[The gene pool of Belgorod oblast population: study of biochemical gene markers in populations of Ukraine and Belarus and the position of the Belgorod population in the Eastern Slavic gene pool system].

PubMed

Lependina, I N; Churnosov, M I; Artamentova, L A; Ishchuk, M A; Tegako, O V; Balanovskaia, E V

2008-04-01

The characteristics of the gene pools of indigenous populations of Ukraine and Belarus have been studied using 28 alleles of 10 loci of biochemical gene markers (HP, GC, TF, PI, C'3, ACP1, GLO1, PGM1, ESD, and 6-PGD). The gene pools of the Russian and Ukrainian indigenous populations of Belgorod oblast (Russia) and the indigenous populations of Ukraine and Belarus have been compared. Cluster analysis, multidimensional scaling, and factor analysis of the obtained data have been used to determine the position of the Belgorod population gene pool in the Eastern Slavic gene pool system.

Genetic diversity, population structure and subdivision of local Balkan pig breeds in Austria, Croatia, Serbia and Bosnia-Herzegovina and its practical value in conservation programs.

PubMed

Druml, Thomas; Salajpal, Kresimir; Dikic, Maria; Urosevic, Miroslav; Grilz-Seger, Gertrud; Baumung, Roswitha

2012-03-01

At present the Croatian Turopolje pig population comprises about 157 breeding animals. In Austria, 324 Turopolje pigs originating from six Croatian founder animals are registered. Multiple bottlenecks have occurred in this population, one major one rather recently and several more older and moderate ones. In addition, it has been subdivided into three subpopulations, one in Austria and two in Croatia, with restricted gene flow. These specificities explain the delicate situation of this endangered Croatian lard-type pig breed. In order to identify candidate breeding animals or gene pools for future conservation breeding programs, we studied the genetic diversity and population structure of this breed using microsatellite data from 197 individuals belonging to five different breeds. The genetic diversity of the Turopolje pig is dramatically low with observed heterozygosities values ranging from 0.38 to 0.57. Split into three populations since 1994, two genetic clusters could be identified: one highly conserved Croatian gene pool in Turopoljski Lug and the"Posavina" gene pool mainly present in the Austrian population. The second Croatian subpopulation in Lonjsko Polje in the Posavina region shows a constant gene flow from the Turopoljski Lug animals. One practical conclusion is that it is necessary to develop a "Posavina" boar line to preserve the "Posavina" gene pool and constitute a corresponding population in Croatia. Animals of the highly inbred herd in Turopoljski Lug should not be crossed with animals of other populations since they represent a specific phenotype-genotype combination. However to increase the genetic diversity of this herd, a program to optimize its sex ratio should be carried out, as was done in the Austrian population where the level of heterozygosity has remained moderate despite its heavy bottleneck in 1994. © 2012 Druml et al; licensee BioMed Central Ltd.

Structural and functional diversity of microbial communities from a lake sediment contaminated with trenbolone, an endocrine-disrupting chemical.

PubMed

Radl, Viviane; Pritsch, Karin; Munch, Jean Charles; Schloter, Michael

2005-09-01

Effects of trenbolone (TBOH), a hormone used in cattle production, on the structure and function of microbial communities in a fresh water sediment from a lake in Southern Germany were studied in a microcosm experiment. The microbial community structure and the total gene pool of the sediment, assessed by 16S rRNA/rDNA and RAPD fingerprint analysis, respectively, were not significantly affected by TBOH. In contrast, the N-acetyl-glucosaminidase activity was almost 50% lower in TBOH treated samples (P<0.05). Also, the substrate utilization potential, measured using the BIOLOG system, was reduced after TBOH treatment. Interestingly, this potential did not recover at the end of the experiment, i.e. 19 days after the addition of the chemical. Repeated application of TBOH did not lead to an additional reduction in the substrate utilization potential. Overall results indicate that microbial community function was more sensitive to TBOH treatment than the community structure and the total gene pool.

Beans in Europe: origin and structure of the European landraces of Phaseolus vulgaris L.

PubMed

Angioi, S A; Rau, D; Attene, G; Nanni, L; Bellucci, E; Logozzo, G; Negri, V; Spagnoletti Zeuli, P L; Papa, R

2010-09-01

This study focuses on the expansion of Phaseolus vulgaris in Europe. The pathways of distribution of beans into and across Europe were very complex, with several introductions from the New World that were combined with direct exchanges between European and other Mediterranean countries. We have analyzed here six chloroplast microsatellite (cpSSR) loci and two unlinked nuclear loci (for phaseolin types and Pv-shatterproof1). We have assessed the genetic structure and level of diversity of a large collection of European landraces of P. vulgaris (307) in comparison to 94 genotypes from the Americas that are representative of the Andean and Mesoamerican gene pools. First, we show that most of the European common bean landraces (67%) are of Andean origin, and that there are no strong differences across European regions for the proportions of the Andean and Mesoamerican gene pools. Moreover, cytoplasmic diversity is evenly distributed across European regions. Secondly, the cytoplasmic bottleneck that was due to the introduction of P. vulgaris into the Old World was very weak or nearly absent. This is in contrast to evidence from nuclear analyses that have suggested a bottleneck of greater intensity. Finally, we estimate that a relatively high proportion of the European bean germplasm (about 44%) was derived from hybridization between the Andean and Mesoamerican gene pools. Moreover, although hybrids are present everywhere in Europe, they show an uneven distribution, with high frequencies in central Europe, and low frequencies in Spain and Italy. On the basis of these data, we suggest that the entire European continent and not only some of the countries therein can be regarded as a secondary diversification center for P. vulgaris. Finally, we outline the relevance of these inter-gene pool hybrids for plant breeding.

Continental-scale assessment of genetic diversity and population structure in quaking aspen (Populus tremuloides)

Treesearch

Colin M. Callahan; Carol A. Rowe; Ronald J. Ryel; John D. Shaw; Michael D. Madritch; Karen E. Mock

2013-01-01

Aspen populations in the south-western portion of the range are consistent with expectations for a historically stable edge, with low within-population diversity, significant geographical population structuring, and little evidence of northward expansion. Structuring within the southwestern cluster may result from distinct gene pools separated during the Pleistocene...

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

Frozen gene pools - A future for species otherwise destined for extinction

USGS Publications Warehouse

Gee, G.F.

1986-01-01

Conclusion: Semen banks and ova and embryo banks can be practical methods to maintain gene pools. Gene pool preservation is desperately needed today due to the rapid decline in number of species and their habitat, a matter that is of concern to.biologists, economists, and politicians worldwide. Techniques are available for the cryopreservation of semen from many animals (and embryos from a few mammals) and adaptations of these techniques to other animals should be possible. A frozen gene pool in conjunction with existing programs makes it possible to preserve gene pools at less cost or in.some cases where no other alternative to extinction existed.

Gene Flow within and between Catchments in the Threatened Riparian Plant Myricaria germanica

PubMed Central

Werth, Silke; Scheidegger, Christoph

2014-01-01

One of the major distinctions of riparian habitats is their linearity. In linear habitats, gene flow is predicted to follow a one-dimensional stepping stone model, characterized by bidirectional gene flow between neighboring populations. Here, we studied the genetic structure of Myricaria germanica, a threatened riparian shrub which is capable of both wind and water dispersal. Our data led us to reject the ‘one catchment – one gene pool’ hypothesis as we found support for two gene pools, rather than four as expected in a study area including four catchments. This result also implies that in the history of the studied populations, dispersal across catchments has occurred. Two contemporary catchment-crossing migration events were detected, albeit between spatially proximate catchments. Allelic richness and inbreeding coefficients differed substantially between gene pools. There was significant isolation by distance, and our data confirmed the one-dimensional stepping-stone model of gene flow. Contemporary migration was bidirectional within the studied catchments, implying that dispersal vectors other than water are important for M. germanica. PMID:24932520

The impact of natural transformation on adaptation in spatially structured bacterial populations.

PubMed

Moradigaravand, Danesh; Engelstädter, Jan

2014-06-20

Recent studies have demonstrated that natural transformation and the formation of highly structured populations in bacteria are interconnected. In spite of growing evidence about this connection, little is known about the dynamics of natural transformation in spatially structured bacterial populations. In this work, we model the interdependency between the dynamics of the bacterial gene pool and those of environmental DNA in space to dissect the effect of transformation on adaptation. Our model reveals that even with only a single locus under consideration, transformation with a free DNA fragment pool results in complex adaptation dynamics that do not emerge in previous models focusing only on the gene shuffling effect of transformation at multiple loci. We demonstrate how spatial restriction on population growth and DNA diffusion in the environment affect the impact of transformation on adaptation. We found that in structured bacterial populations intermediate DNA diffusion rates predominantly cause transformation to impede adaptation by spreading deleterious alleles in the population. Overall, our model highlights distinctive evolutionary consequences of bacterial transformation in spatially restricted compared to planktonic bacterial populations.

A two-generation analysis of pollen pool genetic structure in flowering dogwood, Cornus florida (Cornaceae), in the Missouri Ozarks.

PubMed

Sork, Victoria L; Smouse, Peter E; Apsit, Victoria J; Dyer, Rodney J; Westfall, Robert D

2005-02-01

Anthropogenic landscape change can disrupt gene flow. As part of the Missouri Ozark Forest Ecosystem Project, this study examined whether silvicultural practices influence pollen-mediated gene movement in the insect-pollinated species, Cornus florida L., by comparing pollen pool structure (Φ(st)) among clear-cutting, selective cutting, and uncut regimes with the expectation that pollen movement should be least in the uncut regime. Using a sample of 1500 seedlings-10 each from 150 seed parents (43 in clear-cut, 74 in selective, and 33 in control sites) from six sites (each ranging from 266 to 527 ha), eight allozyme loci were analyzed with a pollen pool structure approach known as TwoGener (Smouse et al., 2001; Evolution 55: 260-271). This analysis revealed that pollen pool structure was less in clear-cut (Φ(C) = 0.090, P < 0.001) than in uncut areas (Φ(U) = 0.174, P < 0.001), with selective-cut intermediate (Φ(S) = 0.125, P < 0.001). These estimates translate into more effective pollen donors (N(ep)) in clear-cut (N(ep) = 5.56) and selective-cut (N(ep) = 4.00) areas than in uncut areas (N(ep) = 2.87). We demonstrate that Φ(C) ≤ Φ(S) ≤ Φ(U), with Φ(C) significantly smaller than Φ(U) (P < 0.034). The findings imply that, as long as a sufficiently large number of seed parents remain to provide adequate reproduction and to avoid a genetic bottleneck in the effective number of mothers, silvicultural management may not negatively affect the effective number of pollen parents, and hence subsequent genetic diversity in Cornus florida.

International efforts to protect the global cotton genetic resources

USDA-ARS?s Scientific Manuscript database

The genetic resources of cotton are classified into five tetraploid species in the primary gene pool, 20 diploid species in the secondary gene pool, and 25 diploid species in the tertiary gene pool. Unlike several globally important grain crops, the Consultative Group on International Agricultural R...

Natural Allelic Diversity, Genetic Structure and Linkage Disequilibrium Pattern in Wild Chickpea

PubMed Central

Kujur, Alice; Das, Shouvik; Badoni, Saurabh; Kumar, Vinod; Singh, Mohar; Bansal, Kailash C.; Tyagi, Akhilesh K.; Parida, Swarup K.

2014-01-01

Characterization of natural allelic diversity and understanding the genetic structure and linkage disequilibrium (LD) pattern in wild germplasm accessions by large-scale genotyping of informative microsatellite and single nucleotide polymorphism (SNP) markers is requisite to facilitate chickpea genetic improvement. Large-scale validation and high-throughput genotyping of genome-wide physically mapped 478 genic and genomic microsatellite markers and 380 transcription factor gene-derived SNP markers using gel-based assay, fluorescent dye-labelled automated fragment analyser and matrix-assisted laser desorption ionization-time of flight (MALDI-TOF) mass array have been performed. Outcome revealed their high genotyping success rate (97.5%) and existence of a high level of natural allelic diversity among 94 wild and cultivated Cicer accessions. High intra- and inter-specific polymorphic potential and wider molecular diversity (11–94%) along with a broader genetic base (13–78%) specifically in the functional genic regions of wild accessions was assayed by mapped markers. It suggested their utility in monitoring introgression and transferring target trait-specific genomic (gene) regions from wild to cultivated gene pool for the genetic enhancement. Distinct species/gene pool-wise differentiation, admixed domestication pattern, and differential genome-wide recombination and LD estimates/decay observed in a six structured population of wild and cultivated accessions using mapped markers further signifies their usefulness in chickpea genetics, genomics and breeding. PMID:25222488

Adaptive genomic divergence under high gene flow between freshwater and brackish-water ecotypes of prickly sculpin (Cottus asper) revealed by Pool-Seq.

PubMed

Dennenmoser, Stefan; Vamosi, Steven M; Nolte, Arne W; Rogers, Sean M

2017-01-01

Understanding the genomic basis of adaptive divergence in the presence of gene flow remains a major challenge in evolutionary biology. In prickly sculpin (Cottus asper), an abundant euryhaline fish in northwestern North America, high genetic connectivity among brackish-water (estuarine) and freshwater (tributary) habitats of coastal rivers does not preclude the build-up of neutral genetic differentiation and emergence of different life history strategies. Because these two habitats present different osmotic niches, we predicted high genetic differentiation at known teleost candidate genes underlying salinity tolerance and osmoregulation. We applied whole-genome sequencing of pooled DNA samples (Pool-Seq) to explore adaptive divergence between two estuarine and two tributary habitats. Paired-end sequence reads were mapped against genomic contigs of European Cottus, and the gene content of candidate regions was explored based on comparisons with the threespine stickleback genome. Genes showing signals of repeated differentiation among brackish-water and freshwater habitats included functions such as ion transport and structural permeability in freshwater gills, which suggests that local adaptation to different osmotic niches might contribute to genomic divergence among habitats. Overall, the presence of both repeated and unique signatures of differentiation across many loci scattered throughout the genome is consistent with polygenic adaptation from standing genetic variation and locally variable selection pressures in the early stages of life history divergence. © 2016 John Wiley & Sons Ltd.

A flexible and economical barcoding approach for highly multiplexed amplicon sequencing of diverse target genes

PubMed Central

Herbold, Craig W.; Pelikan, Claus; Kuzyk, Orest; Hausmann, Bela; Angel, Roey; Berry, David; Loy, Alexander

2015-01-01

High throughput sequencing of phylogenetic and functional gene amplicons provides tremendous insight into the structure and functional potential of complex microbial communities. Here, we introduce a highly adaptable and economical PCR approach to barcoding and pooling libraries of numerous target genes. In this approach, we replace gene- and sequencing platform-specific fusion primers with general, interchangeable barcoding primers, enabling nearly limitless customized barcode-primer combinations. Compared to barcoding with long fusion primers, our multiple-target gene approach is more economical because it overall requires lower number of primers and is based on short primers with generally lower synthesis and purification costs. To highlight our approach, we pooled over 900 different small-subunit rRNA and functional gene amplicon libraries obtained from various environmental or host-associated microbial community samples into a single, paired-end Illumina MiSeq run. Although the amplicon regions ranged in size from approximately 290 to 720 bp, we found no significant systematic sequencing bias related to amplicon length or gene target. Our results indicate that this flexible multiplexing approach produces large, diverse, and high quality sets of amplicon sequence data for modern studies in microbial ecology. PMID:26236305

Next generation haplotyping to decipher nuclear genomic interspecific admixture in Citrus species: analysis of chromosome 2.

PubMed

Curk, Franck; Ancillo, Gema; Garcia-Lor, Andres; Luro, François; Perrier, Xavier; Jacquemoud-Collet, Jean-Pierre; Navarro, Luis; Ollitrault, Patrick

2014-12-29

The most economically important Citrus species originated by natural interspecific hybridization between four ancestral taxa (Citrus reticulata, Citrus maxima, Citrus medica, and Citrus micrantha) and from limited subsequent interspecific recombination as a result of apomixis and vegetative propagation. Such reticulate evolution coupled with vegetative propagation results in mosaic genomes with large chromosome fragments from the basic taxa in frequent interspecific heterozygosity. Modern breeding of these species is hampered by their complex heterozygous genomic structures that determine species phenotype and are broken by sexual hybridisation. Nevertheless, a large amount of diversity is present in the citrus gene pool, and breeding to allow inclusion of desirable traits is of paramount importance. However, the efficient mobilization of citrus biodiversity in innovative breeding schemes requires previous understanding of Citrus origins and genomic structures. Haplotyping of multiple gene fragments along the whole genome is a powerful approach to reveal the admixture genomic structure of current species and to resolve the evolutionary history of the gene pools. In this study, the efficiency of parallel sequencing with 454 methodology to decipher the hybrid structure of modern citrus species was assessed by analysis of 16 gene fragments on chromosome 2. 454 amplicon libraries were established using the Fluidigm array system for 48 genotypes and 16 gene fragments from chromosome 2. Haplotypes were established from the reads of each accession and phylogenetic analyses were performed using the haplotypic data for each gene fragment. The length of 454 reads and the level of differentiation between the ancestral taxa of modern citrus allowed efficient haplotype phylogenetic assignations for 12 of the 16 gene fragments. The analysis of the mixed genomic structure of modern species and cultivars (i) revealed C. maxima introgressions in modern mandarins, (ii) was consistent with previous hypotheses regarding the origin of secondary species, and (iii) provided a new picture of the evolution of chromosome 2. 454 sequencing was an efficient strategy to establish haplotypes with significant phylogenetic assignations in Citrus, providing a new picture of the mixed structure on chromosome 2 in 48 citrus genotypes.

Genetic structure of the invasive tree Ailanthus altissima in eastern United States cities

Treesearch

Preston R. Aldrich; Joseph S. Briguglio; Shyam N. Kapadia; Minesh U. Morker; Ankit Rawal; Preeti Kalra; Cynthia D. Huebner; Gary K. Greer

2010-01-01

Ailanthus altissima is an invasive tree from Asia. It now occurs in most US states, and although primarily an urban weed, it has become a problem in forested areas especially in the eastern states. Little is known about its genetic structure. We explore its naturalized gene pool from 28 populations, mostly of the eastern US where infestations are...

Phylogenetic relationships among Lactuca (Asteraceae) species and related genera based on ITS-1 DNA sequences.

PubMed

Koopman, W J; Guetta, E; van de Wiel, C C; Vosman, B; van den Berg, R G

1998-11-01

Internal transcribed spacer (ITS-1) sequences from 97 accessions representing 23 species of Lactuca and related genera were determined and used to evaluate species relationships of Lactuca sensu lato (s.l.). The ITS-1 phylogenies, calculated using PAUP and PHYLIP, correspond better to the classification of Feráková than to other classifications evaluated, although the inclusion of sect. Lactuca subsect. Cyanicae is not supported. Therefore, exclusion of subsect. Cyanicae from Lactuca sensu Feráková is proposed. The amended genus contains the entire gene pool (sensu Harlan and De Wet) of cultivated lettuce (Lactuca sativa). The position of the species in the amended classification corresponds to their position in the lettuce gene pool. In the ITS-1 phylogenies, a clade with L. sativa, L. serriola, L. dregeana, L. altaica, and L. aculeata represents the primary gene pool. L. virosa and L. saligna, branching off closest to this clade, encompass the secondary gene pool. L. virosa is possibly of hybrid origin. The primary and secondary gene pool species are classified in sect. Lactuca subsect. Lactuca. The species L. quercina, L. viminea, L. sibirica, and L. tatarica, branching off next, represent the tertiary gene pool. They are classified in Lactuca sect. Lactucopsis, sect. Phaenixopus, and sect. Mulgedium, respectively. L. perennis and L. tenerrima, classified in sect. Lactuca subsect. Cyanicae, form clades with species from related genera and are not part of the lettuce gene pool.

Possible ancestral structure in human populations.

PubMed

Plagnol, Vincent; Wall, Jeffrey D

2006-07-01

Determining the evolutionary relationships between fossil hominid groups such as Neanderthals and modern humans has been a question of enduring interest in human evolutionary genetics. Here we present a new method for addressing whether archaic human groups contributed to the modern gene pool (called ancient admixture), using the patterns of variation in contemporary human populations. Our method improves on previous work by explicitly accounting for recent population history before performing the analyses. Using sequence data from the Environmental Genome Project, we find strong evidence for ancient admixture in both a European and a West African population (p approximately 10(-7)), with contributions to the modern gene pool of at least 5%. While Neanderthals form an obvious archaic source population candidate in Europe, there is not yet a clear source population candidate in West Africa.

Phenotypic assessments of peanut nested association mapping (NAM) populations

USDA-ARS?s Scientific Manuscript database

Nested association mapping (NAM) is a valuable innovation and multi-parental mapping population strategy in peanut genetics which increases the power to map quantitative trait loci and assists in extending the gene pool of elite peanut lines. In the peanut research community, two structured mapping ...

Possible effect of biotechnology on plant gene pools in Turkey.

PubMed

Demir, Aynur

2015-01-02

The recent rapid developments in biotechnology have made great contributions to the study of plant gene pools. The application of in vitro methods in freeze storage and DNA protection techniques in fast production studies has made major advances. From that aspect, biotechnology is an indispensable means for the protection of plant gene pools, which includes the insurance of sustainable agriculture and development of species. Besides all the positive developments, one of the primary risks posed by the uncontrolled spreading of genetically modified organisms is the possibility for other non-target organisms to be negatively affected. Genes of plant origin should be given priority in this type of studies by taking into consideration such negative effects that may result in disruption of ecological balance and damage to plant genetic pools. Turkey, due to its ecological conditions and history, has a very important position in terms of plant gene pools. This richness ought to be protected without corrupting its natural quality and natural evolution process in order to provide the sources of species that will be required for future sustainable agricultural applications. Thus, attention should be paid to the use of biotechnological methods, which play an important role especially in the protection and use of local and original plant gene pools.

Possible effect of biotechnology on plant gene pools in Turkey

PubMed Central

Demir, Aynur

2015-01-01

The recent rapid developments in biotechnology have made great contributions to the study of plant gene pools. The application of in vitro methods in freeze storage and DNA protection techniques in fast production studies has made major advances. From that aspect, biotechnology is an indispensable means for the protection of plant gene pools, which includes the insurance of sustainable agriculture and development of species. Besides all the positive developments, one of the primary risks posed by the uncontrolled spreading of genetically modified organisms is the possibility for other non-target organisms to be negatively affected. Genes of plant origin should be given priority in this type of studies by taking into consideration such negative effects that may result in disruption of ecological balance and damage to plant genetic pools. Turkey, due to its ecological conditions and history, has a very important position in terms of plant gene pools. This richness ought to be protected without corrupting its natural quality and natural evolution process in order to provide the sources of species that will be required for future sustainable agricultural applications. Thus, attention should be paid to the use of biotechnological methods, which play an important role especially in the protection and use of local and original plant gene pools. PMID:26019612

Uniparental genetic heritage of belarusians: encounter of rare middle eastern matrilineages with a central European mitochondrial DNA pool.

PubMed

Kushniarevich, Alena; Sivitskaya, Larysa; Danilenko, Nina; Novogrodskii, Tadeush; Tsybovsky, Iosif; Kiseleva, Anna; Kotova, Svetlana; Chaubey, Gyaneshwer; Metspalu, Ene; Sahakyan, Hovhannes; Bahmanimehr, Ardeshir; Reidla, Maere; Rootsi, Siiri; Parik, Jüri; Reisberg, Tuuli; Achilli, Alessandro; Hooshiar Kashani, Baharak; Gandini, Francesca; Olivieri, Anna; Behar, Doron M; Torroni, Antonio; Davydenko, Oleg; Villems, Richard

2013-01-01

Ethnic Belarusians make up more than 80% of the nine and half million people inhabiting the Republic of Belarus. Belarusians together with Ukrainians and Russians represent the East Slavic linguistic group, largest both in numbers and territory, inhabiting East Europe alongside Baltic-, Finno-Permic- and Turkic-speaking people. Till date, only a limited number of low resolution genetic studies have been performed on this population. Therefore, with the phylogeographic analysis of 565 Y-chromosomes and 267 mitochondrial DNAs from six well covered geographic sub-regions of Belarus we strove to complement the existing genetic profile of eastern Europeans. Our results reveal that around 80% of the paternal Belarusian gene pool is composed of R1a, I2a and N1c Y-chromosome haplogroups - a profile which is very similar to the two other eastern European populations - Ukrainians and Russians. The maternal Belarusian gene pool encompasses a full range of West Eurasian haplogroups and agrees well with the genetic structure of central-east European populations. Our data attest that latitudinal gradients characterize the variation of the uniparentally transmitted gene pools of modern Belarusians. In particular, the Y-chromosome reflects movements of people in central-east Europe, starting probably as early as the beginning of the Holocene. Furthermore, the matrilineal legacy of Belarusians retains two rare mitochondrial DNA haplogroups, N1a3 and N3, whose phylogeographies were explored in detail after de novo sequencing of 20 and 13 complete mitogenomes, respectively, from all over Eurasia. Our phylogeographic analyses reveal that two mitochondrial DNA lineages, N3 and N1a3, both of Middle Eastern origin, might mark distinct events of matrilineal gene flow to Europe: during the mid-Holocene period and around the Pleistocene-Holocene transition, respectively.

SNP discovery in common bean by restriction-associated DNA (RAD) sequencing for genetic diversity and population structure analysis.

PubMed

Valdisser, Paula Arielle M R; Pappas, Georgios J; de Menezes, Ivandilson P P; Müller, Bárbara S F; Pereira, Wendell J; Narciso, Marcelo G; Brondani, Claudio; Souza, Thiago L P O; Borba, Tereza C O; Vianello, Rosana P

2016-06-01

Researchers have made great advances into the development and application of genomic approaches for common beans, creating opportunities to driving more real and applicable strategies for sustainable management of the genetic resource towards plant breeding. This work provides useful polymorphic single-nucleotide polymorphisms (SNPs) for high-throughput common bean genotyping developed by RAD (restriction site-associated DNA) sequencing. The RAD tags were generated from DNA pooled from 12 common bean genotypes, including breeding lines of different gene pools and market classes. The aligned sequences identified 23,748 putative RAD-SNPs, of which 3357 were adequate for genotyping; 1032 RAD-SNPs with the highest ADT (assay design tool) score are presented in this article. The RAD-SNPs were structurally annotated in different coding (47.00 %) and non-coding (53.00 %) sequence components of genes. A subset of 384 RAD-SNPs with broad genome distribution was used to genotype a diverse panel of 95 common bean germplasms and revealed a successful amplification rate of 96.6 %, showing 73 % of polymorphic SNPs within the Andean group and 83 % in the Mesoamerican group. A slightly increased He (0.161, n = 21) value was estimated for the Andean gene pool, compared to the Mesoamerican group (0.156, n = 74). For the linkage disequilibrium (LD) analysis, from a group of 580 SNPs (289 RAD-SNPs and 291 BARC-SNPs) genotyped for the same set of genotypes, 70.2 % were in LD, decreasing to 0.10 %in the Andean group and 0.77 % in the Mesoamerican group. Haplotype patterns spanning 310 Mb of the genome (60 %) were characterized in samples from different origins. However, the haplotype frameworks were under-represented for the Andean (7.85 %) and Mesoamerican (5.55 %) gene pools separately. In conclusion, RAD sequencing allowed the discovery of hundreds of useful SNPs for broad genetic analysis of common bean germplasm. From now, this approach provides an excellent panel of molecular tools for whole genome analysis, allowing integrating and better exploring the common bean breeding practices.

De novo assembly and next-generation sequencing to analyse full-length gene variants from codon-barcoded libraries.

PubMed

Cho, Namjin; Hwang, Byungjin; Yoon, Jung-ki; Park, Sangun; Lee, Joongoo; Seo, Han Na; Lee, Jeewon; Huh, Sunghoon; Chung, Jinsoo; Bang, Duhee

2015-09-21

Interpreting epistatic interactions is crucial for understanding evolutionary dynamics of complex genetic systems and unveiling structure and function of genetic pathways. Although high resolution mapping of en masse variant libraries renders molecular biologists to address genotype-phenotype relationships, long-read sequencing technology remains indispensable to assess functional relationship between mutations that lie far apart. Here, we introduce JigsawSeq for multiplexed sequence identification of pooled gene variant libraries by combining a codon-based molecular barcoding strategy and de novo assembly of short-read data. We first validate JigsawSeq on small sub-pools and observed high precision and recall at various experimental settings. With extensive simulations, we then apply JigsawSeq to large-scale gene variant libraries to show that our method can be reliably scaled using next-generation sequencing. JigsawSeq may serve as a rapid screening tool for functional genomics and offer the opportunity to explore evolutionary trajectories of protein variants.

THE ROLES OF METAL IONS IN REGULATION BY RIBOSWITCHES

PubMed Central

2012-01-01

Metal ions are required by all organisms in order to execute an array of essential molecular functions. They play a critical role in many catalytic mechanisms and structural properties. Proper homeostasis of ions is critical; levels that are aberrantly low or high are deleterious to cellular physiology. To maintain stable intracellular pools, metal ion-sensing regulatory (metalloregulatory) proteins couple metal ion concentration fluctuations with expression of genes encoding for cation transport or sequestration. However, these transcriptional-based regulatory strategies are not the only mechanisms by which organisms coordinate metal ions with gene expression. Intriguingly, a few classes of signal-responsive RNA elements have also been discovered to function as metalloregulatory agents. This suggests that RNA-based regulatory strategies can be precisely tuned to intracellular metal ion pools, functionally akin to metalloregulatory proteins. In addition to these metal-sensing regulatory RNAs, there is a yet broader role for metal ions in directly assisting the structural integrity of other signal-responsive regulatory RNA elements. In this chapter, we discuss how the intimate physicochemical relationship between metal ions and nucleic acids is important for the structure and function of metal ion- and metabolite-sensing regulatory RNAs. PMID:22010271

Climate-driven range shifts explain the distribution of extant gene pools and predict future loss of unique lineages in a marine brown alga.

PubMed

Assis, J; Serrão, E A; Claro, B; Perrin, C; Pearson, G A

2014-06-01

The climate-driven dynamics of species ranges is a critical research question in evolutionary ecology. We ask whether present intraspecific diversity is determined by the imprint of past climate. This is an ongoing debate requiring interdisciplinary examination of population genetic pools and persistence patterns across global ranges. Previously, contrasting inferences and predictions have resulted from distinct genomic coverage and/or geographical information. We aim to describe and explain the causes of geographical contrasts in genetic diversity and their consequences for the future baseline of the global genetic pool, by comparing present geographical distribution of genetic diversity and differentiation with predictive species distribution modelling (SDM) during past extremes, present time and future climate scenarios for a brown alga, Fucus vesiculosus. SDM showed that both atmospheric and oceanic variables shape the global distribution of intertidal species, revealing regions of persistence, extinction and expansion during glacial and postglacial periods. These explained the distribution and structure of present genetic diversity, consisting of differentiated genetic pools with maximal diversity in areas of long-term persistence. Most of the present species range comprises postglacial expansion zones and, in contrast to highly dispersive marine organisms, expansions involved only local fronts, leaving distinct genetic pools at rear edges. Besides unravelling a complex phylogeographical history and showing congruence between genetic diversity and persistent distribution zones, supporting the hypothesis of niche conservatism, range shifts and loss of unique genetic diversity at the rear edge were predicted for future climate scenarios, impoverishing the global gene pool. © 2014 John Wiley & Sons Ltd.

Habitat heterogeneity and connectivity shape microbial communities in South American peatlands.

PubMed

Oloo, Felix; Valverde, Angel; Quiroga, María Victoria; Vikram, Surendra; Cowan, Don; Mataloni, Gabriela

2016-05-10

Bacteria play critical roles in peatland ecosystems. However, very little is known of how habitat heterogeneity affects the structure of the bacterial communities in these ecosystems. Here, we used amplicon sequencing of the 16S rRNA and nifH genes to investigate phylogenetic diversity and bacterial community composition in three different sub-Antarctic peat bog aquatic habitats: Sphagnum magellanicum interstitial water, and water from vegetated and non-vegetated pools. Total and putative nitrogen-fixing bacterial communities from Sphagnum interstitial water differed significantly from vegetated and non-vegetated pool communities (which were colonized by the same bacterial populations), probably as a result of differences in water chemistry and biotic interactions. Total bacterial communities from pools contained typically aquatic taxa, and were more dissimilar in composition and less species rich than those from Sphagnum interstitial waters (which were enriched in taxa typically from soils), probably reflecting the reduced connectivity between the former habitats. These results show that bacterial communities in peatland water habitats are highly diverse and structured by multiple concurrent factors.

Epistasis in intra- and inter-gene pool crosses of the common bean.

PubMed

Borel, J C; Ramalho, M A P; Abreu, A F B

2016-02-26

Epistasis has been shown to have an important role in the genetic control of several quantitative traits in the common bean. This study aimed to investigate the occurrence of epistasis in intra- and inter-pool gene crosses of the common bean. Four elite lines adapted to Brazilian conditions were used as parents, two from the Andean gene pool (ESAL 686; BRS Radiante) and two from the Mesoamerican gene pool (BRSMG Majestoso; BRS Valente). Four F2 populations were obtained: "A" (ESAL 686 x BRS Radiante), "B" (BRSMG Majestoso x BRS Valente), "C" (BRS Radiante x BRSMG Majestoso), and "D" (BRS Valente x ESAL 686). A random sample of F2 plants from each population was backcrossed to parents and F1 individuals, according to the triple test cross. Three types of progenies from each population were evaluated in contiguous trials. Seed yield and 100-seed weight were evaluated. Dominance genetic variance was predominant in most cases. However, the estimates of genetic variance may be biased by the occurrence of linkage disequilibrium and epistasis. Epistasis was detected for both traits; however, the occurrence differed among the populations and between the two traits. The results of this study reinforce the hypothesis that epistasis is present in the genetic control of traits in the common bean and suggest that the phenomenon is more frequent in inter-gene pool crosses than in intra-gene pool crosses.

Balancing gene expression without library construction via a reusable sRNA pool.

PubMed

Ghodasara, Amar; Voigt, Christopher A

2017-07-27

Balancing protein expression is critical when optimizing genetic systems. Typically, this requires library construction to vary the genetic parts controlling each gene, which can be expensive and time-consuming. Here, we develop sRNAs corresponding to 15nt 'target' sequences that can be inserted upstream of a gene. The targeted gene can be repressed from 1.6- to 87-fold by controlling sRNA expression using promoters of different strength. A pool is built where six sRNAs are placed under the control of 16 promoters that span a ∼103-fold range of strengths, yielding ∼107 combinations. This pool can simultaneously optimize up to six genes in a system. This requires building only a single system-specific construct by placing a target sequence upstream of each gene and transforming it with the pre-built sRNA pool. The resulting library is screened and the top clone is sequenced to determine the promoter controlling each sRNA, from which the fold-repression of the genes can be inferred. The system is then rebuilt by rationally selecting parts that implement the optimal expression of each gene. We demonstrate the versatility of this approach by using the same pool to optimize a metabolic pathway (β-carotene) and genetic circuit (XNOR logic gate). © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

MafK/NF-E2 p18 is required for beta-globin genes activation by mediating the proximity of LCR and active beta-globin genes in MEL cell line.

PubMed

Du, Mei-Jun; Lv, Xiang; Hao, De-Long; Zhao, Guo-Wei; Wu, Xue-Song; Wu, Feng; Liu, De-Pei; Liang, Chih-Chuan

2008-01-01

Evidences indicate that locus control region (LCR) of beta-globin spatially closes to the downstream active gene promoter to mediate the transcriptional activation by looping. DNA binding proteins may play an important role in the looping formation. NF-E2 is one of the key transcription factors in beta-globin gene transcriptional activation. To shed light on whether NF-E2 is involved in this process, DS19MafKsiRNA cell pools were established by specifically knocked down the expression of MafK/NF-E2 p18, one subunit of NF-E2 heterodimer. In the above cell pools, it was observed that the occupancy efficiency of NF-E2 on beta-globin gene locus and the expression level of beta-globin genes were decreased. H3 acetylation, H3-K4 methylation and the deposition of RNA polymerase II, but not the recruitment of GATA-1, were also found reduced at the beta-globin gene cluster. Chromosome Conformation Capture (3C) assay showed that the cross-linking frequency between the main NF-E2 binding site HS2 and downstream structural genes was reduced compared to the normal cell. This result demonstrated that MafK/NF-E2 p18 recruitment was involved in the physical proximity of LCR and active beta-globin genes upon beta-globin gene transcriptional activation.

Structural and functional partitioning of bread wheat chromosome 3B.

PubMed

Choulet, Frédéric; Alberti, Adriana; Theil, Sébastien; Glover, Natasha; Barbe, Valérie; Daron, Josquin; Pingault, Lise; Sourdille, Pierre; Couloux, Arnaud; Paux, Etienne; Leroy, Philippe; Mangenot, Sophie; Guilhot, Nicolas; Le Gouis, Jacques; Balfourier, Francois; Alaux, Michael; Jamilloux, Véronique; Poulain, Julie; Durand, Céline; Bellec, Arnaud; Gaspin, Christine; Safar, Jan; Dolezel, Jaroslav; Rogers, Jane; Vandepoele, Klaas; Aury, Jean-Marc; Mayer, Klaus; Berges, Hélène; Quesneville, Hadi; Wincker, Patrick; Feuillet, Catherine

2014-07-18

We produced a reference sequence of the 1-gigabase chromosome 3B of hexaploid bread wheat. By sequencing 8452 bacterial artificial chromosomes in pools, we assembled a sequence of 774 megabases carrying 5326 protein-coding genes, 1938 pseudogenes, and 85% of transposable elements. The distribution of structural and functional features along the chromosome revealed partitioning correlated with meiotic recombination. Comparative analyses indicated high wheat-specific inter- and intrachromosomal gene duplication activities that are potential sources of variability for adaption. In addition to providing a better understanding of the organization, function, and evolution of a large and polyploid genome, the availability of a high-quality sequence anchored to genetic maps will accelerate the identification of genes underlying important agronomic traits. Copyright © 2014, American Association for the Advancement of Science.

Population Structure of Barley Landrace Populations and Gene-Flow with Modern Varieties

PubMed Central

Bellucci, Elisa; Bitocchi, Elena; Rau, Domenico; Nanni, Laura; Ferradini, Nicoletta; Giardini, Alessandro; Rodriguez, Monica; Attene, Giovanna; Papa, Roberto

2013-01-01

Landraces are heterogeneous plant varieties that are reproduced by farmers as populations that are subject to both artificial and natural selection. Landraces are distinguished by farmers due to their specific traits, and different farmers often grow different populations of the same landrace. We used simple sequence repeats (SSRs) to analyse 12 barley landrace populations from Sardinia from two collections spanning 10 years. We analysed the population structure, and compared the population diversity of the landraces that were collected at field level (population). We used a representative pool of barley varieties for diversity comparisons and to analyse the effects of gene flow from modern varieties. We found that the Sardinian landraces are a distinct gene pool from those of both two-row and six-row barley varieties. There is also a low, but significant, mean level and population-dependent level of introgression from the modern varieties into the Sardinian landraces. Moreover, we show that the Sardinian landraces have the same level of gene diversity as the representative sample of modern commercial varieties grown in Italy in the last decades, even within population level. Thus, these populations represent crucial sources of germplasm that will be useful for crop improvement and for population genomics studies and association mapping, to identify genes, loci and genome regions responsible for adaptive variations. Our data also suggest that landraces are a source of valuable germplasm for sustainable agriculture in the context of future climate change, and that in-situ conservation strategies based on farmer use can preserve the genetic identity of landraces while allowing adaptation to local environments. PMID:24386303

Linkage disequilibrium and population structure in wild and domesticated populations of Phaseolus vulgaris L.

