structure of an outbred cat stock in China. We selected a set of rich polymorphic microsatellite loci from 131 cat microsatellite loci and 3 Sumatran tiger microsatellite loci using agarose gel electrophoresis. Next, the set of optimized genetic markers was used to analyze the genetic variation in an outbred population of orange tabby cats in China by simple-tandem repeat scanning. Thirty-one loci rich in polymorphisms were selected and the highest allele number in a single locus was 8. Analysis of the orange tabby cat population illustrated that the average observed number of alleles, mean effective allele number, mean Shannon's information index, mean expected heterozygosity, and observed heterozygosity were 3.8387, 2.4027, 0.9787, 0.5565, and 0.5528, respectively. The 31 microsatellite markers used were polymorphic and suitable for analyzing the genetic structure of cats. The population of orange tabby cats was confirmed to be a well-outbred stock.

  12. Isolation and characterisation of 11 polymorphic microsatellite markers in Papaver rhoeas L. (Corn Poppy), a major annual plant species from cultivated areas.

    PubMed

    Kati, Vaya; Corre, Valérie Le; Michel, Séverine; Jaffrelo, Lydia; Poncet, Charles; Délye, Christophe

    2012-12-24

    Papaver rhoeas, an annual plant species in the Papaveraceae family, is part of the biodiversity of agricultural ecosystems and also a noxious agronomic weed. We developed microsatellite markers to study the genetic diversity of P. rhoeas, using an enriched microsatellite library coupled with 454 next-generation sequencing. A total of 13,825 sequences were obtained that yielded 1795 microsatellite loci. After discarding loci with less than six repeats of the microsatellite motif, automated primer design was successful for 598 loci. We tested 74 of these loci for amplification with a total of 97 primer pairs. Thirty loci passed our tests and were subsequently tested for polymorphism using 384 P. rhoeas plants originating from 12 populations from France. Of the 30 loci, 11 showed reliable polymorphism not affected by the presence of null alleles. The number of alleles and the expected heterozygosity ranged from 3 to 7.4 and from 0.27 to 0.73, respectively. A low but significant genetic differentiation among populations was observed (F(ST) = 0.04; p < 0.001). The 11 validated polymorphic microsatellite markers developed in this work will be useful in studies of genetic diversity and population structure of P. rhoeas, assisting in designing management strategies for the control or the conservation of this species.

  13. Microsatellite Variation in Honey Bee (Apis Mellifera L.) Populations: Hierarchical Genetic Structure and Test of the Infinite Allele and Stepwise Mutation Models

    PubMed Central

    Estoup, A.; Garnery, L.; Solignac, M.; Cornuet, J. M.

    1995-01-01

    Samples from nine populations belonging to three African (intermissa, scutellata and capensis) and four European (mellifera, ligustica, carnica and cecropia) Apis mellifera subspecies were scored for seven microsatellite loci. A large amount of genetic variation (between seven and 30 alleles per locus) was detected. Average heterozygosity and average number of alleles were significantly higher in African than in European subspecies, in agreement with larger effective population sizes in Africa. Microsatellite analyses confirmed that A. mellifera evolved in three distinct and deeply differentiated lineages previously detected by morphological and mitochondrial DNA studies. Dendrogram analysis of workers from a given population indicated that super-sisters cluster together when using a sufficient number of microsatellite data whereas half-sisters do not. An index of classification was derived to summarize the clustering of different taxonomic levels in large phylogenetic trees based on individual genotypes. Finally, individual population X loci data were used to test the adequacy of the two alternative mutation models, the infinite allele model (IAM) and the stepwise mutation models. The better fit overall of the IAM probably results from the majority of the microsatellites used including repeats of two or three different length motifs (compound microsatellites). PMID:7498746

  14. Microsatellite variation in honey bee (Apis mellifera L.) populations: hierarchical genetic structure and test of the infinite allele and stepwise mutation models.

    PubMed

    Estoup, A; Garnery, L; Solignac, M; Cornuet, J M

    1995-06-01

    Samples from nine populations belonging to three African (intermissa, scutellata and capensis) and four European (mellifera, ligustica, carnica and cecropia) Apis mellifera subspecies were scored for seven microsatellite loci. A large amount of genetic variation (between seven and 30 alleles per locus) was detected. Average heterozygosity and average number of alleles were significantly higher in African than in European subspecies, in agreement with larger effective population sizes in Africa. Microsatellite analyses confirmed that A. mellifera evolved in three distinct and deeply differentiated lineages previously detected by morphological and mitochondrial DNA studies. Dendrogram analysis of workers from a given population indicated that super-sisters cluster together when using a sufficient number of microsatellite data whereas half-sisters do not. An index of classification was derived to summarize the clustering of different taxonomic levels in large phylogenetic trees based on individual genotypes. Finally, individual population x loci data were used to test the adequacy of the two alternative mutation models, the infinite allele model (IAM) and the stepwise mutation models. The better fit overall of the IAM probably results from the majority of the microsatellites used including repeats of two or three different length motifs (compound microsatellites).

  15. Microsatellites in the tree Foetidia mauritiana (Lecythidaceae) and utility in other Foetidia taxa from the Mascarene Islands1

    PubMed Central

    Martos, Florent; Lebreton, Gérard; Rivière, Eric; Humeau, Laurence; Chevallier, Marie-Hélène

    2016-01-01

    Premise of the study: Polymorphic markers were required for a native tree of the Mascarene Islands, Foetidia mauritiana (Lecythidaceae), to investigate the effects of fragmentation of lowland tropical habitats on tree mating systems and on gene flow. Methods and Results: Using microsatellite enrichment and next-generation sequencing, we identified 13 microsatellite loci (dinucleotide repeats). They were highly polymorphic in 121 trees sampled in the largest three populations on Réunion, revealing 2–17 different alleles per locus. Furthermore, they were found to be polymorphic in conspecific populations on Mauritius and in F. rodriguesiana from Rodrigues. Conclusions: These results indicate the utility of these markers to investigate genetic diversity, mating systems, and gene flow in a genus native to the biodiversity hotspot of Madagascar and the Indian Ocean islands. PMID:27610278

  16. Development and characterization of microsatellite markers for the medicinal plant Smilax brasiliensis (Smilacaceae) and related species1

    PubMed Central

    Martins, Aline R.; Abreu, Aluana G.; Bajay, Miklos M.; Villela, Priscilla M. S.; Batista, Carlos E. A.; Monteiro, Mariza; Alves-Pereira, Alessandro; Figueira, Glyn M.; Pinheiro, José B.; Appezzato-da-Glória, Beatriz; Zucchi, Maria I.

    2013-01-01

    • Premise of the study: A new set of microsatellite or simple sequence repeat (SSR) markers were developed for Smilax brasiliensis, which is popularly known as sarsaparilla and used in folk medicine as a tonic, antirheumatic, and antisyphilitic. Smilax brasiliensis is sold in Brazilian pharmacies, and its origin and effectiveness are not subject to quality control. • Methods and Results: Using a protocol for genomic library enrichment, primer pairs were developed for 26 microsatellite loci and validated in 17 accessions of S. brasiliensis. Thirteen loci were polymorphic and four were monomorphic. The primers successfully amplified alleles in the congeners S. campestris, S. cissoides, S. fluminensis, S. goyazana, S. polyantha, S. quinquenervia, S. rufescens, S. subsessiliflora, and S. syphilitica. • Conclusions: The new SSR markers described herein are informative tools for genetic diversity and gene flow studies in S. brasiliensis and several congeners. PMID:25202555

  17. In- silico exploration of thirty alphavirus genomes for analysis of the simple sequence repeats

    PubMed Central

    Alam, Chaudhary Mashhood; Singh, Avadhesh Kumar; Sharfuddin, Choudhary; Ali, Safdar

    2014-01-01

    The compilation of simple sequence repeats (SSRs) in viruses and its analysis with reference to incidence, distribution and variation would be instrumental in understanding the functional and evolutionary aspects of repeat sequences. Present study encompasses the analysis of SSRs across 30 species of alphaviruses. The full length genome sequences, assessed from NCBI were used for extraction and analysis of repeat sequences using IMEx software. The repeats of different motif sizes (mono- to penta-nucleotide) observed therein exhibited variable incidence across the species. Expectedly, mononucleotide A/T was the most prevalent followed by dinucleotide AG/GA and trinucleotide AAG/GAA in these genomes. The conversion of SSRs to imperfect microsatellite or compound microsatellite (cSSR) is low. cSSR, primarily constituted by variant motifs accounted for up to 12.5% of the SSRs. Interestingly, seven species lacked cSSR in their genomes. However, the SSR and cSSR are predominantly localized to the coding region ORFs for non structural protein and structural proteins. The relative frequencies of different classes of simple and compound microsatellites within and across genomes have been highlighted. PMID:25606453

  18. Genetic variation and evolutionary stability of the FMR1 CGG repeat in six closed human populations

    SciTech Connect

    Eichler, E.E.; Nelson, D.L.