PubMed Central

Rossi, Monica; Bitocchi, Elena; Bellucci, Elisa; Nanni, Laura; Rau, Domenico; Attene, Giovanna; Papa, Roberto

2009-01-01

Together with the knowledge of the population structure, a critical aspect for the planning of association and/or population genomics studies is the level of linkage disequilibrium (LD) that characterizes the species and the population used for such an analysis. We have analyzed the population structure and LD in wild and domesticated populations of Phaseolus vulgaris L. using amplified fragment length polymorphism markers, most of which were genetically mapped in two recombinant inbred populations. Our results reflect the previous knowledge of the occurrence of two major wild gene pools of P. vulgaris, from which two independent domestication events originated, one in the Andes and one in Mesoamerica. The high level of LD in the whole sample was mostly due to the gene pool structure, with a much higher LD in domesticated compared to wild populations. In relation to association studies, our results also suggest that whole-genome-scan approaches are feasible in the common bean. Interestingly, an excess of inter-chromosomal LD was found in the domesticated populations, which suggests an important role for epistatic selection during domestication. Moreover, our results indicate the occurrence of a strong bottleneck in the Andean wild population before domestication, suggesting a Mesoamerican origin of P. vulgaris. Finally, our data support the occurrence of a single domestication event in Mesoamerica, and the same scenario in the Andes. PMID:25567895

Linking Microbial Community and Catabolic Gene Structures during the Adaptation of Three Contaminated Soils under Continuous Long-Term Pollutant Stress

PubMed Central

Lima-Morales, Daiana; Jáuregui, Ruy; Camarinha-Silva, Amelia; Geffers, Robert; Vilchez-Vargas, Ramiro

2016-01-01

Three types of contaminated soil from three geographically different areas were subjected to a constant supply of benzene or benzene/toluene/ethylbenzene/xylenes (BTEX) for a period of 3 months. Different from the soil from Brazil (BRA) and Switzerland (SUI), the Czech Republic (CZE) soil which was previously subjected to intensive in situ bioremediation displayed only negligible changes in community structure. BRA and SUI soil samples showed a clear succession of phylotypes. A rapid response to benzene stress was observed, whereas the response to BTEX pollution was significantly slower. After extended incubation, actinobacterial phylotypes increased in relative abundance, indicating their superior fitness to pollution stress. Commonalities but also differences in the phylotypes were observed. Catabolic gene surveys confirmed the enrichment of actinobacteria by identifying the increase of actinobacterial genes involved in the degradation of pollutants. Proteobacterial phylotypes increased in relative abundance in SUI microcosms after short-term stress with benzene, and catabolic gene surveys indicated enriched metabolic routes. Interestingly, CZE soil, despite staying constant in community structure, showed a change in the catabolic gene structure. This indicates that a highly adapted community, which had to adjust its gene pool to meet novel challenges, has been enriched. PMID:26850298

Habitat heterogeneity and connectivity shape microbial communities in South American peatlands

PubMed Central

Oloo, Felix; Valverde, Angel; Quiroga, María Victoria; Vikram, Surendra; Cowan, Don; Mataloni, Gabriela

2016-01-01

Bacteria play critical roles in peatland ecosystems. However, very little is known of how habitat heterogeneity affects the structure of the bacterial communities in these ecosystems. Here, we used amplicon sequencing of the 16S rRNA and nifH genes to investigate phylogenetic diversity and bacterial community composition in three different sub-Antarctic peat bog aquatic habitats: Sphagnum magellanicum interstitial water, and water from vegetated and non-vegetated pools. Total and putative nitrogen-fixing bacterial communities from Sphagnum interstitial water differed significantly from vegetated and non-vegetated pool communities (which were colonized by the same bacterial populations), probably as a result of differences in water chemistry and biotic interactions. Total bacterial communities from pools contained typically aquatic taxa, and were more dissimilar in composition and less species rich than those from Sphagnum interstitial waters (which were enriched in taxa typically from soils), probably reflecting the reduced connectivity between the former habitats. These results show that bacterial communities in peatland water habitats are highly diverse and structured by multiple concurrent factors. PMID:27162086

Uniparental Genetic Heritage of Belarusians: Encounter of Rare Middle Eastern Matrilineages with a Central European Mitochondrial DNA Pool

PubMed Central

Kushniarevich, Alena; Sivitskaya, Larysa; Danilenko, Nina; Novogrodskii, Tadeush; Tsybovsky, Iosif; Kiseleva, Anna; Kotova, Svetlana; Chaubey, Gyaneshwer; Metspalu, Ene; Sahakyan, Hovhannes; Bahmanimehr, Ardeshir; Reidla, Maere; Rootsi, Siiri; Parik, Jüri; Reisberg, Tuuli; Achilli, Alessandro; Hooshiar Kashani, Baharak; Gandini, Francesca; Olivieri, Anna; Behar, Doron M.; Torroni, Antonio; Davydenko, Oleg; Villems, Richard

2013-01-01

Ethnic Belarusians make up more than 80% of the nine and half million people inhabiting the Republic of Belarus. Belarusians together with Ukrainians and Russians represent the East Slavic linguistic group, largest both in numbers and territory, inhabiting East Europe alongside Baltic-, Finno-Permic- and Turkic-speaking people. Till date, only a limited number of low resolution genetic studies have been performed on this population. Therefore, with the phylogeographic analysis of 565 Y-chromosomes and 267 mitochondrial DNAs from six well covered geographic sub-regions of Belarus we strove to complement the existing genetic profile of eastern Europeans. Our results reveal that around 80% of the paternal Belarusian gene pool is composed of R1a, I2a and N1c Y-chromosome haplogroups – a profile which is very similar to the two other eastern European populations – Ukrainians and Russians. The maternal Belarusian gene pool encompasses a full range of West Eurasian haplogroups and agrees well with the genetic structure of central-east European populations. Our data attest that latitudinal gradients characterize the variation of the uniparentally transmitted gene pools of modern Belarusians. In particular, the Y-chromosome reflects movements of people in central-east Europe, starting probably as early as the beginning of the Holocene. Furthermore, the matrilineal legacy of Belarusians retains two rare mitochondrial DNA haplogroups, N1a3 and N3, whose phylogeographies were explored in detail after de novo sequencing of 20 and 13 complete mitogenomes, respectively, from all over Eurasia. Our phylogeographic analyses reveal that two mitochondrial DNA lineages, N3 and N1a3, both of Middle Eastern origin, might mark distinct events of matrilineal gene flow to Europe: during the mid-Holocene period and around the Pleistocene-Holocene transition, respectively. PMID:23785503

Evaluation of the BD GeneOhm assay using the rotor-gene 6000 platform for rapid detection of methicillin-resistant Staphylococcus aureus from pooled screening swabs.

PubMed

Smith, Melvyn Howard; Hodgson, Julian; Eltringham, Ian Joseph

2010-12-01

As health services move toward universal methicillin-resistant Staphylococcus aureus (MRSA) screening for hospital admissions, the most cost-effective approach is yet to be defined. In this study, one of the largest to date, we evaluated the performance of the BD GeneOhm MRSA assay on the Rotor-Gene 6000 thermal cycler, using samples taken directly from pooled MRSA screens. Results were compared with the same assay performed on the Smart-Cycler II platform and overnight broth culture. Samples yielding discrepant results were subjected to detailed analysis with an in-house PCR and patient note review. A total of 1,428 pooled MRSA screens were tested. Sensitivities and specificities of 85.3% and 95.8% for the Rotor-Gene and 81% and 95.7% for the Smart-Cycler were obtained, compared with broth enrichment. The sensitivity of the BD GeneOhm assay was increased to 100% when the results of in-house PCR and patient note review were taken into account. This study demonstrates that the Rotor-Gene 6000 thermal cycler is a reliable platform for use with the BD GeneOhm assay. It also proves that commercial PCR can be performed direct on pooled samples in selective broth, without the need for overnight incubation.

Recognition of digital characteristics based new improved genetic algorithm

NASA Astrophysics Data System (ADS)

Wang, Meng; Xu, Guoqiang; Lin, Zihao

2017-08-01

In the field of digital signal processing, Estimating the characteristics of signal modulation parameters is an significant research direction. The paper determines the set of eigenvalue which can show the difference of the digital signal modulation based on the deep research of the new improved genetic algorithm. Firstly take them as the best gene pool; secondly, The best gene pool will be changed in the genetic evolvement by selecting, overlapping and eliminating each other; Finally, Adapting the strategy of futher enhance competition and punishment to more optimizer the gene pool and ensure each generation are of high quality gene. The simulation results show that this method not only has the global convergence, stability and faster convergence speed.

Sculpting the Barnyard Gene Pool

ERIC Educational Resources Information Center

Childers, Gina; Wolfe, Kim; Dupree, Alan; Young, Sheila; Caver, Jessica; Quintanilla, Ruby; Thornton, Laura

2016-01-01

Project-based learning (PBL) takes student engagement to a higher level through reflective collaboration, inquiry, critical thinking, problem solving, and personal relevance. This article explains how six high school teachers developed an interconnected, interdisciplinary STEM-focused PBL called "Sculpting the Barnyard Gene Pool." The…

Whole-tree silvic identifications and the microsatellite genetic structure of a red oak species complex in an Indiana old-growth forest

Treesearch

Preston R. Aldrich; George R. Parker; Charles H. Michler; Jeanne Romero-Severson

2003-01-01

The red oaks (Quercus section Lobatae) include important timber species, but we know little about their gene pools. Red oak species can be difficult to identify, possibly because of extensive interspecific hybridization, although most evidence of this is morphological. We used 15 microsatellite loci to examine the genetic...

Evaluation of acetylcholinesterase in an animal model of maple syrup urine disease.

PubMed

Scaini, Giselli; de Rochi, Natália; Jeremias, Isabela C; Deroza, Pedro F; Zugno, Alexandra I; Pereira, Talita C B; Oliveira, Giovanna M T; Kist, Luiza W; Bogo, Maurício R; Schuck, Patrícia F; Ferreira, Gustavo C; Streck, Emilio L

2012-04-01

Maple syrup urine disease is an inherited metabolic disease predominantly characterized by neurological dysfunction. However, the mechanisms underlying the neuropathology of this disease are still not defined. Therefore, the aim of this study was to investigate the effect of acute and chronic administration of a branched-chain amino acids (BCAA) pool (leucine, isoleucine, and valine) on acetylcholinesterase (AChE) activity and gene expression in the brain and serum of rats and to assess if antioxidant treatment prevented the alterations induced by BCAA administration. Our results show that the acute administration of a BCAA pool in 10- and 30-day-old rats increases AChE activity in the cerebral cortex, striatum, hippocampus, and serum. Moreover, chronic administration of the BCAA pool also increases AChE activity in the structures studied, and antioxidant treatment prevents this increase. In addition, we show a significant decrease in the mRNA expression of AChE in the hippocampus following acute administration in 10- and 30-day-old rats. On the other hand, AChE expression increased significantly after chronic administration of the BCAA pool. Interestingly, the antioxidant treatment was able to prevent the increased AChE activity without altering AChE expression. In conclusion, the results from the present study demonstrate a marked increase in AChE activity in all brain structures following the administration of a BCAA pool. Moreover, the increased AChE activity is prevented by the coadministration of N-acetylcysteine and deferoxamine as antioxidants.

Saturation of an intra-gene pool linkage map: toward unified consensus linkage map in common bean

USDA-ARS?s Scientific Manuscript database

Map-based cloning to find genes of interest and marker assisted selection (MAS) requires good genetic maps with high reproducible markers. In this study, we saturated the linkage map of the intra-gene pool population of common bean DOR364×BAT477 (DB) by evaluating 2,706 molecular markers in includin...

Conjugative plasmids: vessels of the communal gene pool

PubMed Central

Norman, Anders; Hansen, Lars H.; Sørensen, Søren J.

2009-01-01

Comparative whole-genome analyses have demonstrated that horizontal gene transfer (HGT) provides a significant contribution to prokaryotic genome innovation. The evolution of specific prokaryotes is therefore tightly linked to the environment in which they live and the communal pool of genes available within that environment. Here we use the term supergenome to describe the set of all genes that a prokaryotic ‘individual’ can draw on within a particular environmental setting. Conjugative plasmids can be considered particularly successful entities within the communal pool, which have enabled HGT over large taxonomic distances. These plasmids are collections of discrete regions of genes that function as ‘backbone modules’ to undertake different aspects of overall plasmid maintenance and propagation. Conjugative plasmids often carry suites of ‘accessory elements’ that contribute adaptive traits to the hosts and, potentially, other resident prokaryotes within specific environmental niches. Insight into the evolution of plasmid modules therefore contributes to our knowledge of gene dissemination and evolution within prokaryotic communities. This communal pool provides the prokaryotes with an important mechanistic framework for obtaining adaptability and functional diversity that alleviates the need for large genomes of specialized ‘private genes’. PMID:19571247

[The Russian gene pool: gene geography of Alu-insertions (ACE, APOA1, B65, PV92 TPA25)].

PubMed

Solov'eva, D S; Balanovskaia, E V; Kuznetsova, M A; Vasinskaia, O A; Frolova, S A; Pocheshkhova, E A; Evseeva, I V; Boldyreva, M N; Balanovskiĭ, O P

2010-01-01

The analysis of five Alu insertion loci (ACE, AP4OA1, B65, PV92, TPA25) has been carried out for the first time in 10 Russian populations (1088 individuals), covered all parts of historical area of the Russian ethnos. Depending on locus, Russian populations exhibit similarity with their western (European populations) or with the eastern (populations of the Ural region) neighbors. Considering frequencies of the studied Alu-insertions, Russian gene pool exhibits low variation: average difference between populations is d = 0.007, whereas on classical markers, mtDNA and Y chromosome heterogeneity of Russian gene pool is essentially higher (0.013, 0.033 and 0.142 respectively). Therefore, this set of five Alu insertions has lower variability on the intra-ethnic level. However in inter-ethnic comparisons the clear pattern was obtained: 13 Eastern European ethnic groups formed three clusters, according with their historical and geographical position--East Slavic, Caucasian and South Ural clusters. The obtained data confirms efficiency of using Alu insertions for studying genetic differentiation and history of a gene pool of the Eastern European populations.

Genetic Structure and Gene Flows within Horses: A Genealogical Study at the French Population Scale

PubMed Central

Pirault, Pauline; Danvy, Sophy; Verrier, Etienne; Leroy, Grégoire

2013-01-01

Since horse breeds constitute populations submitted to variable and multiple outcrossing events, we analyzed the genetic structure and gene flows considering horses raised in France. We used genealogical data, with a reference population of 547,620 horses born in France between 2002 and 2011, grouped according to 55 breed origins. On average, individuals had 6.3 equivalent generations known. Considering different population levels, fixation index decreased from an overall species FIT of 1.37%, to an average of −0.07% when considering the 55 origins, showing that most horse breeds constitute populations without genetic structure. We illustrate the complexity of gene flows existing among horse breeds, a few populations being closed to foreign influence, most, however, being submitted to various levels of introgression. In particular, Thoroughbred and Arab breeds are largely used as introgression sources, since those two populations explain together 26% of founder origins within the overall horse population. When compared with molecular data, breeds with a small level of coancestry also showed low genetic distance; the gene pool of the breeds was probably impacted by their reproducer exchanges. PMID:23630596

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

Marchisio, Mario Andrea, E-mail: marchisio@hit.edu.cn

Published in 2008, Parts & Pools represents one of the first attempts to conceptualize the modular design of bacterial synthetic gene circuits with Standard Biological Parts (DNA segments) and Pools of molecules referred to as common signal carriers (e.g., RNA polymerases and ribosomes). The original framework for modeling bacterial components and designing prokaryotic circuits evolved over the last years and brought, first, to the development of an algorithm for the automatic design of Boolean gene circuits. This is a remarkable achievement since gene digital circuits have a broad range of applications that goes from biosensors for health and environment caremore » to computational devices. More recently, Parts & Pools was enabled to give a proper formal description of eukaryotic biological circuit components. This was possible by employing a rule-based modeling approach, a technique that permits a faithful calculation of all the species and reactions involved in complex systems such as eukaryotic cells and compartments. In this way, Parts & Pools is currently suitable for the visual and modular design of synthetic gene circuits in yeast and mammalian cells too.« less

Genetic connectivity among swarming sites in the wide ranging and recently declining little brown bat (Myotis lucifugus)

PubMed Central

Burns, Lynne E; Frasier, Timothy R; Broders, Hugh G

2014-01-01

Characterizing movement dynamics and spatial aspects of gene flow within a species permits inference on population structuring. As patterns of structuring are products of historical and current demographics and gene flow, assessment of structure through time can yield an understanding of evolutionary dynamics acting on populations that are necessary to inform management. Recent dramatic population declines in hibernating bats in eastern North America from white-nose syndrome have prompted the need for information on movement dynamics for multiple bat species. We characterized population genetic structure of the little brown bat, Myotis lucifugus, at swarming sites in southeastern Canada using 9 nuclear microsatellites and a 292-bp region of the mitochondrial genome. Analyses of FST, ΦST, and Bayesian clustering (STRUCTURE) found weak levels of genetic structure among swarming sites for the nuclear and mitochondrial genome (Global FST = 0.001, P < 0.05, Global ΦST = 0.045, P < 0.01, STRUCTURE K = 1) suggesting high contemporary gene flow. Hierarchical AMOVA also suggests little structuring at a regional (provincial) level. Metrics of nuclear genetic structure were not found to differ between males and females suggesting weak asymmetries in gene flow between the sexes. However, a greater degree of mitochondrial structuring does support male-biased dispersal long term. Demographic analyses were consistent with past population growth and suggest a population expansion occurred from approximately 1250 to 12,500 BP, following Pleistocene deglaciation in the region. Our study suggests high gene flow and thus a high degree of connectivity among bats that visit swarming sites whereby mainland areas of the region may be best considered as one large gene pool for management and conservation. PMID:25505539

Associating quantitative behavioral traits with gene expression in the brain: searching for diamonds in the hay.

PubMed

Reiner-Benaim, Anat; Yekutieli, Daniel; Letwin, Noah E; Elmer, Gregory I; Lee, Norman H; Kafkafi, Neri; Benjamini, Yoav

2007-09-01

Gene expression and phenotypic functionality can best be associated when they are measured quantitatively within the same experiment. The analysis of such a complex experiment is presented, searching for associations between measures of exploratory behavior in mice and gene expression in brain regions. The analysis of such experiments raises several methodological problems. First and foremost, the size of the pool of potential discoveries being screened is enormous yet only few biologically relevant findings are expected, making the problem of multiple testing especially severe. We present solutions based on screening by testing related hypotheses, then testing the hypotheses of interest. In one variant the subset is selected directly, in the other one a tree of hypotheses is tested hierarchical; both variants control the False Discovery Rate (FDR). Other problems in such experiments are in the fact that the level of data aggregation may be different for the quantitative traits (one per animal) and gene expression measurements (pooled across animals); in that the association may not be linear; and in the resolution of interest only few replications exist. We offer solutions to these problems as well. The hierarchical FDR testing strategies presented here can serve beyond the structure of our motivating example study to any complex microarray study. Supplementary data are available at Bioinformatics online.

Understanding genetic control of root system architecture in soybean: Insights into the genetic basis of lateral root number.

PubMed

Prince, Silvas J; Valliyodan, Babu; Ye, Heng; Yang, Ming; Tai, Shuaishuai; Hu, Wushu; Murphy, Mackensie; Durnell, Lorellin A; Song, Li; Joshi, Trupti; Liu, Yang; Van de Velde, Jan; Vandepoele, Klaas; Grover Shannon, J; Nguyen, Henry T

2018-05-10

Developing crops with better root systems is a promising strategy to ensure productivity in both optimum and stress environments. Root system architectural (RSA) traits in 397 soybean accessions were characterized and a high-density single nucleotide polymorphisms (SNP) based genome-wide association study was performed to identify the underlying genes associated with root structure. SNPs associated with root architectural traits specific to landraces and elite germplasm pools were detected. Four loci were detected in landraces for lateral root number (LRN) and distribution of root thickness in diameter class I with a major locus on chromosome 16. This major loci was detected in the coding region of unknown protein, and subsequent analyses demonstrated that root traits are affected with mutated haplotypes of the gene. In elite germplasm pool, three significant SNPs in alanine-glyoxalate aminotransferase, Leucine-Rich Repeat receptor/No apical meristem and unknown functional genes were found to govern multiple traits including root surface area and volume. However, no major loci were detected for LRN in elite germplasm. Nucleotide diversity analysis found evidence of selective sweeps around the landraces LRN gene. Soybean accessions with minor and mutated allelic variants of LRN gene were found to perform better in both water-limited and optimal field conditions. This article is protected by copyright. All rights reserved.

[Book review] Fish Gene Pools: preservation of genetic resources in relation to wild fish stocks, edited by N. Ryman

USGS Publications Warehouse

Simon, R.C.

1984-01-01

Review of: Fish Gene Pools: Preservation of Genetic Resources in Relation to Wild Fish Stocks. Edited by N. Ryman. The Editorial Service/FRN, Box 6710, S-11385, Stockholm, Sweden. 1981. 111 pages. $16.00 (paper).

Sampling and Pooling Methods for Capturing Herd Level Antibiotic Resistance in Swine Feces using qPCR and CFU Approaches

PubMed Central

Mellerup, Anders; Ståhl, Marie

2015-01-01

The aim of this article was to define the sampling level and method combination that captures antibiotic resistance at pig herd level utilizing qPCR antibiotic resistance gene quantification and culture-based quantification of antibiotic resistant coliform indicator bacteria. Fourteen qPCR assays for commonly detected antibiotic resistance genes were developed, and used to quantify antibiotic resistance genes in total DNA from swine fecal samples that were obtained using different sampling and pooling methods. In parallel, the number of antibiotic resistant coliform indicator bacteria was determined in the same swine fecal samples. The results showed that the qPCR assays were capable of detecting differences in antibiotic resistance levels in individual animals that the coliform bacteria colony forming units (CFU) could not. Also, the qPCR assays more accurately quantified antibiotic resistance genes when comparing individual sampling and pooling methods. qPCR on pooled samples was found to be a good representative for the general resistance level in a pig herd compared to the coliform CFU counts. It had significantly reduced relative standard deviations compared to coliform CFU counts in the same samples, and therefore differences in antibiotic resistance levels between samples were more readily detected. To our knowledge, this is the first study to describe sampling and pooling methods for qPCR quantification of antibiotic resistance genes in total DNA extracted from swine feces. PMID:26114765

Sinu Virus, a Novel and divergent Orthomyxovirus Related to Members of the Genus Thogotovirus, Isolated from Mosquitoes in Colombia

PubMed Central

Contreras-Gutiérrez, María Angélica; Nunes, Marcio R.T.; Guzman, Hilda; Uribe, Sandra; Gómez, Juan Carlos Gallego; Vasco, Juan David Suaza; Cardoso, Jedson F.; Popov, Vsevolod L.; Widen, Steven G.; Wood, Thomas G.; Vasilakis, Nikos; Tesh, Robert B.

2016-01-01

The genome and structural organization of a novel insect-specific orthomyxovirus, designated Sinu virus, is described. Sinu virus (SINUV) was isolated in cultures of C6/36 cells from a pool of mosquitoes collected in northwestern Colombia. The virus has six negative-sense ssRNA segments. Genetic analysis of each segment demonstrated the presence of six distinct ORFs encoding the following genes: PB2 (Segment 1), PB1, (Segment 2), PA protein (Segment 3), envelope GP gene (Segment 4), the NP (Segment 5), and M-like gene (Segment 6). Phylogenetically, SINUV appears to be most closed related to viruses in the genus Thogotovirus. PMID:27936462

Questioning the utility of pooling samples in microarray experiments with cell lines.

PubMed

Lusa, L; Cappelletti, V; Gariboldi, M; Ferrario, C; De Cecco, L; Reid, J F; Toffanin, S; Gallus, G; McShane, L M; Daidone, M G; Pierotti, M A

2006-01-01

We describe a microarray experiment using the MCF-7 breast cancer cell line in two different experimental conditions for which the same number of independent pools as the number of individual samples was hybridized on Affymetrix GeneChips. Unexpectedly, when using individual samples, the number of probe sets found to be differentially expressed between treated and untreated cells was about three times greater than that found using pools. These findings indicate that pooling samples in microarray experiments where the biological variability is expected to be small might not be helpful and could even decrease one's ability to identify differentially expressed genes.

Use of a pooled clone method to isolate a novel Bacillus thuringiensis Cry2A toxin with activity against Ostrinia furnacalis.

PubMed

Shu, Changlong; Zhang, Jingtao; Chen, Guihua; Liang, Gemei; He, Kanglai; Crickmore, Neil; Huang, Dafang; Zhang, Jie; Song, Fuping

2013-09-01

A pooled clone method was developed to screen for cry2A genes. This metagenomic method avoids the need to analyse isolated Bacillus thuringiensis strains by performing gene specific PCR on plasmid-enriched DNA prepared from a pooled soil sample. Using this approach the novel holotype gene cry2Ah1 was cloned and characterized. The toxin gene was over-expressed in Escherichia coli Rosetta (DE3) and the expressed toxin accumulated in both the soluble and insoluble fractions. The soluble Cry2Ah1 was found to have a weight loss activity against Ostrinia furnacalis, and a growth inhibitory activity to both Cry1Ac-susceptible and resistant Helicoverpa armigera populations. Copyright © 2013 Elsevier Inc. All rights reserved.

Analyses of Thinopyrum bessarabicum, Th. elongation and Th. junceum chromosomes using EST-SSR markers

USDA-ARS?s Scientific Manuscript database

Wild Thinopyrum grasses serve as important gene pools for forage and cereal crops. Knowledge of their chromosome organizations is pivotal for efficient utilization of this important gene pool in germplasm enhancement programs. EST-SSR markers for Th. bessarabicum, Th. elongatum and Th. junceum chr...

A transcriptome-based assessment of the astrocytic dystrophin-associated complex in the developing human brain.

PubMed

Simon, Matthew J; Murchison, Charles; Iliff, Jeffrey J

2018-02-01

Astrocytes play a critical role in regulating the interface between the cerebral vasculature and the central nervous system. Contributing to this is the astrocytic endfoot domain, a specialized structure that ensheathes the entirety of the vasculature and mediates signaling between endothelial cells, pericytes, and neurons. The astrocytic endfoot has been implicated as a critical element of the glymphatic pathway, and changes in protein expression profiles in this cellular domain are linked to Alzheimer's disease pathology. Despite this, basic physiological properties of this structure remain poorly understood including the developmental timing of its formation, and the protein components that localize there to mediate its functions. Here we use human transcriptome data from male and female subjects across several developmental stages and brain regions to characterize the gene expression profile of the dystrophin-associated complex (DAC), a known structural component of the astrocytic endfoot that supports perivascular localization of the astroglial water channel aquaporin-4. Transcriptomic profiling is also used to define genes exhibiting parallel expression profiles to DAC elements, generating a pool of candidate genes that encode gene products that may contribute to the physiological function of the perivascular astrocytic endfoot domain. We found that several genes encoding transporter proteins are transcriptionally associated with DAC genes. © 2017 Wiley Periodicals, Inc.

Geographical and environmental determinants of the genetic structure of wild barley in southeastern Anatolia.

PubMed

Pournosrat, Reza; Kaya, Selma; Shaaf, Salar; Kilian, Benjamin; Ozkan, Hakan

2018-01-01

Despite the global value of barley, compared to its wild progenitor, genetic variation in this crop has been drastically reduced due to the process of domestication, selection and improvement. In the medium term, this will negatively impact both the vulnerability and yield stability of barley against biotic and abiotic stresses under climate change. Returning to the crop wild relatives (CWR) as sources of new and beneficial alleles is a clear option for enhancing the resilience of diversity and adaptation to climate change. Southeastern Anatolia constitutes an important part of the natural distribution of wild barley in the Fertile Crescent where important crops were initially domesticated. In this study, we investigated genetic diversity in a comprehensive collection of 281 geo-referenced wild barley individuals from 92 collection sites with sample sizes ranging from 1 to 9 individuals per site, collected from southeastern Anatolia and 131 domesticated genotypes from 49 different countries using 40 EST-SSR markers. A total of 375 alleles were detected across entire collection, of which 283 were carried by domesticated genotypes and 316 alleles were present in the wild gene pool. The number of unique alleles in the wild and in the domesticated gene pool was 92 and 59, respectively. The population structure at K = 3 suggested two groups of wild barley namely G1-W consisting wild barley genotypes from the western part and G1-E comprising those mostly from the eastern part of the study area, with a sharp separation from the domesticated gene pool. The geographic and climatic factors jointly showed significant effects on the distribution of wild barley. Using a Latent Factor Mixed Model, we identified four candidate loci potentially involved in adaptation of wild barley to three environmental factors: temperature seasonality, mean temperature of driest quarter, and precipitation of coldest quarter. These loci are probably the targets of genomic regions, with potential roles against abiotic stresses.

Genetic Patterns of Common-Bean Seed Acquisition and Early-Stage Adoption Among Farmer Groups in Western Uganda

PubMed Central

Wilkus, Erin L.; Berny Mier y Teran, Jorge C.; Mukankusi, Clare M.; Gepts, Paul

2018-01-01

Widespread adoption of new varieties can be valuable, especially where improved agricultural production technologies are hard to access. However, as farmers adopt new varieties, in situ population structure and genetic diversity of their seed holdings can change drastically. Consequences of adoption are still poorly understood due to a lack of crop genetic diversity assessments and detailed surveys of farmers' seed management practices. Common bean (Phaseolus vulgaris) is an excellent model for these types of studies, as it has a long history of cultivation among smallholder farmers, exhibits eco-geographic patterns of diversity (e.g., Andean vs. Mesoamerican gene-pools), and has been subjected to post-Columbian dispersal and recent introduction of improved cultivars. The Hoima district of western Uganda additionally provides an excellent social setting for evaluating consequences of adoption because access to improved varieties has varied across farmer groups in this production region. This study establishes a baseline understanding of the common bean diversity found among household producers in Uganda and compares the crop population structure, diversity and consequences of adoption of household producers with different adoption practices. Molecular diversity analysis, based on 4,955 single nucleotide polymorphism (SNP) markers, evaluated a total of 1,156 seed samples that included 196 household samples collected from household producers in the Hoima district, 19 breeder-selected varieties used in participatory breeding activities that had taken place prior to the study in the region, and a global bean germplasm collection. Households that had participated in regional participatory breeding efforts were more likely to adopt new varieties and, consequently, diversify their seed stocks than those that had not participated. Of the three farmer groups that participated in breeding efforts, households from the farmer group with the longest history of bean production were more likely to conserve “Seed Engufu”, a local “Calima”-type variety of the Andean bean gene pool, and, at the same time, introduce rare Mesoamerican gene pool varieties into household seed stocks. PMID:29868053

Genetic Patterns of Common-Bean Seed Acquisition and Early-Stage Adoption Among Farmer Groups in Western Uganda.

PubMed

Wilkus, Erin L; Berny Mier Y Teran, Jorge C; Mukankusi, Clare M; Gepts, Paul

2018-01-01

Widespread adoption of new varieties can be valuable, especially where improved agricultural production technologies are hard to access. However, as farmers adopt new varieties, in situ population structure and genetic diversity of their seed holdings can change drastically. Consequences of adoption are still poorly understood due to a lack of crop genetic diversity assessments and detailed surveys of farmers' seed management practices. Common bean ( Phaseolus vulgaris ) is an excellent model for these types of studies, as it has a long history of cultivation among smallholder farmers, exhibits eco-geographic patterns of diversity (e.g., Andean vs. Mesoamerican gene-pools), and has been subjected to post-Columbian dispersal and recent introduction of improved cultivars. The Hoima district of western Uganda additionally provides an excellent social setting for evaluating consequences of adoption because access to improved varieties has varied across farmer groups in this production region. This study establishes a baseline understanding of the common bean diversity found among household producers in Uganda and compares the crop population structure, diversity and consequences of adoption of household producers with different adoption practices. Molecular diversity analysis, based on 4,955 single nucleotide polymorphism (SNP) markers, evaluated a total of 1,156 seed samples that included 196 household samples collected from household producers in the Hoima district, 19 breeder-selected varieties used in participatory breeding activities that had taken place prior to the study in the region, and a global bean germplasm collection. Households that had participated in regional participatory breeding efforts were more likely to adopt new varieties and, consequently, diversify their seed stocks than those that had not participated. Of the three farmer groups that participated in breeding efforts, households from the farmer group with the longest history of bean production were more likely to conserve "Seed Engufu", a local "Calima"-type variety of the Andean bean gene pool, and, at the same time, introduce rare Mesoamerican gene pool varieties into household seed stocks.

Geographical and environmental determinants of the genetic structure of wild barley in southeastern Anatolia

PubMed Central

Shaaf, Salar; Kilian, Benjamin

2018-01-01

Despite the global value of barley, compared to its wild progenitor, genetic variation in this crop has been drastically reduced due to the process of domestication, selection and improvement. In the medium term, this will negatively impact both the vulnerability and yield stability of barley against biotic and abiotic stresses under climate change. Returning to the crop wild relatives (CWR) as sources of new and beneficial alleles is a clear option for enhancing the resilience of diversity and adaptation to climate change. Southeastern Anatolia constitutes an important part of the natural distribution of wild barley in the Fertile Crescent where important crops were initially domesticated. In this study, we investigated genetic diversity in a comprehensive collection of 281 geo-referenced wild barley individuals from 92 collection sites with sample sizes ranging from 1 to 9 individuals per site, collected from southeastern Anatolia and 131 domesticated genotypes from 49 different countries using 40 EST-SSR markers. A total of 375 alleles were detected across entire collection, of which 283 were carried by domesticated genotypes and 316 alleles were present in the wild gene pool. The number of unique alleles in the wild and in the domesticated gene pool was 92 and 59, respectively. The population structure at K = 3 suggested two groups of wild barley namely G1-W consisting wild barley genotypes from the western part and G1-E comprising those mostly from the eastern part of the study area, with a sharp separation from the domesticated gene pool. The geographic and climatic factors jointly showed significant effects on the distribution of wild barley. Using a Latent Factor Mixed Model, we identified four candidate loci potentially involved in adaptation of wild barley to three environmental factors: temperature seasonality, mean temperature of driest quarter, and precipitation of coldest quarter. These loci are probably the targets of genomic regions, with potential roles against abiotic stresses. PMID:29420597

GPFrontend and GPGraphics: graphical analysis tools for genetic association studies.

PubMed

Uebe, Steffen; Pasutto, Francesca; Krumbiegel, Mandy; Schanze, Denny; Ekici, Arif B; Reis, André

2010-09-21

Most software packages for whole genome association studies are non-graphical, purely text based programs originally designed to run with UNIX-like operating systems. Graphical output is often not intended or supposed to be performed with other command line tools, e.g. gnuplot. Using the Microsoft .NET 2.0 platform and Visual Studio 2005, we have created a graphical software package to analyze data from microarray whole genome association studies, both for a DNA-pooling based approach as well as regular single sample data. Part of this package was made to integrate with GenePool 0.8.2, a previously existing software suite for GNU/Linux systems, which we have modified to run in a Microsoft Windows environment. Further modifications cause it to generate some additional data. This enables GenePool to interact with the .NET parts created by us. The programs we developed are GPFrontend, a graphical user interface and frontend to use GenePool and create metadata files for it, and GPGraphics, a program to further analyze and graphically evaluate output of different WGA analysis programs, among them also GenePool. Our programs enable regular MS Windows users without much experience in bioinformatics to easily visualize whole genome data from a variety of sources.

PTGER4 gene variant rs76523431 is a candidate risk factor for radiological joint damage in rheumatoid arthritis patients: a genetic study of six cohorts.

PubMed

Rodriguez-Rodriguez, Luis; Ivorra-Cortes, Jose; Carmona, F David; Martín, Javier; Balsa, Alejandro; van Steenbergen, Hanna W; van der Helm-van Mil, Annette H M; González-Álvaro, Isidoro; Fernandez-Gutiérrez, Benjamín

2015-11-05

Prostaglandin E receptor 4 (PTGER4) is implicated in immune regulation and bone metabolism. The aim of this study was to analyze its role in radiological joint damage in rheumatoid arthritis (RA). Six independent cohorts of patients with RA of European or North American descent were included, comprising 1789 patients with 5083 sets of X-rays. The Hospital Clínico San Carlos Rheumatoid Arthritis, Princesa Early Arthritis Register Longitudinal study, and Hospital Universitario de La Paz early arthritis (Spain) cohorts were used as discovery cohorts, and the Leiden Early Arthritis Clinic (The Netherlands), Wichita (United States), and National Databank for Rheumatic Diseases (United States and Canada) cohorts as replication cohorts. First, the PTGER4 rs6896969 single-nucleotide polymorphism (SNP) was genotyped using TaqMan assays and available Illumina Immunochip data and studied in the discovery and replication cohorts. Second, the PTGER4 gene and adjacent regions were analyzed using Immunochip genotyping data in the discovery cohorts. On the basis of pooled p values, linkage disequilibrium structure of the region, and location in regions with transcriptional properties, SNPs were selected for replication. The results from discovery, replication, and overall cohorts were pooled using inverse-variance-weighted meta-analysis. Influence of the polymorphisms on the overall radiological damage (constant effect) and on damage progression over time (time-varying effect) was analyzed. The rs6896969 polymorphism showed a significant association with radiological damage in the constant effect pooled analysis of the discovery cohorts, although no significant association was observed in the replication cohorts or the overall pooled analysis. Regarding the analysis of the PTGER4 region, 976 variants were analyzed in the discovery cohorts. From the constant and time-varying effect analyses, 12 and 20 SNPs, respectively, were selected for replication. Only the rs76523431 variant showed a significant association with radiographic progression in the time-varying effect pooled analysis of the discovery, replication, and overall cohorts. The overall pooled effect size was 1.10 (95 % confidence interval 1.05-1.14, p = 2.10 × 10(-5)), meaning that radiographic yearly progression was 10 % greater for each copy of the minor allele. The PTGER4 gene is a candidate risk factor for radiological progression in RA.

Strong influence of regional species pools on continent-wide structuring of local communities.

PubMed

Lessard, Jean-Philippe; Borregaard, Michael K; Fordyce, James A; Rahbek, Carsten; Weiser, Michael D; Dunn, Robert R; Sanders, Nathan J

2012-01-22

There is a long tradition in ecology of evaluating the relative contribution of the regional species pool and local interactions on the structure of local communities. Similarly, a growing number of studies assess the phylogenetic structure of communities, relative to that in the regional species pool, to examine the interplay between broad-scale evolutionary and fine-scale ecological processes. Finally, a renewed interest in the influence of species source pools on communities has shown that the definition of the source pool influences interpretations of patterns of community structure. We use a continent-wide dataset of local ant communities and implement ecologically explicit source pool definitions to examine the relative importance of regional species pools and local interactions for shaping community structure. Then we assess which factors underlie systematic variation in the structure of communities along climatic gradients. We find that the average phylogenetic relatedness of species in ant communities decreases from tropical to temperate regions, but the strength of this relationship depends on the level of ecological realism in the definition of source pools. We conclude that the evolution of climatic niches influences the phylogenetic structure of regional source pools and that the influence of regional source pools on local community structure is strong.

Functional genomics platform for pooled screening and mammalian genetic interaction maps

PubMed Central

Kampmann, Martin; Bassik, Michael C.; Weissman, Jonathan S.

2014-01-01

Systematic genetic interaction maps in microorganisms are powerful tools for identifying functional relationships between genes and defining the function of uncharacterized genes. We have recently implemented this strategy in mammalian cells as a two-stage approach. First, genes of interest are robustly identified in a pooled genome-wide screen using complex shRNA libraries. Second, phenotypes for all pairwise combinations of hit genes are measured in a double-shRNA screen and used to construct a genetic interaction map. Our protocol allows for rapid pooled screening under various conditions without a requirement for robotics, in contrast to arrayed approaches. Each stage of the protocol can be implemented in ~2 weeks, with additional time for analysis and generation of reagents. We discuss considerations for screen design, and present complete experimental procedures as well as a full computational analysis suite for identification of hits in pooled screens and generation of genetic interaction maps. While the protocols outlined here were developed for our original shRNA-based approach, they can be applied more generally, including to CRISPR-based approaches. PMID:24992097

A comprehensive study of the genomic differentiation between temperate Dent and Flint maize.

PubMed

Unterseer, Sandra; Pophaly, Saurabh D; Peis, Regina; Westermeier, Peter; Mayer, Manfred; Seidel, Michael A; Haberer, Georg; Mayer, Klaus F X; Ordas, Bernardo; Pausch, Hubert; Tellier, Aurélien; Bauer, Eva; Schön, Chris-Carolin

2016-07-08

Dent and Flint represent two major germplasm pools exploited in maize breeding. Several traits differentiate the two pools, like cold tolerance, early vigor, and flowering time. A comparative investigation of their genomic architecture relevant for quantitative trait expression has not been reported so far. Understanding the genomic differences between germplasm pools may contribute to a better understanding of the complementarity in heterotic patterns exploited in hybrid breeding and of mechanisms involved in adaptation to different environments. We perform whole-genome screens for signatures of selection specific to temperate Dent and Flint maize by comparing high-density genotyping data of 70 American and European Dent and 66 European Flint inbred lines. We find 2.2 % and 1.4 % of the genes are under selective pressure, respectively, and identify candidate genes associated with agronomic traits known to differ between the two pools. Taking flowering time as an example for the differentiation between Dent and Flint, we investigate candidate genes involved in the flowering network by phenotypic analyses in a Dent-Flint introgression library and find that the Flint haplotypes of the candidates promote earlier flowering. Within the flowering network, the majority of Flint candidates are associated with endogenous pathways in contrast to Dent candidate genes, which are mainly involved in response to environmental factors like light and photoperiod. The diversity patterns of the candidates in a unique panel of more than 900 individuals from 38 European landraces indicate a major contribution of landraces from France, Germany, and Spain to the candidate gene diversity of the Flint elite lines. In this study, we report the investigation of pool-specific differences between temperate Dent and Flint on a genome-wide scale. The identified candidate genes represent a promising source for the functional investigation of pool-specific haplotypes in different genetic backgrounds and for the evaluation of their potential for future crop improvement like the adaptation to specific environments.

Plasmids of Carotenoid-Producing Paracoccus spp. (Alphaproteobacteria) - Structure, Diversity and Evolution

PubMed Central

Maj, Anna; Dziewit, Lukasz; Czarnecki, Jakub; Wlodarczyk, Miroslawa; Baj, Jadwiga; Skrzypczyk, Grazyna; Giersz, Dorota; Bartosik, Dariusz

2013-01-01

Plasmids are components of many bacterial genomes. They enable the spread of a large pool of genetic information via lateral gene transfer. Many bacterial strains contain mega-sized replicons and these are particularly common in Alphaproteobacteria. Considerably less is known about smaller alphaproteobacterial plasmids. We analyzed the genomes of 14 such plasmids residing in 4 multireplicon carotenoid-producing strains of the genus Paracoccus (Alphaproteobacteria): P. aestuarii DSM 19484, P. haeundaensis LG P-21903, P. marcusii DSM 11574 and P. marcusii OS22. Comparative analyses revealed mosaic structures of the plasmids and recombinational shuffling of diverse genetic modules involved in (i) plasmid replication, (ii) stabilization (including toxin-antitoxin systems of the relBE/parDE, tad-ata, higBA, mazEF and toxBA families) and (iii) mobilization for conjugal transfer (encoding relaxases of the MobQ, MobP or MobV families). A common feature of the majority of the plasmids is the presence of AT-rich sequence islets (located downstream of exc1-like genes) containing genes, whose homologs are conserved in the chromosomes of many bacteria (encoding e.g. RelA/SpoT, SMC-like proteins and a retron-type reverse transcriptase). The results of this study have provided insight into the diversity and plasticity of plasmids of Paracoccus spp., and of the entire Alphaproteobacteria. Some of the identified plasmids contain replication systems not described previously in this class of bacteria. The composition of the plasmid genomes revealed frequent transfer of chromosomal genes into plasmids, which significantly enriches the pool of mobile DNA that can participate in lateral transfer. Many strains of Paracoccus spp. have great biotechnological potential, and the plasmid vectors constructed in this study will facilitate genetic studies of these bacteria. PMID:24260361

A High Resolution Genome-Wide Scan for Significant Selective Sweeps: An Application to Pooled Sequence Data in Laying Chickens

PubMed Central

Qanbari, Saber; Strom, Tim M.; Haberer, Georg; Weigend, Steffen; Gheyas, Almas A.; Turner, Frances; Burt, David W.; Preisinger, Rudolf; Gianola, Daniel; Simianer, Henner

2012-01-01

In most studies aimed at localizing footprints of past selection, outliers at tails of the empirical distribution of a given test statistic are assumed to reflect locus-specific selective forces. Significance cutoffs are subjectively determined, rather than being related to a clear set of hypotheses. Here, we define an empirical p-value for the summary statistic by means of a permutation method that uses the observed SNP structure in the real data. To illustrate the methodology, we applied our approach to a panel of 2.9 million autosomal SNPs identified from re-sequencing a pool of 15 individuals from a brown egg layer line. We scanned the genome for local reductions in heterozygosity, suggestive of selective sweeps. We also employed a modified sliding window approach that accounts for gaps in the sequence and increases scanning resolution by moving the overlapping windows by steps of one SNP only, and suggest to call this a “creeping window” strategy. The approach confirmed selective sweeps in the region of previously described candidate genes, i.e. TSHR, PRL, PRLHR, INSR, LEPR, IGF1, and NRAMP1 when used as positive controls. The genome scan revealed 82 distinct regions with strong evidence of selection (genome-wide p-value<0.001), including genes known to be associated with eggshell structure and immune system such as CALB1 and GAL cluster, respectively. A substantial proportion of signals was found in poor gene content regions including the most extreme signal on chromosome 1. The observation of multiple signals in a highly selected layer line of chicken is consistent with the hypothesis that egg production is a complex trait controlled by many genes. PMID:23209582

Emergence and Influence of the Zapatista Social Netwar

DTIC Science & Technology

2001-01-01

Dawkins , Richard, The Selfish Gene , New York: Oxford University Press, 1989. Diaz del Castillo, Bernal, The Conquest of New Spain [1568], Balti- more...as biological bodies are based on units of “self-replicating patterns of information” (p. 329). In his view (p. 192), Just as genes propagate...themselves in the gene pool by leaping from body to body via sperm or eggs, so memes propagate themselves in the meme pool by leaping from brain to brain via

The care and handling of the forest gene pool

Treesearch

Roy R. Silen; Ivan Doig

1976-01-01

What must be the world's most magnificent pool of forest genes has timbered our Pacific slopes.Why else do the tallest firs, pines, spruces, hemlocks, redwoods, and larches all rise along the Pacific Coast of North America? Does their hugeness simply thrust up from our deep soils and mild, rainy climate? From a vantage point of three...

A restoration practitioner's guide to the restoration gene pool concept

Treesearch

Thomas A. Jones; Thomas A. Monaco

2007-01-01

Choosing plant materials for each desired species is often one of the most difficult steps in developing a restoration plan. The Restoration Gene Pool concept was developed to clarify the options available to the ecological restoration practitioner in terms of plant materials. We present a decision-making flowchart incorporating the issues delineated in the Restoration...

High-resolution phylogenetic analysis of southeastern Europe traces major episodes of paternal gene flow among Slavic populations.

PubMed

Pericić, Marijana; Lauc, Lovorka Barać; Klarić, Irena Martinović; Rootsi, Siiri; Janićijevic, Branka; Rudan, Igor; Terzić, Rifet; Colak, Ivanka; Kvesić, Ante; Popović, Dan; Sijacki, Ana; Behluli, Ibrahim; Dordevic, Dobrivoje; Efremovska, Ljudmila; Bajec, Dorde D; Stefanović, Branislav D; Villems, Richard; Rudan, Pavao

2005-10-01

The extent and nature of southeastern Europe (SEE) paternal genetic contribution to the European genetic landscape were explored based on a high-resolution Y chromosome analysis involving 681 males from seven populations in the region. Paternal lineages present in SEE were compared with previously published data from 81 western Eurasian populations and 5,017 Y chromosome samples. The finding that five major haplogroups (E3b1, I1b* (xM26), J2, R1a, and R1b) comprise more than 70% of SEE total genetic variation is consistent with the typical European Y chromosome gene pool. However, distribution of major Y chromosomal lineages and estimated expansion signals clarify the specific role of this region in structuring of European, and particularly Slavic, paternal genetic heritage. Contemporary Slavic paternal gene pool, mostly characterized by the predominance of R1a and I1b* (xM26) and scarcity of E3b1 lineages, is a result of two major prehistoric gene flows with opposite directions: the post-Last Glacial Maximum R1a expansion from east to west, the Younger Dryas-Holocene I1b* (xM26) diffusion out of SEE in addition to subsequent R1a and I1b* (xM26) putative gene flows between eastern Europe and SEE, and a rather weak extent of E3b1 diffusion toward regions nowadays occupied by Slavic-speaking populations.

Identifying Genetic Signatures of Natural Selection Using Pooled Population Sequencing in Picea abies

PubMed Central

Chen, Jun; Källman, Thomas; Ma, Xiao-Fei; Zaina, Giusi; Morgante, Michele; Lascoux, Martin

2016-01-01

The joint inference of selection and past demography remain a costly and demanding task. We used next generation sequencing of two pools of 48 Norway spruce mother trees, one corresponding to the Fennoscandian domain, and the other to the Alpine domain, to assess nucleotide polymorphism at 88 nuclear genes. These genes are candidate genes for phenological traits, and most belong to the photoperiod pathway. Estimates of population genetic summary statistics from the pooled data are similar to previous estimates, suggesting that pooled sequencing is reliable. The nonsynonymous SNPs tended to have both lower frequency differences and lower FST values between the two domains than silent ones. These results suggest the presence of purifying selection. The divergence between the two domains based on synonymous changes was around 5 million yr, a time similar to a recent phylogenetic estimate of 6 million yr, but much larger than earlier estimates based on isozymes. Two approaches, one of them novel and that considers both FST and difference in allele frequencies between the two domains, were used to identify SNPs potentially under diversifying selection. SNPs from around 20 genes were detected, including genes previously identified as main target for selection, such as PaPRR3 and PaGI. PMID:27172202

Identifying Genetic Signatures of Natural Selection Using Pooled Population Sequencing in Picea abies.

PubMed

Chen, Jun; Källman, Thomas; Ma, Xiao-Fei; Zaina, Giusi; Morgante, Michele; Lascoux, Martin

2016-07-07

The joint inference of selection and past demography remain a costly and demanding task. We used next generation sequencing of two pools of 48 Norway spruce mother trees, one corresponding to the Fennoscandian domain, and the other to the Alpine domain, to assess nucleotide polymorphism at 88 nuclear genes. These genes are candidate genes for phenological traits, and most belong to the photoperiod pathway. Estimates of population genetic summary statistics from the pooled data are similar to previous estimates, suggesting that pooled sequencing is reliable. The nonsynonymous SNPs tended to have both lower frequency differences and lower FST values between the two domains than silent ones. These results suggest the presence of purifying selection. The divergence between the two domains based on synonymous changes was around 5 million yr, a time similar to a recent phylogenetic estimate of 6 million yr, but much larger than earlier estimates based on isozymes. Two approaches, one of them novel and that considers both FST and difference in allele frequencies between the two domains, were used to identify SNPs potentially under diversifying selection. SNPs from around 20 genes were detected, including genes previously identified as main target for selection, such as PaPRR3 and PaGI. Copyright © 2016 Chen et al.

Diagnostic value of secreted frizzled-related protein 2 gene promoter hypermethylation in stool for colorectal cancer: A meta-analysis.

PubMed

Zhou, Zhiran; Zhang, Huitian; Lei, Yunxia

2016-10-01

To evaluate the diagnostic value of secreted frizzled-related protein 2 (SFRP2) gene promoter hypermethylation in stool for colorectal cancer (CRC). Open published diagnostic study of SFRP2 gene promoter hypermethylation in stool for CRC detection was electronic searched in the databases of PubMed, EMBASE, Cochrane Library, Web of Science, and China National Knowledge Infrastructure. The data of true positive, false positive false negative, and true negative identified by stool SFRP2 gene hypermethylation was extracted and pooled for diagnostic sensitivity, specificity, and summary receiver operating characteristic (SROC) curve. According to the inclusion and exclusion criteria, we finally included nine publications with 792 cases in the meta-analysis. Thus, the diagnostic sensitivity was aggregated through random effect model. The pooled sensitivity was 0.82 with the corresponding 95% confidence interval (95% CI) of 0.79-0.85; the pooled specificity and its corresponding 95% CI were 0.47 and 0.40-0.53 by the random effect model; we pooled the SROC curve by sensitivity versus specificity according to data published in the nine studies. The area under the SROC curve was 0.70 (95% CI: 0.65-0.73). SFRP2 gene promoter hypermethylation in stool can was a potential biomarker for CRC diagnosis with relative high sensitivity.

Comprehensive analysis of correlation coefficients estimated from pooling heterogeneous microarray data

PubMed Central

2013-01-01

Background The synthesis of information across microarray studies has been performed by combining statistical results of individual studies (as in a mosaic), or by combining data from multiple studies into a large pool to be analyzed as a single data set (as in a melting pot of data). Specific issues relating to data heterogeneity across microarray studies, such as differences within and between labs or differences among experimental conditions, could lead to equivocal results in a melting pot approach. Results We applied statistical theory to determine the specific effect of different means and heteroskedasticity across 19 groups of microarray data on the sign and magnitude of gene-to-gene Pearson correlation coefficients obtained from the pool of 19 groups. We quantified the biases of the pooled coefficients and compared them to the biases of correlations estimated by an effect-size model. Mean differences across the 19 groups were the main factor determining the magnitude and sign of the pooled coefficients, which showed largest values of bias as they approached ±1. Only heteroskedasticity across the pool of 19 groups resulted in less efficient estimations of correlations than did a classical meta-analysis approach of combining correlation coefficients. These results were corroborated by simulation studies involving either mean differences or heteroskedasticity across a pool of N > 2 groups. Conclusions The combination of statistical results is best suited for synthesizing the correlation between expression profiles of a gene pair across several microarray studies. PMID:23822712

Extensive female-mediated gene flow and low phylogeography among seventeen goat breeds in southwest China.

PubMed

Zhao, Wei; Zhong, Tao; Wang, Lin Jie; Li, Li; Zhang, Hong Ping

2014-08-01

Indigenous Chinese goat mtDNA is highly diverse but lacks geographic specificity; however, whether gene flow or gene exchange contributed to this remains unknown. We reanalyzed a consensus fragment of 481 bp in the D-loop region from 339 individuals. The network and neighbor-joining tree revealed three divergent maternal haplogroups (A, B1, and B2) in 17 local breeds. Although high polymorphism resulting in 198 different haplotypes was observed (h = 0.984 ± 0.002; π = 0.0336 ± 0.0008), neither the distribution of haplotypes nor PCA analysis revealed any obvious geographic structure in the local breeds. Extensive gene flow was widely detected among breeds from southwest China. High levels of gene exchange were detected between Qianbei Brown goats and the other breeds, indicating either more contribution or introgression to their gene pools. This study will be helpful in understanding the phylogeography and gene flow among the goat breeds of southwest China.

Multiplexed resequencing analysis to identify rare variants in pooled DNA with barcode indexing using next-generation sequencer.

PubMed

Mitsui, Jun; Fukuda, Yoko; Azuma, Kyo; Tozaki, Hirokazu; Ishiura, Hiroyuki; Takahashi, Yuji; Goto, Jun; Tsuji, Shoji

2010-07-01

We have recently found that multiple rare variants of the glucocerebrosidase gene (GBA) confer a robust risk for Parkinson disease, supporting the 'common disease-multiple rare variants' hypothesis. To develop an efficient method of identifying rare variants in a large number of samples, we applied multiplexed resequencing using a next-generation sequencer to identification of rare variants of GBA. Sixteen sets of pooled DNAs from six pooled DNA samples were prepared. Each set of pooled DNAs was subjected to polymerase chain reaction to amplify the target gene (GBA) covering 6.5 kb, pooled into one tube with barcode indexing, and then subjected to extensive sequence analysis using the SOLiD System. Individual samples were also subjected to direct nucleotide sequence analysis. With the optimization of data processing, we were able to extract all the variants from 96 samples with acceptable rates of false-positive single-nucleotide variants.

Microorganisms Trapped Within Permafrost Ice In The Fox Permafrost Tunnel, Alaska

NASA Astrophysics Data System (ADS)

Katayama, T.; Tanaka, M.; Douglas, T. A.; Cai, Y.; Tomita, F.; Asano, K.; Fukuda, M.

2008-12-01

Several different types of massive ice are common in permafrost. Ice wedges are easily recognized by their shape and foliated structure. They grow syngenetically or epigenetically as a result of repeated cycles of frost cracking followed by the infiltration of snow, melt water, soil or other material into the open frost cracks. Material incorporated into ice wedges becomes frozen and preserved. Pool ice, another massive ice type, is formed by the freezing of water resting on top of frozen thermokarst sediment or melting wedges and is not foliated. The Fox Permafrost Tunnel in Fairbanks was excavated within the discontinuous permafrost zone of central Alaska and it contains permafrost, ice wedges, and pool ice preserved at roughly -3°C. We collected samples from five ice wedges and three pool ice structures in the Fox Permafrost Tunnel. If the microorganisms were incorporated into the ice during its formation, a community analysis of the microorganisms could elucidate the environment in which the ice was formed. Organic material from sediments in the tunnel was radiocarbon-dated between 14,000 and 30,000 years BP. However, it is still not clear when the ice wedges were formed or subsequently deformed because they are only partially exposed and their upper surfaces are above the tunnel walls. The objectives of our study were to determine the biogeochemical conditions during massive ice formation and to analyze the microbial community within the ices by incubation-based and DNA-based analyses. The geochemical profile and the PCR-DGGE band patterns of bacteria among five ice wedge and 3 portions of pool ice samples were markedly different. The DGGE band patterns of fungi were simple with a few bands of fungi or yeast. The dominant bands of ice wedge and pool ice samples were affiliated with the genus Geomyces and Doratomyces, respectively. Phylogenetic analysis using rRNA gene ITS regions indicated isolates of Geomyces spp. from different ice wedges were affiliated with different clusters. The enumeration of fungal colonies among the ice wedge and pool ice samples were also different. These results demonstrate that different massive ice structures had different microbial and geochemical environments or backgrounds when they were formed.

How voluntary prenatal diagnosis and selective abortion increase the abnormal human gene pool.

PubMed

Boss, J A

1990-06-01

It is often assumed that prenatal diagnosis followed by the selective abortion of "defective" fetuses has a positive eugenic effect. Although mandatory selective abortion of "defective" fetuses and, more important, carriers would tend to reduce the number of deleterious genes in the gene pool, the present program of voluntary prenatal diagnosis and selective abortion actually increases the number of deleterious genes. This raises the issue of freedom of choice regarding selective abortion and societal pressure on parents to undergo prenatal testing and to abort their fetus should it have a genetic disorder or be a carrier of one.

Pooled Genome-Wide Analysis to Identify Novel Risk Loci for Pediatric Allergic Asthma

PubMed Central

Ricci, Giampaolo; Astolfi, Annalisa; Remondini, Daniel; Cipriani, Francesca; Formica, Serena; Dondi, Arianna; Pession, Andrea

2011-01-01

Background Genome-wide association studies of pooled DNA samples were shown to be a valuable tool to identify candidate SNPs associated to a phenotype. No such study was up to now applied to childhood allergic asthma, even if the very high complexity of asthma genetics is an appropriate field to explore the potential of pooled GWAS approach. Methodology/Principal Findings We performed a pooled GWAS and individual genotyping in 269 children with allergic respiratory diseases comparing allergic children with and without asthma. We used a modular approach to identify the most significant loci associated with asthma by combining silhouette statistics and physical distance method with cluster-adapted thresholding. We found 97% concordance between pooled GWAS and individual genotyping, with 36 out of 37 top-scoring SNPs significant at individual genotyping level. The most significant SNP is located inside the coding sequence of C5, an already identified asthma susceptibility gene, while the other loci regulate functions that are relevant to bronchial physiopathology, as immune- or inflammation-mediated mechanisms and airway smooth muscle contraction. Integration with gene expression data showed that almost half of the putative susceptibility genes are differentially expressed in experimental asthma mouse models. Conclusion/Significance Combined silhouette statistics and cluster-adapted physical distance threshold analysis of pooled GWAS data is an efficient method to identify candidate SNP associated to asthma development in an allergic pediatric population. PMID:21359210

Single cell transcriptomic analysis of prostate cancer cells.

PubMed

Welty, Christopher J; Coleman, Ilsa; Coleman, Roger; Lakely, Bryce; Xia, Jing; Chen, Shu; Gulati, Roman; Larson, Sandy R; Lange, Paul H; Montgomery, Bruce; Nelson, Peter S; Vessella, Robert L; Morrissey, Colm

2013-02-16

The ability to interrogate circulating tumor cells (CTC) and disseminated tumor cells (DTC) is restricted by the small number detected and isolated (typically <10). To determine if a commercially available technology could provide a transcriptomic profile of a single prostate cancer (PCa) cell, we clonally selected and cultured a single passage of cell cycle synchronized C4-2B PCa cells. Ten sets of single, 5-, or 10-cells were isolated using a micromanipulator under direct visualization with an inverted microscope. Additionally, two groups of 10 individual DTC, each isolated from bone marrow of 2 patients with metastatic PCa were obtained. RNA was amplified using the WT-Ovation™ One-Direct Amplification System. The amplified material was hybridized on a 44K Whole Human Gene Expression Microarray. A high stringency threshold, a mean Alexa Fluor® 3 signal intensity above 300, was used for gene detection. Relative expression levels were validated for select genes using real-time PCR (RT-qPCR). Using this approach, 22,410, 20,423, and 17,009 probes were positive on the arrays from 10-cell pools, 5-cell pools, and single-cells, respectively. The sensitivity and specificity of gene detection on the single-cell analyses were 0.739 and 0.972 respectively when compared to 10-cell pools, and 0.814 and 0.979 respectively when compared to 5-cell pools, demonstrating a low false positive rate. Among 10,000 randomly selected pairs of genes, the Pearson correlation coefficient was 0.875 between the single-cell and 5-cell pools and 0.783 between the single-cell and 10-cell pools. As expected, abundant transcripts in the 5- and 10-cell samples were detected by RT-qPCR in the single-cell isolates, while lower abundance messages were not. Using the same stringency, 16,039 probes were positive on the patient single-cell arrays. Cluster analysis showed that all 10 DTC grouped together within each patient. A transcriptomic profile can be reliably obtained from a single cell using commercially available technology. As expected, fewer amplified genes are detected from a single-cell sample than from pooled-cell samples, however this method can be used to reliably obtain a transcriptomic profile from DTC isolated from the bone marrow of patients with PCa.

Population genomic analysis uncovers environmental stress-driven selection and adaptation of Lentinula edodes population in China.

PubMed

Xiao, Yang; Cheng, Xuanjin; Liu, Jun; Li, Chuang; Nong, Wenyan; Bian, Yinbing; Cheung, Man Kit; Kwan, Hoi Shan

2016-11-10

The elucidation of genome-wide variations could help reveal aspects of divergence, domestication, and adaptation of edible mushrooms. Here, we resequenced the whole genomes of 39 wild and 21 cultivated strains of Chinese Lentinula edodes, the shiitake mushroom. We identified three distinct genetic groups in the Chinese L. edodes population with robust differentiation. Results of phylogenetic and population structure analyses suggest that the cultivated strains and most of the wild trains of L. edodes in China possess different gene pools and two outlier strains show signatures of hybridization between groups. Eighty-four candidate genes contributing to population divergence were detected in outlier analysis, 18 of which are involved in response to environmental stresses. Gene enrichment analysis of group-specific single nucleotide polymorphisms showed that the cultivated strains were genetically diversified in biological processes related to stress response. As the formation of fruiting bodies is a stress-response process, we postulate that environment factors, such as temperature, drove the population divergence of L. edodes in China by natural or artificial selection. We also found phenotypic variations between groups and identified some wild strains that have potential to diversify the genetic pool for improving agricultural traits of L. edodes cultivars in China.

Population genomic analysis uncovers environmental stress-driven selection and adaptation of Lentinula edodes population in China

PubMed Central

Xiao, Yang; Cheng, Xuanjin; Liu, Jun; Li, Chuang; Nong, Wenyan; Bian, Yinbing; Cheung, Man Kit; Kwan, Hoi Shan

2016-01-01

The elucidation of genome-wide variations could help reveal aspects of divergence, domestication, and adaptation of edible mushrooms. Here, we resequenced the whole genomes of 39 wild and 21 cultivated strains of Chinese Lentinula edodes, the shiitake mushroom. We identified three distinct genetic groups in the Chinese L. edodes population with robust differentiation. Results of phylogenetic and population structure analyses suggest that the cultivated strains and most of the wild trains of L. edodes in China possess different gene pools and two outlier strains show signatures of hybridization between groups. Eighty-four candidate genes contributing to population divergence were detected in outlier analysis, 18 of which are involved in response to environmental stresses. Gene enrichment analysis of group-specific single nucleotide polymorphisms showed that the cultivated strains were genetically diversified in biological processes related to stress response. As the formation of fruiting bodies is a stress-response process, we postulate that environment factors, such as temperature, drove the population divergence of L. edodes in China by natural or artificial selection. We also found phenotypic variations between groups and identified some wild strains that have potential to diversify the genetic pool for improving agricultural traits of L. edodes cultivars in China. PMID:27830835

Construction of the BAC Library of Small Abalone (Haliotis diversicolor) for Gene Screening and Genome Characterization.

PubMed

Jiang, Likun; You, Weiwei; Zhang, Xiaojun; Xu, Jian; Jiang, Yanliang; Wang, Kai; Zhao, Zixia; Chen, Baohua; Zhao, Yunfeng; Mahboob, Shahid; Al-Ghanim, Khalid A; Ke, Caihuan; Xu, Peng

2016-02-01

The small abalone (Haliotis diversicolor) is one of the most important aquaculture species in East Asia. To facilitate gene cloning and characterization, genome analysis, and genetic breeding of it, we constructed a large-insert bacterial artificial chromosome (BAC) library, which is an important genetic tool for advanced genetics and genomics research. The small abalone BAC library includes 92,610 clones with an average insert size of 120 Kb, equivalent to approximately 7.6× of the small abalone genome. We set up three-dimensional pools and super pools of 18,432 BAC clones for target gene screening using PCR method. To assess the approach, we screened 12 target genes in these 18,432 BAC clones and identified 16 positive BAC clones. Eight positive BAC clones were then sequenced and assembled with the next generation sequencing platform. The assembled contigs representing these 8 BAC clones spanned 928 Kb of the small abalone genome, providing the first batch of genome sequences for genome evaluation and characterization. The average GC content of small abalone genome was estimated as 40.33%. A total of 21 protein-coding genes, including 7 target genes, were annotated into the 8 BACs, which proved the feasibility of PCR screening approach with three-dimensional pools in small abalone BAC library. One hundred fifty microsatellite loci were also identified from the sequences for marker development in the future. The BAC library and clone pools provided valuable resources and tools for genetic breeding and conservation of H. diversicolor.

From single-cell to cell-pool transcriptomes: stochasticity in gene expression and RNA splicing.

PubMed

Marinov, Georgi K; Williams, Brian A; McCue, Ken; Schroth, Gary P; Gertz, Jason; Myers, Richard M; Wold, Barbara J

2014-03-01

Single-cell RNA-seq mammalian transcriptome studies are at an early stage in uncovering cell-to-cell variation in gene expression, transcript processing and editing, and regulatory module activity. Despite great progress recently, substantial challenges remain, including discriminating biological variation from technical noise. Here we apply the SMART-seq single-cell RNA-seq protocol to study the reference lymphoblastoid cell line GM12878. By using spike-in quantification standards, we estimate the absolute number of RNA molecules per cell for each gene and find significant variation in total mRNA content: between 50,000 and 300,000 transcripts per cell. We directly measure technical stochasticity by a pool/split design and find that there are significant differences in expression between individual cells, over and above technical variation. Specific gene coexpression modules were preferentially expressed in subsets of individual cells, including one enriched for mRNA processing and splicing factors. We assess cell-to-cell variation in alternative splicing and allelic bias and report evidence of significant differences in splice site usage that exceed splice variation in the pool/split comparison. Finally, we show that transcriptomes from small pools of 30-100 cells approach the information content and reproducibility of contemporary RNA-seq from large amounts of input material. Together, our results define an experimental and computational path forward for analyzing gene expression in rare cell types and cell states.

Systematic investigation of the relationship between high myopia and polymorphisms of the MMP2, TIMP2, and TIMP3 genes by a DNA pooling approach.

PubMed

Leung, Kim Hung; Yiu, Wai Chi; Yap, Maurice K H; Ng, Po Wah; Fung, Wai Yan; Sham, Pak Chung; Yip, Shea Ping

2011-06-01

This study examined the relationship between high myopia and three myopia candidate genes--matrix metalloproteinase 2 (MMP2) and tissue inhibitor of metalloproteinase-2 and -3 (TIMP2 and TIMP3)--involved in scleral remodeling. Recruited for the study were unrelated adult Han Chinese who were high myopes (spherical equivalent, ≤ -6.0 D in both eyes; cases) and emmetropes (within ±1.0 D in both eyes; controls). Sample set 1 had 300 cases and 300 controls, and sample set 2 had 356 cases and 354 controls. Forty-nine tag single-nucleotide polymorphisms (SNPs) were selected from these candidate genes. The first stage was an initial screen of six case pools and six control pools constructed from sample set 1, each pool consisting of 50 distinct subjects of the same affection status. In the second stage, positive SNPs from the first stage were confirmed by genotyping individual samples forming the DNA pools. In the third stage, positive SNPs from stage 2 were replicated, with sample set 2 genotyped individually. Of the 49 SNPs screened by DNA pooling, three passed the lenient threshold of P < 0.10 (nested ANOVA) and were followed up by individual genotyping. Of the three SNPs genotyped, two TIMP3 SNPs were found to be significantly associated with high myopia by single-marker or haplotype analysis. However, the initial positive results could not be replicated by sample set 2. MMP2, TIPM2, and TIMP3 genes were not associated with high myopia in this Chinese sample and hence are unlikely to play a major role in the genetic susceptibility to high myopia.

Rapid gene identification in sugar beet using deep sequencing of DNA from phenotypic pools selected from breeding panels.

PubMed

Ries, David; Holtgräwe, Daniela; Viehöver, Prisca; Weisshaar, Bernd

2016-03-15

The combination of bulk segregant analysis (BSA) and next generation sequencing (NGS), also known as mapping by sequencing (MBS), has been shown to significantly accelerate the identification of causal mutations for species with a reference genome sequence. The usual approach is to cross homozygous parents that differ for the monogenic trait to address, to perform deep sequencing of DNA from F2 plants pooled according to their phenotype, and subsequently to analyze the allele frequency distribution based on a marker table for the parents studied. The method has been successfully applied for EMS induced mutations as well as natural variation. Here, we show that pooling genetically diverse breeding lines according to a contrasting phenotype also allows high resolution mapping of the causal gene in a crop species. The test case was the monogenic locus causing red vs. green hypocotyl color in Beta vulgaris (R locus). We determined the allele frequencies of polymorphic sequences using sequence data from two diverging phenotypic pools of 180 B. vulgaris accessions each. A single interval of about 31 kbp among the nine chromosomes was identified which indeed contained the causative mutation. By applying a variation of the mapping by sequencing approach, we demonstrated that phenotype-based pooling of diverse accessions from breeding panels and subsequent direct determination of the allele frequency distribution can be successfully applied for gene identification in a crop species. Our approach made it possible to identify a small interval around the causative gene. Sequencing of parents or individual lines was not necessary. Whenever the appropriate plant material is available, the approach described saves time compared to the generation of an F2 population. In addition, we provide clues for planning similar experiments with regard to pool size and the sequencing depth required.

Selective DNA Pooling for Determination of Linkage between a Molecular Marker and a Quantitative Trait Locus

PubMed Central

Darvasi, A.; Soller, M.

1994-01-01

Selective genotyping is a method to reduce costs in marker-quantitative trait locus (QTL) linkage determination by genotyping only those individuals with extreme, and hence most informative, quantitative trait values. The DNA pooling strategy (termed: ``selective DNA pooling'') takes this one step further by pooling DNA from the selected individuals at each of the two phenotypic extremes, and basing the test for linkage on marker allele frequencies as estimated from the pooled samples only. This can reduce genotyping costs of marker-QTL linkage determination by up to two orders of magnitude. Theoretical analysis of selective DNA pooling shows that for experiments involving backcross, F(2) and half-sib designs, the power of selective DNA pooling for detecting genes with large effect, can be the same as that obtained by individual selective genotyping. Power for detecting genes with small effect, however, was found to decrease strongly with increase in the technical error of estimating allele frequencies in the pooled samples. The effect of technical error, however, can be markedly reduced by replication of technical procedures. It is also shown that a proportion selected of 0.1 at each tail will be appropriate for a wide range of experimental conditions. PMID:7896115

Nuclear microsatellites reveal contrasting patterns of genetic structure between western and southeastern European populations of the common ash (Fraxinus excelsior L.).

PubMed

Heuertz, Myriam; Hausman, Jean-François; Hardy, Olivier J; Vendramin, Giovanni G; Frascaria-Lacoste, Nathalie; Vekemans, Xavier

2004-05-01

To determine extant patterns of population genetic structure in common ash and gain insight into postglacial recolonization processes, we applied multilocus-based Bayesian approaches to data from 36 European populations genotyped at five nuclear microsatellite loci. We identified two contrasting patterns in terms of population genetic structure: (1) a large area from the British Isles to Lithuania throughout central Europe constituted effectively a single deme, whereas (2) strong genetic differentiation occurred over short distances in Sweden and southeastern Europe. Concomitant geographical variation was observed in estimates of allelic richness and genetic diversity, which were lowest in populations from southeastern Europe, that is, in regions close to putative ice age refuges, but high in western and central Europe, that is, in more recently recolonized areas. We suggest that in southeastern Europe, restricted postglacial gene flow caused by a rapid expansion of refuge populations in a mountainous topography is responsible for the observed strong genetic structure. In contrast, admixture of previously differentiated gene pools and high gene flow at the onset of postglacial recolonization of western and central Europe would have homogenized the genetic structure and raised the levels of genetic diversity above values in the refuges.

Microbial contributions to coupled arsenic and sulfur cycling in the acid-sulfide hot spring Champagne Pool, New Zealand.

PubMed

Hug, Katrin; Maher, William A; Stott, Matthew B; Krikowa, Frank; Foster, Simon; Moreau, John W

2014-01-01

Acid-sulfide hot springs are analogs of early Earth geothermal systems where microbial metal(loid) resistance likely first evolved. Arsenic is a metalloid enriched in the acid-sulfide hot spring Champagne Pool (Waiotapu, New Zealand). Arsenic speciation in Champagne Pool follows reaction paths not yet fully understood with respect to biotic contributions and coupling to biogeochemical sulfur cycling. Here we present quantitative arsenic speciation from Champagne Pool, finding arsenite dominant in the pool, rim and outflow channel (55-75% total arsenic), and dithio- and trithioarsenates ubiquitously present as 18-25% total arsenic. In the outflow channel, dimethylmonothioarsenate comprised ≤9% total arsenic, while on the outflow terrace thioarsenates were present at 55% total arsenic. We also quantified sulfide, thiosulfate, sulfate and elemental sulfur, finding sulfide and sulfate as major species in the pool and outflow terrace, respectively. Elemental sulfur concentration reached a maximum at the terrace. Phylogenetic analysis of 16S rRNA genes from metagenomic sequencing revealed the dominance of Sulfurihydrogenibium at all sites and an increased archaeal population at the rim and outflow channel. Several phylotypes were found closely related to known sulfur- and sulfide-oxidizers, as well as sulfur- and sulfate-reducers. Bioinformatic analysis revealed genes underpinning sulfur redox transformations, consistent with sulfur speciation data, and illustrating a microbial role in sulfur-dependent transformation of arsenite to thioarsenate. Metagenomic analysis also revealed genes encoding for arsenate reductase at all sites, reflecting the ubiquity of thioarsenate and a need for microbial arsenate resistance despite anoxic conditions. Absence of the arsenite oxidase gene, aio, at all sites suggests prioritization of arsenite detoxification over coupling to energy conservation. Finally, detection of methyl arsenic in the outflow channel, in conjunction with increased sequences from Aquificaceae, supports a role for methyltransferase in thermophilic arsenic resistance. Our study highlights microbial contributions to coupled arsenic and sulfur cycling at Champagne Pool, with implications for understanding the evolution of microbial arsenic resistance in sulfidic geothermal systems.

[Structure of the gene pool of eastern Ukrainians from Y-chromosome haplogroups].

PubMed

Khar'kov, V N; Stepanov, V A; Borinskaia, S A; Kozhekbaeva, Zh M; Gusar, V A; Grechanina, E Ia; Puzyrev, V P; Khusnutdinova, E K; Iankovskiĭ, N K

2004-03-01

Y chromosomes from representative sample of Eastern Ukrainians (94 individuals) were analyzed for composition and frequencies of haplogroups, defined by 11 biallelic loci located in non-recombining part of the chromosome (SRY1532, YAP, 92R7, DYF155S2, 12f2, Tat, M9, M17, M25, M89, and M56). In the Ukrainian gene, pool six haplogroups were revealed: E, F (including G and I), J, N3, P, and R1a1. These haplogroups were earlier detected in a study of Y-chromosome diversity on the territory of Europe as a whole. The major haplogroup in the Ukrainian gene pool, haplogroup R1a1 (earlier designated HG3), accounted for about 44% of all Y chromosomes in the sample examined. This haplogroup is thought to mark the migration patterns of the early Indo-Europeans and is associated with the distribution of the Kurgan archaeological culture. The second major haplogroup is haplogroup F (21.3%), which is a combination of the lineages differing by the time of appearance. Haplogroup P found with the frequency of 9.6%, represents the genetic contribution of the population originating from the ancient autochthonous population of Europe. Haplogroups J and E (11.7 and 4.2%, respectively) mark the migration patterns of the Middle-Eastern agriculturists during the Neolithic. The presence of the N3 lineage (9.6%) is likely explained by a contribution of the assimilated Finno-Ugric tribes. The data on the composition and frequencies of Y-chromosome haplogroups in the sample studied substantially supplement the existing picture of the male lineage distribution in the Eastern Slav population.

Genetic structure in the Sherpa and neighboring Nepalese populations.

PubMed

Cole, Amy M; Cox, Sean; Jeong, Choongwon; Petousi, Nayia; Aryal, Dhana R; Droma, Yunden; Hanaoka, Masayuki; Ota, Masao; Kobayashi, Nobumitsu; Gasparini, Paolo; Montgomery, Hugh; Robbins, Peter; Di Rienzo, Anna; Cavalleri, Gianpiero L

2017-01-19

We set out to describe the fine-scale population structure across the Eastern region of Nepal. To date there is relatively little known about the genetic structure of the Sherpa residing in Nepal and their genetic relationship with the Nepalese. We assembled dense genotype data from a total of 1245 individuals representing Nepal and a variety of different populations resident across the greater Himalayan region including Tibet, China, India, Pakistan, Kazakhstan, Uzbekistan, Tajikistan and Kirghizstan. We performed analysis of principal components, admixture and homozygosity. We identified clear substructure across populations resident in the Himalayan arc, with genetic structure broadly mirroring geographical features of the region. Ethnic subgroups within Nepal show distinct genetic structure, on both admixture and principal component analysis. We detected differential proportions of ancestry from northern Himalayan populations across Nepalese subgroups, with the Nepalese Rai, Magar and Tamang carrying the greatest proportions of Tibetan ancestry. We show that populations dwelling on the Himalayan plateau have had a clear impact on the Northern Indian gene pool. We illustrate how the Sherpa are a remarkably isolated population, with little gene flow from surrounding Nepalese populations.

Gene expression changes in blood RNA after swimming in a chlorinated pool.

PubMed

Salas, Lucas A; Font-Ribera, Laia; Bustamante, Mariona; Sumoy, Lauro; Grimalt, Joan O; Bonnin, Sarah; Aguilar, Maria; Mattlin, Heidi; Hummel, Manuela; Ferrer, Anna; Kogevinas, Manolis; Villanueva, Cristina M

2017-08-01

Exposure to disinfection by-products (DBP) such as trihalomethanes (THM) in swimming pools has been linked to adverse health effects in humans, but their biological mechanisms are unclear. We evaluated short-term changes in blood gene expression of adult recreational swimmers after swimming in a chlorinated pool. Volunteers swam 40min in an indoor chlorinated pool. Blood samples were drawn and four THM (chloroform, bromodichloromethane, dibromochloromethane and bromoform) were measured in exhaled breath before and after swimming. Intensity of physical activity was measured as metabolic equivalents (METs). Gene expression in whole blood mRNA was evaluated using IlluminaHumanHT-12v3 Expression-BeadChip. Linear mixed models were used to evaluate the relationship between gene expression changes and THM exposure. Thirty-seven before-after pairs were analyzed. The median increase from baseline to after swimming were: 0.7 to 2.3 for MET, and 1.4 to 7.1μg/m 3 for exhaled total THM (sum of the four THM). Exhaled THM increased on average 0.94μg/m 3 per 1 MET. While 1643 probes were differentially expressed post-exposure. Of them, 189 were also associated with exhaled levels of individual/total THM or MET after False Discovery Rate. The observed associations with the exhaled THM were low to moderate (Log-fold change range: -0.17 to 0.15). In conclusion, we identified short-term gene expression changes associated with swimming in a pool that were minor in magnitude and their biological meaning was unspecific. The high collinearity between exhaled THM levels and intensity of physical activity precluded mutually adjusted models with both covariates. These exploratory results should be validated in future studies. Copyright © 2017. Published by Elsevier B.V.

Combined sequence and sequence-structure-based methods for analyzing RAAS gene SNPs: a computational approach.