    1996-07-12

    In an attempt to understand the allelic diversity and mutability of the human FMR1 CGG repeat, we have analyzed the AGG substructure of this locus within six genetically-closed populations (Mbuti pygmy, Baka pygmy, R. surui, Karitiana, Mayan, and Hutterite). Most alleles (61/92 or 66%) possessed two AGG interspersions occurring with a periodicity of one AGG every nine or ten CGG repeats, indicating that this pattern is highly conserved in all human populations. Significant differences in allele distribution were observed among the populations for rare variants possessing fewer or more AGG interruptions than the canonical FMR1 CGG repeat sequence. Comparisons of expected heterozygosity of the FMR1 CGG repeat locus with 30 other microsatellite loci, demonstrated remarkably similar levels of polymorphism within each population, suggesting that most FMR1 CGG repeat alleles mutate at rates indistinguishable from other microsatellite loci. A single allele (1 out of 92) was identified with a large uninterrupted tract of pure repeats (42 pure CGG triplets). Retrospective pedigree analysis indicated that this allele had been transmitted unstably. Although such alleles mutate rapidly and likely represent evolving premutations, our analysis suggests that in spite of the estimated frequency of their occurrence, these unstable alleles do not significantly alter the expected heterozygosity of the FMR1 CGG repeat in the human population. 45 refs., 1 fig., 2 tabs.

  19. Multicentric epidemiological study of Aspergillus fumigatus isolates by multilocus enzyme electrophoresis.

    PubMed Central

    Rodriguez, E; De Meeüs, T; Mallie, M; Renaud, F; Symoens, F; Mondon, P; Piens, M A; Lebeau, B; Viviani, M A; Grillot, R; Nolard, N; Chapuis, F; Tortorano, A M; Bastide, J M

    1996-01-01

    The genotypes of 63 isolates of Aspergillus fumigatus obtained from three hospitals in different geographical areas and of eight culture collection strains were determined by multilocus enzyme electrophoresis. Twelve of the 17 enzymatic loci studied were polymorphic, giving rise to 48 different electrophoretic types. The existence of fixed multilocus genotypes, significant heterozygote deficits and excesses at the different loci, and linkage disequilibria within subpopulations strongly suggests a clonal reproduction mode for A. fumigatus. Numerical analysis of the comparison and disposition of the different electrophoretic types demonstrates a significant genetic differentiation between the three sampling sites. However, no correlation could be found between geographical distances and genetic differentiation. On account of the multiple discriminatory markers, multilocus enzyme electrophoresis typing seems to be a very powerful tool for epidemiological and reproductive mode studies of A. fumigatus. PMID:8880520

  20. Development of novel microsatellite DNA markers by cross-amplification and analysis of genetic variation in gerbils.

    PubMed

    Du, Xiaoyan; Chen, Zhenwen; Li, Wei; Tan, Yuanqing; Lu, Jing; Zhu, Xiangdong; Zhao, Taiyun; Dong, Gang; Zeng, Lin

    2010-01-01

    The objectives of this study are to establish microsatellite loci for the Mongolian gerbil based on mouse microsatellite DNA sequences and to investigate genetic variation in the laboratory gerbil (Capital Medical University, CMU) and 2 wild gerbil populations (from Yin Chuan city [YIN] and the Hohehot Municipality [HOH]). In total, 536 mouse microsatellite markers were chosen to identify polymorphic dinucleotide repeat loci in the gerbil by cross-amplification. Of these markers, 313 (58.39%) have been discretely amplified from the CMU laboratory gerbil and been sequenced. Of the 313 sequenced markers, 130 were confirmed as simple sequence repeat (SSR) loci in the gerbil. In total, 6 of those newly identified loci plus 6 identified in previous reports were used to estimate the genetic polymorphism for 30 laboratory gerbils and 54 wild gerbils (27 each of the HOH and YIN groups). A total of 29 alleles were observed in the 3 populations, and 11 of 12 loci (91.67%) are polymorphic markers. Nei's standard genetic distances of 0.0592 (CMU vs. HOH) and 0.1033 (CMU vs. YIN) were observed. The averages of observed versus expected heterozygosity are 0.5231/0.4008, 0.5051/0.3882, and 0.4825/0.3665 for the YIN, HOH, and CMU populations, respectively. These results show that cross-amplification using mouse microsatellite primers is an efficient way to identify gerbil SSR loci. By using these 12 selected markers, we have demonstrated that genetic variation level within the CMU population is higher than that has been reported previously and are comparable with the levels found in 2 wild populations.

  1. Microsatellite instability typing in serum and tissue of patients with colorectal cancer: comparing real time PCR with hybridization probe and high-performance liquid chromatography.

    PubMed

    Mokarram, P; Rismanchi, M; Alizadeh Naeeni, M; Mirab Samiee, S; Paryan, M; Alipour, A; Honardar, Z; Kavousipour, S; Naghibalhossaini, F; Mostafavi-Pour, Z; Monabati, A; Hosseni, S V; Shamsdin, S A

    2014-05-01

    Allelic variation of BAT-25 (a 25-repeat quasimonomorphic poly T) and BAT-26 (a 26-repeat quasimonomorphic polyA) loci as two mononucleotide microsatellite markers, were analyzed with high-performance liquid chromatography (HPLC) compared with Real-Time PCR using hybridization probes. BAT-26 and BAT-25 markers were used to determine an appropriate screening technique with high sensitivity and specificity to diagnose microsatellite instability (MSI) status in patients with colorectal cancer (CRC). One of the pathways in colorectal tumor genesis is microsatellite instability (MSI+). MSI is detected in about 15% of all CRCs; 3% are of these are associated with Lynch syndrome and the other 12% are caused by sporadic. Colorectal tumors with MSI have distinctive features compared with microsatellite stable tumors. Due to the high percentage of MSI+ CRC in Iran, screening of this type of CRC is imperative. Two markers were analyzed in tissues and sera of 44 normal volunteers and tumor and matched normal mucosal tissues as well as sera of 44 patients with sporadic CRC. The sensitivity and specificity of BAT-26 with real time PCR method (Hybridization probe) were 100% in comparison with sequencing method as the gold standard, while HPLC had a lower sensitivity and specificity. According to HPLC data, BAT-26 was more sensitive than BAT-25 in identifying MSI tumors. Therefore, MSI typing using the BAT-26 hybridization probe method compared to HPLC could be considered as an accurate method for diagnosing MSI in CRC tumors but not in serum circulating DNAs.

  2. Novel microsatellite markers for the oriental fruit moth Grapholita molesta (Lepidoptera: Tortricidae) and effects of null alleles on population genetics analyses.

    PubMed

    Song, W; Cao, L-J; Wang, Y-Z; Li, B-Y; Wei, S-J

    2016-11-07

    The oriental fruit moth (OFM) Grapholita molesta (Lepidoptera: Tortricidae) is an important economic pest of stone and pome fruits worldwide. We sequenced the OFM genome using next-generation sequencing and characterized the microsatellite distribution. In total, 56,674 microsatellites were identified, with 11,584 loci suitable for primer design. Twenty-seven polymorphic microsatellites, including 24 loci with trinucleotide repeat and three with pentanucleotide repeat, were validated in 95 individuals from four natural populations. The allele numbers ranged from 4 to 40, with an average value of 13.7 per locus. A high frequency of null alleles was observed in most loci developed for the OFM. Three marker panels, all of the loci, nine loci with the lowest null allele frequencies, and nine loci with the highest null allele frequencies, were established for population genetics analyses. The null allele influenced estimations of genetic diversity parameters but not the OFM's genetic structure. Both a STRUCTURE analysis and a discriminant analysis of principal components, using the three marker panels, divided the four natural populations into three groups. However, more individuals were incorrectly assigned by the STRUCTURE analysis when the marker panel with the highest null allele frequency was used compared with the other two panels. Our study provides empirical research on the effects of null alleles on population genetics analyses. The microsatellites developed will be valuable markers for genetic studies of the OFM.

  3. A multilocus linkage disequilibrium measure based on mutual information theory and its applications.

    PubMed

    Zhang, Lei; Liu, Jianfeng; Deng, Hong-Wen

    2009-12-01

    Evaluating the patterns of linkage disequilibrium (LD) is important for association mapping study as well as for studying the genomic architecture of human genome (e.g., haplotype block structures). Commonly used bi-allelic pairwise measures for assessing LD between two loci, such as r(2) and D', may not make full and efficient use of modern multilocus data. Though extended to multilocus scenarios, their performance is still questionable. Meanwhile, most existing measures for an entire multilocus region, such as normalized entropy difference, do not consider existence of LD heterogeneity across the region under investigation. Additionally, these existing multilocus measures cannot handle distant regions where long-range LD patterns may exist. In this study, we proposed a novel multilocus LD measure developed based on mutual information theory. Our proposed measure described LD pattern between two chromosome regions each of which may consist of multiple loci (including multi-allele loci). As such, the proposed measure can better characterize LD patterns between two arbitrary regions. As potential applications, we developed algorithms on the proposed measure for partitioning haplotype blocks and for selecting haplotype tagging SNPs (htSNPs), which were helpful for follow-up association tests. The results on both simulated and empirical data showed that our LD measure had distinct advantages over pairwise and other multilocus measures. First, our measure was more robust, and can capture comprehensively the LD information between neighboring as well as disjointed regions. Second, haplotype blocks were better described via our proposed measure. Furthermore, association tests with htSNPs from the proposed algorithm had improved power over tests on single markers and on haplotypes.

  4. Longitudinal prevalence and molecular typing of Escherichia coli O157:H7 using multiple-locus variable-number tandem-repeats analysis and pulsed field gel electrophoresis in a range cattle herd in California

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Objectives –(1) Identify the seasonal pattern and risk factors for Escherichia coli O157:H7 in feces in range cattle in California, (2) Determine strain diversity and transition over time using Multi-Locus Variable-Number Tandem Repeat Analysis (MLVA) and Pulsed Field Gel Electrophoresis (PFGE) Samp...