PubMed

Singh, Kh Dhanachandra; Karthikeyan, Muthusamy

2014-12-01

The renin-angiotensin-aldosterone system (RAAS) plays a key role in the regulation of blood pressure (BP). Mutations on the genes that encode components of the RAAS have played a significant role in genetic susceptibility to hypertension and have been intensively scrutinized. The identification of such probably causal mutations not only provides insight into the RAAS but may also serve as antihypertensive therapeutic targets and diagnostic markers. The methods for analyzing the SNPs from the huge dataset of SNPs, containing both functional and neutral SNPs is challenging by the experimental approach on every SNPs to determine their biological significance. To explore the functional significance of genetic mutation (SNPs), we adopted combined sequence and sequence-structure-based SNP analysis algorithm. Out of 3864 SNPs reported in dbSNP, we found 108 missense SNPs in the coding region and remaining in the non-coding region. In this study, we are reporting only those SNPs in coding region to be deleterious when three or more tools are predicted to be deleterious and which have high RMSD from the native structure. Based on these analyses, we have identified two SNPs of REN gene, eight SNPs of AGT gene, three SNPs of ACE gene, two SNPs of AT1R gene, three SNPs of CYP11B2 gene and three SNPs of CMA1 gene in the coding region were found to be deleterious. Further this type of study will be helpful in reducing the cost and time for identification of potential SNP and also helpful in selecting potential SNP for experimental study out of SNP pool.

Protocols for Biotechnological Interventions in Improvement of Vanilla (Vanilla planifolia Andrews.).

PubMed

Divakaran, Minoo; Babu, K Nirmal; Peter, K V

2016-01-01

Vanilla (Vanilla planifolia Andrews (syn. V. fragrans Salisb.), a native of Central America, is the primary source of natural vanillin and plays a major role in the global economy. The gene pool of vanilla is threatened by deforestation and overcollection that has resulted in disappearance of natural habitats and wild species. Continuous vegetative propagation and lack of natural seed set and sufficient variations in the gene pool hamper crop improvement programs. In vitro techniques, one of the key tools of plant biotechnology, can be employed for overcoming specific problems, viz. production of disease-free clones, inducing somaclonal variations, developing hybrids, gene pool conservation, incorporating desired traits by distant hybridization, genetic engineering, etc. However, realization of these objectives necessitates standardization of protocols. This chapter describes the various protocols optimized for crop improvement in Vanilla species.

Impact of selective logging on inbreeding and gene dispersal in an Amazonian tree population of Carapa guianensis Aubl.

PubMed

Cloutier, D; Kanashiro, M; Ciampi, A Y; Schoen, D J

2007-02-01

Selective logging may impact patterns of genetic diversity within populations of harvested forest tree species by increasing distances separating conspecific trees, and modifying physical and biotic features of the forest habitat. We measured levels of gene diversity, inbreeding, pollen dispersal and spatial genetic structure (SGS) of an Amazonian insect-pollinated Carapa guianensis population before and after commercial selective logging. Similar levels of gene diversity and allelic richness were found before and after logging in both the adult and the seed generations. Pre- and post-harvest outcrossing rates were high, and not significantly different from one another. We found no significant levels of biparental inbreeding either before or after logging. Low levels of pollen pool differentiation were found, and the pre- vs. post-harvest difference was not significant. Pollen dispersal distance estimates averaged between 75 m and 265 m before logging, and between 76 m and 268 m after logging, depending on the value of tree density and the dispersal model used. There were weak and similar levels of differentiation of allele frequencies in the adults and in the pollen pool, before and after logging occurred, as well as weak and similar pre- and post-harvest levels of SGS among adult trees. The large neighbourhood sizes estimated suggest high historical levels of gene flow. Overall our results indicate that there is no clear short-term genetic impact of selective logging on this population of C. guianensis.

High-Resolution Analysis of Human Y-Chromosome Variation Shows a Sharp Discontinuity and Limited Gene Flow between Northwestern Africa and the Iberian Peninsula

PubMed Central

Bosch, Elena; Calafell, Francesc; Comas, David; Oefner, Peter J.; Underhill, Peter A.; Bertranpetit, Jaume

2001-01-01

In the present study we have analyzed 44 Y-chromosome biallelic polymorphisms in population samples from northwestern (NW) Africa and the Iberian Peninsula, which allowed us to place each chromosome unequivocally in a phylogenetic tree based on >150 polymorphisms. The most striking results are that contemporary NW African and Iberian populations were found to have originated from distinctly different patrilineages and that the Strait of Gibraltar seems to have acted as a strong (although not complete) barrier to gene flow. In NW African populations, an Upper Paleolithic colonization that probably had its origin in eastern Africa contributed 75% of the current gene pool. In comparison, ∼78% of contemporary Iberian Y chromosomes originated in an Upper Paleolithic expansion from western Asia, along the northern rim of the Mediterranean basin. Smaller contributions to these gene pools (constituting 13% of Y chromosomes in NW Africa and 10% of Y chromosomes in Iberia) came from the Middle East during the Neolithic and, during subsequent gene flow, from Sub-Saharan to NW Africa. Finally, bidirectional gene flow across the Strait of Gibraltar has been detected: the genetic contribution of European Y chromosomes to the NW African gene pool is estimated at 4%, and NW African populations may have contributed 7% of Iberian Y chromosomes. The Islamic rule of Spain, which began in a.d. 711 and lasted almost 8 centuries, left only a minor contribution to the current Iberian Y-chromosome pool. The high-resolution analysis of the Y chromosome allows us to separate successive migratory components and to precisely quantify each historical layer. PMID:11254456

A novel lineage of myoviruses infecting cyanobacteria is widespread in the oceans.

PubMed

Sabehi, Gazalah; Shaulov, Lihi; Silver, David H; Yanai, Itai; Harel, Amnon; Lindell, Debbie

2012-02-07

Viruses infecting bacteria (phages) are thought to greatly impact microbial population dynamics as well as the genome diversity and evolution of their hosts. Here we report on the discovery of a novel lineage of tailed dsDNA phages belonging to the family Myoviridae and describe its first representative, S-TIM5, that infects the ubiquitous marine cyanobacterium, Synechococcus. The genome of this phage encodes an entirely unique set of structural proteins not found in any currently known phage, indicating that it uses lineage-specific genes for virion morphogenesis and represents a previously unknown lineage of myoviruses. Furthermore, among its distinctive collection of replication and DNA metabolism genes, it carries a mitochondrial-like DNA polymerase gene, providing strong evidence for the bacteriophage origin of the mitochondrial DNA polymerase. S-TIM5 also encodes an array of bacterial-like metabolism genes commonly found in phages infecting cyanobacteria including photosynthesis, carbon metabolism and phosphorus acquisition genes. This suggests a common gene pool and gene swapping of cyanophage-specific genes among different phage lineages despite distinct sets of structural and replication genes. All cytosines following purine nucleotides are methylated in the S-TIM5 genome, constituting a unique methylation pattern that likely protects the genome from nuclease degradation. This phage is abundant in the Red Sea and S-TIM5 gene homologs are widespread in the oceans. This unusual phage type is thus likely to be an important player in the oceans, impacting the population dynamics and evolution of their primary producing cyanobacterial hosts.

A new family of β-helix proteins with similarities to the polysaccharide lyases

DOE PAGES

Close, Devin W.; D'Angelo, Sara; Bradbury, Andrew R. M.

2014-09-27

Microorganisms that degrade biomass produce diverse assortments of carbohydrate-active enzymes and binding modules. Despite tremendous advances in the genomic sequencing of these organisms, many genes do not have an ascribed function owing to low sequence identity to genes that have been annotated. Consequently, biochemical and structural characterization of genes with unknown function is required to complement the rapidly growing pool of genomic sequencing data. A protein with previously unknown function (Cthe_2159) was recently isolated in a genome-wide screen using phage display to identify cellulose-binding protein domains from the biomass-degrading bacterium Clostridium thermocellum. Here, the crystal structure of Cthe_2159 is presentedmore » and it is shown that it is a unique right-handed parallel β-helix protein. Despite very low sequence identity to known β-helix or carbohydrate-active proteins, Cthe_2159 displays structural features that are very similar to those of polysaccharide lyase (PL) families 1, 3, 6 and 9. Cthe_2159 is conserved across bacteria and some archaea and is a member of the domain of unknown function family DUF4353. This suggests that Cthe_2159 is the first representative of a previously unknown family of cellulose and/or acid-sugar binding β-helix proteins that share structural similarities with PLs. More importantly, these results demonstrate how functional annotation by biochemical and structural analysis remains a critical tool in the characterization of new gene products.« less

A new family of β-helix proteins with similarities to the polysaccharide lyases

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

Close, Devin W.; D'Angelo, Sara; Bradbury, Andrew R. M.

Microorganisms that degrade biomass produce diverse assortments of carbohydrate-active enzymes and binding modules. Despite tremendous advances in the genomic sequencing of these organisms, many genes do not have an ascribed function owing to low sequence identity to genes that have been annotated. Consequently, biochemical and structural characterization of genes with unknown function is required to complement the rapidly growing pool of genomic sequencing data. A protein with previously unknown function (Cthe_2159) was recently isolated in a genome-wide screen using phage display to identify cellulose-binding protein domains from the biomass-degrading bacterium Clostridium thermocellum. Here, the crystal structure of Cthe_2159 is presentedmore » and it is shown that it is a unique right-handed parallel β-helix protein. Despite very low sequence identity to known β-helix or carbohydrate-active proteins, Cthe_2159 displays structural features that are very similar to those of polysaccharide lyase (PL) families 1, 3, 6 and 9. Cthe_2159 is conserved across bacteria and some archaea and is a member of the domain of unknown function family DUF4353. This suggests that Cthe_2159 is the first representative of a previously unknown family of cellulose and/or acid-sugar binding β-helix proteins that share structural similarities with PLs. More importantly, these results demonstrate how functional annotation by biochemical and structural analysis remains a critical tool in the characterization of new gene products.« less

The complete mitochondrial genome of the sandbar shark Carcharhinus plumbeus.

PubMed

Blower, Dean C; Ovenden, Jennifer R

2016-01-01

The sandbar shark, Carcharhinus plumbeus, a major representative species in shark fisheries worldwide is now considered vulnerable to overfishing. A pool of 774,234 Roche 454 shotgun sequences from one individual were assembled into a 16,706 bp mitogenome with 33× average coverage depth. It comprised 13 protein coding genes, 22 transfer RNA's, 2 ribosomal genes and 2 non-coding regions, typical of a vertebrate mitogenome. As expected for sharks, an A-T nucleotide bias was evident. This adds to rapidly growing number of mitogenome assemblies for the economically important Carcharhinidae family. The C. plumbeus mitogenome will assist researchers, fisheries and conservation managers interested in shark molecular systematics, phylogeography, conservation genetics, population and stock structure.

Clinicopathologic features of patients with non-small cell lung cancer harboring the EML4-ALK fusion gene: a meta-analysis.

PubMed

Wang, Ying; Wang, Shumin; Xu, Shiguang; Qu, Jiaqi; Liu, Bo

2014-01-01

The frequencies of EML4-ALK fusion gene in non-small cell lung cancer (NSCLC) with different clinicopathologic features described by previous studies are inconsistent. The key demographic and pathologic features associated with EML4-ALK fusion gene have not been definitively established. This meta-analysis was conducted to compare the frequency of the EML4-ALK fusion gene in patients with different clinicopathologic features and to identify an enriched population of patients with NSCLC harboring EML4-ALK fusion gene. The Pubmed and Embase databases for all studies on EML4-ALK fusion gene in NSCLC patients were searched up to July 2014. A criteria list and exclusion criteria were established to screen the studies. The frequency of the EML4-ALK fusion gene and the clinicopathologic features, including smoking status, pathologic type, gender, and EGFR status were abstracted. Seventeen articles consisting of 4511 NSCLC cases were included in this meta-analysis. A significant lower EML4-ALK fusion gene positive rate was associated with smokers (pooled OR = 0.40, 95% CI = 0.30-0.54, P<0.00001). A significantly higher EML4-ALK fusion gene positivity rate was associated with adenocarcinomas (pooled OR = 2.53, 95% CI = 1.66-3.86, P<0.0001) and female (pooled OR = 0.61, 95% CI = 0.41-0.90, P = 0.01). We found that a significantly lower EML4-ALK fusion gene positivity rate was associated with EGFR mutation (pooled OR = 0.07, 95% CI = 0.03-0.19, P<0.00001). No publication bias was observed in any meta-analysis (all P value of Egger's test >0.05); however, because of the small sample size, no results were in the meta-analysis regarding EGFR gene status. This meta-analysis revealed that the EML4-ALK fusion gene is highly correlated with a never/light smoking history, female and the pathologic type of adenocarcinoma, and is largely mutually exclusive of EGFR.

Clinicopathologic Features of Patients with Non-Small Cell Lung Cancer Harboring the EML4-ALK Fusion Gene: A Meta-Analysis

PubMed Central

Wang, Shumin; Xu, Shiguang; Qu, Jiaqi

2014-01-01

Background The frequencies of EML4-ALK fusion gene in non-small cell lung cancer (NSCLC) with different clinicopathologic features described by previous studies are inconsistent. The key demographic and pathologic features associated with EML4-ALK fusion gene have not been definitively established. This meta-analysis was conducted to compare the frequency of the EML4-ALK fusion gene in patients with different clinicopathologic features and to identify an enriched population of patients with NSCLC harboring EML4-ALK fusion gene. Methods The Pubmed and Embase databases for all studies on EML4-ALK fusion gene in NSCLC patients were searched up to July 2014. A criteria list and exclusion criteria were established to screen the studies. The frequency of the EML4-ALK fusion gene and the clinicopathologic features, including smoking status, pathologic type, gender, and EGFR status were abstracted. Results Seventeen articles consisting of 4511 NSCLC cases were included in this meta-analysis. A significant lower EML4-ALK fusion gene positive rate was associated with smokers (pooled OR = 0.40, 95% CI = 0.30–0.54, P<0.00001). A significantly higher EML4-ALK fusion gene positivity rate was associated with adenocarcinomas (pooled OR = 2.53, 95% CI = 1.66–3.86, P<0.0001) and female (pooled OR = 0.61, 95% CI = 0.41–0.90, P = 0.01). We found that a significantly lower EML4-ALK fusion gene positivity rate was associated with EGFR mutation (pooled OR = 0.07, 95% CI = 0.03–0.19, P<0.00001). No publication bias was observed in any meta-analysis (all P value of Egger's test >0.05); however, because of the small sample size, no results were in the meta-analysis regarding EGFR gene status. Conclusion This meta-analysis revealed that the EML4-ALK fusion gene is highly correlated with a never/light smoking history, female and the pathologic type of adenocarcinoma, and is largely mutually exclusive of EGFR. PMID:25360721

A computational proposal for designing structured RNA pools for in vitro selection of RNAs.

PubMed

Kim, Namhee; Gan, Hin Hark; Schlick, Tamar

2007-04-01

Although in vitro selection technology is a versatile experimental tool for discovering novel synthetic RNA molecules, finding complex RNA molecules is difficult because most RNAs identified from random sequence pools are simple motifs, consistent with recent computational analysis of such sequence pools. Thus, enriching in vitro selection pools with complex structures could increase the probability of discovering novel RNAs. Here we develop an approach for engineering sequence pools that links RNA sequence space regions with corresponding structural distributions via a "mixing matrix" approach combined with a graph theory analysis. We define five classes of mixing matrices motivated by covariance mutations in RNA; these constructs define nucleotide transition rates and are applied to chosen starting sequences to yield specific nonrandom pools. We examine the coverage of sequence space as a function of the mixing matrix and starting sequence via clustering analysis. We show that, in contrast to random sequences, which are associated only with a local region of sequence space, our designed pools, including a structured pool for GTP aptamers, can target specific motifs. It follows that experimental synthesis of designed pools can benefit from using optimized starting sequences, mixing matrices, and pool fractions associated with each of our constructed pools as a guide. Automation of our approach could provide practical tools for pool design applications for in vitro selection of RNAs and related problems.

Genome-wide analysis of the structural genes regulating defense phenylpropanoid metabolism in Populus

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

Tschaplinski, Timothy J; Tsai, Chung-Jui; Harding, Scott A

Salicin-based phenolic glycosides, hydroxycinnamate derivatives and flavonoid-derived condensed tannins comprise up to one-third of Populus leaf dry mass. Genes regulating the abundance and chemical diversity of these substances have not been comprehensively analysed in tree species exhibiting this metabolically demanding level of phenolic metabolism. Here, shikimate-phenylpropanoid pathway genes thought to give rise to these phenolic products were annotated from the Populus genome, their expression assessed by semiquantitative or quantitative reverse transcription polymerase chain reaction (PCR), and metabolic evidence for function presented. Unlike Arabidopsis, Populus leaves accumulate an array of hydroxycinnamoyl-quinate esters, which is consistent with broadened function of the expandedmore » hydroxycinnamoyl-CoA transferase gene family. Greater flavonoid pathway diversity is also represented, and flavonoid gene families are larger. Consistent with expanded pathway function, most of these genes were upregulated during wound-stimulated condensed tannin synthesis in leaves. The suite of Populus genes regulating phenylpropanoid product accumulation should have important application in managing phenolic carbon pools in relation to climate change and global carbon cycling.« less

Exome Pool-Seq in neurodevelopmental disorders.

PubMed

Popp, Bernt; Ekici, Arif B; Thiel, Christian T; Hoyer, Juliane; Wiesener, Antje; Kraus, Cornelia; Reis, André; Zweier, Christiane

2017-12-01

High throughput sequencing has greatly advanced disease gene identification, especially in heterogeneous entities. Despite falling costs this is still an expensive and laborious technique, particularly when studying large cohorts. To address this problem we applied Exome Pool-Seq as an economic and fast screening technology in neurodevelopmental disorders (NDDs). Sequencing of 96 individuals can be performed in eight pools of 12 samples on less than one Illumina sequencer lane. In a pilot study with 96 cases we identified 27 variants, likely or possibly affecting function. Twenty five of these were identified in 923 established NDD genes (based on SysID database, status November 2016) (ACTB, AHDC1, ANKRD11, ATP6V1B2, ATRX, CASK, CHD8, GNAS, IFIH1, KCNQ2, KMT2A, KRAS, MAOA, MED12, MED13L, RIT1, SETD5, SIN3A, TCF4, TRAPPC11, TUBA1A, WAC, ZBTB18, ZMYND11), two in 543 (SysID) candidate genes (ZNF292, BPTF), and additionally a de novo loss-of-function variant in LRRC7, not previously implicated in NDDs. Most of them were confirmed to be de novo, but we also identified X-linked or autosomal-dominantly or autosomal-recessively inherited variants. With a detection rate of 28%, Exome Pool-Seq achieves comparable results to individual exome analyses but reduces costs by >85%. Compared with other large scale approaches using Molecular Inversion Probes (MIP) or gene panels, it allows flexible re-analysis of data. Exome Pool-Seq is thus well suited for large-scale, cost-efficient and flexible screening in characterized but heterogeneous entities like NDDs.

Registry in a tube: multiplexed pools of retrievable parts for genetic design space exploration

PubMed Central

Woodruff, Lauren B. A.; Gorochowski, Thomas E.; Roehner, Nicholas; Densmore, Douglas; Gordon, D. Benjamin; Nicol, Robert

2017-01-01

Abstract Genetic designs can consist of dozens of genes and hundreds of genetic parts. After evaluating a design, it is desirable to implement changes without the cost and burden of starting the construction process from scratch. Here, we report a two-step process where a large design space is divided into deep pools of composite parts, from which individuals are retrieved and assembled to build a final construct. The pools are built via multiplexed assembly and sequenced using next-generation sequencing. Each pool consists of ∼20 Mb of up to 5000 unique and sequence-verified composite parts that are barcoded for retrieval by PCR. This approach is applied to a 16-gene nitrogen fixation pathway, which is broken into pools containing a total of 55 848 composite parts (71.0 Mb). The pools encompass an enormous design space (1043 possible 23 kb constructs), from which an algorithm-guided 192-member 4.5 Mb library is built. Next, all 1030 possible genetic circuits based on 10 repressors (NOR/NOT gates) are encoded in pools where each repressor is fused to all permutations of input promoters. These demonstrate that multiplexing can be applied to encompass entire design spaces from which individuals can be accessed and evaluated. PMID:28007941

Modular design of synthetic gene circuits with biological parts and pools.

PubMed

Marchisio, Mario Andrea

2015-01-01

Synthetic gene circuits can be designed in an electronic fashion by displaying their basic components-Standard Biological Parts and Pools of molecules-on the computer screen and connecting them with hypothetical wires. This procedure, achieved by our add-on for the software ProMoT, was successfully applied to bacterial circuits. Recently, we have extended this design-methodology to eukaryotic cells. Here, highly complex components such as promoters and Pools of mRNA contain hundreds of species and reactions whose calculation demands a rule-based modeling approach. We showed how to build such complex modules via the joint employment of the software BioNetGen (rule-based modeling) and ProMoT (modularization). In this chapter, we illustrate how to utilize our computational tool for synthetic biology with the in silico implementation of a simple eukaryotic gene circuit that performs the logic AND operation.

Phylogenomics of Reichenowia parasitica, an Alphaproteobacterial Endosymbiont of the Freshwater Leech Placobdella parasitica

PubMed Central

Kvist, Sebastian; Narechania, Apurva; Oceguera-Figueroa, Alejandro; Fuks, Bella; Siddall, Mark E.

2011-01-01

Although several commensal alphaproteobacteria form close relationships with plant hosts where they aid in (e.g.,) nitrogen fixation and nodulation, only a few inhabit animal hosts. Among these, Reichenowia picta, R. ornata and R. parasitica, are currently the only known mutualistic, alphaproteobacterial endosymbionts to inhabit leeches. These bacteria are harbored in the epithelial cells of the mycetomal structures of their freshwater leech hosts, Placobdella spp., and these structures have no other obvious function than housing bacterial symbionts. However, the function of the bacterial symbionts has remained unclear. Here, we focused both on exploring the genomic makeup of R. parasitica and on performing a robust phylogenetic analysis, based on more data than previous hypotheses, to test its position among related bacteria. We sequenced a combined pool of host and symbiont DNA from 36 pairs of mycetomes and performed an in silico separation of the different DNA pools through subtractive scaffolding. The bacterial contigs were compared to 50 annotated bacterial genomes and the genome of the freshwater leech Helobdella robusta using a BLASTn protocol. Further, amino acid sequences inferred from the contigs were used as queries against the 50 bacterial genomes to establish orthology. A total of 358 orthologous genes were used for the phylogenetic analyses. In part, results suggest that R. parasitica possesses genes coding for proteins related to nitrogen fixation, iron/vitamin B translocation and plasmid survival. Our results also indicate that R. parasitica interacts with its host in part by transmembrane signaling and that several of its genes show orthology across Rhizobiaceae. The phylogenetic analyses support the nesting of R. parasitica within the Rhizobiaceae, as sister to a group containing Agrobacterium and Rhizobium species. PMID:22132238

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

PubMed

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

2016-05-01

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

Detecting differentially expressed genes in heterogeneous diseases using half Student's t-test.

PubMed

Hsu, Chun-Lun; Lee, Wen-Chung

2010-12-01

Microarray technology provides information about hundreds and thousands of gene-expression data in a single experiment. To search for disease-related genes, researchers test for those genes that are differentially expressed between the case subjects and the control subjects. The authors propose a new test, the 'half Student's t-test', specifically for detecting differentially expressed genes in heterogeneous diseases. Monte-Carlo simulation shows that the test maintains the nominal α level quite well for both normal and non-normal distributions. Power of the half Student's t is higher than that of the conventional 'pooled' Student's t when there is heterogeneity in the disease under study. The power gain by using the half Student's t can reach ∼10% when the standard deviation of the case group is 50% larger than that of the control group. Application to a colon cancer data reveals that when the false discovery rate (FDR) is controlled at 0.05, the half Student's t can detect 344 differentially expressed genes, whereas the pooled Student's t can detect only 65 genes. Or alternatively, if only 50 genes are to be selected, the FDR for the pooled Student's t has to be set at 0.0320 (false positive rate of ∼3%), but for the half Student's t, it can be at as low as 0.0001 (false positive rate of about one per ten thousands). The half Student's t-test is to be recommended for the detection of differentially expressed genes in heterogeneous diseases.

Cloning of the Pseudomonas aeruginosa outer membrane porin protein P gene: evidence for a linked region of DNA homology.

PubMed Central

Siehnel, R J; Worobec, E A; Hancock, R E

1988-01-01

The gene encoding the outer membrane phosphate-selective porin protein P from Pseudomonas aeruginosa was cloned into Escherichia coli. The protein product was expressed and transported to the outer membrane of an E. coli phoE mutant and assembled into functional trimers. Expression of a product of the correct molecular weight was confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot (immunoblot) analysis, using polyclonal antibodies to protein P monomer and trimer forms. Protein P trimers were partially purified from the E. coli clone and shown to form channels with the same conductance as those formed by protein P from P. aeruginosa. The location and orientation of the protein P-encoding (oprP) gene on the cloned DNA was identified by three methods: (i) mapping the insertion point of transposon Tn501 in a previously isolated P. aeruginosa protein P-deficient mutant; (ii) hybridization of restriction fragments from the cloned DNA to an oligonucleotide pool synthesized on the basis of the amino-terminal protein sequence of protein P; and (iii) fusion of a PstI fragment of the cloned DNA to the amino terminus of the beta-galactosidase gene of pUC8, producing a fusion protein that contained protein P-antigenic epitopes. Structural analysis of the cloned DNA and P. aeruginosa chromosomal DNA revealed the presence of two adjacent PstI fragments which cross-hybridized, suggesting a possible gene duplication. The P-related (PR) region hybridized to the oligonucleotide pool described above. When the PstI fragment which contained the PR region was fused to the beta-galactosidase gene of pUC8, a fusion protein was produced which reacted with a protein P-specific antiserum. However, the restriction endonuclease patterns of the PR region and the oprP gene differed significantly beyond the amino-terminal one-third of the two genes. Images PMID:2834340

Exact Length Distribution of Filamentous Structures Assembled from a Finite Pool of Subunits.

PubMed

Harbage, David; Kondev, Jané

2016-07-07

Self-assembling filamentous structures made of protein subunits are ubiquitous in cell biology. These structures are often highly dynamic, with subunits in a continuous state of flux, binding to and falling off of filaments. In spite of this constant turnover of their molecular parts, many cellular structures seem to maintain a well-defined size over time, which is often required for their proper functioning. One widely discussed mechanism of size regulation involves the cell maintaining a finite pool of protein subunits available for assembly. This finite pool mechanism can control the length of a single filament by having assembly proceed until the pool of free subunits is depleted to the point when assembly and disassembly are balanced. Still, this leaves open the question of whether the same mechanism can provide size control for multiple filamentous structures that are assembled from a common pool of protein subunits, as is often the case in cells. We address this question by solving the steady-state master equation governing the stochastic assembly and disassembly of multifilament structures made from a shared finite pool of subunits. We find that, while the total number of subunits within a multifilament structure is well-defined, individual filaments within the structure have a wide, power-law distribution of lengths. We also compute the phase diagram for two multifilament structures competing for the same pool of subunits and identify conditions for coexistence when both have a well-defined size. These predictions can be tested in cell experiments in which the size of the subunit pool or the number of filament nucleators is tuned.

Comparative analysis of grapevine whole-genome gene predictions, functional annotation, categorization and integration of the predicted gene sequences

PubMed Central

2012-01-01

Background The first draft assembly and gene prediction of the grapevine genome (8X base coverage) was made available to the scientific community in 2007, and functional annotation was developed on this gene prediction. Since then additional Sanger sequences were added to the 8X sequences pool and a new version of the genomic sequence with superior base coverage (12X) was produced. Results In order to more efficiently annotate the function of the genes predicted in the new assembly, it is important to build on as much of the previous work as possible, by transferring 8X annotation of the genome to the 12X version. The 8X and 12X assemblies and gene predictions of the grapevine genome were compared to answer the question, “Can we uniquely map 8X predicted genes to 12X predicted genes?” The results show that while the assemblies and gene structure predictions are too different to make a complete mapping between them, most genes (18,725) showed a one-to-one relationship between 8X predicted genes and the last version of 12X predicted genes. In addition, reshuffled genomic sequence structures appeared. These highlight regions of the genome where the gene predictions need to be taken with caution. Based on the new grapevine gene functional annotation and in-depth functional categorization, twenty eight new molecular networks have been created for VitisNet while the existing networks were updated. Conclusions The outcomes of this study provide a functional annotation of the 12X genes, an update of VitisNet, the system of the grapevine molecular networks, and a new functional categorization of genes. Data are available at the VitisNet website (http://www.sdstate.edu/ps/research/vitis/pathways.cfm). PMID:22554261

Strong population genetic structuring in an annual fish, Nothobranchius furzeri, suggests multiple savannah refugia in southern Mozambique.

PubMed

Bartáková, Veronika; Reichard, Martin; Janko, Karel; Polačik, Matej; Blažek, Radim; Reichwald, Kathrin; Cellerino, Alessandro; Bryja, Josef

2013-09-12

Intraspecific genetic variation of African fauna has been significantly affected by pronounced climatic fluctuations in Plio-Pleistocene, but, with the exception of large mammals, very limited empirical data on diversity of natural populations are available for savanna-dwelling animals. Nothobranchius furzeri is an annual fish from south-eastern Africa, inhabiting discrete temporary savannah pools outside main river alluvia. Their dispersal is limited and population processes affecting its genetic structure are likely a combination of those affecting terrestrial and aquatic taxa. N. furzeri is a model taxon in ageing research and several populations of known geographical origin are used in laboratory studies. Here, we analysed the genetic structure, diversity, historical demography and temporal patterns of divergence in natural populations of N. furzeri across its entire distribution range. Genetic structure and historical demography of N. furzeri were analysed using a combination of mitochondrial (partial cytochrome b sequences, 687 bp) and nuclear (13 microsatellites) markers in 693 fish from 36 populations. Genetic markers consistently demonstrated strong population structuring and suggested two main genetic groups associated with river basins. The split was dated to the Pliocene (>2 Mya). The northern group inhabits savannah pools across the basin of the intermittent river Chefu in south-western Mozambique and eastern Zimbabwe. The southern group (from southernmost Mozambique) is subdivided, with the River Limpopo forming a barrier (maximum divergence time 1 Mya). A strong habitat fragmentation (isolated temporary pools) is reflected in significant genetic structuring even between adjacent pools, with a major influence of genetic drift and significant isolation-by-distance. Analysis of historical demography revealed that the expansion of both groups is ongoing, supported by frequent founder effects in marginal parts of the range and evidence of secondary contact between Chefu and Limpopo populations. We demonstrated: (1) ancient (pre-Pleistocene) divergence between the two main N. furzeri lineages, their recent secondary contact and lack of reproductive isolation; (2) important genetic structuring attributed to the fragmented nature of their environment and isolation-by-distance, suggesting that dispersal is limited, occurs over short distances and is not directly associated with river routes; (3) an apparent role of the River Limpopo as a barrier to dispersal and gene flow.

Meta-analysis of six genes (BDNF, DRD1, DRD3, DRD4, GRIN2B and MAOA) involved in neuroplasticity and the risk for alcohol dependence.

PubMed

Forero, Diego A; López-León, Sandra; Shin, Hyoung Doo; Park, Byung Lae; Kim, Dai-Jin

2015-04-01

Alcohol-related problems have a large impact on human health, accounting for around 4% of deaths and 4.5% of disability-adjusted life-years around the world. Genetic factors could explain a significant fraction of the risk for alcohol dependence (AD). Recent meta-analyses have found significant pooled odds ratios (ORs) for variants in the ADH1B, ADH1C, DRD2 and HTR2A genes. In the present study, we carried out a meta-analysis of common variants in 6 candidate genes involved in neurotransmission and neuroplasticity: BDNF, DRD1, DRD3, DRD4, GRIN2B and MAOA. We carried out a systematic search for published association studies that analyzed the genes of interest. Relevant articles were retrieved and demographic and genetic data were extracted. Pooled ORs were calculated using a random-effects model using the Meta-Analyst program. Dominant, recessive and allelic models were tested and analyses were also stratified by ethnicity. Forty two published studies were included in the current meta-analysis: BDNF-rs6265 (nine studies), DRD1-rs4532 (four studies), DRD3-rs6280 (eleven studies), DRD4-VNTR (seven studies), GRIN2B-rs1806201 (three studies) and MAOA-uVNTR (eight studies). We did not find significant pooled ORs for any of the six genes, under different models and stratifying for ethnicity. In terms of the number of candidate genes included, this is one of the most comprehensive meta-analyses for genetics of AD. Pooled ORs did not support consistent associations with any of the six candidate genes tested. Future studies of novel genes of functional relevance and meta-analyses of quantitative endophenotypes could identify further susceptibility molecular factors for AD. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

New genes from non-coding sequence: the role of de novo protein-coding genes in eukaryotic evolutionary innovation.

PubMed

McLysaght, Aoife; Guerzoni, Daniele

2015-09-26

The origin of novel protein-coding genes de novo was once considered so improbable as to be impossible. In less than a decade, and especially in the last five years, this view has been overturned by extensive evidence from diverse eukaryotic lineages. There is now evidence that this mechanism has contributed a significant number of genes to genomes of organisms as diverse as Saccharomyces, Drosophila, Plasmodium, Arabidopisis and human. From simple beginnings, these genes have in some instances acquired complex structure, regulated expression and important functional roles. New genes are often thought of as dispensable late additions; however, some recent de novo genes in human can play a role in disease. Rather than an extremely rare occurrence, it is now evident that there is a relatively constant trickle of proto-genes released into the testing ground of natural selection. It is currently unknown whether de novo genes arise primarily through an 'RNA-first' or 'ORF-first' pathway. Either way, evolutionary tinkering with this pool of genetic potential may have been a significant player in the origins of lineage-specific traits and adaptations. © 2015 The Authors.

Antibiotic Administration Routes Significantly Influence the Levels of Antibiotic Resistance in Gut Microbiota

PubMed Central

Zhang, Lu; Huang, Ying; Zhou, Yang; Buckley, Timothy

2013-01-01

This study examined the impact of oral exposure to antibiotic-resistant bacteria and antibiotic administration methods on antibiotic resistance (AR) gene pools and the profile of resistant bacteria in host gastrointestinal (GI) tracts using C57BL/6J mice with natural gut microbiota. Mice inoculated with a mixture of tet(M)-carrying Enterococcus spp. or blaCMY-2-carrying Escherichia coli were treated with different doses of tetracycline hydrochloride (Tet) or ampicillin sodium (Amp) and delivered via either feed or intravenous (i.v.) injection. Quantitative PCR assessment of mouse fecal samples revealed that (i) AR gene pools were below the detection limit in mice without prior inoculation of AR gene carriers regardless of subsequent exposure to corresponding antibiotics; (ii) oral exposure to high doses of Tet and Amp in mice inoculated with AR gene carriers led to rapid enrichment of corresponding AR gene pools in feces; (iii) significantly less or delayed development of AR in the GI tract of the AR carrier-inoculated mice was observed when the same doses of antibiotics were administered via i.v. injection rather than oral administration; and (iv) antibiotic dosage, and maybe the excretion route, affected AR in the GI tract. The shift of dominant AR bacterial populations in the gut microbiota was consistent with the dynamics of AR gene pools. The emergence of endogenous resistant bacteria in the gut microbiota corresponding to drug exposure was also observed. Together, these data suggest that oral administration of antibiotics has a prominent effect on AR amplification and development in gut microbiota, which may be minimized by alternative drug administration approaches, as illustrated by i.v. injection in this study and proper drug selection. PMID:23689712

Antibiotic administration routes significantly influence the levels of antibiotic resistance in gut microbiota.

PubMed

Zhang, Lu; Huang, Ying; Zhou, Yang; Buckley, Timothy; Wang, Hua H

2013-08-01

This study examined the impact of oral exposure to antibiotic-resistant bacteria and antibiotic administration methods on antibiotic resistance (AR) gene pools and the profile of resistant bacteria in host gastrointestinal (GI) tracts using C57BL/6J mice with natural gut microbiota. Mice inoculated with a mixture of tet(M)-carrying Enterococcus spp. or blaCMY-2-carrying Escherichia coli were treated with different doses of tetracycline hydrochloride (Tet) or ampicillin sodium (Amp) and delivered via either feed or intravenous (i.v.) injection. Quantitative PCR assessment of mouse fecal samples revealed that (i) AR gene pools were below the detection limit in mice without prior inoculation of AR gene carriers regardless of subsequent exposure to corresponding antibiotics; (ii) oral exposure to high doses of Tet and Amp in mice inoculated with AR gene carriers led to rapid enrichment of corresponding AR gene pools in feces; (iii) significantly less or delayed development of AR in the GI tract of the AR carrier-inoculated mice was observed when the same doses of antibiotics were administered via i.v. injection rather than oral administration; and (iv) antibiotic dosage, and maybe the excretion route, affected AR in the GI tract. The shift of dominant AR bacterial populations in the gut microbiota was consistent with the dynamics of AR gene pools. The emergence of endogenous resistant bacteria in the gut microbiota corresponding to drug exposure was also observed. Together, these data suggest that oral administration of antibiotics has a prominent effect on AR amplification and development in gut microbiota, which may be minimized by alternative drug administration approaches, as illustrated by i.v. injection in this study and proper drug selection.

Whole Genome Association Study in a Homogenous Population in Shandong Peninsula of China Reveals JARID2 as a Susceptibility Gene for Schizophrenia

PubMed Central

Liu, Yang; Chen, Gang; Norton, Nadine; Liu, Wenmin; Zhu, Haining; Zhou, Peng; Luan, Meng; Yang, Shulin; Chen, Xing; Carroll, Liam; Williams, Nigel M.; O'Donovan, Michael C.; Kirov, George; Owen, Michael J.

2009-01-01

DNA pooling can provide an economic and efficient way to detect susceptibility loci to complex diseases. We carried out a genome screen with 400 microsatellite markers spaced at approximately 10 cm in two DNA pools consisting of 119 schizophrenia (SZ) patients and 119 controls recruited from a homogenous population in the Chang Le area of the Shandong peninsula of China. Association of D6S289, a dinucleotide repeat polymorphism in the JARID2 gene with SZ, was found and confirmed by individual genotyping (X2 = 17.89; P = .047). In order to refine the signal, we genotyped 14 single nucleotide polymorphisms (SNPs) covering JARID2 and the neighboring gene, DNTBP1, in an extended sample of 309 cases and 309 controls from Shandong peninsula (including the samples from the pools). However, rs2235258 and rs9654600 in JARID2 showed association in allelic, genotypic and haplotypic tests with SZ patients from Chang Le area. This was not replicates in the extended sample, we conclude that JARID2 could be a susceptibility gene for SZ. PMID:19884986

Hidden Dangers.

ERIC Educational Resources Information Center

Greven, Erik D. W.; Knox, Winthrop S.

1999-01-01

Discusses each of the major system components of a maintenance survey that can provide pool facilities with the information needed to properly maintain aging pools. Components examined include mechanical and structural systems; pool structure; and the filtration, circulation, and chemical treatment systems. (GR)

Origin and evolution of spliceosomal introns

PubMed Central

2012-01-01

Evolution of exon-intron structure of eukaryotic genes has been a matter of long-standing, intensive debate. The introns-early concept, later rebranded ‘introns first’ held that protein-coding genes were interrupted by numerous introns even at the earliest stages of life's evolution and that introns played a major role in the origin of proteins by facilitating recombination of sequences coding for small protein/peptide modules. The introns-late concept held that introns emerged only in eukaryotes and new introns have been accumulating continuously throughout eukaryotic evolution. Analysis of orthologous genes from completely sequenced eukaryotic genomes revealed numerous shared intron positions in orthologous genes from animals and plants and even between animals, plants and protists, suggesting that many ancestral introns have persisted since the last eukaryotic common ancestor (LECA). Reconstructions of intron gain and loss using the growing collection of genomes of diverse eukaryotes and increasingly advanced probabilistic models convincingly show that the LECA and the ancestors of each eukaryotic supergroup had intron-rich genes, with intron densities comparable to those in the most intron-rich modern genomes such as those of vertebrates. The subsequent evolution in most lineages of eukaryotes involved primarily loss of introns, with only a few episodes of substantial intron gain that might have accompanied major evolutionary innovations such as the origin of metazoa. The original invasion of self-splicing Group II introns, presumably originating from the mitochondrial endosymbiont, into the genome of the emerging eukaryote might have been a key factor of eukaryogenesis that in particular triggered the origin of endomembranes and the nucleus. Conversely, splicing errors gave rise to alternative splicing, a major contribution to the biological complexity of multicellular eukaryotes. There is no indication that any prokaryote has ever possessed a spliceosome or introns in protein-coding genes, other than relatively rare mobile self-splicing introns. Thus, the introns-first scenario is not supported by any evidence but exon-intron structure of protein-coding genes appears to have evolved concomitantly with the eukaryotic cell, and introns were a major factor of evolution throughout the history of eukaryotes. This article was reviewed by I. King Jordan, Manuel Irimia (nominated by Anthony Poole), Tobias Mourier (nominated by Anthony Poole), and Fyodor Kondrashov. For the complete reports, see the Reviewers’ Reports section. PMID:22507701

Convergent evolution of adenosine aptamers spanning bacterial, human, and random sequences revealed by structure-based bioinformatics and genomic SELEX

PubMed Central

Vu, Michael M. K.; Jameson, Nora E.; Masuda, Stuart J.; Lin, Dana; Larralde-Ridaura, Rosa; Lupták, Andrej

2012-01-01

SUMMARY Aptamers are structured macromolecules in vitro evolved to bind molecular targets, whereas in nature they form the ligand-binding domains of riboswitches. Adenosine aptamers of a single structural family were isolated several times from random pools but they have not been identified in genomic sequences. We used two unbiased methods, structure-based bioinformatics and human genome-based in vitro selection, to identify aptamers that form the same adenosine-binding structure in a bacterium, and several vertebrates, including humans. Two of the human aptamers map to introns of RAB3C and FGD3 genes. The RAB3C aptamer binds ATP with dissociation constants about ten times lower than physiological ATP concentration, while the minimal FGD3 aptamer binds ATP only co-transcriptionally. PMID:23102219

Association between maternal, fetal and paternal MTHFR gene C677T and A1298C polymorphisms and risk of recurrent pregnancy loss: a comprehensive evaluation.