  5. A power system for a microsatellite

    NASA Astrophysics Data System (ADS)

    Olsson, Dan

    1993-01-01

    A power system for a microsatellite was designed for the Space Technology Research Vehicle (STRV-1) mainly aimed at technology demonstration experiments and studies of radiation effects and planned to be launched into geostationary transfer orbit. The low average solar array power (25 W) and the small volume allocation of the power system electronics (2.8 dm cubed) became a challenge when trying to produce a failure tolerant power system. The power system elements: shunt regulator, battery discharge regulator, power distribution, solar array, battery, and their management and protection are described. Power system performance and growth potential are also discussed. After several simplification iteractions, a modular power electronics system with a high degree of failure tolerance, with a mass less than 2.5 kg and an internal power consumption less than 3 W was developed.

  6. Microsatellite markers for Russian olive (Elaeagnus angustifolia; Elaeagnaceae)1

    PubMed Central

    Gaskin, John F.; Hufbauer, Ruth A.; Bogdanowicz, Steven M.

    2013-01-01

    • Premise of the study: Microsatellite markers were developed for the plant species Elaeagnus angustifolia to assist in future investigations of genetic variability in its native and invasive ranges and the precise origins of the United States/Canada invasion. • Methods and Results: Eleven polymorphic microsatellite markers were developed. The number of alleles observed for each locus ranged from three to 11. • Conclusions: These microsatellites have sufficient potential variability to define population structure and origins of the Russian olive invasion. PMID:25202584

  7. Successful genotyping of microsatellites in the woolly mammoth.

    PubMed

    Ishida, Yasuko; Roca, Alfred L; Fratpietro, Stephen; Greenwood, Alex D

    2012-01-01

    Genetic analyses using ancient DNA from Pleistocene and early Holocene fossils have largely relied on mitochondrial DNA (mtDNA) sequences. Among woolly mammoths, Mammuthus primigenius, mtDNA analyses have identified 2 distinct clades (I and II) that diverged 1-2 Ma. Here, we establish that microsatellite markers can be effective on Pleistocene samples, successfully genotyping woolly mammoth specimens at 2 loci. Although significant differentiation at the 2 microsatellite loci was not detected between 16 clade I and 4 clade II woolly mammoths, our results demonstrate that the nuclear population structure of Pleistocene species can be examined using fast-evolving nuclear microsatellite markers.

  8. Pooled Genotyping of Microsatellite Markers in Parent–Offspring Trios

    PubMed Central

    Kirov, George; Williams, Nigel; Sham, Pak; Craddock, Nick; Owen, Michael J.

    2000-01-01

    We studied the extent to which genotyping of simple sequence repeat polymorphisms (SSRs) in pooled DNA samples can be used to predict differences in allele frequencies between parents and their affected offspring. We also developed a simple method of correction for the effects of stutter and differential amplification on the analysis of SSRs in pooled DNA samples based on widely available software. We genotyped individually eight polymorphic microsatellite markers in 110 parent–offspring trios affected with bipolar affective disorder (BP). Analysis of pooled DNA samples predicted very accurately the differences in individual allele frequency distributions between children and their parents. The mean error was <1% (range 0%–3.2%) when marker-specific corrections for stutter and differential amplification were performed. We show that if an individual allele is significantly preferentially transmitted from parents to affected offspring, the difference in the frequency of that allele would be sufficiently large to be detected with pooling in most situations. We propose recommendations for disequilibrium mapping with pooling in which both case-control samples and trios are used in an initial screen and markers are genotyped individually only if they satisfy very relaxed criteria for statistical significance. The use of case-control samples should reduce the false-negative rate as the differences in allele frequencies between cases and controls are twice as high in the presence of the same genetic effect. The use of trios will confirm or reject any suggested differences, thus reducing the false-positive rate that can be created by hidden population stratification. PMID:10645955

  9. Comparative Microsatellite Typing of New World Leishmania infantum Reveals Low Heterogeneity among Populations and Its Recent Old World Origin

    PubMed Central

    Kuhls, Katrin; Alam, Mohammad Zahangir; Cupolillo, Elisa; Ferreira, Gabriel Eduardo M.; Mauricio, Isabel L.; Oddone, Rolando; Feliciangeli, M. Dora; Wirth, Thierry; Miles, Michael A.; Schönian, Gabriele

    2011-01-01

    Leishmania infantum (syn. L. chagasi) is the causative agent of visceral leishmaniasis (VL) in the New World (NW) with endemic regions extending from southern USA to northern Argentina. The two hypotheses about the origin of VL in the NW suggest (1) recent importation of L. infantum from the Old World (OW), or (2) an indigenous origin and a distinct taxonomic rank for the NW parasite. Multilocus microsatellite typing was applied in a survey of 98 L. infantum isolates from different NW foci. The microsatellite profiles obtained were compared to those of 308 L. infantum and 20 L. donovani strains from OW countries previously assigned to well-defined populations. Two main populations were identified for both NW and OW L. infantum. Most of the NW strains belonged to population 1, which corresponded to the OW MON-1 population. However, the NW population was much more homogeneous. A second, more heterogeneous, population comprised most Caribbean strains and corresponded to the OW non-MON-1 population. All Brazilian L. infantum strains belonged to population 1, although they represented 61% of the sample and originated from 9 states. Population analysis including the OW L. infantum populations indicated that the NW strains were more similar to MON-1 and non-MON-1 sub-populations of L. infantum from southwest Europe, than to any other OW sub-population. Moreover, similarity between NW and Southwest European L. infantum was higher than between OW L. infantum from distinct parts of the Mediterranean region, Middle East and Central Asia. No correlation was found between NW L. infantum genotypes and clinical picture or host background. This study represents the first continent-wide analysis of NW L. infantum population structure. It confirmed that the agent of VL in the NW is L. infantum and that the parasite has been recently imported multiple times to the NW from southwest Europe. PMID:21666787

  10. Novel polymorphic microsatellite loci for distinguishing rock bass (Ambloplites rupestris), Roanoke bass (Ambloplites cavifrons), and their hybrids.

    PubMed

    Eschenroeder, Jackman C; Roberts, James H

    2016-10-01

    The rock bass (Ambloplites rupestris) is a popular sport-fish native to the Mississippi and Great Lakes basins of North America. The species has been widely introduced outside its native range, including into Atlantic-slope streams of Virginia where it may hybridize with an imperiled, similar-looking congener, the Roanoke bass (Ambloplites cavifrons). In this study, we identified and evaluated novel molecular markers to facilitate identification of these species and study the extent of hybridization. Using molecular libraries developed from A. rupestris, we identified a suite of candidate nuclear microsatellite loci, synthesized primer sets, and tested these markers for amplification and polymorphism in populations of both species. We then calculated standard diversity statistics within and differentiation statistics between species, the latter providing an indication of marker power for distinguishing the species and their hybrids. Additionally, we evaluated our efficiency for identifying hybrids by classifying simulated genotypes of known ancestry. Eleven loci were polymorphic (2-22 alleles per locus) and reliably amplified in both species. Multilocus genetic differentiation between A. cavifrons and A. rupestris was quite high (F ST  = 0.66; D LR  = 19.3), indicating the high statistical power of this marker set for species and hybrid identification. Analyses of simulated data suggested these markers reliably distinguish between hybrids and non-hybrids, as well as between F1 hybrids and backcrossed individuals. This panel of 11 loci should prove useful for understanding patterns of hybridization between A. rupestris and A. cavifrons. As the first microsatellite markers developed for Ambloplites, these markers also should prove broadly useful for population genetic studies of this genus.

  11. Use of multilocus sequence typing for the investigation of colonisation by Candida albicans in intensive care unit patients.

    PubMed

    Cliff, P R; Sandoe, J A T; Heritage, J; Barton, R C

    2008-05-01

    A prospective study was performed to determine the prevalence of candidal colonisation on the general intensive care unit at a large teaching hospital. Colonisation with Candida spp. was found to be common, occurring in 79% of patients on the unit. C. albicans was the commonest species, colonising 64% of patients, followed by C. glabrata (18%) and C. parapsilosis (14%). Most of the members of staff tested carried Candida spp. at some point, although carriage appeared to be transient. C. parapsilosis was the most commonly isolated species from staff hands, whereas C. albicans was the most commonly isolated species from the mouth. The molecular epidemiology of C. albicans was investigated using Ca3 typing and multilocus sequence typing (MLST). MLST proved to be a reproducible typing method and a useful tool for the investigation of the molecular epidemiology of C. albicans. The results of the molecular typing provided evidence for the presence of an endemic strain on the unit, which was isolated repeatedly from patients and staff. This finding suggests horizontal transmission of C. albicans on the unit though it may also reflect the relative frequency of C. albicans strain types colonising patients on admission. This study has important implications for the epidemiology of systemic candidal infections.

  12. Analysis of the genetic distribution among members of Clostridium botulinum group I using a novel multilocus sequence typing (MLST) assay.

    PubMed

    Olsen, Jaran S; Scholz, Holger; Fillo, Silvia; Ramisse, Vincent; Lista, Florigio; Trømborg, Anette K; Aarskaug, Tone; Thrane, Ingjerd; Blatny, Janet M

    2014-01-01

    Clostridium botulinum is the etiological agent of botulism. Due to food-borne poisoning and the potential use of the extremely toxic botulinum neurotoxin (BoNT) from C. botulinum in bioterror or biocrime related actions, reliable high resolution typing methods for discriminating C. botulinum strains are needed. Partial sequencing of the adk, atpH, gyrB, proC, rpoD and spo0A genes from 51 various C. botulinum/sporogenes isolates was performed, resulting in 37 different sequence types (STs). Analysis of the sequence data revealed a genetic distribution in five larger clusters with a loose correlation to the BoNT serotypes. The developed MLST assay had a slightly lower resolution ability when compared to the MLVA (multilocus variable number of tandem repeat analysis), but the two methods resulted in similar subclusters of the strains possessing the BoNT serotypes A, B and F. The current work presents the development of a novel MLST assay useful for genotyping C. botulinum related to basic phylogenetic research and trace-back analysis in microbial forensic studies.

  13. Assignment of Staphylococcus Isolates to Groups by spa Typing, SmaI Macrorestriction Analysis, and Multilocus Sequence Typing

    PubMed Central

    Strommenger, B.; Kettlitz, C.; Weniger, T.; Harmsen, D.; Friedrich, A. W.; Witte, W.

    2006-01-01

    The implementation of the new clustering algorithm Based Upon Repeat Pattern (BURP) into the Ridom StaphType software tool enables clustering based on spa typing data for Staphylococcus aureus. We compared clustering results obtained by spa typing/BURP to those obtained by currently well-established methods, i.e., SmaI macrorestriction analysis and multilocus sequence typing/eBURST. A total of 99 clinical S. aureus strains, including MRSA and representing major clonal lineages associated with important kinds of infections which have been prevalent in Germany and Central Europe during the last 10 years, were used for comparison. SmaI macrorestriction analysis revealed the highest discriminatory power, and clustering results for all three methods resulted in concordance values ranging from 96.8% between the two sequence-based methods to 93.4% between spa typing/BURP and SmaI macrorestriction/cluster analysis. The results of this study indicate that spa typing, together with BURP clustering, is a useful tool in S. aureus epidemiology, especially because of ease of use and the advantages of unambiguous sequence analysis as well as reproducibility and exchange of typing data. PMID:16825376

  14. Multilocus phylogeny and recent rapid radiation of the viviparous sea snakes (Elapidae: Hydrophiinae).

    PubMed

    Sanders, Kate L; Lee, Michael S Y; Mumpuni; Bertozzi, Terry; Rasmussen, Arne R

    2013-03-01

    The viviparous sea snakes (Hydrophiinae: Hydrophiini) comprise a young but morphologically and ecologically diverse clade distributed throughout the Indo-Pacific. Despite presenting a very promising model for marine diversification studies, many relationships among the 62 species and 16 genera in Hydrophiini remain unresolved. Here, we extend previous taxonomic and genomic sampling for Hydrophiini using three mitochondrial fragments and five nuclear loci for multiple individuals of 39 species in 15 genera. Our results highlight many of the impediments to inferring phylogenies in recent rapid radiations, including low variation at all five nuclear markers, and conflicting relationships supported by mitochondrial and nuclear trees. However, concatenated Bayesian and likelihood analyses, and a multilocus coalescent tree, recovered concordant support for primary clades and several previously unresolved inter-specific groupings. The Aipysurus group is monophyletic, with egg-eating specialists forming separate, early-diverging lineages. All three monotypic semi-aquatic genera (Ephalophis, Parahydrophis and Hydrelaps) are robustly placed as early diverging lineages along the branch leading to the Hydrophis group, with Ephalophis recovered as sister to Parahydrophis. The molecular phylogeny implies extensive evolutionary convergence in feeding adaptations within the Hydrophis group, especially the repeated evolution of small-headed (microcephalic) forms. Microcephalophis (Hydrophis) gracilis is robustly recovered as a relatively distant sister lineage to all other sampled Hydrophis group species, here termed the 'core Hydrophis group'. Within the 'core Hydrophis group', Hydrophis is recovered as broadly paraphyletic, with several other genera nested within it (Pelamis, Enhydrina, Astrotia, Thalassophina, Acalyptophis, Kerilia, Lapemis, Disteira). Instead of erecting multiple new genera, we recommend dismantling the latter (mostly monotypic) genera and recognising a single

  15. Comparison of a retrotransposon-based marker with microsatellite markers for discriminating accessions of Vitis vinifera.

    PubMed

    Sant'Ana, G C; Ferreira, J L; Rocha, H S; Borém, A; Pasqual, M; Cançado, G M A

    2012-05-21

    Identification and knowledge concerning genetic diversity are fundamental for efficient management and use of grapevine germplasm. Recently, new types of molecular markers have been developed, such as retrotransposon-based markers. Because of their multilocus pattern, retrotransposon-based markers might be able to differentiate grapevine accessions with just one pair of primers. In order to evaluate the efficiency of this type of marker, we compared retrotransposon marker Tvv1 with seven microsatellite markers frequently used for genotyping of the genus Vitis (VVMD7, VVMD25, VVMD5, VVMD27, VVMD31, VVS2, and VZAG62). The reference population that we used consisted of 26 accessions of Vitis, including seven European varieties of Vitis vinifera, four North American varieties and hybrids of Vitis labrusca, and 15 rootstock hybrids obtained from crosses of several Vitis species. Individually, the Tvv1 and the group of seven SSR markers were capable of distinguishing all accessions except 'White Niagara' compared to 'Red Niagara'. Using the Structure software, the retrotransposon marker Tvv1 generated two clusters: one with V. vinifera plus North American varieties and the other comprising rootstocks. The seven SSR markers generated five clusters: V. vinifera, the North American varieties, and three groups of rootstock hybrids. The percentages of variation explained by the first two components in the principal coordinate analysis were 65.21 (Tvv1) and 50.42 (SSR markers) while the Mantel correlation between the distance matrixes generated by the two types of markers was 42.5%. We conclude that the Tvv1 marker is useful for DNA fingerprinting, but it lacks efficiency for discrimination of structured groups.

  16. Genetic Fingerprinting Using Microsatellite Markers in a Multiplex PCR Reaction: A Compilation of Methodological Approaches from Primer Design to Detection Systems.

    PubMed

    Krüger, Jacqueline; Schleinitz, Dorit

    2017-01-01

    Microsatellites are polymorphic DNA loci comprising repeated sequence motifs of two to five base pairs which are dispersed throughout the genome. Genotyping of microsatellites is a widely accepted tool for diagnostic and research purposes such as forensic investigations and parentage testing, but also in clinics (e.g. monitoring of bone marrow transplantation), as well as for the agriculture and food industries. The co-amplification of several short tandem repeat (STR) systems in a multiplex reaction with simultaneous detection helps to obtain more information from a DNA sample where its availability may be limited. Here, we introduce and describe this commonly used genotyping technique, providing an overview on available resources on STRs, multiplex design, and analysis.

  17. Characterization and Transferable Utility of Microsatellite Markers in the Wild and Cultivated Arachis Species

    PubMed Central

    Huang, Li; Wu, Bei; Zhao, Jiaojiao; Li, Haitao; Chen, Weigang; Zheng, Yanli; Ren, Xiaoping; Chen, Yuning; Zhou, Xiaojing; Lei, Yong; Liao, Boshou; Jiang, Huifang

    2016-01-01

    Microsatellite or simple sequence repeat (SSR) is one of the most widely distributed molecular markers that have been widely utilized to assess genetic diversity and genetic mapping for important traits in plants. However, the understanding of microsatellite characteristics in Arachis species and the currently available amount of high-quality SSR markers remain limited. In this study, we identified 16,435 genome survey sequences SSRs (GSS-SSRs) and 40,199 expressed sequence tag SSRs (EST-SSRs) in Arachis hypogaea and its wild relative species using the publicly available sequence data. The GSS-SSRs had a density of 159.9–239.8 SSRs/Mb for wild Arachis and 1,015.8 SSR/Mb for cultivated Arachis, whereas the EST-SSRs had the density of 173.5–384.4 SSR/Mb and 250.9 SSRs/Mb for wild and cultivated Arachis, respectively. The trinucleotide SSRs were predominant across Arachis species, except that the dinucleotide accounted for most in A. hypogaea GSSs. From Arachis GSS-SSR and EST-SSR sequences, we developed 2,589 novel SSR markers that showed a high polymorphism in six diverse A. hypogaea accessions. A genetic linkage map that contained 540 novel SSR loci and 105 anchor SSR loci was constructed by case of a recombinant inbred lines F6 population. A subset of 82 randomly selected SSR markers were used to screen 39 wild and 22 cultivated Arachis accessions, which revealed a high transferability of the novel SSRs across Arachis species. Our results provided informative clues to investigate microsatellite patterns across A. hypogaea and its wild relative species and potentially facilitate the germplasm evaluation and gene mapping in Arachis species. PMID:27243460

  18. Characterization of microsatellites and gene contents from genome shotgun sequences of mungbean (Vigna radiata (L.) Wilczek)