PubMed

Yang, Yi; Luo, Yunyao; Yuan, Jing; Tang, Yidan; Xiong, Lang; Xu, MangMang; Rao, XuDong; Liu, Hao

2016-06-01

Numerous studies have investigated the associations between methylenetetrahydrofolate reductase (MTHFR) gene C677T and A1298C polymorphisms and risk of recurrent pregnancy loss (RPL); however, the results remain controversial. The aim of this study is to drive a more precise estimation of association between MTHFR gene polymorphisms and risk of RPL. We searched PubMed, EMBASE, Cochrane library, Web of Science and China Knowledge Resource Integrated Database for papers on MTHFR gene C677T and A1298C polymorphisms and RPL risk. The pooled odds ratios (ORs) with 95 % confidence intervals (CIs) were used to assess the strength of association in the homozygous model, heterozygous model, dominant model, recessive model and an additive model. The software STATA (Version 13.0) was used for statistical analysis. Overall, 57 articles were included in the final meta-analysis. In maternal group the MTHFR C677T polymorphism showed pooled odds ratios for the homozygous comparison [OR = 2.285, 95 % CI (1.702, 3.067)] and the MTHFR A1298C polymorphism showed pooled odds ratios for recessive model [OR = 1.594, 95 % CI (1.136, 2.238)]. In fetal group the MTHFR C677T polymorphism showed pooled odds ratios for dominant model [OR = 1.037, 95 % CI (0.567, 1.894)] and the MTHFR A1298C polymorphism showed pooled odds ratios for dominant model [OR = 1.495, 95 % CI (1.102, 2.026)]. In summary, the results of our meta-analysis indicate that maternal and paternal MTHFR gene C677T and A1298C polymorphisms are associated with RPL. We also observed a significant association between fetal MTHFR A1298C polymorphism and RPL but not C677T.

Association between SLC2A9 (GLUT9) gene polymorphisms and gout susceptibility: an updated meta-analysis.

PubMed

Zhang, Xu; Yang, Xiao; Wang, Mengmeng; Li, Xiaona; Xia, Qing; Xu, Shengqian; Xu, Jianhua; Cai, Guoqi; Wang, Li; Xin, Lihong; Zou, Yanfeng; Pan, Faming

2016-08-01

The relationship between the SLC2A9 (solute carrier family 2, member 9) gene polymorphisms and gout was still inconsistent among the individual genetic association studies. Therefore, this present research was aimed to systematically evaluate the association between SLC2A9 gene polymorphisms and gout susceptibility. Relevant studies were enrolled by searching databases systematically. The pooled odds ratios (ORs) with 95 % confidence intervals (CIs) were used to assess the associations. The heterogeneity between each of the studies was calculated by using the Q statistic methods, and Begg's funnel plot and Egger's tests were performed to evaluate publication bias. A total of 13 studies investigated four single nucleotide polymorphisms (SNPs) in SLC2A9 were included. In this study, we found that the allele C of rs3733591 was higher in patients than in controls in both all-pooled population [C vs. T: OR (95 % CI) = 1.432 (1.213-1.691)] and Asians-pooled population [C vs. T: OR (95 % CI) = 1.583 (1.365-1.835)]. The allele frequency C of s6449213 was lower in the gout patients than in controls in both all-pooled population and Caucasians-pooled population. Additionally, the allele frequency T of rs16890979 and the allele frequency C of rs1014290 were lower in gout patients than in controls. This study demonstrated that the genetic susceptibility for gout is associated with the SLC2A9 gene polymorphisms. Four of them except for the rs3733591 are protective SNPs in Caucasians, and rs16890979 and rs1014290 are protective SNPs in both Caucasians and Asians, while rs3733591 may be susceptibility SNP in Asians.

Structural Insight Into the Role of Mutual Polymorphism and Conservatism in the Contact Zone of the NFR5-K1 Heterodimer With the Nod Factor.

PubMed

Igolkina, A A; Porozov, Yu B; Chizhevskaya, E P; Andronov, E E

2018-01-01

Sandwich-like docking configurations of the heterodimeric complex of NFR5 and K1 Vicia sativa receptor-like kinases together with the putative ligand, Nod factor (NF) of Rhizobium leguminosarum bv. viciae , were modeled and two of the most probable configurations were assessed through the analysis of the mutual polymorphisms and conservatism. We carried out this analysis based on the hypothesis that in a contact zone of two docked components (proteins or ligands) the population polymorphism or conservatism is mutual, i.e., the variation in one component has a reflected variation in the other component. The population material of 30 wild-growing V. sativa (leaf pieces) was collected from a large field (uncultivated for the past 25-years) and pooled; form this pool, 100 randomly selected cloned fragments of NFR5 gene and 100 of K1 gene were sequenced by the Sanger method. Congruence between population trees of NFR5 and K1 haplotypes allowed us to select two respective haplotypes, build their 3D structures, and perform protein-protein docking. In a separate simulation, the protein-ligand docking between NFR5 and NF was carried out. We merged the results of the two docking experiments and extracted NFR5-NF-K1 complexes, in which NF was located within the cavity between two receptors. Molecular dynamics simulations indicated two out of six complexes as stable. Regions of mutual polymorphism in the contact zone of one complex overlapped with known NF structural variations produced by R. leguminosarum bv. viciae . A total of 74% of the contact zone of another complex contained mutually polymorphic and conservative areas. Common traits of the obtained two stable structures allowed us to hypothesize the functional role of three-domain structure of plant LysM-RLKs in their heteromers.

Local structuring factors of invertebrate communities in ephemeral freshwater rock pools and the influence of more permanent water bodies in the region

USGS Publications Warehouse

Jocque, M.; Graham, T.; Brendonck, L.

2007-01-01

We used three isolated clusters of small ephemeral rock pools on a sandstone flat in Utah to test the importance of local structuring processes on aquatic invertebrate communities. In the three clusters we characterized all ephemeral rock pools (total: 27) for their morphometry, and monitored their water quality, hydrology and community assemblage during a full hydrocycle. In each cluster we also sampled a set of more permanent interconnected freshwater systems positioned in a wash, draining the water from each cluster of rock pools. This design allowed additional testing for the potential role of more permanent water bodies in the region as source populations for the active dispersers and the effect on the community structure in the rock pools. Species richness and community composition in the rock pools correlated with level of permanence and the ammonia concentration. The length of the rock pool inundation cycle shaped community structure, most probably by inhibiting colonization by some taxa (e.g. tadpoles and insect larvae) through developmental constraints. The gradient in ammonia concentrations probably reflects differences in primary production. The more permanent water bodies in each wash differed both environmentally and in community composition from the connected set of rock pools. A limited set of active dispersers was observed in the rock pools. Our findings indicate that aquatic invertebrate communities in the ephemeral rock pools are mainly structured through habitat permanence, possibly linked with biotic interactions and primary production. ?? 2007 Springer Science+Business Media B.V.

Registry in a tube: multiplexed pools of retrievable parts for genetic design space exploration.

PubMed

Woodruff, Lauren B A; Gorochowski, Thomas E; Roehner, Nicholas; Mikkelsen, Tarjei S; Densmore, Douglas; Gordon, D Benjamin; Nicol, Robert; Voigt, Christopher A

2017-02-17

Genetic designs can consist of dozens of genes and hundreds of genetic parts. After evaluating a design, it is desirable to implement changes without the cost and burden of starting the construction process from scratch. Here, we report a two-step process where a large design space is divided into deep pools of composite parts, from which individuals are retrieved and assembled to build a final construct. The pools are built via multiplexed assembly and sequenced using next-generation sequencing. Each pool consists of ∼20 Mb of up to 5000 unique and sequence-verified composite parts that are barcoded for retrieval by PCR. This approach is applied to a 16-gene nitrogen fixation pathway, which is broken into pools containing a total of 55 848 composite parts (71.0 Mb). The pools encompass an enormous design space (1043 possible 23 kb constructs), from which an algorithm-guided 192-member 4.5 Mb library is built. Next, all 1030 possible genetic circuits based on 10 repressors (NOR/NOT gates) are encoded in pools where each repressor is fused to all permutations of input promoters. These demonstrate that multiplexing can be applied to encompass entire design spaces from which individuals can be accessed and evaluated. © The Author(s) 2016. Published by Oxford University Press on behalf of Nucleic Acids Research.

Whole genome comparison between table and wine grapes reveals a comprehensive catalog of structural variants

PubMed Central

2014-01-01

Background Grapevine (Vitis vinifera L.) is the most important Mediterranean fruit crop, used to produce both wine and spirits as well as table grape and raisins. Wine and table grape cultivars represent two divergent germplasm pools with different origins and domestication history, as well as differential characteristics for berry size, cluster architecture and berry chemical profile, among others. ‘Sultanina’ plays a pivotal role in modern table grape breeding providing the main source of seedlessness. This cultivar is also one of the most planted for fresh consumption and raisins production. Given its importance, we sequenced it and implemented a novel strategy for the de novo assembly of its highly heterozygous genome. Results Our approach produced a draft genome of 466 Mb, recovering 82% of the genes present in the grapevine reference genome; in addition, we identified 240 novel genes. A large number of structural variants and SNPs were identified. Among them, 45 (21 SNPs and 24 INDELs) were experimentally confirmed in ‘Sultanina’ and six SNPs in other 23 table grape varieties. Transposable elements corresponded to ca. 80% of the repetitive sequences involved in structural variants and more than 2,000 genes were affected in their structure by these variants. Some of these genes are likely involved in embryo development, suggesting that they may contribute to seedlessness, a key trait for table grapes. Conclusions This work produced the first structural variants and SNPs catalog for grapevine, constituting a novel and very powerful tool for genomic studies in this key fruit crop, particularly useful to support marker assisted breeding in table grapes. PMID:24397443

High-density SNP assay development for genetic analysis in maritime pine (Pinus pinaster).

PubMed

Plomion, C; Bartholomé, J; Lesur, I; Boury, C; Rodríguez-Quilón, I; Lagraulet, H; Ehrenmann, F; Bouffier, L; Gion, J M; Grivet, D; de Miguel, M; de María, N; Cervera, M T; Bagnoli, F; Isik, F; Vendramin, G G; González-Martínez, S C

2016-03-01

Maritime pine provides essential ecosystem services in the south-western Mediterranean basin, where it covers around 4 million ha. Its scattered distribution over a range of environmental conditions makes it an ideal forest tree species for studies of local adaptation and evolutionary responses to climatic change. Highly multiplexed single nucleotide polymorphism (SNP) genotyping arrays are increasingly used to study genetic variation in living organisms and for practical applications in plant and animal breeding and genetic resource conservation. We developed a 9k Illumina Infinium SNP array and genotyped maritime pine trees from (i) a three-generation inbred (F2) pedigree, (ii) the French breeding population and (iii) natural populations from Portugal and the French Atlantic coast. A large proportion of the exploitable SNPs (2052/8410, i.e. 24.4%) segregated in the mapping population and could be mapped, providing the densest ever gene-based linkage map for this species. Based on 5016 SNPs, natural and breeding populations from the French gene pool exhibited similar level of genetic diversity. Population genetics and structure analyses based on 3981 SNP markers common to the Portuguese and French gene pools revealed high levels of differentiation, leading to the identification of a set of highly differentiated SNPs that could be used for seed provenance certification. Finally, we discuss how the validated SNPs could facilitate the identification of ecologically and economically relevant genes in this species, improving our understanding of the demography and selective forces shaping its natural genetic diversity, and providing support for new breeding strategies. © 2015 John Wiley & Sons Ltd.

Pooled Enrichment Sequencing Identifies Diversity and Evolutionary Pressures at NLR Resistance Genes within a Wild Tomato Population

PubMed Central

Stam, Remco; Scheikl, Daniela; Tellier, Aurélien

2016-01-01

Nod-like receptors (NLRs) are nucleotide-binding domain and leucine-rich repeats containing proteins that are important in plant resistance signaling. Many of the known pathogen resistance (R) genes in plants are NLRs and they can recognize pathogen molecules directly or indirectly. As such, divergence and copy number variants at these genes are found to be high between species. Within populations, positive and balancing selection are to be expected if plants coevolve with their pathogens. In order to understand the complexity of R-gene coevolution in wild nonmodel species, it is necessary to identify the full range of NLRs and infer their evolutionary history. Here we investigate and reveal polymorphism occurring at 220 NLR genes within one population of the partially selfing wild tomato species Solanum pennellii. We use a combination of enrichment sequencing and pooling ten individuals, to specifically sequence NLR genes in a resource and cost-effective manner. We focus on the effects which different mapping and single nucleotide polymorphism calling software and settings have on calling polymorphisms in customized pooled samples. Our results are accurately verified using Sanger sequencing of polymorphic gene fragments. Our results indicate that some NLRs, namely 13 out of 220, have maintained polymorphism within our S. pennellii population. These genes show a wide range of πN/πS ratios and differing site frequency spectra. We compare our observed rate of heterozygosity with expectations for this selfing and bottlenecked population. We conclude that our method enables us to pinpoint NLR genes which have experienced natural selection in their habitat. PMID:27189991

Rapid Creation and Quantitative Monitoring of High Coverage shRNA Libraries

PubMed Central

Bassik, Michael C.; Lebbink, Robert Jan; Churchman, L. Stirling; Ingolia, Nicholas T.; Patena, Weronika; LeProust, Emily M.; Schuldiner, Maya; Weissman, Jonathan S.; McManus, Michael T.

2009-01-01

Short hairpin RNA (shRNA) libraries are limited by the low efficacy of many shRNAs, giving false negatives, and off-target effects, giving false positives. Here we present a strategy for rapidly creating expanded shRNA pools (∼30 shRNAs/gene) that are analyzed by deep-sequencing (EXPAND). This approach enables identification of multiple effective target-specific shRNAs from a complex pool, allowing a rigorous statistical evaluation of whether a gene is a true hit. PMID:19448642

MISSION LentiPlex pooled shRNA library screening in mammalian cells.

PubMed

Coussens, Matthew J; Corman, Courtney; Fischer, Ashley L; Sago, Jack; Swarthout, John

2011-12-21

RNA interference (RNAi) is an intrinsic cellular mechanism for the regulation of gene expression. Harnessing the innate power of this system enables us to knockdown gene expression levels in loss of gene function studies. There are two main methods for performing RNAi. The first is the use of small interfering RNAs (siRNAs) that are chemically synthesized, and the second utilizes short-hairpin RNAs (shRNAs) encoded within plasmids. The latter can be transfected into cells directly or packaged into replication incompetent lentiviral particles. The main advantages of using lentiviral shRNAs is the ease of introduction into a wide variety of cell types, their ability to stably integrate into the genome for long term gene knockdown and selection, and their efficacy in conducting high-throughput loss of function screens. To facilitate this we have created the LentiPlex pooled shRNA library. The MISSION LentiPlex Human shRNA Pooled Library is a genome-wide lentiviral pool produced using a proprietary process. The library consists of over 75,000 shRNA constructs from the TRC collection targeting 15,000+ human genes. Each library is tested for shRNA representation before product release to ensure robust library coverage. The library is provided in a ready-to-use lentiviral format at titers of at least 5 x 10(8) TU/ml via p24 assay and is pre-divided into ten subpools of approximately 8,000 shRNA constructs each. Amplification and sequencing primers are also provided for downstream target identification. Previous studies established a synergistic antitumor activity of TRAIL when combined with Paclitaxel in A549 cells, a human lung carcinoma cell line. In this study we demonstrate the application of a pooled LentiPlex shRNA library to rapidly conduct a positive selection screen for genes involved in the cytotoxicity of A549 cells when exposed to TRAIL and Paclitaxel. One barrier often encountered with high-throughput screens is the cost and difficulty in deconvolution; we also detail a cost-effective polyclonal approach utilizing traditional sequencing.

Hydroxyurea-Mediated Cytotoxicity Without Inhibition of Ribonucleotide Reductase.

PubMed

Liew, Li Phing; Lim, Zun Yi; Cohen, Matan; Kong, Ziqing; Marjavaara, Lisette; Chabes, Andrei; Bell, Stephen D

2016-11-01

In many organisms, hydroxyurea (HU) inhibits class I ribonucleotide reductase, leading to lowered cellular pools of deoxyribonucleoside triphosphates. The reduced levels for DNA precursors is believed to cause replication fork stalling. Upon treatment of the hyperthermophilic archaeon Sulfolobus solfataricus with HU, we observe dose-dependent cell cycle arrest, accumulation of DNA double-strand breaks, stalled replication forks, and elevated levels of recombination structures. However, Sulfolobus has a HU-insensitive class II ribonucleotide reductase, and we reveal that HU treatment does not significantly impact cellular DNA precursor pools. Profiling of protein and transcript levels reveals modulation of a specific subset of replication initiation and cell division genes. Notably, the selective loss of the regulatory subunit of the primase correlates with cessation of replication initiation and stalling of replication forks. Furthermore, we find evidence for a detoxification response induced by HU treatment. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

A 3,000-loci transcription map of chromosome 3B unravels the structural and functional features of gene islands in hexaploid wheat.

PubMed

Rustenholz, Camille; Choulet, Frédéric; Laugier, Christel; Safár, Jan; Simková, Hana; Dolezel, Jaroslav; Magni, Federica; Scalabrin, Simone; Cattonaro, Federica; Vautrin, Sonia; Bellec, Arnaud; Bergès, Hélène; Feuillet, Catherine; Paux, Etienne

2011-12-01

To improve our understanding of the organization and regulation of the wheat (Triticum aestivum) gene space, we established a transcription map of a wheat chromosome (3B) by hybridizing a newly developed wheat expression microarray with bacterial artificial chromosome pools from a new version of the 3B physical map as well as with cDNA probes derived from 15 RNA samples. Mapping data for almost 3,000 genes showed that the gene space spans the whole chromosome 3B with a 2-fold increase of gene density toward the telomeres due to an increase in the number of genes in islands. Comparative analyses with rice (Oryza sativa) and Brachypodium distachyon revealed that these gene islands are composed mainly of genes likely originating from interchromosomal gene duplications. Gene Ontology and expression profile analyses for the 3,000 genes located along the chromosome revealed that the gene islands are enriched significantly in genes sharing the same function or expression profile, thereby suggesting that genes in islands acquired shared regulation during evolution. Only a small fraction of these clusters of cofunctional and coexpressed genes was conserved with rice and B. distachyon, indicating a recent origin. Finally, genes with the same expression profiles in remote islands (coregulation islands) were identified suggesting long-distance regulation of gene expression along the chromosomes in wheat.

Genetic Association and Gene-Gene Interaction Analyses in African American Dialysis Patients With Nondiabetic Nephropathy

PubMed Central

Bostrom, Meredith A.; Kao, W.H. Linda; Li, Man; Abboud, Hanna E.; Adler, Sharon G.; Iyengar, Sudha K.; Kimmel, Paul L.; Hanson, Robert L.; Nicholas, Susanne B.; Rasooly, Rebekah S.; Sedor, John R.; Coresh, Josef; Kohn, Orly F.; Leehey, David J.; Thornley-Brown, Denyse; Bottinger, Erwin P.; Lipkowitz, Michael S.; Meoni, Lucy A.; Klag, Michael J.; Lu, Lingyi; Hicks, Pamela J.; Langefeld, Carl D.; Parekh, Rulan S.; Bowden, Donald W.; Freedman, Barry I.

2011-01-01

Background African Americans (AAs) have increased susceptibility to non-diabetic nephropathy relative to European Americans. Study Design Follow-up of a pooled genome-wide association study (GWAS) in AA dialysis patients with nondiabetic nephropathy; novel gene-gene interaction analyses. Setting & Participants Wake Forest sample: 962 AA nondiabetic nephropathy cases; 931 non-nephropathy controls. Replication sample: 668 Family Investigation of Nephropathy and Diabetes (FIND) AA nondiabetic nephropathy cases; 804 non-nephropathy controls. Predictors Individual genotyping of top 1420 pooled GWAS-associated single nucleotide polymorphisms (SNPs) and 54 SNPs in six nephropathy susceptibility genes. Outcomes APOL1 genetic association and additional candidate susceptibility loci interacting with, or independently from, APOL1. Results The strongest GWAS associations included two non-coding APOL1 SNPs, rs2239785 (odds ratio [OR], 0.33; dominant; p = 5.9 × 10−24) and rs136148 (OR, 0.54; additive; p = 1.1 × 10−7) with replication in FIND (p = 5.0 × 10−21 and 1.9 × 10−05, respectively). Rs2239785 remained significantly associated after controlling for the APOL1 G1 and G2 coding variants. Additional top hits included a CFH SNP(OR from meta-analysis in above 3367 AA cases and controls, 0.81; additive; p = 6.8 × 10−4). The 1420 SNPs were tested for interaction with APOL1 G1 and G2 variants. Several interactive SNPs were detected, the most significant was rs16854341 in the podocin gene (NPHS2) (p = 0.0001). Limitations Non-pooled GWAS have not been performed in AA nondiabetic nephropathy. Conclusions This follow-up of a pooled GWAS provides additional and independent evidence that APOL1 variants contribute to nondiabetic nephropathy in AAs and identified additional associated and interactive non-diabetic nephropathy susceptibility genes. PMID:22119407

Does a drop in the bucket make a splash? Assessing the impact of antibiotic use on plants.

PubMed

McManus, Patricia S

2014-06-01

Antibiotics are applied to plants to prevent bacterial diseases, although the diversity of antibiotics and total amounts used are dwarfed by antibiotic use in animal agriculture. Nevertheless, the release of antibiotics into the open environment during crop treatment draws scrutiny for its potential impact on the global pool of resistance genes. The main use of antibiotics on plants is application of streptomycin to prevent fire blight, a serious disease of apple and pear trees. A series of recent studies identified and quantified antibiotic resistance genes and profiled bacterial communities in apple orchard plots that were or were not sprayed with streptomycin. While the specific objectives and methods varied, the results of these studies suggest that streptomycin application for fire blight control does not influence bacterial community structure or increase the abundance of resistance genes in orchards. Copyright © 2014 Elsevier Ltd. All rights reserved.

Gene expression profiling of pre-eclamptic placentae by RNA sequencing.

PubMed

Kaartokallio, Tea; Cervera, Alejandra; Kyllönen, Anjuska; Laivuori, Krista; Kere, Juha; Laivuori, Hannele

2015-09-21

Pre-eclampsia is a common and complex pregnancy disorder that often involves impaired placental development. In order to identify altered gene expression in pre-eclamptic placenta, we sequenced placental transcriptomes of nine pre-eclamptic and nine healthy pregnant women in pools of three. The differential gene expression was tested both by including all the pools in the analysis and by excluding some of the pools based on phenotypic characteristics. From these analyses, we identified altogether 53 differently expressed genes, a subset of which was validated by qPCR in 20 cases and 19 controls. Furthermore, we conducted pathway and functional analyses which revealed disturbed vascular function and immunological balance in pre-eclamptic placenta. Some of the genes identified in our study have been reported by numerous microarray studies (BHLHE40, FSTL3, HK2, HTRA4, LEP, PVRL4, SASH1, SIGLEC6), but many have been implicated in only few studies or have not previously been linked to pre-eclampsia (ARMS2, BTNL9, CCSAP, DIO2, FER1L4, HPSE, LOC100129345, LYN, MYO7B, NCMAP, NDRG1, NRIP1, PLIN2, SBSPON, SERPINB9, SH3BP5, TET3, TPBG, ZNF175). Several of the molecules produced by these genes may have a role in the pathogenesis of pre-eclampsia, and some could qualify as biomarkers for prediction or detection of this pregnancy complication.

Gene expression profiling of pre-eclamptic placentae by RNA sequencing

PubMed Central

Kaartokallio, Tea; Cervera, Alejandra; Kyllönen, Anjuska; Laivuori, Krista; Laivuori, Hannele; Heinonen, Seppo; Kajantie, Eero; Kere, Juha; Kivinen, Katja; Pouta, Anneli

2015-01-01

Pre-eclampsia is a common and complex pregnancy disorder that often involves impaired placental development. In order to identify altered gene expression in pre-eclamptic placenta, we sequenced placental transcriptomes of nine pre-eclamptic and nine healthy pregnant women in pools of three. The differential gene expression was tested both by including all the pools in the analysis and by excluding some of the pools based on phenotypic characteristics. From these analyses, we identified altogether 53 differently expressed genes, a subset of which was validated by qPCR in 20 cases and 19 controls. Furthermore, we conducted pathway and functional analyses which revealed disturbed vascular function and immunological balance in pre-eclamptic placenta. Some of the genes identified in our study have been reported by numerous microarray studies (BHLHE40, FSTL3, HK2, HTRA4, LEP, PVRL4, SASH1, SIGLEC6), but many have been implicated in only few studies or have not previously been linked to pre-eclampsia (ARMS2, BTNL9, CCSAP, DIO2, FER1L4, HPSE, LOC100129345, LYN, MYO7B, NCMAP, NDRG1, NRIP1, PLIN2, SBSPON, SERPINB9, SH3BP5, TET3, TPBG, ZNF175). Several of the molecules produced by these genes may have a role in the pathogenesis of pre-eclampsia, and some could qualify as biomarkers for prediction or detection of this pregnancy complication. PMID:26388242

A defect in the thymidine kinase 2 gene causing isolated mitochondrial myopathy without mtDNA depletion.

PubMed

Leshinsky-Silver, E; Michelson, M; Cohen, S; Ginsberg, M; Sadeh, M; Barash, V; Lerman-Sagie, T; Lev, D

2008-07-01

Isolated mitochondrial myopathies (IMM) are either due to primary defects in mtDNA, in nuclear genes that control mtDNA abundance and structure such as thymidine kinase 2 (TK2), or due to CoQ deficiency. Defects in the TK2 gene have been found to be associated with mtDNA depletion attributed to a depleted mitochondrial dNTP pool in non-dividing cells. We report an unusual case of IMM, homozygous for the H90N mutation in the TK2 gene but unlike other cases with the same mutation, does not demonstrate mtDNA depletion. The patient's clinical course is relatively mild and a muscle biopsy showed ragged red muscle fibers with a mild decrease in complexes I and an increase in complexes IV and II activities. This report extends the phenotypic expression of TK2 defects and suggests that all patients who present with an IMM even with normal quantities of mtDNA should be screened for TK2 mutations.

Koolpinyah and Yata viruses: two newly recognised ephemeroviruses from tropical regions of Australia and Africa.

PubMed

Blasdell, Kim R; Widen, Steven G; Diviney, Sinéad M; Firth, Cadhla; Wood, Thomas G; Guzman, Hilda; Holmes, Edward C; Tesh, Robert B; Vasilakis, Nikos; Walker, Peter J

2014-12-05

Koolpinyah virus (KOOLV) isolated from healthy Australian cattle and Yata virus (YATV) isolated from a pool of Mansonia uniformis mosquitoes in the Central African Republic have been tentatively identified as rhabdoviruses. KOOLV was shown previously to be related antigenically to kotonkon virus, an ephemerovirus that has caused an ephemeral fever-like illness in cattle in Nigeria, but YATV failed to react antigenically with any other virus tested. Here we report the complete genome sequences of KOOLV (16,133 nt) and YATV (14,479 nt). Each has a complex genome organisation, with multiple genes, including a second non-structural glycoprotein (GNS) gene and a viroporin (α1) gene, between the G and L genes as is characteristic of ephemeroviruses. Based on an analysis of genome organisation, sequence identity and cross-neutralisation, we demonstrate that both KOOLV and YATV should be classified as two new species in the genus Ephemerovirus. Crown Copyright © 2014. Published by Elsevier B.V. All rights reserved.

Contemporary gene flow and mating system of Arabis alpina in a Central European alpine landscape

PubMed Central

Buehler, D.; Graf, R.; Holderegger, R.; Gugerli, F.

2012-01-01

Background and Aims Gene flow is important in counteracting the divergence of populations but also in spreading genes among populations. However, contemporary gene flow is not well understood across alpine landscapes. The aim of this study was to estimate contemporary gene flow through pollen and to examine the realized mating system in the alpine perennial plant, Arabis alpina (Brassicaceae). Methods An entire sub-alpine to alpine landscape of 2 km2 was exhaustively sampled in the Swiss Alps. Eighteen nuclear microsatellite loci were used to genotype 595 individuals and 499 offspring from 49 maternal plants. Contemporary gene flow by pollen was estimated from paternity analysis, matching the genotypes of maternal plants and offspring to the pool of likely father plants. Realized mating patterns and genetic structure were also estimated. Key Results Paternity analysis revealed several long-distance gene flow events (≤1 km). However, most outcrossing pollen was dispersed close to the mother plants, and 84 % of all offspring were selfed. Individuals that were spatially close were more related than by chance and were also more likely to be connected by pollen dispersal. Conclusions In the alpine landscape studied, genetic structure occurred on small spatial scales as expected for alpine plants. However, gene flow also covered large distances. This makes it plausible for alpine plants to spread beneficial alleles at least via pollen across landscapes at a short time scale. Thus, gene flow potentially facilitates rapid adaptation in A. alpina likely to be required under ongoing climate change. PMID:22492332

Rapid protein production from stable CHO cell pools using plasmid vector and the cumate gene-switch.

PubMed

Poulain, Adeline; Perret, Sylvie; Malenfant, Félix; Mullick, Alaka; Massie, Bernard; Durocher, Yves

2017-08-10

To rapidly produce large amounts of recombinant proteins, the generation of stable Chinese Hamster Ovary (CHO) cell pools represents a useful alternative to large-scale transient gene expression (TGE). We have developed a cell line (CHO BRI/rcTA ) allowing the inducible expression of recombinant proteins, based on the cumate gene switch. After the identification of optimal plasmid DNA topology (supercoiled vs linearized plasmid) for PEIpro™ mediated transfection and of optimal conditions for methionine sulfoximine (MSX) selection, we were able to generate CHO BRI/rcTA pools producing high levels of recombinant proteins. Volumetric productivities of up to 900mg/L were reproducibly achieved for a Fc fusion protein and up to 350mg/L for an antibody after 14days post-induction in non-optimized fed-batch cultures. In addition, we show that CHO pool volumetric productivities are not affected by a freeze-thaw cycle or following maintenance in culture for over one month in the presence of MSX. Finally, we demonstrate that volumetric protein production with the CR5 cumate-inducible promoter is three- to four-fold higher than with the human CMV or hybrid EF1α-HTLV constitutive promoters. These results suggest that the cumate-inducible CHO BRI/rcTA stable pool platform is a powerful and robust system for the rapid production of gram amounts of recombinant proteins. Crown Copyright © 2017. Published by Elsevier B.V. All rights reserved.

Prevalence of parvovirus in Minnesota turkeys.

PubMed

Sharafeldin, T A; Singh, A; Abdel-Glil, M Y; Mor, S K; Porter, R E; Goyal, S M

2017-02-01

Poult enteritis syndrome (PES) is characterized by enteritis and decreased body weight gain in growing turkey poults between one d and 7 wk of age. Another syndrome called light turkey syndrome (LTS) causes a decrease in body weight of adult tom turkeys in Minnesota leading to huge economic losses. Reovirus, rotavirus, and astrovirus have been found in LTS and PES flocks in Minnesota. We tested 80 fecal pools collected from four LTS flocks and 35 fecal pools from non-LTS flocks for the presence of parvovirus. In addition, 116 fecal and meconium samples from turkeys submitted to the Minnesota Veterinary Diagnostic Laboratory (MVDL) also were tested. The samples were tested by PCR using primers for the non-structural 1 (NS1) gene of parvovirus. Of the 80 samples from LTS flocks, 41 were positive for parvovirus while 20 of 35 samples from non-LTS flocks were positive. The prevalence of parvovirus in fecal samples submitted to MVDL was relatively low; only five of the 116 pools were positive. The partial NS1 gene sequences from LTS and non-LTS samples showed 98 to 100% nt identity except for one divergent turkey parvovirus (TuPV) strain that revealed 90% identity and clustered with chicken-like parvoviruses. The presence of this divergent strain suggests circulation of a recombinant strain of TuPV in Minnesota turkeys. Our results indicate that TuPVs are circulating in both LTS and non-LTS flocks of turkeys in Minnesota, and further experimental studies are indicated to study the role of TuPV in LTS. © 2016 Poultry Science Association Inc.

Large Deletions at the SHOX Locus in the Pseudoautosomal Region Are Associated with Skeletal Atavism in Shetland Ponies

PubMed Central

Rafati, Nima; Andersson, Lisa S.; Mikko, Sofia; Feng, Chungang; Raudsepp, Terje; Pettersson, Jessica; Janecka, Jan; Wattle, Ove; Ameur, Adam; Thyreen, Gunilla; Eberth, John; Huddleston, John; Malig, Maika; Bailey, Ernest; Eichler, Evan E.; Dalin, Göran; Chowdary, Bhanu; Andersson, Leif; Lindgren, Gabriella; Rubin, Carl-Johan

2016-01-01

Skeletal atavism in Shetland ponies is a heritable disorder characterized by abnormal growth of the ulna and fibula that extend the carpal and tarsal joints, respectively. This causes abnormal skeletal structure and impaired movements, and affected foals are usually killed. In order to identify the causal mutation we subjected six confirmed Swedish cases and a DNA pool consisting of 21 control individuals to whole genome resequencing. We screened for polymorphisms where the cases and the control pool were fixed for opposite alleles and observed this signature for only 25 SNPs, most of which were scattered on genome assembly unassigned scaffolds. Read depth analysis at these loci revealed homozygosity or compound heterozygosity for two partially overlapping large deletions in the pseudoautosomal region (PAR) of chromosome X/Y in cases but not in the control pool. One of these deletions removes the entire coding region of the SHOX gene and both deletions remove parts of the CRLF2 gene located downstream of SHOX. The horse reference assembly of the PAR is highly fragmented, and in order to characterize this region we sequenced bacterial artificial chromosome (BAC) clones by single-molecule real-time (SMRT) sequencing technology. This considerably improved the assembly and enabled size estimations of the two deletions to 160−180 kb and 60−80 kb, respectively. Complete association between the presence of these deletions and disease status was verified in eight other affected horses. The result of the present study is consistent with previous studies in humans showing crucial importance of SHOX for normal skeletal development. PMID:27207956

Predicting the activity of drugs for a group of imidazopyridine anticoccidial compounds.

PubMed

Si, Hongzong; Lian, Ning; Yuan, Shuping; Fu, Aiping; Duan, Yun-Bo; Zhang, Kejun; Yao, Xiaojun

2009-10-01

Gene expression programming (GEP) is a novel machine learning technique. The GEP is used to build nonlinear quantitative structure-activity relationship model for the prediction of the IC(50) for the imidazopyridine anticoccidial compounds. This model is based on descriptors which are calculated from the molecular structure. Four descriptors are selected from the descriptors' pool by heuristic method (HM) to build multivariable linear model. The GEP method produced a nonlinear quantitative model with a correlation coefficient and a mean error of 0.96 and 0.24 for the training set, 0.91 and 0.52 for the test set, respectively. It is shown that the GEP predicted results are in good agreement with experimental ones.

Origin of year-long bean (Phaseolus dumosus Macfady, Fabaceae) from reticulated hybridization events between multiple Phaseolus species.

PubMed

Mina-Vargas, Angela M; McKeown, Peter C; Flanagan, Nicola S; Debouck, Daniel G; Kilian, Andrzej; Hodkinson, Trevor R; Spillane, Charles

2016-08-06

Improved understanding of the secondary gene pools of crops is essential for advancing genetic gain in breeding programmes. Common bean, Phaseolus vulgaris, is a staple crop with several wild relatives in its secondary gene pool. The year-long bean, P. dumosus, an important crop in Guatemala, is considered particularly closely related to P. vulgaris and a potential source of novel variation. However, the genetic diversity and relationship to other Phaseolus species of P. dumosus remain unclear. We conducted the first comprehensive investigation of P. dumosus genetic diversity using both nuclear and chloroplast genome markers. Our nuclear marker set included over 700 markers present within the Phaseolus DArT (Diversity Arrays Technology) array, which we applied to P. dumosus and other relatives of P. vulgaris (including every secondary gene pool species: P. acutifolius, P. albescens, P. coccineus and P. costaricensis). Phaseolus dumosus arose from hybridization of P. vulgaris and P. coccineus, followed by at least two later hybridizations with sympatric congener populations. Existing P. dumosus collections have low genetic diversity. The under-utilized crop P. dumosus has a complex hybrid origin. Further sampling in the region in which it arose may uncover additional germplasm for introgressing favourable traits into crops within the P. vulgaris gene pool. © The Author 2016. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Bacteria from Animals as a Pool of Antimicrobial Resistance Genes

PubMed Central

Argudín, Maria Angeles; Deplano, Ariane; Meghraoui, Alaeddine; Dodémont, Magali; Heinrichs, Amelie; Denis, Olivier; Nonhoff, Claire; Roisin, Sandrine

2017-01-01

Antimicrobial agents are used in both veterinary and human medicine. The intensive use of antimicrobials in animals may promote the fixation of antimicrobial resistance genes in bacteria, which may be zoonotic or capable to transfer these genes to human-adapted pathogens or to human gut microbiota via direct contact, food or the environment. This review summarizes the current knowledge of the use of antimicrobial agents in animal health and explores the role of bacteria from animals as a pool of antimicrobial resistance genes for human bacteria. This review focused in relevant examples within the ESC(K)APE (Enterococcus faecium, Staphylococcus aureus, Clostridium difficile (Klebsiella pneumoniae), Acinetobacter baumannii, Pseudomonas aeruginosa, and Enterobacteriaceae) group of bacterial pathogens that are the leading cause of nosocomial infections throughout the world. PMID:28587316

Is variation in susceptibility to Phytophthora ramorum correlated with population genetic structure in coast live oak (Quercus agrifolia)?

PubMed

Dodd, Richard S; Hüberli, Daniel; Douhovnikoff, Vlad; Harnik, Tamar Y; Afzal-Rafii, Zara; Garbelotto, Matteo

2005-01-01

California coastal woodlands are suffering severe disease and mortality as a result of infection from Phytophthora ramorum. Quercus agrifolia is one of the major woodland species at risk. This study investigated within- and among-population variation in host susceptibility to inoculation with P. ramorum and compared this with population genetic structure using molecular markers. Susceptibility was assessed using a branch-cutting inoculation test. Trees were selected from seven natural populations in California. Amplified fragment length polymorphism molecular markers were analysed for all trees used in the trials. Lesion sizes varied quantitatively among individuals within populations, with up to an eightfold difference. There was little support for population differences in susceptibility. Molecular structure also showed a strong within-population, and weaker among-population, pattern of variation. Our data suggest that susceptibility of Q. agrifolia to P. ramorum is variable and is under the control of several gene loci. This variation exists within populations, so that less susceptible local genotypes may provide the gene pool for regeneration of woodlands where mortality is high.

Genetic diversity and structure of Lolium perenne ssp. multiflorum in California vineyards and orchards indicate potential for spread of herbicide resistance via gene flow.