    PubMed Central

    2009-01-01

    Background Mungbean is an important economical crop in Asia. However, genomic research has lagged behind other crop species due to the lack of polymorphic DNA markers found in this crop. The objective of this work is to develop and characterize microsatellite or simple sequence repeat (SSR) markers from genome shotgun sequencing of mungbean. Result We have generated and characterized a total of 470,024 genome shotgun sequences covering 100.5 Mb of the mungbean (Vigna radiata (L.) Wilczek) genome using 454 sequencing technology. We identified 1,493 SSR motifs that could be used as potential molecular markers. Among 192 tested primer pairs in 17 mungbean accessions, 60 loci revealed polymorphism with polymorphic information content (PIC) values ranging from 0.0555 to 0.6907 with an average of 0.2594. Majority of microsatellite markers were transferable in Vigna species, whereas transferability rates were only 22.90% and 24.43% in Phaseolus vulgaris and Glycine max, respectively. We also used 16 SSR loci to evaluate phylogenetic relationship of 35 genotypes of the Asian Vigna group. The genome survey sequences were further analyzed to search for gene content. The evidence suggested 1,542 gene fragments have been sequence tagged, that fell within intersected existing gene models and shared sequence homology with other proteins in the database. Furthermore, potential microRNAs that could regulate developmental stages and environmental responses were discovered from this dataset. Conclusion In this report, we provided evidence of generating remarkable levels of diverse microsatellite markers and gene content from high throughput genome shotgun sequencing of the mungbean genomic DNA. The markers could be used in germplasm analysis, accessing genetic diversity and linkage mapping of mungbean. PMID:19930676

  19. Genome-wide survey and analysis of microsatellites in nematodes, with a focus on the plant-parasitic species Meloidogyne incognita

    PubMed Central

    2010-01-01

    Background Microsatellites are the most popular source of molecular markers for studying population genetic variation in eukaryotes. However, few data are currently available about their genomic distribution and abundance across the phylum Nematoda. The recent completion of the genomes of several nematode species, including Meloidogyne incognita, a major agricultural pest worldwide, now opens the way for a comparative survey and analysis of microsatellites in these organisms. Results Using MsatFinder, the total numbers of 1-6 bp perfect microsatellites detected in the complete genomes of five nematode species (Brugia malayi, Caenorhabditis elegans, M. hapla, M. incognita, Pristionchus pacificus) ranged from 2,842 to 61,547, and covered from 0.09 to 1.20% of the nematode genomes. Under our search criteria, the most common repeat motifs for each length class varied according to the different nematode species considered, with no obvious relation to the AT-richness of their genomes. Overall, (AT)n, (AG)n and (CT)n were the three most frequent dinucleotide microsatellite motifs found in the five genomes considered. Except for two motifs in P. pacificus, all the most frequent trinucleotide motifs were AT-rich, with (AAT)n and (ATT)n being the only common to the five nematode species. A particular attention was paid to the microsatellite content of the plant-parasitic species M. incognita. In this species, a repertoire of 4,880 microsatellite loci was identified, from which 2,183 appeared suitable to design markers for population genetic studies. Interestingly, 1,094 microsatellites were identified in 801 predicted protein-coding regions, 99% of them being trinucleotides. When compared against the InterPro domain database, 497 of these CDS were successfully annotated, and further assigned to Gene Ontology terms. Conclusions Contrasted patterns of microsatellite abundance and diversity were characterized in five nematode genomes, even in the case of two closely related

  20. Phylogeny and Strain Typing of Escherichia coli, Inferred from Variation at Mononucleotide Repeat Loci

    PubMed Central

    Diamant, Eran; Palti, Yniv; Gur-Arie, Riva; Cohen, Helit; Hallerman, Eric M.; Kashi, Yechezkel

    2004-01-01

    Multilocus sequencing of housekeeping genes has been used previously for bacterial strain typing and for inferring evolutionary relationships among strains of Escherichia coli. In this study, we used shorter intergenic sequences that contained simple sequence repeats (SSRs) of repeating mononucleotide motifs (mononucleotide repeats [MNRs]) to infer the phylogeny of pathogenic and commensal E. coli strains. Seven noncoding loci (four MNRs and three non-SSRs) were sequenced in 27 strains, including enterohemorrhagic (six isolates of O157:H7), enteropathogenic, enterotoxigenic, B, and K-12 strains. The four MNRs were also sequenced in 20 representative strains of the E. coli reference (ECOR) collection. Sequence polymorphism was significantly higher at the MNR loci, including the flanking sequences, indicating a higher mutation rate in the sequences flanking the MNR tracts. The four MNR loci were amplifiable by PCR in the standard ECOR A, B1, and D groups, but only one (yaiN) in the B2 group was amplified, which is consistent with previous studies that suggested that B2 is the most ancient group. High sequence compatibility was found between the four MNR loci, indicating that they are in the same clonal frame. The phylogenetic trees that were constructed from the sequence data were in good agreement with those of previous studies that used multilocus enzyme electrophoresis. The results demonstrate that MNR loci are useful for inferring phylogenetic relationships and provide much higher sequence variation than housekeeping genes. Therefore, the use of MNR loci for multilocus sequence typing should prove efficient for clinical diagnostics, epidemiology, and evolutionary study of bacteria. PMID:15066845

  1. CHARACTERIZATION OF MICROSATELLITE LOCI IN SCHOENOPLECTUS AMERICANUS (CYPERACEAE)

    EPA Science Inventory

    Schoenoplectus americanus is a model organism for studying ecological and ecosystem responses of salt marsh plant communities to global climate change. Here we characterize 16 microsatellite loci in S. americanus to facilitate studies on the genetic basis of phenotypic responses...

  2. A Novel Approach for Mining Polymorphic Microsatellite Markers In Silico

    PubMed Central

    Hoffman, Joseph I.; Nichols, Hazel J.

    2011-01-01

    An important emerging application of high-throughput 454 sequencing is the isolation of molecular markers such as microsatellites from genomic DNA. However, few studies have developed microsatellites from cDNA despite the added potential for targeting candidate genes. Moreover, to develop microsatellites usually requires the evaluation of numerous primer pairs for polymorphism in the focal species. This can be time-consuming and wasteful, particularly for taxa with low genetic diversity where the majority of primers often yield monomorphic polymerase chain reaction (PCR) products. Transcriptome assemblies provide a convenient solution, functional annotation of transcripts allowing markers to be targeted towards candidate genes, while high sequence coverage in principle permits the assessment of variability in silico. Consequently, we evaluated fifty primer pairs designed to amplify microsatellites, primarily residing within transcripts related to immunity and growth, identified from an Antarctic fur seal (Arctocephalus gazella) transcriptome assembly. In silico visualization was used to classify each microsatellite as being either polymorphic or monomorphic and to quantify the number of distinct length variants, each taken to represent a different allele. The majority of loci (n = 36, 76.0%) yielded interpretable PCR products, 23 of which were polymorphic in a sample of 24 fur seal individuals. Loci that appeared variable in silico were significantly more likely to yield polymorphic PCR products, even after controlling for microsatellite length measured in silico. We also found a significant positive relationship between inferred and observed allele number. This study not only demonstrates the feasibility of generating modest panels of microsatellites targeted towards specific classes of gene, but also suggests that in silico microsatellite variability may provide a useful proxy for PCR product polymorphism. PMID:21853104

  3. Development of microsatellite markers for Hancornia speciosa Gomes (Apocynaceae).

    PubMed

    Rodrigues, A J L; Yamaguishi, A T; Chaves, L J; Coelho, A S G; Lima, J S; Telles, M P C

    2015-07-03

    Herein, we describe 34 microsatellite loci developed using an enrichment genomic library for the tree species Hancornia speciosa Gomes (Apocynaceae). Thirty-five individuals were genotyped using 34 primers to analyze the polymorphisms at each locus. The number of alleles per locus ranged from 4 to 20. The average number of alleles was 8.11, and the expected heterozygosity ranged from 0.62 to 0.94. These microsatellite primers will be useful in population genetics studies for this species.

  4. Rapid isolation of microsatellite DNAs and identification of polymorphic mitochondrial DNA regions in the fish rotan (Perccottus glenii) invading European Russia

    USGS Publications Warehouse

    King, Timothy L.; Eackles, Michael S.; Reshetnikov, Andrey N.

    2015-01-01

    Human-mediated translocations and subsequent large-scale colonization by the invasive fish rotan (Perccottus glenii Dybowski, 1877; Perciformes, Odontobutidae), also known as Amur or Chinese sleeper, has resulted in dramatic transformations of small lentic ecosystems. However, no detailed genetic information exists on population structure, levels of effective movement, or relatedness among geographic populations of P. glenii within the European part of the range. We used massively parallel genomic DNA shotgun sequencing on the semiconductor-based Ion Torrent Personal Genome Machine (PGM) sequencing platform to identify nuclear microsatellite and mitochondrial DNA sequences in P. glenii from European Russia. Here we describe the characterization of nine nuclear microsatellite loci, ascertain levels of allelic diversity, heterozygosity, and demographic status of P. glenii collected from Ilev, Russia, one of several initial introduction points in European Russia. In addition, we mapped sequence reads to the complete P. glenii mitochondrial DNA sequence to identify polymorphic regions. Nuclear microsatellite markers developed for P. glenii yielded sufficient genetic diversity to: (1) produce unique multilocus genotypes; (2) elucidate structure among geographic populations; and (3) provide unique perspectives for analysis of population sizes and historical demographics. Among 4.9 million filtered P. glenii Ion Torrent PGM sequence reads, 11,304 mapped to the mitochondrial genome (NC_020350). This resulted in 100 % coverage of this genome to a mean coverage depth of 102X. A total of 130 variable sites were observed between the publicly available genome from China and the studied composite mitochondrial genome. Among these, 82 were diagnostic and monomorphic between the mitochondrial genomes and distributed among 15 genome regions. The polymorphic sites (N = 48) were distributed among 11 mitochondrial genome regions. Our results also indicate that sequence reads generated

  5. Multilocus sequence typing of ‘Candidatus Liberibacter solanacearum’ isolates in North America and New Zealand.

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A panel of 10 multilocus sequence typing (MLST) markers for‘Candidatus Liberibacter solanacearum’ was developed. Using this marker system, genetic relationships among ‘Ca. L. solanacearum’ strains in North America (USA and Mexico) and New Zealand were characterized. MLST analysis differentiated 59 s...