PubMed

Karn, Elizabeth; Jasieniuk, Marie

2017-07-01

Management of agroecosystems with herbicides imposes strong selection pressures on weedy plants leading to the evolution of resistance against those herbicides. Resistance to glyphosate in populations of Lolium perenne L. ssp. multiflorum is increasingly common in California, USA, causing economic losses and the loss of effective management tools. To gain insights into the recent evolution of glyphosate resistance in L. perenne in perennial cropping systems of northwest California and to inform management, we investigated the frequency of glyphosate resistance and the genetic diversity and structure of 14 populations. The sampled populations contained frequencies of resistant plants ranging from 10% to 89%. Analyses of neutral genetic variation using microsatellite markers indicated very high genetic diversity within all populations regardless of resistance frequency. Genetic variation was distributed predominantly among individuals within populations rather than among populations or sampled counties, as would be expected for a wide-ranging outcrossing weed species. Bayesian clustering analysis provided evidence of population structuring with extensive admixture between two genetic clusters or gene pools. High genetic diversity and admixture, and low differentiation between populations, strongly suggest the potential for spread of resistance through gene flow and the need for management that limits seed and pollen dispersal in L. perenne .

From the ORFeome concept to highly comprehensive, full-genome screening libraries.

PubMed

Rid, Raphaela; Abdel-Hadi, Omar; Maier, Richard; Wagner, Martin; Hundsberger, Harald; Hintner, Helmut; Bauer, Johann; Onder, Kamil

2013-02-01

Recombination-based cloning techniques have in recent times facilitated the establishment of genome-scale single-gene ORFeome repositories. Their further handling and downstream application in systematic fashion is, however, practically impeded because of logistical plus economic challenges. At this juncture, simultaneously transferring entire gene collections in compiled pool format could represent an advanced compromise between systematic ORFeome (an organism's entire set of protein-encoding open reading frames) projects and traditional random library approaches, but has not yet been considered in great detail. In our endeavor to merge the comprehensiveness of ORFeomes with a basically simple, streamlined, and easily executable single-tube design, we have here produced five different pooled screening-ready libraries for both Staphylococcus aureus and Homo sapiens. By evaluating the parallel transfer efficiencies of differentially sized genes from initial polymerase chain reaction (PCR) product amplification to entry and final destination library construction via quantitative real-time PCR, we found that the complexity of the gene population is fairly stably maintained once an entry resource has been successfully established, and that no apparent size-selection bias loss of large inserts takes place. Recombinational transfer processes are hence robust enough for straightforwardly achieving such pooled screening libraries.

Microbial life in Champagne Pool, a geothermal spring in Waiotapu, New Zealand.

PubMed

Hetzer, Adrian; Morgan, Hugh W; McDonald, Ian R; Daughney, Christopher J

2007-07-01

Surveys of Champagne Pool, one of New Zealand's largest terrestrial hot springs and rich in arsenic ions and compounds, have been restricted to geological and geochemical descriptions, and a few microbiological studies applying culture-independent methods. In the current investigation, a combination of culture and culture-independent approaches were chosen to determine microbial density and diversity in Champagne Pool. Recovered total DNA and adenosine 5'-triphosphate (ATP) content of spring water revealed relatively low values compared to other geothermal springs within New Zealand and are in good agreement with low cell numbers of 5.6 +/- 0.5 x 10(6) cells/ml obtained for Champagne Pool water samples by 4',6-diamidino-2-phenylindole (DAPI) staining. Denaturing Gradient Gel Electrophoresis (DGGE) and 16S rRNA (small-subunit ribosomal nucleic acid) gene clone library analyses of environmental DNA indicated the abundance of Sulfurihydrogenibium, Sulfolobus, and Thermofilum-like populations in Champagne Pool. From these results, media were selected to target the enrichment of hydrogen-oxidizing and sulfur-dependent microorganisms. Three isolates were successfully obtained having 16S rRNA gene sequences with similarities of approximately 98% to Thermoanaerobacter tengcongensis, 94% to Sulfurihydrogenibium azorense, and 99% to Thermococcus waiotapuensis, respectively.

Pooled Enrichment Sequencing Identifies Diversity and Evolutionary Pressures at NLR Resistance Genes within a Wild Tomato Population.

PubMed

Stam, Remco; Scheikl, Daniela; Tellier, Aurélien

2016-06-02

Nod-like receptors (NLRs) are nucleotide-binding domain and leucine-rich repeats containing proteins that are important in plant resistance signaling. Many of the known pathogen resistance (R) genes in plants are NLRs and they can recognize pathogen molecules directly or indirectly. As such, divergence and copy number variants at these genes are found to be high between species. Within populations, positive and balancing selection are to be expected if plants coevolve with their pathogens. In order to understand the complexity of R-gene coevolution in wild nonmodel species, it is necessary to identify the full range of NLRs and infer their evolutionary history. Here we investigate and reveal polymorphism occurring at 220 NLR genes within one population of the partially selfing wild tomato species Solanum pennellii. We use a combination of enrichment sequencing and pooling ten individuals, to specifically sequence NLR genes in a resource and cost-effective manner. We focus on the effects which different mapping and single nucleotide polymorphism calling software and settings have on calling polymorphisms in customized pooled samples. Our results are accurately verified using Sanger sequencing of polymorphic gene fragments. Our results indicate that some NLRs, namely 13 out of 220, have maintained polymorphism within our S. pennellii population. These genes show a wide range of πN/πS ratios and differing site frequency spectra. We compare our observed rate of heterozygosity with expectations for this selfing and bottlenecked population. We conclude that our method enables us to pinpoint NLR genes which have experienced natural selection in their habitat. © The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

Gene coexpression measures in large heterogeneous samples using count statistics.

PubMed

Wang, Y X Rachel; Waterman, Michael S; Huang, Haiyan

2014-11-18

With the advent of high-throughput technologies making large-scale gene expression data readily available, developing appropriate computational tools to process these data and distill insights into systems biology has been an important part of the "big data" challenge. Gene coexpression is one of the earliest techniques developed that is still widely in use for functional annotation, pathway analysis, and, most importantly, the reconstruction of gene regulatory networks, based on gene expression data. However, most coexpression measures do not specifically account for local features in expression profiles. For example, it is very likely that the patterns of gene association may change or only exist in a subset of the samples, especially when the samples are pooled from a range of experiments. We propose two new gene coexpression statistics based on counting local patterns of gene expression ranks to take into account the potentially diverse nature of gene interactions. In particular, one of our statistics is designed for time-course data with local dependence structures, such as time series coupled over a subregion of the time domain. We provide asymptotic analysis of their distributions and power, and evaluate their performance against a wide range of existing coexpression measures on simulated and real data. Our new statistics are fast to compute, robust against outliers, and show comparable and often better general performance.

Structure and function of conjugative pili: monoclonal antibodies as probes for structural variants of F pili.

PubMed Central

Grossman, T H; Frost, L S; Silverman, P M

1990-01-01

The lac-tra operon fusion plasmid pTG801 contains the known F plasmid DNA transfer (tra) genes required by Escherichia coli to elaborate functional F pili (T. Grossman and P. M. Silverman, J. Bacteriol. 171:650-656, 1989). Here, we show that these pili are actually structural variants of normal F pili and that the F plasmid must contain additional genes that affect pilus structure and function. We confirmed a previous report that two monoclonal antibodies that recognize epitopes at and near the amino terminus of F pilin do not decorate the sides of normal F pili, as determined by immunogold electron microscopy. However, both antibodies laterally decorated pTG801 pili. The epitope for one of the antibodies has been shown to include the amino-terminal acetyl group of F pilin, which must therefore also be present on pTG801 pilin. Normal antibody staining was restored to pTG801 pili when cells contained, in addition to pTG801, the compatible plasmid pRS31, which must therefore include at least one gene affecting F-pilus structure. One candidate, traD, was excluded as the sole such gene, since traD+ derivatives of a pTG801 strain still elaborated pili that could be laterally decorated with antibody. Moreover, although traD alone restored RNA bacteriophage R17 infectivity to pTG801 cells, as expected, it did not mimic pRS31 in restoring to pTG801 pili other characteristics of normal F pili. We conclude that pRS31 contains as yet uncharacterized genes required for elaboration of structurally normal F pili. Finally, we identified vesicular material, especially abundant in cultures of pTG801 transformants, that stained heavily with the anti-F-pilin monoclonal antibodies. This material may reflect the inner membrane pool of F pilin. Images FIG. 3 FIG. 4 FIG. 5 PMID:1689713

Large woody debris input and its influence on channel structure in agricultural lands of Southeast Brazil.

PubMed

de Paula, Felipe Rossetti; Ferraz, Silvio Frosini de Barros; Gerhard, Pedro; Vettorazzi, Carlos Alberto; Ferreira, Anderson

2011-10-01

Riparian forests are important for the structure and functioning of stream ecosystems, providing structural components such as large woody debris (LWD). Changes in these forests will cause modifications in the LWD input to streams, affecting their structure. In order to assess the influence of riparian forests changes in LWD supply, 15 catchments (third and fourth order) with riparian forests at different conservation levels were selected for sampling. In each catchment we quantified the abundance, volume and diameter of LWD in stream channels; the number, area and volume of pools formed by LWD and basal area and tree diameter of riparian forest. We found that riparian forests were at a secondary successional stage with predominantly young trees (diameter at breast height <10 cm) in all studied streams. Results showed that basal area and diameter of riparian forest differed between the stream groups (forested and non-forested), but tree density did not differ between groups. Differences were also observed in LWD abundance, volume, frequency of LWD pools with subunits and area and volume of LWD pools. LWD diameter, LWD that form pools diameter and frequency of LWD pools without subunits did not differ between stream groups. Regression analyses showed that LWD abundance and volume, and frequency of LWD pools (with and without subunits) were positively related with the proportion of riparian forest. LWD diameter was not correlated to riparian tree diameter. The frequency of LWD pools was correlated to the abundance and volume of LWD, but characteristics of these pools (area and volume) were not correlated to the diameter of LWD that formed the pools. These results show that alterations in riparian forest cause modifications in the LWD abundance and volume in the stream channel, affecting mainly the structural complexity of these ecosystems (reduction in the number and structural characteristics of LWD pools). Our results also demonstrate that riparian forest conservation actions must consider not only its extension, but also successional stage to guarantee the quantity and quality of LWD necessary to enable the structuring of stream channels.

Population structure and genetic diversity in a commercial maize breeding program assessed with SSR and SNP markers.

PubMed

Van Inghelandt, Delphine; Melchinger, Albrecht E; Lebreton, Claude; Stich, Benjamin

2010-05-01

Information about the genetic diversity and population structure in elite breeding material is of fundamental importance for the improvement of crops. The objectives of our study were to (a) examine the population structure and the genetic diversity in elite maize germplasm based on simple sequence repeat (SSR) markers, (b) compare these results with those obtained from single nucleotide polymorphism (SNP) markers, and (c) compare the coancestry coefficient calculated from pedigree records with genetic distance estimates calculated from SSR and SNP markers. Our study was based on 1,537 elite maize inbred lines genotyped with 359 SSR and 8,244 SNP markers. The average number of alleles per locus, of group specific alleles, and the gene diversity (D) were higher for SSRs than for SNPs. Modified Roger's distance (MRD) estimates and membership probabilities of the STRUCTURE matrices were higher for SSR than for SNP markers but the germplasm organization in four heterotic pools was consistent with STRUCTURE results based on SSRs and SNPs. MRD estimates calculated for the two marker systems were highly correlated (0.87). Our results suggested that the same conclusions regarding the structure and the diversity of heterotic pools could be drawn from both markers types. Furthermore, although our results suggested that the ratio of the number of SSRs and SNPs required to obtain MRD or D estimates with similar precision is not constant across the various precision levels, we propose that between 7 and 11 times more SNPs than SSRs should be used for analyzing population structure and genetic diversity.

Final Technical Report to DOE for the Award DE-SC0004601

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

Zhou, Jizhong

Understanding the responses, adaptations and feedback mechanisms of biological communities to climate change is critical to project future state of earth and climate systems. Although significant amount of knowledge is available on the feedback responses of aboveground communities to climate change, little is known about the responses of belowground microbial communities due to the challenges in analyzing soil microbial community structure. Thus the goal overall goal of this study is to provide system-level, predictive mechanistic understanding of the temperature sensitivity of soil carbon (C) decomposition to climate warming by using cutting-edge integrated metagenomic technologies. Towards this goal, the following fourmore » objectives will be pursued: (i) To determine phylogenetic composition and metabolic diversity of microbial communities in the temperate grassland and tundra ecosystems; (ii) To delineate the responses of microbial community structure, functions and activities to climate change in the temperate grassland and tundra ecosystems; (iii) To determine the temperature sensitivity of microbial respiration in soils with different mixtures of labile versus recalcitrant C, and the underlying microbiological basis for temperature sensitivity of these pools; and (iv) To synthesize all experimental data for revealing microbial control of ecosystem carbon processes in responses to climate change. We have achieved our goals for all four proposed objectives. First, we determined the phylogenetic composition and metabolic diversity of microbial communities in the temperate grassland and tundra ecosystems. For this objective, we have developed a novel phasing amplicon sequencing (PAS) approach for MiSeq sequencing of amplicons. This approach has been used for sequencing various phylogenetic and functional genes related to ecosystem functioning. A comprehensive functional gene array (e.g., GeoChip 5.0) has also been developed and used for soil microbial community analysis in this study. In addition, shot-gun metagenome sequencing along with the above approaches have been used to understand the phylogenetic and functional diversity, composition, and structure of soil microbial communities in both temperature grassland and tundra ecosystems. Second, we determined the response of soil microbial communities to climate warming in both temperate grassland and tundra ecosystems using various methods. Our major findings are: (i) Microorganisms are very rapid to respond to climate warming in the tundra ecosystem, AK, which is vulnerable, too. (ii) Climate warming also significantly shifted the metabolic diversity, composition and structure of microbial communities, and key metabolic pathways related to carbon turnover, such as cellulose degradation (~13%) and CO2 production (~10%), and to nitrogen cycling, including denitrification (~12%) were enriched by warming. (iii) Warming also altered the expression patterns of microbial functional genes important to ecosystem functioning and stability through GeoChip and metatranscriptomic analysis of soil microbial communities at the OK site. Third, we analyzed temperature sensitivity of C decomposition to climate warming for both AK and OK soils through laboratory incubations. Key results include: (i) Alaska tundra soils showed that after one year of incubation, CT in the top 15 cm could be as high as 25% and 15% of the initial soil C content at 25°C and 15°C incubations, respectively. (ii) analysis of 456 incubated soil samples with 16S rRNA gene, ITS and GeoChip hybridization showed that warming shifted the phylogenretic and functional diversity, composition, structure and metabolic potential of soil microbial communities, and at different stages of incubation, key populations and functional genes significantly changed along with soil substrate changes. Functional gene diversity and functional genes for degrading labile C components decrease along incubation when labile C components are exhausting, but the genes related to degrading recalcitrant C increase. These molecular data will be directly used for modeling. Fourth, we have developed novel approaches to integrate and model experimental data to understand microbial control of ecosystem C processes in response to climate change. We compared different methods to calculate Q10 for estimating temperature sensitivity, and new approaches for Q10 calculation and molecular ecological network analysis were also developed. Using those newly developed approaches, our result indicated that Q10s increased with the recalcitrance of C pools, suggesting that longer incubation studies are needed in order to assess the temperature sensitivity of slower C pools, especially at low temperature regimes. This project has been very productive, resulting in 42 papers published or in press, 4 submitted, and 13 in preparation.« less

Mining a database of single amplified genomes from Red Sea brine pool extremophiles—improving reliability of gene function prediction using a profile and pattern matching algorithm (PPMA)

PubMed Central

Grötzinger, Stefan W.; Alam, Intikhab; Ba Alawi, Wail; Bajic, Vladimir B.; Stingl, Ulrich; Eppinger, Jörg

2014-01-01

Reliable functional annotation of genomic data is the key-step in the discovery of novel enzymes. Intrinsic sequencing data quality problems of single amplified genomes (SAGs) and poor homology of novel extremophile's genomes pose significant challenges for the attribution of functions to the coding sequences identified. The anoxic deep-sea brine pools of the Red Sea are a promising source of novel enzymes with unique evolutionary adaptation. Sequencing data from Red Sea brine pool cultures and SAGs are annotated and stored in the Integrated Data Warehouse of Microbial Genomes (INDIGO) data warehouse. Low sequence homology of annotated genes (no similarity for 35% of these genes) may translate into false positives when searching for specific functions. The Profile and Pattern Matching (PPM) strategy described here was developed to eliminate false positive annotations of enzyme function before progressing to labor-intensive hyper-saline gene expression and characterization. It utilizes InterPro-derived Gene Ontology (GO)-terms (which represent enzyme function profiles) and annotated relevant PROSITE IDs (which are linked to an amino acid consensus pattern). The PPM algorithm was tested on 15 protein families, which were selected based on scientific and commercial potential. An initial list of 2577 enzyme commission (E.C.) numbers was translated into 171 GO-terms and 49 consensus patterns. A subset of INDIGO-sequences consisting of 58 SAGs from six different taxons of bacteria and archaea were selected from six different brine pool environments. Those SAGs code for 74,516 genes, which were independently scanned for the GO-terms (profile filter) and PROSITE IDs (pattern filter). Following stringent reliability filtering, the non-redundant hits (106 profile hits and 147 pattern hits) are classified as reliable, if at least two relevant descriptors (GO-terms and/or consensus patterns) are present. Scripts for annotation, as well as for the PPM algorithm, are available through the INDIGO website. PMID:24778629

Construction of CRISPR Libraries for Functional Screening.

PubMed

Carstens, Carsten P; Felts, Katherine A; Johns, Sarah E

2018-01-01

Identification of gene function has been aided by the ability to generate targeted gene knockouts or transcriptional repression using the CRISPR/CAS9 system. Using pooled libraries of guide RNA expression vectors that direct CAS9 to a specific genomic site allows identification of genes that are either enriched or depleted in response to a selection scheme, thus linking the affected gene to the chosen phenotype. The quality of the data generated by the screening is dependent on the quality of the guide RNA delivery library with regards to error rates and especially evenness of distribution of the guides. Here, we describe a method for constructing complex plasmid libraries based on pooled designed oligomers with high representation and tight distributions. The procedure allows construction of plasmid libraries of >60,000 members with a 95th/5th percentile ratio of less than 3.5.

AST: an automated sequence-sampling method for improving the taxonomic diversity of gene phylogenetic trees.

PubMed

Zhou, Chan; Mao, Fenglou; Yin, Yanbin; Huang, Jinling; Gogarten, Johann Peter; Xu, Ying

2014-01-01

A challenge in phylogenetic inference of gene trees is how to properly sample a large pool of homologous sequences to derive a good representative subset of sequences. Such a need arises in various applications, e.g. when (1) accuracy-oriented phylogenetic reconstruction methods may not be able to deal with a large pool of sequences due to their high demand in computing resources; (2) applications analyzing a collection of gene trees may prefer to use trees with fewer operational taxonomic units (OTUs), for instance for the detection of horizontal gene transfer events by identifying phylogenetic conflicts; and (3) the pool of available sequences is biased towards extensively studied species. In the past, the creation of subsamples often relied on manual selection. Here we present an Automated sequence-Sampling method for improving the Taxonomic diversity of gene phylogenetic trees, AST, to obtain representative sequences that maximize the taxonomic diversity of the sampled sequences. To demonstrate the effectiveness of AST, we have tested it to solve four problems, namely, inference of the evolutionary histories of the small ribosomal subunit protein S5 of E. coli, 16 S ribosomal RNAs and glycosyl-transferase gene family 8, and a study of ancient horizontal gene transfers from bacteria to plants. Our results show that the resolution of our computational results is almost as good as that of manual inference by domain experts, hence making the tool generally useful to phylogenetic studies by non-phylogeny specialists. The program is available at http://csbl.bmb.uga.edu/~zhouchan/AST.php.

AST: An Automated Sequence-Sampling Method for Improving the Taxonomic Diversity of Gene Phylogenetic Trees

PubMed Central

Zhou, Chan; Mao, Fenglou; Yin, Yanbin; Huang, Jinling; Gogarten, Johann Peter; Xu, Ying

2014-01-01

A challenge in phylogenetic inference of gene trees is how to properly sample a large pool of homologous sequences to derive a good representative subset of sequences. Such a need arises in various applications, e.g. when (1) accuracy-oriented phylogenetic reconstruction methods may not be able to deal with a large pool of sequences due to their high demand in computing resources; (2) applications analyzing a collection of gene trees may prefer to use trees with fewer operational taxonomic units (OTUs), for instance for the detection of horizontal gene transfer events by identifying phylogenetic conflicts; and (3) the pool of available sequences is biased towards extensively studied species. In the past, the creation of subsamples often relied on manual selection. Here we present an Automated sequence-Sampling method for improving the Taxonomic diversity of gene phylogenetic trees, AST, to obtain representative sequences that maximize the taxonomic diversity of the sampled sequences. To demonstrate the effectiveness of AST, we have tested it to solve four problems, namely, inference of the evolutionary histories of the small ribosomal subunit protein S5 of E. coli, 16 S ribosomal RNAs and glycosyl-transferase gene family 8, and a study of ancient horizontal gene transfers from bacteria to plants. Our results show that the resolution of our computational results is almost as good as that of manual inference by domain experts, hence making the tool generally useful to phylogenetic studies by non-phylogeny specialists. The program is available at http://csbl.bmb.uga.edu/~zhouchan/AST.php. PMID:24892935

Domestication to Crop Improvement: Genetic Resources for Sorghum and Saccharum (Andropogoneae)

PubMed Central

Dillon, Sally L.; Shapter, Frances M.; Henry, Robert J.; Cordeiro, Giovanni; Izquierdo, Liz; Lee, L. Slade

2007-01-01

Background Both sorghum (Sorghum bicolor) and sugarcane (Saccharum officinarum) are members of the Andropogoneae tribe in the Poaceae and are each other's closest relatives amongst cultivated plants. Both are relatively recent domesticates and comparatively little of the genetic potential of these taxa and their wild relatives has been captured by breeding programmes to date. This review assesses the genetic gains made by plant breeders since domestication and the progress in the characterization of genetic resources and their utilization in crop improvement for these two related species. Genetic Resources The genome of sorghum has recently been sequenced providing a great boost to our knowledge of the evolution of grass genomes and the wealth of diversity within S. bicolor taxa. Molecular analysis of the Sorghum genus has identified close relatives of S. bicolor with novel traits, endosperm structure and composition that may be used to expand the cultivated gene pool. Mutant populations (including TILLING populations) provide a useful addition to genetic resources for this species. Sugarcane is a complex polyploid with a large and variable number of copies of each gene. The wild relatives of sugarcane represent a reservoir of genetic diversity for use in sugarcane improvement. Techniques for quantitative molecular analysis of gene or allele copy number in this genetically complex crop have been developed. SNP discovery and mapping in sugarcane has been advanced by the development of high-throughput techniques for ecoTILLING in sugarcane. Genetic linkage maps of the sugarcane genome are being improved for use in breeding selection. The improvement of both sorghum and sugarcane will be accelerated by the incorporation of more diverse germplasm into the domesticated gene pools using molecular tools and the improved knowledge of these genomes. PMID:17766842

Capturing neutral and adaptive genetic diversity for conservation in a highly structured tree species.

PubMed

Rodríguez-Quilón, Isabel; Santos-Del-Blanco, Luis; Serra-Varela, María Jesús; Koskela, Jarkko; González-Martínez, Santiago C; Alía, Ricardo

2016-10-01

Preserving intraspecific genetic diversity is essential for long-term forest sustainability in a climate change scenario. Despite that, genetic information is largely neglected in conservation planning, and how conservation units should be defined is still heatedly debated. Here, we use maritime pine (Pinus pinaster Ait.), an outcrossing long-lived tree with a highly fragmented distribution in the Mediterranean biodiversity hotspot, to prove the importance of accounting for genetic variation, of both neutral molecular markers and quantitative traits, to define useful conservation units. Six gene pools associated to distinct evolutionary histories were identified within the species using 12 microsatellites and 266 single nucleotide polymorphisms (SNPs). In addition, height and survival standing variation, their genetic control, and plasticity were assessed in a multisite clonal common garden experiment (16 544 trees). We found high levels of quantitative genetic differentiation within previously defined neutral gene pools. Subsequent cluster analysis and post hoc trait distribution comparisons allowed us to define 10 genetically homogeneous population groups with high evolutionary potential. They constitute the minimum number of units to be represented in a maritime pine dynamic conservation program. Our results uphold that the identification of conservation units below the species level should account for key neutral and adaptive components of genetic diversity, especially in species with strong population structure and complex evolutionary histories. The environmental zonation approach currently used by the pan-European genetic conservation strategy for forest trees would be largely improved by gradually integrating molecular and quantitative trait information, as data become available. © 2016 by the Ecological Society of America.

The congruence between matrilineal genetic (mtDNA) and geographic diversity of Iranians and the territorial populations

PubMed Central

Bahmanimehr, Ardeshir; Eskandari, Ghafar; Nikmanesh, Fatemeh

2015-01-01

Objective(s): From the ancient era, emergence of Agriculture in the connecting region of Mesopotamia and the Iranian plateau at the foothills of the Zagros Mountains, made Iranian gene pool as an important source of populating the region. It has differentiated the population spread and different language groups. In order to trace the maternal genetic affinity between Iranians and other populations of the area and to establish the place of Iranians in a broad framework of ethnically and linguistically diverse groups of Middle Eastern and South Asian populations, a comparative study of territorial groups was designed and used in the population statistical analysis. Materials and Methods: Mix of 616 samples was sequenced for complete mtDNA or hyper variable regions in this study. A published dataset of neighboring populations was used as a comparison in the Iranian matrilineal lineage study based on mtDNA haplogroups. Results: Statistical analyses data, demonstrate a close genetic structure of all Iranian populations, thus suggesting their origin from a common maternal ancestral gene pool and show that the diverse maternal genetic structure does not reflect population differentiation in the region in their language. Conclusion: In the aggregate of the eastward spreads of proto-Elamo-Dravidian language from the Southwest region of Iran, the Elam province, a reasonable degree of homogeneity has been observed among Iranians in this study. The approach will facilitate our perception of the more detailed relationship of the ethnic groups living in Iran with the other ancient peoples of the area, testing linguistic hypothesis and population movements. PMID:25810873

Novel Two-Step Hierarchical Screening of Mutant Pools Reveals Mutants under Selection in Chicks

PubMed Central

Yang, Hee-Jeong; Bogomolnaya, Lydia M.; Elfenbein, Johanna R.; Endicott-Yazdani, Tiana; Reynolds, M. Megan; Porwollik, Steffen; Cheng, Pui; Xia, Xiao-Qin

2016-01-01

Contaminated chicken/egg products are major sources of human salmonellosis, yet the strategies used by Salmonella to colonize chickens are poorly understood. We applied a novel two-step hierarchical procedure to identify new genes important for colonization and persistence of Salmonella enterica serotype Typhimurium in chickens. A library of 182 S. Typhimurium mutants each containing a targeted deletion of a group of contiguous genes (for a total of 2,069 genes deleted) was used to identify regions under selection at 1, 3, and 9 days postinfection in chicks. Mutants in 11 regions were under selection at all assayed times (colonization mutants), and mutants in 15 regions were under selection only at day 9 (persistence mutants). We assembled a pool of 92 mutants, each deleted for a single gene, representing nearly all genes in nine regions under selection. Twelve single gene deletion mutants were under selection in this assay, and we confirmed 6 of 9 of these candidate mutants via competitive infections and complementation analysis in chicks. STM0580, STM1295, STM1297, STM3612, STM3615, and STM3734 are needed for Salmonella to colonize and persist in chicks and were not previously associated with this ability. One of these key genes, STM1297 (selD), is required for anaerobic growth and supports the ability to utilize formate under these conditions, suggesting that metabolism of formate is important during infection. We report a hierarchical screening strategy to interrogate large portions of the genome during infection of animals using pools of mutants of low complexity. Using this strategy, we identified six genes not previously known to be needed during infection in chicks, and one of these (STM1297) suggests an important role for formate metabolism during infection. PMID:26857572

Eco-engineered rock pools: a concrete solution to biodiversity loss and urban sprawl in the marine environment

NASA Astrophysics Data System (ADS)

Firth, Louise B.; Browne, Keith A.; Knights, Antony M.; Hawkins, Stephen J.; Nash, Róisín

2016-09-01

In coastal habitats artificial structures typically support lower biodiversity and can support greater numbers of non-native and opportunistic species than natural rocky reefs. Eco-engineering experiments are typically trialed to succeed; but arguably as much is learnt from failure than from success. Our goal was to trial a generic, cost effective, eco-engineering technique that could be incorporated into rock armouring anywhere in the world. Artificial rock pools were created from manipulated concrete between boulders on the exposed and sheltered sides of a causeway. Experimental treatments were installed in locations where they were expected to fail and compared to controls installed in locations in which they were expected to succeed. Control pools were created lower on the structure where they were immersed on every tidal cycle; experimental pools were created above mean high water spring tide which were only immersed on spring tides. We hypothesised that lower and exposed pools would support significantly higher taxon and functional diversity than upper and sheltered pools. The concrete pools survived the severe winter storms of 2013/14. After 12 months, non-destructive sampling revealed significantly higher mean taxon and functional richness in lower pools than upper pools on the exposed side only. After 24 months the sheltered pools had become inundated with sediments, thus failing to function as rock pools as intended. Destructive sampling on the exposed side revealed significantly higher mean functional richness in lower than upper pools. However, a surprisingly high number of taxa colonised the upper pools leading to no significant difference in mean taxon richness among shore heights. A high number of rare taxa in the lower pools led to total taxon richness being almost twice that of upper pools. These findings highlight that even when expected to fail concrete pools supported diverse assemblages, thus representing an affordable, replicable means of enhancing biodiversity on a variety of artificial structures.

Gene Flow Among Different Teosinte Taxa and Into the Domesticated Maize Gene Pool

USDA-ARS?s Scientific Manuscript database

Maize (Zea mays ssp. mays) was domesticated from one wild species ancestor, the Balsas teosinte (Zea mays ssp. parviglumis) about 9000 years ago. Higher levels of gene diversity are found in teosinte taxa compared to maize following domestication and selection bottlenecks. Diversity in maize can b...

Geomorphic and hydraulic Analyses of In-stream Step-pool Structures (I)

NASA Astrophysics Data System (ADS)

Kuo, W. C.; Hu, Y. L.; Wang, H. W.

2016-12-01

Longitudinal stair-like structures, such as alternating steps and pools, are found commonly in steep mountain streams. In a way to mimic the natural characteristics, many implementations of constructing artificial step structures have long been found in field practice to stabilize streambeds and enhance aquatic systems. To better understand how constructed step-pool systems form and function, this paper discusses the hydraulic and geomorphic factors based on flume experiments, and further compare to our field observations in Chijiawan Creek in Taiwan. We constructed a 2.9-m-long, 0.15-wide, and 0.3-m-high acrylic walled recirculating channel and conducted experiments to understand the formation, hydraulic features, and channel stability of step-pools of scenarios considering different channel slopes, discharges, feeding sediments. The results indicated that the keystones played a crucial role in stabilizing step-pool structures. The grain sizes of keystones from the experiments ranged approximately from one-third to one-tenth of channel width, while those from the field observations were about one-ninth to two-ninths. While the experimental discharge increased from 0.0012 cms to 0.006 cms, the flow transformed from nappe flow to skimming flow and the difference of average velocity between steps and pools reduced 30%. Besides, experiments showed that the step-pool structures failed immediately after keystones destroyed at a sediment transportation rate about 1.5 times of sediment feeding rate. It highlights the step-pools and channel stability is highly related to keystones. We further found the step-pools were buried at the experimental cases with coarse sediment fed upstream, similar to our field observations in Chijiawan Creek, with an approximately 1 3m deposition after Typhoon Soudelor, a 5-year event. The results obtained in this study would serve as a basis for ongoing discussions on how constructed step-pool structures would function and fail. More efforts of field investigations, flume experiments, and field experiments in helping developing specific recommendations and providing scientific insights for not only in Taiwan but around the world are still in need.

A Transcriptomic Analysis of Cave, Surface, and Hybrid Isopod Crustaceans of the Species Asellus aquaticus

PubMed Central

Stahl, Bethany A.; Gross, Joshua B.; Speiser, Daniel I.; Oakley, Todd H.; Patel, Nipam H.; Gould, Douglas B.; Protas, Meredith E.

2015-01-01

Cave animals, compared to surface-dwelling relatives, tend to have reduced eyes and pigment, longer appendages, and enhanced mechanosensory structures. Pressing questions include how certain cave-related traits are gained and lost, and if they originate through the same or different genetic programs in independent lineages. An excellent system for exploring these questions is the isopod, Asellus aquaticus. This species includes multiple cave and surface populations that have numerous morphological differences between them. A key feature is that hybrids between cave and surface individuals are viable, which enables genetic crosses and linkage analyses. Here, we advance this system by analyzing single animal transcriptomes of Asellus aquaticus. We use high throughput sequencing of non-normalized cDNA derived from the head of a surface-dwelling male, the head of a cave-dwelling male, the head of a hybrid male (produced by crossing a surface individual with a cave individual), and a pooled sample of surface embryos and hatchlings. Assembling reads from surface and cave head RNA pools yielded an integrated transcriptome comprised of 23,984 contigs. Using this integrated assembly as a reference transcriptome, we aligned reads from surface-, cave- and hybrid- head tissue and pooled surface embryos and hatchlings. Our approach identified 742 SNPs and placed four new candidate genes to an existing linkage map for A. aquaticus. In addition, we examined SNPs for allele-specific expression differences in the hybrid individual. All of these resources will facilitate identification of genes and associated changes responsible for cave adaptation in A. aquaticus and, in concert with analyses of other species, will inform our understanding of the evolutionary processes accompanying adaptation to the subterranean environment. PMID:26462237

Hypoxia promotes proliferation of human myogenic satellite cells: a potential benefactor in tissue engineering of skeletal muscle.

PubMed

Koning, Merel; Werker, Paul M N; van Luyn, Marja J A; Harmsen, Martin C

2011-07-01

Facial paralysis is a physically, psychologically, and socially disabling condition. Innovative treatment strategies based on regenerative medicine, in particular tissue engineering of skeletal muscle, are promising for treatment of patients with facial paralysis. The natural source for tissue-engineered muscle would be muscle stem cells, that is, human satellite cells (SC). In vivo, SC respond to hypoxic, ischemic muscle damage by activation, proliferation, differentiation to myotubes, and maturation to muscle fibers, while maintaining their reserve pool of SC. Therefore, our hypothesis is that hypoxia improves proliferation and differentiation of SC. During tissue engineering, a three-dimensional construct, or implanting SC in vivo, SC will encounter hypoxic environments. Thus, we set out to test our hypothesis on SC in vitro. During the first five passages, hypoxically cultured SC proliferated faster than their counterparts under normoxia. Moreover, also at higher passages, a switch from normoxia to hypoxia enhanced proliferation of SC. Hypoxia did not affect the expression of SC markers desmin and NCAM. However, the average surface expression per cell of NCAM was downregulated by hypoxia, and it also downregulated the gene expression of NCAM. The gene expression of the myogenic transcription factors PAX7, MYF5, and MYOD was upregulated by hypoxia. Moreover, gene expression of structural proteins α-sarcomeric actin, and myosins MYL1 and MYL3 was upregulated by hypoxia during differentiation. This indicates that hypoxia promotes a promyogenic shift in SC. Finally, Pax7 expression was not influenced by hypoxia and maintained in a subset of mononucleated cells, whereas these cells were devoid of structural muscle proteins. This suggests that during myogenesis in vitro, at least part of the SC adopt a quiescent, that is, reserve cells, phenotype. In conclusion, tissue engineering under hypoxic conditions would seem favorable in terms of myogenic proliferation, while maintaining the quiescent SC pool.

Ulk4 Regulates Neural Stem Cell Pool.

PubMed

Liu, Min; Guan, Zhenlong; Shen, Qin; Flinter, Frances; Domínguez, Laura; Ahn, Joo Wook; Collier, David A; O'Brien, Timothy; Shen, Sanbing

2016-09-01

The size of neural stem cell (NSC) pool at birth determines the starting point of adult neurogenesis. Aberrant neurogenesis is associated with major mental illness, in which ULK4 is proposed as a rare risk factor. Little is known about factors regulating the NSC pool, or function of the ULK4. Here, we showed that Ulk4(tm1a/tm1a) mice displayed a dramatically reduced NSC pool at birth. Ulk4 was expressed in a cell cycle-dependent manner and peaked in G2/M phases. Targeted disruption of the Ulk4 perturbed mid-neurogenesis and significantly reduced cerebral cortex in postnatal mice. Pathway analyses of dysregulated genes in Ulk4(tm1a/tm1a) mice revealed Ulk4 as a key regulator of cell cycle and NSC proliferation, partially through regulation of the Wnt signaling. In addition, we identified hemizygous deletion of ULK4 gene in 1.2/1,000 patients with pleiotropic symptoms including severe language delay and learning difficulties. ULK4, therefore, may significantly contribute to neurodevelopmental, neuropsychiatric, and neurodegenerative disorders. Stem Cells 2016;34:2318-2331. © 2016 AlphaMed Press.

Two endornaviruses show differential infection patterns between gene pools of Phaseolus vulgaris.

PubMed

Khankhum, Surasak; Valverde, Rodrigo A; Pastor-Corrales, Marcial A; Osorno, Juan M; Sabanadzovic, Sead

2015-04-01

We investigated the occurrence of two plant endornaviruses, Phaseolus vulgaris endornavirus 1 and Phaseolus vulgaris endornavirus 2, in breeding lines, cultivars, landraces, and wild genotypes of common bean (Phaseolus vulgaris) collected from the two centers of common bean domestication: Mesoamerica and the Andes. The two endornaviruses were detected in many genotypes of Mesoamerican origin but rarely in genotypes of Andean origin. The results suggest that these two endornaviruses were introduced into the Mesoamerican modern genotypes during common bean domestication and provide more evidence for the existence of two divergent gene pools of common bean.

Tetraploid Wheat Landraces in the Mediterranean Basin: Taxonomy, Evolution and Genetic Diversity

PubMed Central

Oliveira, Hugo R.; Campana, Michael G.; Jones, Huw; Hunt, Harriet V.; Leigh, Fiona; Redhouse, David I.; Lister, Diane L.; Jones, Martin K.

2012-01-01

The geographic distribution of genetic diversity and the population structure of tetraploid wheat landraces in the Mediterranean basin has received relatively little attention. This is complicated by the lack of consensus concerning the taxonomy of tetraploid wheats and by unresolved questions regarding the domestication and spread of naked wheats. These knowledge gaps hinder crop diversity conservation efforts and plant breeding programmes. We investigated genetic diversity and population structure in tetraploid wheats (wild emmer, emmer, rivet and durum) using nuclear and chloroplast simple sequence repeats, functional variations and insertion site-based polymorphisms. Emmer and wild emmer constitute a genetically distinct population from durum and rivet, the latter seeming to share a common gene pool. Our population structure and genetic diversity data suggest a dynamic history of introduction and extinction of genotypes in the Mediterranean fields. PMID:22615891

Investigating energy-based pool structure selection in the structure ensemble modeling with experimental distance constraints: The example from a multidomain protein Pub1.

PubMed

Zhu, Guanhua; Liu, Wei; Bao, Chenglong; Tong, Dudu; Ji, Hui; Shen, Zuowei; Yang, Daiwen; Lu, Lanyuan

2018-05-01

The structural variations of multidomain proteins with flexible parts mediate many biological processes, and a structure ensemble can be determined by selecting a weighted combination of representative structures from a simulated structure pool, producing the best fit to experimental constraints such as interatomic distance. In this study, a hybrid structure-based and physics-based atomistic force field with an efficient sampling strategy is adopted to simulate a model di-domain protein against experimental paramagnetic relaxation enhancement (PRE) data that correspond to distance constraints. The molecular dynamics simulations produce a wide range of conformations depicted on a protein energy landscape. Subsequently, a conformational ensemble recovered with low-energy structures and the minimum-size restraint is identified in good agreement with experimental PRE rates, and the result is also supported by chemical shift perturbations and small-angle X-ray scattering data. It is illustrated that the regularizations of energy and ensemble-size prevent an arbitrary interpretation of protein conformations. Moreover, energy is found to serve as a critical control to refine the structure pool and prevent data overfitting, because the absence of energy regularization exposes ensemble construction to the noise from high-energy structures and causes a more ambiguous representation of protein conformations. Finally, we perform structure-ensemble optimizations with a topology-based structure pool, to enhance the understanding on the ensemble results from different sources of pool candidates. © 2018 Wiley Periodicals, Inc.

Cellular zinc fluxes and the regulation of apoptosis/gene-directed cell death.