  6. Multilocus and Multitrait Measures of Differentiation for Gene Markers and Phenotypic Traits

    PubMed Central

    Kremer, A.; Zanetto, A.; Ducousso, A.

    1997-01-01

    Multilocus measures of differentiation taking into account gametic disequilibrium are developed. Even if coupling and repulsion heterozygotes cannot be separated at the multilocus level, a method is given to calculate a composite measure of differentiation (CF(st)) at the zygotic level, which accounts for allelic associations combining both gametic and nongametic effects. Mean and maximum differentiations may be relevant when multilocus measures are computed. Maximum differentiation is the highest eigenvalue of the F(st) matrix, whereas mean differentiation corresponds to the mean value of all eigenvalues of the F(st) matrix. Gametic disequilibrium has a stronger effect on maximum differentiation than on mean differentiation and takes into account the anisotropy that may exist between within- and between-population components of disequilibria. Multilocus mean and maximum differentiation are calculated for a set of 81 Quercus petraea (sessile oak) populations assessed with eight allozyme loci and two phenotypic traits (bud burst and height growth). The results indicate that maximum differentiation increases as more loci (traits) are considered whereas mean differentiation remains constant or decreases. Phenotypic traits exhibit higher population differentiation than allozymes. The applications and uses of mean and maximum differentiations are further discussed. PMID:9093871

  7. Novel fusarium head blight pathogens from Nepal and Louisiana revealed by multilocus genealogical concordance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    This study was conducted to assess evolutionary relationships, species diversity, and trichothecene toxin potential of five Fusarium graminearum complex (FGSC) isolates identified as genetically novel during prior Fusarium head blight (FHB) surveys in Nepal and Louisiana. Results of a multilocus gen...

  8. Analysis of Multilocus Sequence Typing for Identification of Leptospira Isolates in Brazil ▿

    PubMed Central

    Romero, E. C.; Blanco, R. M.; Galloway, R. L.

    2011-01-01

    A collection of 101 Leptospira isolates was tested by multilocus sequence typing (MLST) and by traditional serotyping. MLST divided the isolates into 4 sequence types (STs), while serotyping classified them into 6 serogroups. Two isolates failed to generate products for some genes by MLST. MLST was less discriminatory than serotyping for uncommonly occurring isolates from humans in Brazil. PMID:21880969

  9. Microsatellite genotyping reveals a signature in breast cancer exomes.

    PubMed

    McIver, L J; Fonville, N C; Karunasena, E; Garner, H R

    2014-06-01

    Genomic instability at microsatellite loci is a hallmark of many cancers, including breast cancer. However, much of the genomic variation and many of the hereditary components responsible for breast cancer remain undetected. We hypothesized that variation at microsatellites could provide additional genomic markers for breast cancer risk assessment. A total of 1,345 germline and tumor DNA samples from individuals diagnosed with breast cancer, exome sequenced as part of The Cancer Genome Atlas, were analyzed for microsatellite variation. The comparison group for our analysis, representing healthy individuals, consisted of 249 females which were exome sequenced as part of the 1,000 Genomes Project. We applied our microsatellite-based genotyping pipeline to identify 55 microsatellite loci that can distinguish between the germline of individuals diagnosed with breast cancer and healthy individuals with a sensitivity of 88.4 % and a specificity of 77.1 %. Further, we identified additional microsatellite loci that are potentially useful for distinguishing between breast cancer subtypes, revealing a possible fifth subtype. These findings are of clinical interest as possible risk diagnostics and reveal genes that may be of potential therapeutic value, including genes previously not associated with breast cancer.

  10. Genome fingerprinting by simple sequence repeat (SSR)-anchored polymerase chain reaction amplification

    SciTech Connect

    Zietkiewicz, E.; Labuda, D. ); Rafalski, A. )

    1994-03-15

    Simple sequence repeats (SSR), or microsatellites, are ubiquitous in eukaryotic genomes. Here the authors demonstrate the utility of microsatellite-directed DNA fingerprinting by polymerase chain reaction (PCR) amplification of the interrepeat region. No sequencing is required to design the oligonucleotide primers. They tested primers anchored at 3[prime] or 5[prime] termini of the (A)[sub n] repeats, extended into the flanking sequence by 2 to 4 nucleotide residues [3[prime]-anchored primers: (CA)[sub 8]RG, (CA)[sub 8]RY, and (CA)[sub 7]RTCY; and 5[prime]-anchored primers: BDB(CA)[sub 7]C, DBDA(CA)[sub 7], VHVG(TG)[sub 7] and HVH(TG)[sub 7]T]. Radioactively labeled amplification products were analyzed by electrophoresis, revealing information on multiple genomic loci in a single gel lane. Complex, species-specific patterns were obtained from a variety of eukaryotic taxa. Intraspecies polymorphisms were also observed and shown to segregate as Mendelian markers. Inter-SSR PCR provides a novel fingerprinting approach applicable for taxonomic and phylogenetic comparisons and as a mapping tool in a wide range of organisms. This application of (CA)[sub n] repeats may be extended to different microsatellites and other common dispersed elements. 24 refs., 6 figs.

  11. Repeating the Past

    NASA Astrophysics Data System (ADS)

    Moore, John W.

    1998-05-01

    As part of the celebration of the Journal 's 75th year, we are scanning each Journal issue from 25, 50, and 74 years ago. Many of the ideas and practices described are so similar to present-day "innovations" that George Santayana's adage (1) "Those who cannot remember the past are condemned to repeat it" comes to mind. But perhaps "condemned" is too strong - sometimes it may be valuable to repeat something that was done long ago. One example comes from the earliest days of the Division of Chemical Education and of the Journal.

  12. Characterization of Iranian nonaflatoxigenic strains of Aspergillus flavus based on microsatellite-primed PCR

    PubMed Central

    Houshyarfard, Mahmoud; Rouhani, Hamid; Falahati-Rastegar, Mahrokh; Malekzadeh-Shafaroudi, Saeid; Mahdikhani-Moghaddam, Esmat

    2015-01-01

    Out of fifty-two Iranian nonaflatoxigenic strains of Aspergillus flavus,collected from various substrates (soil and kernel) and sources (peanut, corn and pistachio), fifteen representatives were selected according to their different geographical origins (six provinces: Guilan and Golestan, Ardebil, Fars, Kerman and Semnan) and vegetative compatibility groups (VCGs, IR1 to IR15) for microsatellite-primed PCR analysis. Two inter-simple sequence repeat (ISSR) primers AFMPP and AFM13 were used to determine polymorphism and the relationship among strain isolates. A. flavus isolates were identified by their morphologies and their identities were confirmed by PCR amplification using the specific primer pair ITS1 and ITS4. The results revealed variations in the percentages of polymorphisms. In the ISSR analysis, primers AFMPP and AFM13 generated a total of 18 and 23 amplicons among the fungal strains, out of which 12 (66.7%) and 22 (95.7%) were polymorphic, respectively. Cluster analysis of the ISSR data was carried out using 1 D DNA gel image analysis. The two dendrograms obtained through these markers showed six different clusterings of testing nonaflatoxigenic A. flavus L strains, but we noticed that some clusters were different in some cases. The microsatellite-primed PCR data revealed that the Iranian nonaflatoxigenic isolates of A. flavus were not clustered according to their origins and sources. This study is the first to characterize Iranian nonaflatoxigenic isolates of A. flavus using ISSR markers. PMID:27843995

  13. Microsatellite markers isolated from Cabomba aquatica s.l. (Cabombaceae) from an enriched genomic library1

    PubMed Central

    Barbosa, Tiago D. M.; Trad, Rafaela J.; Bajay, Miklos M.; Amaral, Maria C. E.