PubMed

Truong-Tran, A Q; Ho, L H; Chai, F; Zalewski, P D

2000-05-01

The maintenance of discrete subcellular pools of zinc (Zn) is critical for the functional and structural integrity of cells. Among the important biological processes influenced by Zn is apoptosis, a process that is important in cellular homeostasis (an important cellular homeostatic process). It has also been identified as a major mechanism contributing to cell death in response to toxins and in disease, offering hope that novel therapies that target apoptotic pathways may be developed. Because Zn levels in the body can be increased in a relatively nontoxic manner, it may be possible to prevent or ameliorate degenerative disorders that are associated with high rates of apoptotic cell death. This review begins with brief introductions that address, first, the cellular biology of Zn, especially the critical labile Zn pools, and, second, the phenomenon of apoptosis. We then review the evidence relating Zn to apoptosis and address three major hypotheses: (1) that a specific pool or pools of intracellular labile Zn regulates apoptosis; (2) that systemic changes in Zn levels in the body, due to dietary factors, altered physiological states or disease, can influence cell susceptibility to apoptosis, and (3) that this altered susceptibility to apoptosis contributes to pathophysiological changes in the body. Other key issues are the identity of the molecular targets of Zn in the apoptotic cascade, the types of cells and tissues most susceptible to Zn-regulated apoptosis, the role of Zn as a coordinate regulator of mitosis and apoptosis and the apparent release of tightly bound intracellular pools of Zn during the later stages of apoptosis. This review concludes with a section highlighting areas of priority for future studies.

Revealing the transcriptomic complexity of switchgrass by PacBio long-read sequencing.

PubMed

Zuo, Chunman; Blow, Matthew; Sreedasyam, Avinash; Kuo, Rita C; Ramamoorthy, Govindarajan Kunde; Torres-Jerez, Ivone; Li, Guifen; Wang, Mei; Dilworth, David; Barry, Kerrie; Udvardi, Michael; Schmutz, Jeremy; Tang, Yuhong; Xu, Ying

2018-01-01

Switchgrass ( Panicum virgatum L.) is an important bioenergy crop widely used for lignocellulosic research. While extensive transcriptomic analyses have been conducted on this species using short read-based sequencing techniques, very little has been reliably derived regarding alternatively spliced (AS) transcripts. We present an analysis of transcriptomes of six switchgrass tissue types pooled together, sequenced using Pacific Biosciences (PacBio) single-molecular long-read technology. Our analysis identified 105,419 unique transcripts covering 43,570 known genes and 8795 previously unknown genes. 45,168 are novel transcripts of known genes. A total of 60,096 AS transcripts are identified, 45,628 being novel. We have also predicted 1549 transcripts of genes involved in cell wall construction and remodeling, 639 being novel transcripts of known cell wall genes. Most of the predicted transcripts are validated against Illumina-based short reads. Specifically, 96% of the splice junction sites in all the unique transcripts are validated by at least five Illumina reads. Comparisons between genes derived from our identified transcripts and the current genome annotation revealed that among the gene set predicted by both analyses, 16,640 have different exon-intron structures. Overall, substantial amount of new information is derived from the PacBio RNA data regarding both the transcriptome and the genome of switchgrass.

Identification of mutations, genotype-phenotype correlation and prenatal diagnosis of maple syrup urine disease in Indian patients.

PubMed

Gupta, Deepti; Bijarnia-Mahay, Sunita; Saxena, Renu; Kohli, Sudha; Dua-Puri, Ratna; Verma, Jyotsna; Thomas, E; Shigematsu, Yosuke; Yamaguchi, Seiji; Deb, Roumi; Verma, Ishwar Chander

2015-09-01

Maple syrup urine disease (MSUD) is caused by mutations in genes BCKDHA, BCKDHB, DBT encoding E1α, E1β, and E2 subunits of enzyme complex, branched-chain alpha-ketoacid dehydrogenase (BCKDH). BCKDH participates in catabolism of branched-chain amino acids (BCAAs) - leucine, isoleucine and valine in the energy production pathway. Deficiency or defect in the enzyme complex causes accumulation of BCAAs and keto-acids leading to toxicity. Twenty-four patients with MSUD were enrolled in the study for molecular characterization and genotype-phenotype correlation. Molecular studies were carried out by sequencing of the 3 genes by Sanger method. Bioinformatics tools were employed to classify novel variations into pathogenic or benign. The predicted effects of novel changes on protein structure were elucidated by 3D modeling. Mutations were detected in 22 of 24 patients (11, 7 and 4 in BCKDHB, BCKDHA and DBT genes, respectively). Twenty mutations including 11 novel mutations were identified. Protein modeling in novel mutations showed alteration of structure and function of these subunits. Mutations, c.1065 delT (BCKDHB gene) and c.939G > C (DBT gene) were noted to be recurrent, identified in 6 of 22 alleles and 5 of 8 alleles, respectively. Two-third patients were of neonatal classical phenotype (16 of 24). BCKDHB gene mutations were present in 10 of these 16 patients. Prenatal diagnoses were performed in 4 families. Consanguinity was noted in 37.5% families. Although no obvious genotype-phenotype correlation could be found in our study, most cases with mutation in BCKDHB gene presented in neonatal period. Large number of novel mutations underlines the heterogeneity and distinctness of gene pool from India. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

Three sympatric clusters of the malaria vector Anopheles culicifacies E (Diptera: Culicidae) detected in Sri Lanka.

PubMed

Harischandra, Iresha Nilmini; Dassanayake, Ranil Samantha; De Silva, Bambaranda Gammacharige Don Nissanka Kolitha

2016-01-04

The disease re-emergence threat from the major malaria vector in Sri Lanka, Anopheles culicifacies, is currently increasing. To predict malaria vector dynamics, knowledge of population genetics and gene flow is required, but this information is unavailable for Sri Lanka. This study was carried out to determine the population structure of An. culicifacies E in Sri Lanka. Eight microsatellite markers were used to examine An. culicifacies E collected from six sites in Sri Lanka during 2010-2012. Standard population genetic tests and analyses, genetic differentiation, Hardy-Weinberg equilibrium, linkage disequilibrium, Bayesian cluster analysis, AMOVA, SAMOVA and isolation-by-distance were conducted using five polymorphic loci. Five microsatellite loci were highly polymorphic with high allelic richness. Hardy-Weinberg Equilibrium (HWE) was significantly rejected for four loci with positive F(IS) values in the pooled population (p < 0.0100). Three loci showed high deviations in all sites except Kataragama, which was in agreement with HWE for all loci except one locus (p < 0.0016). Observed heterozygosity was less than the expected values for all sites except Kataragama, where reported negative F(IS) values indicated a heterozygosity excess. Genetic differentiation was observed for all sampling site pairs and was not supported by the isolation by distance model. Bayesian clustering analysis identified the presence of three sympatric clusters (gene pools) in the studied population. Significant genetic differentiation was detected in cluster pairs with low gene flow and isolation by distance was not detected between clusters. Furthermore, the results suggested the presence of a barrier to gene flow that divided the populations into two parts with the central hill region of Sri Lanka as the dividing line. Three sympatric clusters were detected among An. culicifacies E specimens isolated in Sri Lanka. There was no effect of geographic distance on genetic differentiation and the central mountain ranges in Sri Lanka appeared to be a barrier to gene flow.

A Preliminary Study of Genetic Variation in Populations of Monstera adansonii var. klotzschiana (Araceae) from North-East Brazil, Estimated with AFLP Molecular Markers

PubMed Central

Andrade, I. M.; Mayo, S. J.; van den Berg, C.; Fay, M. F.; Chester, M.; Lexer, C.; Kirkup, D.

2007-01-01

Background and Aims This study sought genetic evidence of long-term isolation in populations of Monstera adansonii var. klotzschiana (Araceae), a herbaceous, probably outbreeding, humid forest hemi-epiphyte, in the brejo forests of Ceará (north-east Brazil), and clarification of their relationships with populations in Amazonia and the Atlantic forest of Brazil. Methods Within-population genetic diversity and between-population dissimilarity were estimated using AFLP molecular markers in 75 individuals from eight populations located in Ceará, the Brazilian Atlantic Forest and Amazonia. Key Results The populations showed a clinal pattern of weak genetic differentiation over a large geographical region (FST = 0·1896). A strong correlation between genetic and geographical distance (Mantel test: r = 0·6903, P = 0·002) suggests a historical pattern of isolation by distance. Genetic structure analysis revealed at least two distinct gene pools in the data. The two isolated Ceará populations are significantly different from each other (pairwise ΦPT = 0·137, P = 0·003) and as diverse (Nei's gene diversity, average He = 0·1832, 0·1706) as those in the Atlantic and Amazon forest regions. The population in southern Brazil is less diverse (Nei's gene diversity, average He = 0·127) than the rest. The Ceará populations are related to those of the Atlantic forest rather than those from Amazonia (AMOVA, among-groups variation = 11·95 %, P = 0·037). Conclusions The gene pools detected within an overall pattern of clinal variation suggest distinct episodes of gene flow, possibly correlated with past humid forest expansions. The Ceará populations show no evidence of erosion of genetic diversity, although this was expected because of their isolation. Their genetic differentiation and relatively high diversity reinforce the importance of conserving the endangered brejo forests. PMID:17823112

Evidence for a common gene pool and frequent recombinational exchange of the tbpBA operon in Mannheimia haemolytica, Mannheimia glucosida and Bibersteinia trehalosi

PubMed Central

Lee, Inkyoung; Davies, Robert L.

2012-01-01

SUMMARY The tbpBA operon was sequenced in 42 representative isolates of Mannheimia haemolytica (32), Mannheimia glucosida (6) and Bibersteinia trehalosi (4). A total of 27 tbpB and 20 tbpA alleles were identified whilst the tbpBA operon was represented by 28 unique alleles that could be assigned to seven classes. There were 1566 (34.8% variation) polymorphic nucleotide sites and 482 (32.1% variation) variable inferred amino acid positions among the 42 tbpBA sequences. The tbpBA operons of serotype A2 M. haemolytica isolates are, with one exception, substantially more diverse than those of the other M. haemolytica serotypes and most likely have a different ancestral origin. The tbpBA phylogeny has been severely disrupted by numerous small- and large-scale intragenic recombination events. In addition, assortative (entire gene) recombination events, involving either the entire tbpBA operon or the individual tbpB and tbpA genes, have played a major role in shaping tbpBA structure and it’s distribution in the three species. Our findings indicate that a common gene pool exists for tbpBA in M. haemolytica, M. glucosida and B. trehalosi. In particular, B. trehalosi, M. glucosida and ovine M. haemolytica isolates share a large portion of the tbpA gene and this probably reflects selection for a conserved TbpA protein that provides effective iron-uptake in sheep. Bovine and ovine serotype A2 lineages have very different tbpBA alleles. Bovine-like tbpBA alleles have been partially, or completely, replaced by ovine-like tbpBA alleles in ovine serotype A2 isolates suggesting that different transferrin receptors are required by serotype A2 isolates for optimum iron uptake in cattle and sheep. Conversely, the tbpBA alleles of bovine-pathogenic serotype A1 and A6 isolates are very similar to those of closely related ovine isolates suggesting a recent and common evolutionary origin. PMID:20884693

Genomes in Turmoil: Frugality Drives Microbial Community Structure in Extremely Acidic Environments

NASA Astrophysics Data System (ADS)

Holmes, D. S.

2016-12-01

Extremely acidic environments (To gain insight into these issues, we have conducted deep bioinformatic analyses, including metabolic reconstruction of key assimilatory pathways, phylogenomics and network scrutiny of >160 genomes of acidophiles, including representatives from Archaea, Bacteria and Eukarya and at least ten metagenomes of acidic environments [Cardenas JP, et al. pp 179-197 in Acidophiles, eds R. Quatrini and D. B. Johnson, Caister Academic Press, UK (2016)]. Results yielded valuable insights into cellular processes, including carbon and nitrogen management and energy production, linking biogeochemical processes to organismal physiology. They also provided insight into the evolutionary forces that shape the genomic structure of members of acidophile communities. Niche partitioning can explain diversity patterns in rapidly changing acidic environments such as bioleaching heaps. However, in spatially and temporally homogeneous acidic environments genome flux appears to provide deeper insight into the composition and evolution of acidic consortia. Acidophiles have undergone genome streamlining by gene loss promoting mutual coexistence of species that exploit complementarity use of scarce resources consistent with the Black Queen hypothesis [Morris JJ et al. mBio 3: e00036-12 (2012)]. Acidophiles also have a large pool of accessory genes (the microbial super-genome) that can be accessed by horizontal gene transfer. This further promotes dependency relationships as drivers of community structure and the evolution of keystone species. Acknowledgements: Fondecyt 1130683; Basal CCTE PFB16

Development of innovative techniques and principles that may be used as models to improve plant performance. Progress report

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

Hanna, W.W.; Burton, G.W.

1984-06-01

Dominant immunity to rust (Puccinia) and Piricularia leaf spot, a new stable A/sub 1/ cytoplasm and yield genes have been discovered in a single accession of P. monodii, a subspecies of pearl millet Pennisetum americanum. The useful genes were transferred to the best pearl millet inbreds using a backcrossing and screening system enhanced by controlling daylength, temperature, and breaking seed dormancy. Genes for fertility restoration, plant morphology, head characteristics and grain yield were transferred from the A' genome of P. purpureum (secondary gene pool) to pearl millet. This was made possible by doubling the chromosome number of the sterile triploidmore » between pearl millet and P. purpureum to produce fertile hexaploids. A number of hexaploids were backcrossed to pearl millet to find hexaploids that produced A' gametes to produce AA' 2n = 14 pearl millet plants. The value of the genes on the A' genome were not recognized before the AA' plants were produced because the B genome masked the characteristics on the A' genome in P. purpureum. In the tertiary gene pool, male and female fertile apomictic BC/sub 1/ plants were produced between pearl millet and P. orientale or P. squamulatum by manipulating chromosome levels or by producing trispecific hybrids. The development of fertile backcross plants in this gene pool is encouraging for further germplasm transfer research with emphasis on apomixis. The cytoplasm of P. schweinfurthii has been transferred to pearl millet. Mutations for early maturity and daylength insensitivity induced with mutagens have resulted in the development and release of early maturing inbreds Tift 23A/sub 1/E/sub 1/ and Tift 23B/sub 1/E/sub 1/.« less

Environmental genomics of "Haloquadratum walsbyi" in a saltern crystallizer indicates a large pool of accessory genes in an otherwise coherent species

PubMed Central

Legault, Boris A; Lopez-Lopez, Arantxa; Alba-Casado, Jose Carlos; Doolittle, W Ford; Bolhuis, Henk; Rodriguez-Valera, Francisco; Papke, R Thane

2006-01-01

Background Mature saturated brine (crystallizers) communities are largely dominated (>80% of cells) by the square halophilic archaeon "Haloquadratum walsbyi". The recent cultivation of the strain HBSQ001 and thesequencing of its genome allows comparison with the metagenome of this taxonomically simplified environment. Similar studies carried out in other extreme environments have revealed very little diversity in gene content among the cell lineages present. Results The metagenome of the microbial community of a crystallizer pond has been analyzed by end sequencing a 2000 clone fosmid library and comparing the sequences obtained with the genome sequence of "Haloquadratum walsbyi". The genome of the sequenced strain was retrieved nearly complete within this environmental DNA library. However, many ORF's that could be ascribed to the "Haloquadratum" metapopulation by common genome characteristics or scaffolding to the strain genome were not present in the specific sequenced isolate. Particularly, three regions of the sequenced genome were associated with multiple rearrangements and the presence of different genes from the metapopulation. Many transposition and phage related genes were found within this pool which, together with the associated atypical GC content in these areas, supports lateral gene transfer mediated by these elements as the most probable genetic cause of this variability. Additionally, these sequences were highly enriched in putative regulatory and signal transduction functions. Conclusion These results point to a large pan-genome (total gene repertoire of the genus/species) even in this highly specialized extremophile and at a single geographic location. The extensive gene repertoire is what might be expected of a population that exploits a diverse nutrient pool, resulting from the degradation of biomass produced at lower salinities. PMID:16820057

Genetic Structure of Anopheles (Nyssorhynchus) marajoara (Diptera: Culicidae) in Colombia

PubMed Central

Brochero, Helena; Li, Cong; Wilkerson, Richard; Conn, Jan E.; Ruiz-García, Manuel

2010-01-01

Five Anopheles marajoara Galvão and Damasceno populations, representing diverse ecological conditions, were sampled throughout Colombia and analyzed using nine hypervariable DNA microsatellite loci. The overall genetic diversity (H = 0.58) was lower than that determined for some Brazilian populations using the same markers. The Caquetá population (Colombia) had the lowest gene diversity (H = 0.48), and it was the only population at Hardy–Weinberg equilibrium. Hardy–Weinberg disequilibrium in the remaining four populations was probably caused by the Wahlund effect. The assignment analyses showed two incompletely isolated gene pools separated by the Eastern Andean cordillera. However, other possible geographical barriers (rivers and other mountains) did not play any role in the moderate genetic heterogeneity found among these populations (FST = 0.069). These results are noteworthy, because this species is a putative malaria vector in Colombia. PMID:20810825

Shedding genomic light on Aristotle's lantern.

PubMed

Sodergren, Erica; Shen, Yufeng; Song, Xingzhi; Zhang, Lan; Gibbs, Richard A; Weinstock, George M

2006-12-01

Sea urchins have proved fascinating to biologists since the time of Aristotle who compared the appearance of their bony mouth structure to a lantern in The History of Animals. Throughout modern times it has been a model system for research in developmental biology. Now, the genome of the sea urchin Strongylocentrotus purpuratus is the first echinoderm genome to be sequenced. A high quality draft sequence assembly was produced using the Atlas assembler to combine whole genome shotgun sequences with sequences from a collection of BACs selected to form a minimal tiling path along the genome. A formidable challenge was presented by the high degree of heterozygosity between the two haplotypes of the selected male representative of this marine organism. This was overcome by use of the BAC tiling path backbone, in which each BAC represents a single haplotype, as well as by improvements in the Atlas software. Another innovation introduced in this project was the sequencing of pools of tiling path BACs rather than individual BAC sequencing. The Clone-Array Pooled Shotgun Strategy greatly reduced the cost and time devoted to preparing shotgun libraries from BAC clones. The genome sequence was analyzed with several gene prediction methods to produce a comprehensive gene list that was then manually refined and annotated by a volunteer team of sea urchin experts. This latter annotation community edited over 9000 gene models and uncovered many unexpected aspects of the sea urchin genetic content impacting transcriptional regulation, immunology, sensory perception, and an organism's development. Analysis of the basic deuterostome genetic complement supports the sea urchin's role as a model system for deuterostome and, by extension, chordate development.

Population structure and genetic diversity in a commercial maize breeding program assessed with SSR and SNP markers

PubMed Central

Van Inghelandt, Delphine; Melchinger, Albrecht E.; Lebreton, Claude

2010-01-01

Information about the genetic diversity and population structure in elite breeding material is of fundamental importance for the improvement of crops. The objectives of our study were to (a) examine the population structure and the genetic diversity in elite maize germplasm based on simple sequence repeat (SSR) markers, (b) compare these results with those obtained from single nucleotide polymorphism (SNP) markers, and (c) compare the coancestry coefficient calculated from pedigree records with genetic distance estimates calculated from SSR and SNP markers. Our study was based on 1,537 elite maize inbred lines genotyped with 359 SSR and 8,244 SNP markers. The average number of alleles per locus, of group specific alleles, and the gene diversity (D) were higher for SSRs than for SNPs. Modified Roger’s distance (MRD) estimates and membership probabilities of the STRUCTURE matrices were higher for SSR than for SNP markers but the germplasm organization in four heterotic pools was consistent with STRUCTURE results based on SSRs and SNPs. MRD estimates calculated for the two marker systems were highly correlated (0.87). Our results suggested that the same conclusions regarding the structure and the diversity of heterotic pools could be drawn from both markers types. Furthermore, although our results suggested that the ratio of the number of SSRs and SNPs required to obtain MRD or D estimates with similar precision is not constant across the various precision levels, we propose that between 7 and 11 times more SNPs than SSRs should be used for analyzing population structure and genetic diversity. Electronic supplementary material The online version of this article (doi:10.1007/s00122-009-1256-2) contains supplementary material, which is available to authorized users. PMID:20063144

Prevalence of sulfonamide-resistant bacteria, resistance genes and integron-associated horizontal gene transfer in natural water bodies and soils adjacent to a swine feedlot in northern Taiwan.

PubMed

Hsu, Jih-Tay; Chen, Chia-Yang; Young, Chu-Wen; Chao, Wei-Liang; Li, Mao-Hao; Liu, Yung-Hsin; Lin, Chu-Ming; Ying, Chingwen

2014-07-30

Antibiotics are commonly used in swine feed to treat and prevent disease, as well as to promote growth. Antibiotics released into the environment via wastewater could accelerate the emergence of antibiotic-resistant bacteria and resistance genes in the surrounding environment. In this study, we quantified the occurrence of sulfonamides, sulfonamide-resistant microorganisms and resistance genes in the wastewater from a swine farm in northern Taiwan and its surrounding natural water bodies and soils. Sulfonamide levels were similar in the receiving downstream and upstream river water. However, the prevalence of sulfonamide-resistant bacteria and resistance genes, as analyzed by cultivation-dependent and -independent molecular approaches, was significantly greater in the downstream compared to the upstream river water samples. Barcoded-pyrosequencing revealed a highly diverse bacterial community structure in each sample. However, the sequence identity of the sulfonamide resistance gene sul1 in the wastewater and downstream environment samples was nearly identical (99-100%). The sul1 gene, which is genetically linked to class 1 integrons, was dominant in the downstream water bodies and soils. In conclusion, the increased prevalence of sulfonamide resistance genes in the wastewater from a swine farm, independent of the persistent presence of sulfonamides, could be a potential source of resistant gene pools in the surrounding environment. Copyright © 2014 Elsevier B.V. All rights reserved.

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

Barker, J. Jr.; Tenenbaum, B.; Woolf, F.

This paper focuses on the governance and regulation of power pools outside the United States. The current governance and regulatory arrangements for four power pools, as developed in pool documents and government regulations and laws, are compared and contrasted. The power pools analyzed are located in England and Wales, Australia, Canada, and Scandinavia. Topics discussed in relation to these pools are the effects of structure on governance, how each pool has dealt with a number of basic governance decisions, how the pools monitor the markets, ways in which regulators and other institutions control pools, and self-governance issues.

The Fate of Spermatogonial Stem Cells in the Cryptorchid Testes of RXFP2 Deficient Mice

PubMed Central

Ferguson, Lydia; How, Javier J.; Agoulnik, Alexander I.

2013-01-01

The environmental niche of the spermatogonial stem cell pool is critical to ensure the continued generation of the germ cell population. To study the consequences of an aberrant testicular environment in cryptorchidism we used a mouse model with a deletion of Rxfp2 gene resulting in a high intra-abdominal testicular position. Mutant males were infertile with the gross morphology of the cryptorchid testis progressively deteriorating with age. Few spermatogonia were identifiable in 12 month old cryptorchid testes. Gene expression analysis showed no difference between mutant and control testes at postnatal day 10. In three month old males a decrease in expression of spermatogonial stem cell (SSC) markers Id4, Nanos2, and Ret was shown. The direct counting of ID4+ cells supported a significant decrease of SSCs. In contrast, the expression of Plzf, a marker for undifferentiated and differentiating spermatogonia was not reduced, and the number of PLZF+ cells in the cryptorchid testis was higher in three month old testes, but equal to control in six month old mutants. The PLZF+ cells did not show a higher rate of apoptosis in cryptorchid testis. The expression of the Sertoli cell FGF2 gene required for SSC maintenance was significantly reduced in mutant testis. Based on these findings we propose that the deregulation of somatic and germ cell genes in the cryptorchid testis, directs the SSCs towards the differentiation pathway. This leads to a depletion of the SSC pool and an increase in the number of PLZF+ spermatogonial cells, which too, eventually decreases with the exhaustion of the stem cell pool. Such a dynamic suggests that an early correction of cryptorchidism is critical for the retention of the SSC pool. PMID:24098584

Detection of new HLA-DPB1 alleles generated by interallelic gene conversion using PCR amplification of DPB1 second exon sequences from sperm

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

Erlich, H.; Zangenberg, G.; Bugawan, T.

The rate at which allelic diversity at the HLA class I and class II loci evolves has been the subject of considerable controversy as have the mechanisms which generate new alleles. The patchwork pattern of polymorphism, particularly within the second exon of the HLA-DPB1 locus where the polymorphic sequence motifs are localized to 6 discrete regions, is consistent with the hypothesis that much of the allelic sequence variation may have been generated by segmental exchange (gene conversion). To measure the rate of new DPB1 variant generation, we have developed a strategy in which DPB1 second exon sequences are amplified frommore » pools of FACS-sorted sperm (n=50) from a heterozygous sperm donor. Pools of sperm from these heterozygous individuals are amplified with an allele-specific primer for one allele and analyzed with sequence-specific oligonucleotide probes (SSOP) complementary to the other allele. This screening procedure, which is capable of detecting a single variant molecule in a pool of parental alleles, allows the identification of new variants that have been generated by recombination and/or gene conversion between the two parental alleles. To control for potential PCR artifacts, the same screening procedure was carried out with mixtures of sperm from DPB1 *0301/*0301 and DPB1 *0401/ 0401 individuals. Pools containing putative new variants DPB1 alleles were analyzed further by cloning into M13 and sequencing the M13 clones. Our current estimate is that about 1/10,000 sperm from these heterozygous individuals represents a new DPB1 allele generated by micro-gene conversion within the second exon.« less

The post-vaccine microevolution of invasive Streptococcus pneumoniae

PubMed Central

Cremers, Amelieke J. H.; Mobegi, Fredrick M.; de Jonge, Marien I.; van Hijum, Sacha A. F. T.; Meis, Jacques F.; Hermans, Peter W. M.; Ferwerda, Gerben; Bentley, Stephen D.; Zomer, Aldert L.

2015-01-01

The 7-valent pneumococcal conjugated vaccine (PCV7) has affected the genetic population of Streptococcus pneumoniae in pediatric carriage. Little is known however about pneumococcal population genomics in adult invasive pneumococcal disease (IPD) under vaccine pressure. We sequenced and serotyped 349 strains of S. pneumoniae isolated from IPD patients in Nijmegen between 2001 and 2011. Introduction of PCV7 in the Dutch National Immunization Program in 2006 preluded substantial alterations in the IPD population structure caused by serotype replacement. No evidence could be found for vaccine induced capsular switches. We observed that after a temporary bottleneck in gene diversity after the introduction of PCV7, the accessory gene pool re-expanded mainly by genes already circulating pre-PCV7. In the post-vaccine genomic population a number of genes changed frequency, certain genes became overrepresented in vaccine serotypes, while others shifted towards non-vaccine serotypes. Whether these dynamics in the invasive pneumococcal population have truly contributed to invasiveness and manifestations of disease remains to be further elucidated. We suggest the use of whole genome sequencing for surveillance of pneumococcal population dynamics that could give a prospect on the course of disease, facilitating effective prevention and management of IPD. PMID:26492862

Premature Mobility of Boulders in Constructed Step-pool River Structures in the Carmel River, CA: The Role of Fish-centric Design Constraints, and Flow on Structural Stability

NASA Astrophysics Data System (ADS)

Smith, D. P.; Chow, K.; Luna, L.

2017-12-01

The 32 m tall San Clemente Dam (Carmel River, CA) was removed in 2015 to eliminate seismic risk and to improve fish passage for all life stages of steelhead (O. mykiss). Reservoir sediment was sequestered in place, rather than released, and a new 1000 m long channel/floodplain system was constructed to circumvent the stored sediment. The channel comprised a 250 m long, meandering low-gradient reach and a 750 m reach with alternating step-pool sections, plane beds, and resting pools. The floodprone surfaces were compacted, wrapped in geotechnical fabric and vegetated. This study analyzes the geomorphic evolution of the new channel system during its first two years of service based upon detailed field inspection, SfM photogrammetry, orthophoto analysis, and 2d hydraulic modeling. A significant proportion of the step-pool structures experienced premature mobility and several reaches of engineered stream banks were eroded in the first year. Individual, six-tonne boulders were mobilized despite experiencing less than the 3 yr flow. The channel and floodplain were fully repaired following the first year. Strong flows (two 10-yr floods and a 30-yr flood) during the second year catastrophically altered the constructed channel and floodplain. While the low-gradient reach remained intact, each of the original step-pool structures was either completely mobilized and destroyed, buried by gravel, or bypassed by the subsequent channel. Despite the overall structural failure of the constructed channel, the new channel does not block steelhead migration, and can be serendipitously considered an ecological success. Step-pool design was constrained by a fish-centric requirement that steps be 1 ft tall or less. Some constructed "resting pools" filled rather than transport sediment. Using fish-centric constraints in the design, rather than strictly fluvial geomorphic principles may have contributed to early failure of the step-pool structures and other parts of the system.

Spatial genetic structuring of baobab (Adansonia digitata, Malvaceae) in the traditional agroforestry systems of West Africa.

PubMed

Kyndt, Tina; Assogbadjo, Achille E; Hardy, Olivier J; Glele Kakaï, Romain; Sinsin, Brice; Van Damme, Patrick; Gheysen, Godelieve

2009-05-01

This study evaluates the spatial genetic structure of baobab (Adansonia digitata) populations from West African agroforestry systems at different geographical scales using AFLP fingerprints. Eleven populations from four countries (Benin, Ghana, Burkina Faso, and Senegal) had comparable levels of genetic diversity, although the two populations in the extreme west (Senegal) had less diversity. Pairwise F(ST) ranged from 0.02 to 0.28 and increased with geographic distance, even at a regional scale. Gene pools detected by Bayesian clustering seem to be a byproduct of the isolation-by-distance pattern rather than representing actual discrete entities. The organization of genetic diversity appears to result essentially from spatially restricted gene flow, with some influences of human seed exchange. Despite the potential for relatively long-distance pollen and seed dispersal by bats within populations, statistically significant spatial genetic structuring within populations (SGS) was detected and gave a mean indirect estimate of neighborhood size of ca. 45. This study demonstrated that relatively high levels of genetic structuring are present in baobab at both large and within-population level, which was unexpected in regard to its dispersal by bats and the influence of human exchange of seeds. Implications of these results for the conservation of baobab populations are discussed.

“Every Gene Is Everywhere but the Environment Selects”: Global Geolocalization of Gene Sharing in Environmental Samples through Network Analysis

PubMed Central

Fondi, Marco; Karkman, Antti; Tamminen, Manu V.; Bosi, Emanuele; Virta, Marko; Fani, Renato; Alm, Eric; McInerney, James O.

2016-01-01

The spatial distribution of microbes on our planet is famously formulated in the Baas Becking hypothesis as “everything is everywhere but the environment selects.” While this hypothesis does not strictly rule out patterns caused by geographical effects on ecology and historical founder effects, it does propose that the remarkable dispersal potential of microbes leads to distributions generally shaped by environmental factors rather than geographical distance. By constructing sequence similarity networks from uncultured environmental samples, we show that microbial gene pool distributions are not influenced nearly as much by geography as ecology, thus extending the Bass Becking hypothesis from whole organisms to microbial genes. We find that gene pools are shaped by their broad ecological niche (such as sea water, fresh water, host, and airborne). We find that freshwater habitats act as a gene exchange bridge between otherwise disconnected habitats. Finally, certain antibiotic resistance genes deviate from the general trend of habitat specificity by exhibiting a high degree of cross-habitat mobility. The strong cross-habitat mobility of antibiotic resistance genes is a cause for concern and provides a paradigmatic example of the rate by which genes colonize new habitats when new selective forces emerge. PMID:27190206

Distinct effects of struvite and biochar amendment on the class 1 integron antibiotic resistance gene cassettes in phyllosphere and rhizosphere.

PubMed

An, Xin-Li; Chen, Qing-Lin; Zhu, Dong; Su, Jian-Qiang

2018-08-01

Struvite recovered from wastewater is promising for recycling phosphorus into soil as fertilizers. However, struvite application may prompt the proliferation of antibiotic resistance in soil and plant. This study examined the impacts of struvite application and biochar amendment on integrons abundance and gene cassette contexts in rhizosphere soil and phyllosphere using quantitative PCR and clone library analysis. Microcosm experiments revealed that class 1 integron was the most prevalent in all samples, with higher concentration and higher relative abundance in rhizosphere than those in phyllosphere. The majority of resistance gene cassettes were associated with genes encoding resistance to aminoglycosides, beta-lactams and chloramphenicols. Struvite application significantly increased the genetic diversity of antibiotic resistance gene cassettes in both rhizosphere and phyllosphere. However, biochar amendment attenuated the increasing effect of struvite application exerting on the class 1 integron antibiotic resistance gene cassette pool in phyllosphere. These findings highlighted human activities to be the source of integron gene cassette pool and raised the possibility of using biochar amendment as an alternative mean for mitigating antibiotic resistance in environments. Copyright © 2018 Elsevier B.V. All rights reserved.

Serial analysis of gene expression (SAGE) in bovine trypanotolerance: preliminary results

PubMed Central

2003-01-01

In Africa, trypanosomosis is a tsetse-transmitted disease which represents the most important constraint to livestock production. Several indigenous West African taurine (Bos taurus) breeds, such as the Longhorn (N'Dama) cattle are well known to control trypanosome infections. This genetic ability named "trypanotolerance" results from various biological mechanisms under multigenic control. The methodologies used so far have not succeeded in identifying the complete pool of genes involved in trypanotolerance. New post genomic biotechnologies such as transcriptome analyses are efficient in characterising the pool of genes involved in the expression of specific biological functions. We used the serial analysis of gene expression (SAGE) technique to construct, from Peripheral Blood Mononuclear Cells of an N'Dama cow, 2 total mRNA transcript libraries, at day 0 of a Trypanosoma congolense experimental infection and at day 10 post-infection, corresponding to the peak of parasitaemia. Bioinformatic comparisons in the bovine genomic databases allowed the identification of 187 up- and down- regulated genes, EST and unknown functional genes. Identification of the genes involved in trypanotolerance will allow to set up specific microarray sets for further metabolic and pharmacological studies and to design field marker-assisted selection by introgression programmes. PMID:12927079

Serial analysis of gene expression (SAGE) in bovine trypanotolerance: preliminary results.

PubMed

Berthier, David; Quéré, Ronan; Thevenon, Sophie; Belemsaga, Désiré; Piquemal, David; Marti, Jacques; Maillard, Jean-Charles

2003-01-01

In Africa, trypanosomosis is a tsetse-transmitted disease which represents the most important constraint to livestock production. Several indigenous West African taurine Bos taurus) breeds, such as the Longhorn (N'Dama) cattle are well known to control trypanosome infections. This genetic ability named "trypanotolerance" results from various biological mechanisms under multigenic control. The methodologies used so far have not succeeded in identifying the complete pool of genes involved in trypanotolerance. New post genomic biotechnologies such as transcriptome analyses are efficient in characterising the pool of genes involved in the expression of specific biological functions. We used the serial analysis of gene expression (SAGE) technique to construct, from Peripheral Blood Mononuclear Cells of an N'Dama cow, 2 total mRNA transcript libraries, at day 0 of a Trypanosoma congolense experimental infection and at day 10 post-infection, corresponding to the peak of parasitaemia. Bioinformatic comparisons in the bovine genomic databases allowed the identification of 187 up- and down- regulated genes, EST and unknown functional genes. Identification of the genes involved in trypanotolerance will allow to set up specific microarray sets for further metabolic and pharmacological studies and to design field marker-assisted selection by introgression programmes.

Balinese Y-chromosome perspective on the peopling of Indonesia: genetic contributions from pre-neolithic hunter-gatherers, Austronesian farmers, and Indian traders.

PubMed

Karafet, Tatiana M; Lansing, J S; Redd, Alan J; Reznikova, Svetlana; Watkins, Joseph C; Surata, S P K; Arthawiguna, W A; Mayer, Laura; Bamshad, Michael; Jorde, Lynn B; Hammer, Michael F

2005-02-01

The island of Bali lies near the center of the southern chain of islands in the Indonesian archipelago, which served as a stepping-stone for early migrations of hunter-gatherers to Melanesia and Australia and for more recent migrations of Austronesian farmers from mainland Southeast Asia to the Pacific. Bali is the only Indonesian island with a population that currently practices the Hindu religion and preserves various other Indian cultural, linguistic, and artistic traditions (Lansing 1983). Here, we examine genetic variation on the Y chromosomes of 551 Balinese men to investigate the relative contributions of Austronesian farmers and pre-Neolithic hunter-gatherers to the contemporary Balinese paternal gene pool and to test the hypothesis of recent paternal gene flow from the Indian subcontinent. Seventy-one Y-chromosome binary polymorphisms (single nucleotide polymorphisms, SNPs) and 10 Y-chromosome-linked short tandem repeats (STRs) were genotyped on a sample of 1,989 Y chromosomes from 20 populations representing Indonesia (including Bali), southern China, Southeast Asia, South Asia, the Near East, and Oceania. SNP genotyping revealed 22 Balinese lineages, 3 of which (O-M95, O-M119, and O-M122) account for nearly 83.7% of Balinese Y chromosomes. Phylogeographic analyses suggest that all three major Y-chromosome haplogroups migrated to Bali with the arrival of Austronesian speakers; however, STR diversity patterns associated with these haplogroups are complex and may be explained by multiple waves of Austronesian expansion to Indonesia by different routes. Approximately 2.2% of contemporary Balinese Y chromosomes (i.e., K-M9*, K-M230, and M lineages) may represent the pre-Neolithic component of the Indonesian paternal gene pool. In contrast, eight other haplogroups (e.g., within H, J, L, and R), making up approximately 12% of the Balinese paternal gene pool, appear to have migrated to Bali from India. These results indicate that the Austronesian expansion had a profound effect on the composition of the Balinese paternal gene pool and that cultural transmission from India to Bali was accompanied by substantial levels of gene flow.

Pooled-DNA sequencing identifies genomic regions of selection in Nigerian isolates of Plasmodium falciparum.

PubMed

Oyebola, Kolapo M; Idowu, Emmanuel T; Olukosi, Yetunde A; Awolola, Taiwo S; Amambua-Ngwa, Alfred

2017-06-29

The burden of falciparum malaria is especially high in sub-Saharan Africa. Differences in pressure from host immunity and antimalarial drugs lead to adaptive changes responsible for high level of genetic variations within and between the parasite populations. Population-specific genetic studies to survey for genes under positive or balancing selection resulting from drug pressure or host immunity will allow for refinement of interventions. We performed a pooled sequencing (pool-seq) of the genomes of 100 Plasmodium falciparum isolates from Nigeria. We explored allele-frequency based neutrality test (Tajima's D) and integrated haplotype score (iHS) to identify genes under selection. Fourteen shared iHS regions that had at least 2 SNPs with a score > 2.5 were identified. These regions code for genes that were likely to have been under strong directional selection. Two of these genes were the chloroquine resistance transporter (CRT) on chromosome 7 and the multidrug resistance 1 (MDR1) on chromosome 5. There was a weak signature of selection in the dihydrofolate reductase (DHFR) gene on chromosome 4 and MDR5 genes on chromosome 13, with only 2 and 3 SNPs respectively identified within the iHS window. We observed strong selection pressure attributable to continued chloroquine and sulfadoxine-pyrimethamine use despite their official proscription for the treatment of uncomplicated malaria. There was also a major selective sweep on chromosome 6 which had 32 SNPs within the shared iHS region. Tajima's D of circumsporozoite protein (CSP), erythrocyte-binding antigen (EBA-175), merozoite surface proteins - MSP3 and MSP7, merozoite surface protein duffy binding-like (MSPDBL2) and serine repeat antigen (SERA-5) were 1.38, 1.29, 0.73, 0.84 and 0.21, respectively. We have demonstrated the use of pool-seq to understand genomic patterns of selection and variability in P. falciparum from Nigeria, which bears the highest burden of infections. This investigation identified known genomic signatures of selection from drug pressure and host immunity. This is evidence that P. falciparum populations explore common adaptive strategies that can be targeted for the development of new interventions.

Process-Based Species Pools Reveal the Hidden Signature of Biotic Interactions Amid the Influence of Temperature Filtering.