    2015-01-01

    Premise of the study: Microsatellite primers were designed for the submersed aquatic plant Cabomba aquatica s.l. (Cabombaceae) and characterized to estimate genetic diversity parameters. Methods and Results: Using a selective hybridization method, we designed and tested 30 simple sequence repeat loci using two natural populations of C. aquatica s.l., resulting in 13 amplifiable loci. Twelve loci were polymorphic, and alleles per locus ranged from two to four across the 49 C. aquatica s.l. individuals. Observed heterozygosity, expected heterozygosity, and fixation index varied from 0.0 to 1.0, 0.0 to 0.5, and −1.0 to −0.0667, respectively, for the Manaus population and from 0.0 to 1.0, 0.0 to 0.6, and −1.0 to 0.4643 for the Viruá population. Conclusions: The developed markers will be used in further taxonomic and population studies within Cabomba. This set of microsatellite primers represents the first report on rapid molecular markers in the genus. PMID:26649271

  14. CmMDb: a versatile database for Cucumis melo microsatellite markers and other horticulture crop research.

    PubMed

    Bhawna; Chaduvula, Pavan K; Bonthala, Venkata S; Manjusha, Verma; Siddiq, Ebrahimali A; Polumetla, Ananda K; Prasad, Gajula M N V

    2015-01-01

    Cucumis melo L. that belongs to Cucurbitaceae family ranks among one of the highest valued horticulture crops being cultivated across the globe. Besides its economical and medicinal importance, Cucumis melo L. is a valuable resource and model system for the evolutionary studies of cucurbit family. However, very limited numbers of molecular markers were reported for Cucumis melo L. so far that limits the pace of functional genomic research in melon and other similar horticulture crops. We developed the first whole genome based microsatellite DNA marker database of Cucumis melo L. and comprehensive web resource that aids in variety identification and physical mapping of Cucurbitaceae family. The Cucumis melo L. microsatellite database (CmMDb: http://65.181.125.102/cmmdb2/index.html) encompasses 39,072 SSR markers along with its motif repeat, motif length, motif sequence, marker ID, motif type and chromosomal locations. The database is featured with novel automated primer designing facility to meet the needs of wet lab researchers. CmMDb is a freely available web resource that facilitates the researchers to select the most appropriate markers for marker-assisted selection in melons and to improve breeding strategies.

  15. Genetic relationships in the peregrine falcon (Falco peregrinus) analysed by microsatellite DNA markers.

    PubMed

    Nesje, M; Røed, K H; Lifjeld, J T; Lindberg, P; Steen, O F

    2000-01-01

    Microsatellite DNA markers were developed from a peregrine falcon (Falco peregrinus) and genetic relationships among peregrine falcons in southern Norway were analysed using the markers. The genomic DNA library was screened for the presence of dinucleotide microsatellite repeats. Twelve loci revealed polymorphism through the initial analysis of 24 unrelated peregrine falcons, and Mendelian inheritance was confirmed in two peregrine falcon families bred in captivity. The estimated mean probability of identical genotypes in two unrelated individuals was 3 x 10-8, and the combined exclusion probability for parentage testing was 0.99 and 0.94 for one or both parents unknown, respectively. The markers were used to investigate the parentage of peregrine broods from the same nest site from different breeding seasons, and subsequently the nest-site fidelity of the breeding peregrines. High nest-site fidelity was found by studying pairwise comparisons of relatedness (rxy) estimates among chicks at six nest sites from three different breeding seasons. Cross-species amplifications showed that most loci also appeared to amplify polymorphic products in the gyrfalcon (F. rusticolus), merlin (F. columbarius), hobby (F. subbuteo) and kestrel (F. tinnunculus), demonstrating that the loci will provide powerful genetic markers in these falcons too.

  16. A novel microsatellite (STR) marker for forensic identification of big cats in India.

    PubMed

    Singh, Anju; Gaur, Ajay; Shailaja, K; Satyare Bala, B; Singh, Lalji

    2004-05-10

    India is the home to five of the eight majestic big cats of the world. The three major big cats namely, lion, tiger, and leopard are listed in the Schedule I of the Indian Wildlife Protection Act, 1972. Apart from the severe loss of the habitat, these are continuously facing the danger of extinction mainly due to poaching and hunting for their body parts, which are being greatly valued by apothecaries marketing traditional Chinese medicines. With the advent of polymerase chain reaction (PCR), DNA-based markers have emerged as major tools in the arena of wildlife forensics. Microsatellites (short tandem repeats, STRs) are markers of choice because of their polymorphic and co-dominant nature. These strictly follow the Mendelian inheritance and are highly reproducible. We have identified a new microsatellite (STR) locus Ple 46, which shows amplification in a species-specific manner (size of STR) in all the members of the family felidae studied here. This PCR-based, non-invasive method opens a new avenue to forensic identification of big cats.

  17. CmMDb: A Versatile Database for Cucumis melo Microsatellite Markers and Other Horticulture Crop Research

    PubMed Central

    Bhawna; Chaduvula, Pavan K.; Bonthala, Venkata S.; Manjusha, Verma; Siddiq, Ebrahimali A.; Polumetla, Ananda K.; Prasad, Gajula M. N. V.

    2015-01-01

    Cucumis melo L. that belongs to Cucurbitaceae family ranks among one of the highest valued horticulture crops being cultivated across the globe. Besides its economical and medicinal importance, Cucumis melo L. is a valuable resource and model system for the evolutionary studies of cucurbit family. However, very limited numbers of molecular markers were reported for Cucumis melo L. so far that limits the pace of functional genomic research in melon and other similar horticulture crops. We developed the first whole genome based microsatellite DNA marker database of Cucumis melo L. and comprehensive web resource that aids in variety identification and physical mapping of Cucurbitaceae family. The Cucumis melo L. microsatellite database (CmMDb: http://65.181.125.102/cmmdb2/index.html) encompasses 39,072 SSR markers along with its motif repeat, motif length, motif sequence, marker ID, motif type and chromosomal locations. The database is featured with novel automated primer designing facility to meet the needs of wet lab researchers. CmMDb is a freely available web resource that facilitates the researchers to select the most appropriate markers for marker-assisted selection in melons and to improve breeding strategies. PMID:25885062

  18. Microsatellite markers derived from Quercus mongolica var. crispula (Fagaceae) inner bark expressed sequence tags.

    PubMed

    Ueno, Saneyoshi; Taguchi, Yuriko; Tsumura, Yoshihiko

    2008-04-01

    In reforestation programs the genetic composition and diversity of populations that could be used as sources of planting material needs to be carefully considered to maximize the chances of successful establishment. For such purposes genetic analyses that include the identification of functional genes are required. In this study, we constructed a cDNA library from inner bark of Quercus mongolica (which is widely distributed in Japan) and collected 3385 ESTs. After constructing 2140 unigenes, 274 microsatellites were found within them. The most frequent microsatellite had AG motif (48%) and the next most common was AAG motif (12%). There were no CG repeats in the unigenes. In total, 20 EST-SSR markers were developed, polymorphisms of which were described by using eight individuals from eight populations over the species' distributional range. The number of alleles per locus (Na) and observed heterozygosity (H(o)) ranged from 2 to 12, and from 0.25 to 1.00, respectively. Cross-species amplification was successful for 19 loci in eight individuals of Q. serrata and for 20 loci in eight individuals of Q. dentata, with values of Na and H(o) comparable to those of Q. mongolica. The EST-SSR markers characterized in this study should facilitate the analysis of genetic diversity in future studies.

  19. Reduced microsatellite heterozygosity in island endemics supports the role of long-term effective population size in avian microsatellite diversity

    PubMed Central

    2011-01-01

    Because of the high mutation rate of microsatellites, polymorphism at microsatellite loci might be predicted to reflect the effective population size over a time span of about 10,000 years and thus to be associated with biogeographic factors impacting species on that time frame. This prediction was tested by comparing heterozygosity at microsatellite loci from 294 bird species, including 58 species endemic to oceanic islands. Controlling statistically for phylogenetic effects, mean heterozygosity was significantly reduced in oceanic island endemics compared to other species. There was also an effect of current endangerment, statistically independent of the effect of island endemicity. These results support the hypothesis that long-term effective population size can be an important causative factor behind differences among species with respect to microsatellite heterozygosity. PMID:21104002

  20. Validating the use of colouration patterns for individual recognition in the worm pipefish using a novel set of microsatellite markers.

    PubMed

    Monteiro, N M; Silva, R M; Cunha, M; Antunes, A; Jones, A G; Vieira, M N

    2014-01-01

    In studies of behaviour, ecology and evolution, identification of individual organisms can be an invaluable tool, capable of unravelling otherwise cryptic information regarding group structure, movement patterns, population size and mating strategies. The use of natural markings is arguably the least invasive method for identification. However, to be truly useful natural markings must be sufficiently variable to allow for unique identification, while being stable enough to permit long-term studies. Non-invasive marking techniques are especially important in fishes of the Family Syngnathidae (pipefishes, seahorses and seadragons), as many of these taxa are of conservation concern or used extensively in studies of sexual selection. Here, we assessed the reliability of natural markings as a character for individual identification in a wild population of Nerophis lumbriciformis by comparing results from natural markings to individual genetic assignments based on eight novel microsatellite loci. We also established a minimally invasive method based on epithelial cell swabbing to sample DNA. All pipefish used in the validation of natural markings, independently of sex or time between recaptures, were individually recognized through facial colouration patterns. Their identities were verified by the observation of the same multilocus genotype at every sampling event for each individual that was identified on the basis of natural markings. Successful recaptures of previously swabbed pipefish indicated that this process probably did not induce an elevated rate of mortality. Also, the recapture of newly pregnant males showed that swabbing did not affect reproductive behaviour.

  1. Microsatellite diversity and genetic structure among common bean (Phaseolus vulgaris L.) landraces in Brazil, a secondary center of diversity

    PubMed Central

    Burle, Marília Lobo; Fonseca, Jaime Roberto; Kami, James A.

    2010-01-01

    Brazil is the largest producer and consumer of common bean (Phaseolus vulgaris L.), which is the most important source of human dietary protein in that country. This study assessed the genetic diversity and the structure of a sample of 279 geo-referenced common bean landraces from Brazil, using molecular markers. Sixty-seven microsatellite markers spread over the 11 linkage groups of the common bean genome, as well as Phaseolin, PvTFL1y, APA and four SCAR markers were used. As expected, the sample showed lower genetic diversity compared to the diversity in the primary center of diversification. Andean and Mesoamerican gene pools were both present but the latter gene pool was four times more frequent than the former. The two gene pools could be clearly distinguished; limited admixture was observed between these groups. The Mesoamerican group consisted of two sub-populations, with a high level of admixture between them leading to a large proportion of stabilized hybrids not observed in the centers of domestication. Thus, Brazil can be considered a secondary center of diversification of common bean. A high degree of genome-wide multilocus associations even among unlinked loci was observed, confirming the high level of structure in the sample and suggesting that association mapping should be conducted in separate Andean and Mesoamerican Brazilian samples. Electronic supplementary material The online version of this article (doi:10.1007/s00122-010-1350-5) contains supplementary material, which is available to authorized users. PMID:20502861

  2. A comparative analysis of distribution and conservation of microsatellites in the transcripts of sequenced Fusarium species and development of genic-SSR markers for polymorphism analysis.

    PubMed

    Mahfooz, Sahil; Srivastava, Arpita; Srivastava, Alok K; Arora, Dilip K

    2015-09-01

    We used an in silico approach to survey and compare microsatellites in transcript sequences of four sequenced members of genus Fusarium. G + C content of transcripts was found to be positively correlated with the frequency of SSRs. Our analysis revealed that, in all the four transcript sequences studied, the occurrence, relative abundance and density of microsatellites varied and was not influenced by transcript sizes. No correlation between relative abundance and transcript sizes was observed. The relative abundance and density of microsatellites were highest in the transcripts of Fusarium solani when compared with F. graminearum, F. verticillioides and F. oxysporum. The maximum frequency of SSRs among all four sequence sets was of trinucleotide repeats (67.8%), whereas the dinucleotide repeat represents <1%. Among all classes of repeats, 36.5% motifs were found conserved within Fusarium species. In order to study polymorphism within Fusarium isolates, 11 polymorphic genic-SSR markers were developed. Of the 11 markers, 5 were from F. oxysporum and remaining 6 belongs to F. solani. SSR markers from F. oxysporum were found to be more polymorphic (38%) as compared to F. solani (26%). Eleven polymorphic markers obtained in this study clearly demonstrate the utility of newly developed SSR markers in establishing genetic relationships among different isolates of Fusarium.

  3. Microsatellite (SSR) amplification by PCR usually led to polymorphic bands: Evidence which shows replication slippage occurs in extend or nascent DNA strands

    PubMed Central

    Hosseinzadeh-Colagar, Abasalt; Haghighatnia, Mohammad Javad; Amiri, Zahra; Mohadjerani, Maryam; Tafrihi, Majid

    2016-01-01

    Microsatellites or simple sequence repeats (SSRs) are very effective molecular markers in population genetics, genome mapping, taxonomic study and other large-scale studies. Variation in number of tandem repeats within microsatellite refers to simple sequence length polymorphism (SSLP); but there are a few studies that are showed SSRs replication slippage may be occurred during in vitro amplification which are produced ‘stutter products’ differing in length from the main products. The purpose of this study is introducing a reliable method to realize SSRs replication slippage. At first, three unique primers designed to amplify SSRs loci in the great gerbil (Rhombomys opimus) by PCR. Crush and soak method used to isolate interesting DNA bands from polyacrylamide gel. PCR products analyzed using by sequencing methods. Our study has been shown that Taq DNA polymerase slipped during microsatellite in vitro amplification which led to insertion or deletion of repeats in sense or antisense DNA strands. It is produced amplified fragments with various lengths in gel electrophoresis showed as ‘stutter bands’. Thus, in population studies by SSRs markers recommend that replication slippage effects and stutter bands have been considered. PMID:28097170

  4. Microsatellite (SSR) amplification by PCR usually led to polymorphic bands: Evidence which shows replication slippage occurs in extend or nascent DNA strands.

    PubMed

    Hosseinzadeh-Colagar, Abasalt; Haghighatnia, Mohammad Javad; Amiri, Zahra; Mohadjerani, Maryam; Tafrihi, Majid

    2016-09-01

    Microsatellites or simple sequence repeats (SSRs) are very effective molecular markers in population genetics, genome mapping, taxonomic study and other large-scale studies. Variation in number of tandem repeats within microsatellite refers to simple sequence length polymorphism (SSLP); but there are a few studies that are showed SSRs replication slippage may be occurred during in vitro amplification which are produced 'stutter products' differing in length from the main products. The purpose of this study is introducing a reliable method to realize SSRs replication slippage. At first, three unique primers designed to amplify SSRs loci in the great gerbil (Rhombomys opimus) by PCR. Crush and soak method used to isolate interesting DNA bands from polyacrylamide gel. PCR products analyzed using by sequencing methods. Our study has been shown that Taq DNA polymerase slipped during microsatellite in vitro amplification which led to insertion or deletion of repeats in sense or antisense DNA strands. It is produced amplified fragments with various lengths in gel electrophoresis showed as 'stutter bands'. Thus, in population studies by SSRs markers recommend that replication slippage effects and stutter bands have been considered.

  5. De novo development and characterization of polymorphic microsatellite markers in a schilbid catfish, Silonia silondia (Hamilton, 1822) and their validation for population genetic studies.

    PubMed

    Mandal, Sangeeta; Jena, J K; Singh, Rajeev K; Mohindra, Vindhya; Lakra, W S; Deshmukhe, Geetanjali; Pathak, Abhinav; Lal, Kuldeep K

    2016-02-01

    The stock characterization of wild populations of Silonia silondia is important for its scientific management. At present, the information on genetic parameters of S. silondia is very limited. The species-specific microsatellite markers were developed in current study. The validated markers were used to genotype individuals from four distant rivers. To develop de novo microsatellite loci, an enriched genomic library was constructed for S. silondia using affinity-capture approach. The markers were validated for utility in population genetics. A total number of 76 individuals from four natural riverine populations were used to generate data for population analysis. The screening of isolated repeat sequences yielded eleven novel polymorphic microsatellite loci. The microsatellite loci exhibited high level of polymorphism, with 6-24 alleles per locus and the PIC value ranged from 0.604 to 0.927. The observed (Ho) and expected (He) heterozygosities ranged from 0.081 to 0.84 and 0.66 to 0.938, respectively. The AMOVA analysis indicated significant genetic differentiation among riverine populations (overall FST = 0.075; P < 0.0001) with maximum variation (92.5%) within populations. Cross-priming assessment revealed successful amplification (35-38 %) of heterologous loci in four related species viz. Clupisoma garua, C. taakree, Ailia coila and Eutropiichthys vacha. The results demonstrated that these de novo polymorphic microsatellite loci are promising for population genetic variation and diversity studies in S. silondia. Cross-priming results indicated that these primers can help to get polymorphic microsatellite loci in the related catfish species of family Schilbidae.

  6. Characterization of 13 microsatellites for Lahontan cutthroat trout (Oncorhynchus clarki henshawi) and cross-amplification in six other salmonids.

    PubMed

    Robinson, M L; Kirchoff, V S; Peacock, M M

    2009-01-01

    Thirteen newly developed tri- and tetranucleotide repeat microsatellite markers were developed for Lahontan cutthroat trout (Oncorhynchus clarki henshawi), a threatened subspecies endemic to the Lahontan hydrographic basin in the western USA. These loci are highly polymorphic with five to 30 alleles per locus and observed heterozygosities ranging from 0.4 to 0.7. Cross-species amplification of these markers was most successful in the closely related rainbow trout, Oncorhynchus mykiss, with only three loci amplifying in brown trout, Salmo trutta. Nonoverlapping allelic distributions for many of these loci among the six salmonid species screened suggest these markers may be useful for hybrid determination.

  7. Assessment of genetic diversity and relationships among wild and cultivated Tunisian plums (Prunus spp) using random amplified microsatellite polymorphism markers.

    PubMed

    Ben Tamarzizt, H; Ben Mustapha, S; Baraket, G; Abdallah, D; Salhi-Hannachi, A

    2015-03-20

    The usefulness of random amplified microsatellite polymorphism markers to study the genetic diversity and relationships among cultivars belonging to Prunus salicina and P. domestica and their wild relatives (P. insititia and P. spinosa) was investigated. A total of 226 of 234 bands were polymorphic (96.58%). The 226 random amplified microsatellite polymorphism markers were screened using 15 random amplified polymorphic DNA and inter-simple sequence repeat primers combinations for 54 Tunisian plum accessions. The percentage of polymorphic bands (96.58%), the resolving power of primers values (135.70), and the polymorphic information content demonstrated the efficiency of the primers used in this study. The genetic distances between accessions ranged from 0.18 to 0.79 with a mean of 0.24, suggesting a high level of genetic diversity at the intra- and interspecific levels. The unweighted pair group with arithmetic mean dendrogram and principal component analysis discriminated cultivars efficiently and illustrated relationships and divergence between spontaneous, locally cultivated, and introduced plum types. These procedures showed continuous variation that occurs independently of the status of the species and geographical origin of the plums. In this study, random amplified microsatellite polymorphism was found to be as a reliable molecular marker for fingerprinting and for examining the diversity study of the plum and its relatives.

  8. Isolation and characterization of microsatellite loci for the isopod crustacean Armadillidium vulgare and transferability in terrestrial isopods.

    PubMed

    Giraud, Isabelle; Valette, Victorien; Bech, Nicolas; Grandjean, Fré