PubMed

Lessard, Jean-Philippe; Weinstein, Ben G; Borregaard, Michael K; Marske, Katharine A; Martin, Danny R; McGuire, Jimmy A; Parra, Juan L; Rahbek, Carsten; Graham, Catherine H

2016-01-01

A persistent challenge in ecology is to tease apart the influence of multiple processes acting simultaneously and interacting in complex ways to shape the structure of species assemblages. We implement a heuristic approach that relies on explicitly defining species pools and permits assessment of the relative influence of the main processes thought to shape assemblage structure: environmental filtering, dispersal limitations, and biotic interactions. We illustrate our approach using data on the assemblage composition and geographic distribution of hummingbirds, a comprehensive phylogeny and morphological traits. The implementation of several process-based species pool definitions in null models suggests that temperature-but not precipitation or dispersal limitation-acts as the main regional filter of assemblage structure. Incorporating this environmental filter directly into the definition of assemblage-specific species pools revealed an otherwise hidden pattern of phylogenetic evenness, indicating that biotic interactions might further influence hummingbird assemblage structure. Such hidden patterns of assemblage structure call for a reexamination of a multitude of phylogenetic- and trait-based studies that did not explicitly consider potentially important processes in their definition of the species pool. Our heuristic approach provides a transparent way to explore patterns and refine interpretations of the underlying causes of assemblage structure.

Y-chromosome diversity in modern Bulgarians: new clues about their ancestry.

PubMed

Karachanak, Sena; Grugni, Viola; Fornarino, Simona; Nesheva, Desislava; Al-Zahery, Nadia; Battaglia, Vincenza; Carossa, Valeria; Yordanov, Yordan; Torroni, Antonio; Galabov, Angel S; Toncheva, Draga; Semino, Ornella

2013-01-01

To better define the structure and origin of the Bulgarian paternal gene pool, we have examined the Y-chromosome variation in 808 Bulgarian males. The analysis was performed by high-resolution genotyping of biallelic markers and by analyzing the STR variation within the most informative haplogroups. We found that the Y-chromosome gene pool in modern Bulgarians is primarily represented by Western Eurasian haplogroups with ∼ 40% belonging to haplogroups E-V13 and I-M423, and 20% to R-M17. Haplogroups common in the Middle East (J and G) and in South Western Asia (R-L23*) occur at frequencies of 19% and 5%, respectively. Haplogroups C, N and Q, distinctive for Altaic and Central Asian Turkic-speaking populations, occur at the negligible frequency of only 1.5%. Principal Component analyses group Bulgarians with European populations, apart from Central Asian Turkic-speaking groups and South Western Asia populations. Within the country, the genetic variation is structured in Western, Central and Eastern Bulgaria indicating that the Balkan Mountains have been permeable to human movements. The lineage analysis provided the following interesting results: (i) R-L23* is present in Eastern Bulgaria since the post glacial period; (ii) haplogroup E-V13 has a Mesolithic age in Bulgaria from where it expanded after the arrival of farming; (iii) haplogroup J-M241 probably reflects the Neolithic westward expansion of farmers from the earliest sites along the Black Sea. On the whole, in light of the most recent historical studies, which indicate a substantial proto-Bulgarian input to the contemporary Bulgarian people, our data suggest that a common paternal ancestry between the proto-Bulgarians and the Altaic and Central Asian Turkic-speaking populations either did not exist or was negligible.

Association of SSR markers with contents of fatty acids in olive oil and genetic diversity analysis of an olive core collection.

PubMed

Ipek, M; Ipek, A; Seker, M; Gul, M K

2015-03-27

The purpose of this research was to characterize an olive core collection using some agronomic characters and simple sequence repeat (SSR) markers and to determine SSR markers associated with the content of fatty acids in olive oil. SSR marker analysis demonstrated the presence of a high amount of genetic variation between the olive cultivars analyzed. A UPGMA dendrogram demonstrated that olive cultivars did not cluster on the basis of their geographic origin. Fatty acid components of olive oil in these cultivars were determined. The results also showed that there was a great amount of variation between the olive cultivars in terms of fatty acid composition. For example, oleic acid content ranged from 57.76 to 76.9% with standard deviation of 5.10%. Significant correlations between fatty acids of olive oil were observed. For instance, a very high negative correlation (-0.812) between oleic and linoleic acids was detected. A structured association analysis between the content of fatty acids in olive oil and SSR markers was performed. STRUCTURE analysis assigned olive cultivars to two gene pools (K = 2). Assignment of olive cultivars to these gene pools was not based on geographical origin. Association between fatty acid traits and SSR markers was evaluated using the general linear model of TASSEL. Significant associations were determined between five SSR markers and stearic, oleic, linoleic, and linolenic acids of olive oil. Very high associations (P < 0.001) between ssrOeUA-DCA14 and stearic acid and between GAPU71B and oleic acid indicated that these markers could be used for marker-assisted selection in olive.

Structural differences between cell matrix and culture medium N-linked oligosaccharides on heavy proteochondroitin sulfate proteoglycan (PCS-H)

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

Cioffi, L.; Conrad, H.E.

1986-05-01

Tibial chondrocytes were labeled metabolically with /sup 3/H-man and the PCS-H was isolated from the culture medium (CM) and the cell matrix (Ma) pools. Equal amounts of /sup 3/H were incorporated into the PCS-H of the CM and Ma pools. The PCS-H pools were digested with thermolysin, Chondroitinase, and then N-glycanase, and the N-linked oligosaccharides were chromatographed on Con-A Sepharose. The ratios of complex to high mannose oligosaccharides for the CM and Ma were 6.1 and 2.6, respectively. More than 60% of the complex CM N-linked oligosaccharides were charged species whereas only 40% of the Ma N-linked oligosaccharides were charged.more » The oligosaccharides were analyzed by HPLC. Both complex and high mannose oligosaccharides found in the PCS-H of the CM and Ma pools were mixtures of identical structures but the amounts of each structure in the two pools showed marked differences. These observations indicate that distinct PCS-H species are found in the CM and Ma pools.« less

Nuclear Calcium Signaling Controls Expression of a Large Gene Pool: Identification of a Gene Program for Acquired Neuroprotection Induced by Synaptic Activity

PubMed Central

Zhang, Sheng-Jia; Zou, Ming; Lu, Li; Lau, David; Ditzel, Désirée A. W.; Delucinge-Vivier, Celine; Aso, Yoshinori; Descombes, Patrick; Bading, Hilmar

2009-01-01

Synaptic activity can boost neuroprotection through a mechanism that requires synapse-to-nucleus communication and calcium signals in the cell nucleus. Here we show that in hippocampal neurons nuclear calcium is one of the most potent signals in neuronal gene expression. The induction or repression of 185 neuronal activity-regulated genes is dependent upon nuclear calcium signaling. The nuclear calcium-regulated gene pool contains a genomic program that mediates synaptic activity-induced, acquired neuroprotection. The core set of neuroprotective genes consists of 9 principal components, termed Activity-regulated Inhibitor of Death (AID) genes, and includes Atf3, Btg2, GADD45β, GADD45γ, Inhibin β-A, Interferon activated gene 202B, Npas4, Nr4a1, and Serpinb2, which strongly promote survival of cultured hippocampal neurons. Several AID genes provide neuroprotection through a common process that renders mitochondria more resistant to cellular stress and toxic insults. Stereotaxic delivery of AID gene-expressing recombinant adeno-associated viruses to the hippocampus confers protection in vivo against seizure-induced brain damage. Thus, treatments that enhance nuclear calcium signaling or supplement AID genes represent novel therapies to combat neurodegenerative conditions and neuronal cell loss caused by synaptic dysfunction, which may be accompanied by a deregulation of calcium signal initiation and/or propagation to the cell nucleus. PMID:19680447

Cellular choline and glycine betaine pools impact osmoprotection and phospholipase C production in Pseudomonas aeruginosa.

PubMed

Fitzsimmons, Liam F; Hampel, Ken J; Wargo, Matthew J

2012-09-01

Choline is abundantly produced by eukaryotes and plays an important role as a precursor of the osmoprotectant glycine betaine. In Pseudomonas aeruginosa, glycine betaine has additional roles as a nutrient source and an inducer of the hemolytic phospholipase C, PlcH. The multiple functions for glycine betaine suggested that the cytoplasmic pool of glycine betaine is regulated in P. aeruginosa. We used (13)C nuclear magnetic resonance ((13)C-NMR) to demonstrate that P. aeruginosa maintains both choline and glycine betaine pools under a variety of conditions, in contrast to the transient glycine betaine pool reported for most bacteria. We were able to experimentally manipulate the choline and glycine betaine pools by overexpression of the cognate catabolic genes. Depletion of either the choline or glycine betaine pool reduced phospholipase production, a result unexpected for choline depletion. Depletion of the glycine betaine pool, but not the choline pool, inhibited growth under conditions of high salt with glucose as the primary carbon source. Depletion of the choline pool inhibited growth under high-salt conditions with choline as the sole carbon source, suggesting a role for the choline pool under these conditions. Here we have described the presence of a choline pool in P. aeruginosa and other pseudomonads that, with the glycine betaine pool, regulates osmoprotection and phospholipase production and impacts growth under high-salt conditions. These findings suggest that the levels of both pools are actively maintained and that perturbation of either pool impacts P. aeruginosa physiology.

Cellular Choline and Glycine Betaine Pools Impact Osmoprotection and Phospholipase C Production in Pseudomonas aeruginosa

PubMed Central

Fitzsimmons, Liam F.; Hampel, Ken J.

2012-01-01

Choline is abundantly produced by eukaryotes and plays an important role as a precursor of the osmoprotectant glycine betaine. In Pseudomonas aeruginosa, glycine betaine has additional roles as a nutrient source and an inducer of the hemolytic phospholipase C, PlcH. The multiple functions for glycine betaine suggested that the cytoplasmic pool of glycine betaine is regulated in P. aeruginosa. We used 13C nuclear magnetic resonance (13C-NMR) to demonstrate that P. aeruginosa maintains both choline and glycine betaine pools under a variety of conditions, in contrast to the transient glycine betaine pool reported for most bacteria. We were able to experimentally manipulate the choline and glycine betaine pools by overexpression of the cognate catabolic genes. Depletion of either the choline or glycine betaine pool reduced phospholipase production, a result unexpected for choline depletion. Depletion of the glycine betaine pool, but not the choline pool, inhibited growth under conditions of high salt with glucose as the primary carbon source. Depletion of the choline pool inhibited growth under high-salt conditions with choline as the sole carbon source, suggesting a role for the choline pool under these conditions. Here we have described the presence of a choline pool in P. aeruginosa and other pseudomonads that, with the glycine betaine pool, regulates osmoprotection and phospholipase production and impacts growth under high-salt conditions. These findings suggest that the levels of both pools are actively maintained and that perturbation of either pool impacts P. aeruginosa physiology. PMID:22753069

ENCoRE: an efficient software for CRISPR screens identifies new players in extrinsic apoptosis.

PubMed

Trümbach, Dietrich; Pfeiffer, Susanne; Poppe, Manuel; Scherb, Hagen; Doll, Sebastian; Wurst, Wolfgang; Schick, Joel A

2017-11-25

As CRISPR/Cas9 mediated screens with pooled guide libraries in somatic cells become increasingly established, an unmet need for rapid and accurate companion informatics tools has emerged. We have developed a lightweight and efficient software to easily manipulate large raw next generation sequencing datasets derived from such screens into informative relational context with graphical support. The advantages of the software entitled ENCoRE (Easy NGS-to-Gene CRISPR REsults) include a simple graphical workflow, platform independence, local and fast multithreaded processing, data pre-processing and gene mapping with custom library import. We demonstrate the capabilities of ENCoRE to interrogate results from a pooled CRISPR cellular viability screen following Tumor Necrosis Factor-alpha challenge. The results not only identified stereotypical players in extrinsic apoptotic signaling but two as yet uncharacterized members of the extrinsic apoptotic cascade, Smg7 and Ces2a. We further validated and characterized cell lines containing mutations in these genes against a panel of cell death stimuli and involvement in p53 signaling. In summary, this software enables bench scientists with sensitive data or without access to informatic cores to rapidly interpret results from large scale experiments resulting from pooled CRISPR/Cas9 library screens.

The role of the megagametophyte in maintaining loblolly pine (Pinus taeda L.) seedling arginase gene expression in vitro.

PubMed

Todd, Christopher D; Gifford, David J

2002-05-01

Following loblolly pine (Pinus taeda L.) seed germination, storage-protein breakdown in the megagametophyte and in the seedling results in a large increase in the seedling's free amino acid pool. A substantial portion of both the storage proteins and the amino acid pool is arginine, a very efficient nitrogen-storage compound. Free arginine is hydrolyzed in the seedling by the enzyme arginase (EC 3.5.3.1), which is under strong developmental control. At present, regulation of arginase in conifers is not well understood. Here we report the utilization of an in vitro culture system to address the separate impacts of the seedling and megagametophyte tissues on arginase enzyme activity, protein levels and patterns of gene expression. We also describe the generation of an anti-arginase antibody prepared from a histidine-tagged loblolly pine arginase fusion protein expressed in Escherichia coli. Our results indicate that arginase gene expression in the seedling is initiated by the seedling itself and then maintained or up-regulated by the megagametophyte. The contribution of storage-protein breakdown and the free amino acid pool, particularly arginine, in this regulation is also addressed.

Genetics of resistance to the geminivirus, Bean dwarf mosaic virus, and the role of the hypersensitive response in common bean.

PubMed

Seo, Y-S; Gepts, P; Gilbertson, R L

2004-03-01

Bean dwarf mosaic virus (BDMV) is a single-stranded DNA virus (genus: Begomovirus, family: Geminiviridae) that infects common bean ( Phaseolus vulgaris L.) and causes stunted plant growth, and mosaic and mottle symptoms in leaves. BDMV shows differential pathogenicity in common bean, infecting germplasm of the Andean gene pool (e.g., the snap bean cultivar Topcrop), but not that of the Middle American gene pool (e.g., the pinto bean cultivar Othello). Resistance to BDMV in Othello is associated with development of a hypersensitive response (HR) in vascular (phloem) tissues. In this study, Middle American germplasm representing the four recognized races (i.e., Durango, Guatemala, Jalisco, and Mesoamerica) and the parents of Othello were inoculated with BDMV and a BDMV-green fluorescent protein (GFP) reporter. All genotypes showed partial or complete resistance to BDMV and BDMV-GFP, indicating the widespread distribution of resistance in the Middle American gene pool. A number of BDMV-resistant germplasm did not show the HR, indicating it is not correlated with resistance. In the F(1), F(2), and F(3) of reciprocal crosses between Othello and Topcrop, a single dominant allele, Bdm, conferred BDMV resistance.

Nanobiotechnologies in the System of Farm Animals' Gene Pool Preservation

NASA Astrophysics Data System (ADS)

Kovtun, S. I.; Galagan, N. P.; Shcherbak, O. V.; Klymenko, N. Y.; Osypchuk, O. S.

It has been determined that sperm viability and mobility upon long storage in liquid nitrogen is being reduced. This issue is relevant for gene pool objects' preservation and for reproductive medicine. The use of nanomaterials (NM) in reproductive biotechnology can expand to the methodology of the rational gene pool preservation technology, especially of valuable biomaterial. Use of nanoparticles, based on ultrafine silica (UFS) with different agents located on its surface gives a positive effect on the thawed sperm mobility, increasing its activity and survival rate and leading to increased fertilization ability. After thawing, the sperm of bulls had an average activity level of 47 %. This activity index of gametes in control was being maintained for 30 min. In the experimental groups after 30 min, the most active gametes were those with UFS + sucrose (52 %). A lower activity compared with the control and UFS + sucrose had gametes with UFS + D-galactosamine. Sperm activity with UFS + D-galactosamine decreased by 14 % compared with the control and by 19 % compared to the UFS + sucrose. So, the feasibility of adding NM with UFS + sucrose at a concentration of 0.001 % to thawed bull sperm that has been cryopreserved for a long time was proved.

Transcriptional programming during cell wall maturation in the expanding Arabidopsis stem.

PubMed

Hall, Hardy; Ellis, Brian

2013-01-25

Plant cell walls are complex dynamic structures that play a vital role in coordinating the directional growth of plant tissues. The rapid elongation of the inflorescence stem in the model plant Arabidopsis thaliana is accompanied by radical changes in cell wall structure and chemistry, but analysis of the underlying mechanisms and identification of the genes that are involved has been hampered by difficulties in accurately sampling discrete developmental states along the developing stem. By creating stem growth kinematic profiles for individual expanding Arabidopsis stems we have been able to harvest and pool developmentally-matched tissue samples, and to use these for comparative analysis of global transcript profiles at four distinct phases of stem growth: the period of elongation rate increase, the point of maximum growth rate, the point of stem growth cessation and the fully matured stem. The resulting profiles identify numerous genes whose expression is affected as the stem tissues pass through these defined growth transitions, including both novel loci and genes identified in earlier studies. Of particular note is the preponderance of highly active genes associated with secondary cell wall deposition in the region of stem growth cessation, and of genes associated with defence and stress responses in the fully mature stem. The use of growth kinematic profiling to create tissue samples that are accurately positioned along the expansion growth continuum of Arabidopsis inflorescence stems establishes a new standard for transcript profiling analyses of such tissues. The resulting expression profiles identify a substantial number of genes whose expression is correlated for the first time with rapid cell wall extension and subsequent fortification, and thus provide an important new resource for plant biologists interested in gene discovery related to plant biomass accumulation.

Allele mining in the gene pool of wild Solanum species for homologues of late blight resistance gene RB/Rpi-blb1

USDA-ARS?s Scientific Manuscript database

Solanum bulbocastanum comprising a CC-NBS-LRR gene RB/Rpi-blb1 confers broad-spectrum resistance to Phytophthora infestans and is currently employed in potato breeding for durable late blight (LB) resistance. Genomes of several Solanum species were reported to contain RB homologues with confirmed b...

Large-scale gene expression profiling data for the model moss Physcomitrella patens aid understanding of developmental progression, culture and stress conditions.

PubMed

Hiss, Manuel; Laule, Oliver; Meskauskiene, Rasa M; Arif, Muhammad A; Decker, Eva L; Erxleben, Anika; Frank, Wolfgang; Hanke, Sebastian T; Lang, Daniel; Martin, Anja; Neu, Christina; Reski, Ralf; Richardt, Sandra; Schallenberg-Rüdinger, Mareike; Szövényi, Peter; Tiko, Theodhor; Wiedemann, Gertrud; Wolf, Luise; Zimmermann, Philip; Rensing, Stefan A

2014-08-01

The moss Physcomitrella patens is an important model organism for studying plant evolution, development, physiology and biotechnology. Here we have generated microarray gene expression data covering the principal developmental stages, culture forms and some environmental/stress conditions. Example analyses of developmental stages and growth conditions as well as abiotic stress treatments demonstrate that (i) growth stage is dominant over culture conditions, (ii) liquid culture is not stressful for the plant, (iii) low pH might aid protoplastation by reduced expression of cell wall structure genes, (iv) largely the same gene pool mediates response to dehydration and rehydration, and (v) AP2/EREBP transcription factors play important roles in stress response reactions. With regard to the AP2 gene family, phylogenetic analysis and comparison with Arabidopsis thaliana shows commonalities as well as uniquely expressed family members under drought, light perturbations and protoplastation. Gene expression profiles for P. patens are available for the scientific community via the easy-to-use tool at https://www.genevestigator.com. By providing large-scale expression profiles, the usability of this model organism is further enhanced, for example by enabling selection of control genes for quantitative real-time PCR. Now, gene expression levels across a broad range of conditions can be accessed online for P. patens. © 2014 The Authors The Plant Journal © 2014 John Wiley & Sons Ltd.

Next generation semiconductor based-sequencing of a nutrigenetics target gene (GPR120) and association with growth rate in Italian Large White pigs.

PubMed

Fontanesi, Luca; Bertolini, Francesca; Scotti, Emilio; Schiavo, Giuseppina; Colombo, Michela; Trevisi, Paolo; Ribani, Anisa; Buttazzoni, Luca; Russo, Vincenzo; Dall'Olio, Stefania

2015-01-01

The GPR120 gene (also known as FFAR4 or O3FAR1) encodes for a functional omega-3 fatty acid receptor/sensor that mediates potent insulin sensitizing effects by repressing macrophage-induced tissue inflammation. For its functional role, GPR120 could be considered a potential target gene in animal nutrigenetics. In this work we resequenced the porcine GPR120 gene by high throughput Ion Torrent semiconductor sequencing of amplified fragments obtained from 8 DNA pools derived, on the whole, from 153 pigs of different breeds/populations (two Italian Large White pools, Italian Duroc, Italian Landrace, Casertana, Pietrain, Meishan, and wild boars). Three single nucleotide polymorphisms (SNPs), two synonymous substitutions and one in the putative 3'-untranslated region (g.114765469C > T), were identified and their allele frequencies were estimated by sequencing reads count. The g.114765469C > T SNP was also genotyped by PCR-RFLP confirming estimated frequency in Italian Large White pools. Then, this SNP was analyzed in two Italian Large White cohorts using a selective genotyping approach based on extreme and divergent pigs for back fat thickness (BFT) estimated breeding value (EBV) and average daily gain (ADG) EBV. Significant differences of allele and genotype frequencies distribution was observed between the extreme ADG-EBV groups (P < 0.001) whereas this marker was not associated with BFT-EBV.

Genes expressed during the development and ripening of watermelon fruit.

PubMed

Levi, A; Davis, A; Hernandez, A; Wechter, P; Thimmapuram, J; Trebitsh, T; Tadmor, Y; Katzir, N; Portnoy, V; King, S

2006-11-01

A normalized cDNA library was constructed using watermelon flesh mRNA from three distinct developmental time-points and was subtracted by hybridization with leaf cDNA. Random cDNA clones of the watermelon flesh subtraction library were sequenced from the 5' end in order to identify potentially informative genes associated with fruit setting, development, and ripening. One-thousand and forty-six 5'-end sequences (expressed sequence tags; ESTs) were assembled into 832 non-redundant sequences, designated as "EST-unigenes". Of these 832 "EST-unigenes", 254 ( approximately 30%) have no significant homology to sequences published so far for other plant species. Additionally, 168 "EST-unigenes" ( approximately 20%) correspond to genes with unknown function, whereas 410 "EST-unigenes" ( approximately 50%) correspond to genes with known function in other plant species. These "EST-unigenes" are mainly associated with metabolism, membrane transport, cytoskeleton synthesis and structure, cell wall formation and cell division, signal transduction, nucleic acid binding and transcription factors, defense and stress response, and secondary metabolism. This study provides the scientific community with novel genetic information for watermelon as well as an expanded pool of genes associated with fruit development in watermelon. These genes will be useful targets in future genetic and functional genomic studies of watermelon and its development.

Colonia Tovar: the history of a semi-isolated Venezuelan population of German ancestry described by HLA class I genes.

PubMed

Gendzekhadze, K; Montagnani, S; Ogando, V; Balbas, O; Mendez-Castellano, H; Layrisse, Z

2003-11-01

The history of Colonia Tovar is very complex, being the home of descendants of only a small fraction of immigrants arriving to the South American continent from a specific region of Germany, with a restricted number of founders, small population size and consanguineous mating, experiencing isolation for 100 years, with later migrations, a low rate of population growth and a high mean number of children per couple. How complex is its genetic structure? Do the highly polymorphic HLA genes reflect its history and confirm the story of this population described by other genes? Several studies have been made in this population, but we describe for the first time the HLA Class I variability in the population of Colonia Tovar using PCR-SSOP. Random genetic drift, founder effect and gene flow could explain the HLA allele and haplotype frequencies observed in this population but alleles at the class I loci were insufficient to identify the German origin of the community established through history. This agrees with findings obtained testing other genetic systems (ACP, AK, ESD, G6PD, GLO, PGM, PGD, ALB, CP, HP, TF), but the HLA-typing results indicate that the original gene pool has been diluted due to gene flow from the surrounding Mestizo population.

Advances in Arachis genomics for peanut improvement

USDA-ARS?s Scientific Manuscript database

Peanut genomics is very challenging due to its inherent problem of genetic architecture. Blockage of gene flow from diploid wild relatives to tetraploid cultivated peanut, recent polyploidization combined with self pollination and narrow genetic base of primary gene pool resulted in low genetic dive...

Population genetic structure of a California endemic Branchiopod, Branchinecta sandiegonensis

USGS Publications Warehouse

Davies, Cathleen P.; Simovich, Marie A.; Hathaway, Stacie A.

1997-01-01

Branchinecta sandiegonensis (Crustacea: Anostraca) is a narrow range endemic fairy shrimp discontinuously distributed in ephemeral pools on coastal mesas in San Diego County, USA. Ten populations across the range of the species were subjected to allozyme analysis for eleven loci. The species exhibits low variability (P95 =9.1–45.5) and one third of the loci tested did not conform to Hardy-Weinberg equilibrium expectations. The species also exhibited a high degree of genetic differentiation between populations. F ST values (fixation index) for most pairs of populations were above 0.25 (0.036–0.889).Low genetic variability and high genetic structure may result from low gene flow and founder effects due to habitat fragmentation and the lack of potential vectors for cyst dispersal. The unpredictable rainfall of the region also creates potential for variable population sizes which could affect structure and variability.

Magic Pools: Parallel Assessment of Transposon Delivery Vectors in Bacteria

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

Liu, Hualan; Price, Morgan N.; Waters, Robert Jordan

Transposon mutagenesis coupled to next-generation sequencing (TnSeq) is a powerful approach for discovering the functions of bacterial genes. However, the development of a suitable TnSeq strategy for a given bacterium can be costly and time-consuming. To meet this challenge, we describe a part-based strategy for constructing libraries of hundreds of transposon delivery vectors, which we term “magic pools.” Within a magic pool, each transposon vector has a different combination of upstream sequences (promoters and ribosome binding sites) and antibiotic resistance markers as well as a random DNA barcode sequence, which allows the tracking of each vector during mutagenesis experiments. Tomore » identify an efficient vector for a given bacterium, we mutagenize it with a magic pool and sequence the resulting insertions; we then use this efficient vector to generate a large mutant library. We used the magic pool strategy to construct transposon mutant libraries in five genera of bacteria, including three genera of the phylumBacteroidetes. IMPORTANCEMolecular genetics is indispensable for interrogating the physiology of bacteria. However, the development of a functional genetic system for any given bacterium can be time-consuming. Here, we present a streamlined approach for identifying an effective transposon mutagenesis system for a new bacterium. Our strategy first involves the construction of hundreds of different transposon vector variants, which we term a “magic pool.” The efficacy of each vector in a magic pool is monitored in parallel using a unique DNA barcode that is introduced into each vector design. Using archived DNA “parts,” we next reassemble an effective vector for making a whole-genome transposon mutant library that is suitable for large-scale interrogation of gene function using competitive growth assays. Here, we demonstrate the utility of the magic pool system to make mutant libraries in five genera of bacteria.« less

Magic Pools: Parallel Assessment of Transposon Delivery Vectors in Bacteria

DOE PAGES

Liu, Hualan; Price, Morgan N.; Waters, Robert Jordan; ...

2018-01-16

Transposon mutagenesis coupled to next-generation sequencing (TnSeq) is a powerful approach for discovering the functions of bacterial genes. However, the development of a suitable TnSeq strategy for a given bacterium can be costly and time-consuming. To meet this challenge, we describe a part-based strategy for constructing libraries of hundreds of transposon delivery vectors, which we term “magic pools.” Within a magic pool, each transposon vector has a different combination of upstream sequences (promoters and ribosome binding sites) and antibiotic resistance markers as well as a random DNA barcode sequence, which allows the tracking of each vector during mutagenesis experiments. Tomore » identify an efficient vector for a given bacterium, we mutagenize it with a magic pool and sequence the resulting insertions; we then use this efficient vector to generate a large mutant library. We used the magic pool strategy to construct transposon mutant libraries in five genera of bacteria, including three genera of the phylumBacteroidetes. IMPORTANCEMolecular genetics is indispensable for interrogating the physiology of bacteria. However, the development of a functional genetic system for any given bacterium can be time-consuming. Here, we present a streamlined approach for identifying an effective transposon mutagenesis system for a new bacterium. Our strategy first involves the construction of hundreds of different transposon vector variants, which we term a “magic pool.” The efficacy of each vector in a magic pool is monitored in parallel using a unique DNA barcode that is introduced into each vector design. Using archived DNA “parts,” we next reassemble an effective vector for making a whole-genome transposon mutant library that is suitable for large-scale interrogation of gene function using competitive growth assays. Here, we demonstrate the utility of the magic pool system to make mutant libraries in five genera of bacteria.« less

Molecular polymorphisms of the ABO locus as informative markers of ancestry in Central Argentina.

PubMed

Tavella, María Pía; García, Angelina; Pauro, Maia; Demarchi, Darío A; Nores, Rodrigo

2017-07-08

The aim of this study was to investigate the distribution of molecular polymorphisms of the ABO gene in four population samples from the province of Córdoba, in Central Argentina, and to compare them with other worldwide populations. A total of 110 buccal swab samples from autochthonous individuals of Córdoba were typified. Molecular characterization of the allelic variants was performed by the analysis of exons 6 and 7 of the ABO gene using PCR-RFLP analysis. Additionally, the Native American AIM O1v542 was characterized by direct sequencing. The four Córdoba populations did not show significant geographic structure, although the frequency of the O1v542 haplotype, detected in all the populations studied, ranged from 0.019 to 0.222. The principal component analysis based on O allele distribution showed that the populations from Córdoba clustered close to the admixed populations of Santiago and Mexico City, and at intermediate distances between European and Native American populations, while being distant from the African population. The results demonstrate that the analysis of the ABO system constitutes a useful tool for the study of the genetic structure and evolutionary history of human populations, reflecting accurately the relative contribution of parental continental contribution to the gene pool of admixed populations. © 2017 Wiley Periodicals, Inc.

High-throughput, pooled sequencing identifies mutations in NUBPL and FOXRED1 in human complex I deficiency

PubMed Central

Calvo, Sarah E; Tucker, Elena J; Compton, Alison G; Kirby, Denise M; Crawford, Gabriel; Burtt, Noel P; Rivas, Manuel A; Guiducci, Candace; Bruno, Damien L; Goldberger, Olga A; Redman, Michelle C; Wiltshire, Esko; Wilson, Callum J; Altshuler, David; Gabriel, Stacey B; Daly, Mark J; Thorburn, David R; Mootha, Vamsi K

2010-01-01

Discovering the molecular basis of mitochondrial respiratory chain disease is challenging given the large number of both mitochondrial and nuclear genes involved. We report a strategy of focused candidate gene prediction, high-throughput sequencing, and experimental validation to uncover the molecular basis of mitochondrial complex I (CI) disorders. We created five pools of DNA from a cohort of 103 patients and then performed deep sequencing of 103 candidate genes to spotlight 151 rare variants predicted to impact protein function. We used confirmatory experiments to establish genetic diagnoses in 22% of previously unsolved cases, and discovered that defects in NUBPL and FOXRED1 can cause CI deficiency. Our study illustrates how large-scale sequencing, coupled with functional prediction and experimental validation, can reveal novel disease-causing mutations in individual patients. PMID:20818383

Ion Torrent sequencing as a tool for mutation discovery in the flax (Linum usitatissimum L.) genome.

PubMed

Galindo-González, Leonardo; Pinzón-Latorre, David; Bergen, Erik A; Jensen, Dustin C; Deyholos, Michael K

2015-01-01

Detection of induced mutations is valuable for inferring gene function and for developing novel germplasm for crop improvement. Many reverse genetics approaches have been developed to identify mutations in genes of interest within a mutagenized population, including some approaches that rely on next-generation sequencing (e.g. exome capture, whole genome resequencing). As an alternative to these genome or exome-scale methods, we sought to develop a scalable and efficient method for detection of induced mutations that could be applied to a small number of target genes, using Ion Torrent technology. We developed this method in flax (Linum usitatissimum), to demonstrate its utility in a crop species. We used an amplicon-based approach in which DNA samples from an ethyl methanesulfonate (EMS)-mutagenized population were pooled and used as template in PCR reactions to amplify a region of each gene of interest. Barcodes were incorporated during PCR, and the pooled amplicons were sequenced using an Ion Torrent PGM. A pilot experiment with known SNPs showed that they could be detected at a frequency > 0.3% within the pools. We then selected eight genes for which we wanted to discover novel mutations, and applied our approach to screen 768 individuals from the EMS population, using either the Ion 314 or Ion 316 chips. Out of 29 potential mutations identified after processing the NGS reads, 16 mutations were confirmed using Sanger sequencing. The methodology presented here demonstrates the utility of Ion Torrent technology in detecting mutation variants in specific genome regions for large populations of a species such as flax. The methodology could be scaled-up to test >100 genes using the higher capacity chips now available from Ion Torrent.

Sex Bias and Maternal Contribution to Gene Expression Divergence in Drosophila Blastoderm Embryos

PubMed Central

Paris, Mathilde; Villalta, Jacqueline E.; Eisen, Michael B.; Lott, Susan E.

2015-01-01

Early embryogenesis is a unique developmental stage where genetic control of development is handed off from mother to zygote. Yet the contribution of this transition to the evolution of gene expression is poorly understood. Here we study two aspects of gene expression specific to early embryogenesis in Drosophila: sex-biased gene expression prior to the onset of canonical X chromosomal dosage compensation, and the contribution of maternally supplied mRNAs. We sequenced mRNAs from individual unfertilized eggs and precisely staged and sexed blastoderm embryos, and compared levels between D. melanogaster, D. yakuba, D. pseudoobscura and D. virilis. First, we find that mRNA content is highly conserved for a given stage and that studies relying on pooled embryos likely systematically overstate the degree of gene expression divergence. Unlike studies done on larvae and adults where most species show a larger proportion of genes with male-biased expression, we find that transcripts in Drosophila embryos are largely female-biased in all species, likely due to incomplete dosage compensation prior to the activation of the canonical dosage compensation mechanism. The divergence of sex-biased gene expression across species is observed to be often due to lineage-specific decrease of expression; the most drastic example of which is the overall reduction of male expression from the neo-X chromosome in D. pseudoobscura, leading to a pervasive female-bias on this chromosome. We see no evidence for a faster evolution of expression on the X chromosome in embryos (no “faster-X” effect), unlike in adults, and contrary to a previous study on pooled non-sexed embryos. Finally, we find that most genes are conserved in regard to their maternal or zygotic origin of transcription, and present evidence that differences in maternal contribution to the blastoderm transcript pool may be due to species-specific divergence of transcript degradation rates. PMID:26485701

Genomic Footprints in Selected and Unselected Beef Cattle Breeds in Korea.

PubMed

Lim, Dajeong; Strucken, Eva M; Choi, Bong Hwan; Chai, Han Ha; Cho, Yong Min; Jang, Gul Won; Kim, Tae-Hun; Gondro, Cedric; Lee, Seung Hwan

2016-01-01

Korean Hanwoo cattle have been subjected to intensive artificial selection over the past four decades to improve meat production traits. Another three cattle varieties very closely related to Hanwoo reside in Korea (Jeju Black and Brindle) and in China (Yanbian). These breeds have not been part of a breeding scheme to improve production traits. Here, we compare the selected Hanwoo against these similar but presumed to be unselected populations to identify genomic regions that have been under recent selection pressure due to the breeding program. Rsb statistics were used to contrast the genomes of Hanwoo versus a pooled sample of the three unselected population (UN). We identified 37 significant SNPs (FDR corrected) in the HW/UN comparison and 21 known protein coding genes were within 1 MB to the identified SNPs. These genes were previously reported to affect traits important for meat production (14 genes), reproduction including mammary gland development (3 genes), coat color (2 genes), and genes affecting behavioral traits in a broader sense (2 genes). We subsequently sequenced (Illumina HiSeq 2000 platform) 10 individuals of the brown Hanwoo and the Chinese Yanbian to identify SNPs within the candidate genomic regions. Based on allele frequency differences, haplotype structures, and literature research, we singled out one non-synonymous SNP in the APP gene (APP: c.569C>T, Ala199Val) and predicted the mutational effect on the protein structure. We found that protein-protein interactions might be impaired due to increased exposed hydrophobic surfaces of the mutated protein. The APP gene has also been reported to affect meat tenderness in pigs and obesity in humans. Meat tenderness has been linked to intramuscular fat content, which is one of the main breeding goals for brown Hanwoo, potentially supporting a causal influence of the herein described nsSNP in the APP gene.

RESISTANCE OF EARTH BANK AGAINST TSUNAMI AND STRUCTURE OF DUG POOL FORMED BY TSUNAMI IN THE 2011 OFF THE PACIFIC COAST OF TOHOKU EARTHQUAKE

NASA Astrophysics Data System (ADS)

Tokida, Ken-Ichi; Tanimoto, Ryusuke

In the 2011 Off the Pacific Coast of Tohoku Earthquake, very huge damages of civil engineering structures etc. occurred by tsunami strikes along the rias coast and plane coast of the Pacific Ocean. For the urgent repair and the future reconstruction of these structures, the fundamental damage characteristics of these structures should be clarified by the field surveys from which effective lessons can be expected. In this paper, several important lessons on the resistance characteristics of 13 earth structures such as river dykes and sand banks which are obtained from the field surveys conducted by the authors are indicated. Because many dug pools were formed by the tsunami overflow at the backside of earth embankments and sea walls in this earthquake, the fundamental characteristics of the 10 dug pools are investigated thorough the field survey to estimate the effects of the dug pools quantitatively in the future. Furthermore through the field survey conducted at the representative site named Idoura, the scale and waterbed conditions of the natural canals and the strength characteristics of the river dykes and the base ground neighboring the natural canals are measured in detail and discussed. These fundamental lessons and data on the earth banks and dug pools will be able to be used to simulate the effects of the dug pools and to discuss the artificial canals as one of the hard countermeasures to reduce the tsunami height and/or force.

Comparing the Performance of Three Land Models in Global C Cycle Simulations: A Detailed Structural Analysis: Structural Analysis of Land Models

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

Rafique, Rashid; Xia, Jianyang; Hararuk, Oleksandra

Land models are valuable tools to understand the dynamics of global carbon (C) cycle. Various models have been developed and used for predictions of future C dynamics but uncertainties still exist. Diagnosing the models’ behaviors in terms of structures can help to narrow down the uncertainties in prediction of C dynamics. In this study three widely used land surface models, namely CSIRO’s Atmosphere Biosphere Land Exchange (CABLE) with 9 C pools, Community Land Model (version 3.5) combined with Carnegie-Ames-Stanford Approach (CLM-CASA) with 12 C pools and Community Land Model (version 4) (CLM4) with 26 C pools were driven by themore » observed meteorological forcing. The simulated C storage and residence time were used for analysis. The C storage and residence time were computed globally for all individual soil and plant pools, as well as net primary productivity (NPP) and its allocation to different plant components’ based on these models. Remotely sensed NPP and statistically derived HWSD, and GLC2000 datasets were used as a reference to evaluate the performance of these models. Results showed that CABLE exhibited better agreement with referenced C storage and residence time for plant and soil pools, as compared with CLM-CASA and CLM4. CABLE had longer bulk residence time for soil C pools and stored more C in roots, whereas, CLM-CASA and CLM4 stored more C in woody pools due to differential NPP allocation. Overall, these results indicate that the differences in C storage and residence times in three models are largely due to the differences in their fundamental structures (number of C pools), NPP allocation and C transfer rates. Our results have implications in model development and provide a general framework to explain the bias/uncertainties in simulation of C storage and residence times from the perspectives of model structures.« less

Integrated platform for genome-wide screening and construction of high-density genetic interaction maps in mammalian cells

PubMed Central

Kampmann, Martin; Bassik, Michael C.; Weissman, Jonathan S.

2013-01-01

A major challenge of the postgenomic era is to understand how human genes function together in normal and disease states. In microorganisms, high-density genetic interaction (GI) maps are a powerful tool to elucidate gene functions and pathways. We have developed an integrated methodology based on pooled shRNA screening in mammalian cells for genome-wide identification of genes with relevant phenotypes and systematic mapping of all GIs among them. We recently demonstrated the potential of this approach in an application to pathways controlling the susceptibility of human cells to the toxin ricin. Here we present the complete quantitative framework underlying our strategy, including experimental design, derivation of quantitative phenotypes from pooled screens, robust identification of hit genes using ultra-complex shRNA libraries, parallel measurement of tens of thousands of GIs from a single double-shRNA experiment, and construction of GI maps. We describe the general applicability of our strategy. Our pooled approach enables rapid screening of the same shRNA library in different cell lines and under different conditions to determine a range of different phenotypes. We illustrate this strategy here for single- and double-shRNA libraries. We compare the roles of genes for susceptibility to ricin and Shiga toxin in different human cell lines and reveal both toxin-specific and cell line-specific pathways. We also present GI maps based on growth and ricin-resistance phenotypes, and we demonstrate how such a comparative GI mapping strategy enables functional dissection of physical complexes and context-dependent pathways. PMID:23739767

Variation in chromosome constitution of the Xiaoyan series partial amphiploids and its relations to stripe rust and stem rust resistance

USDA-ARS?s Scientific Manuscript database

In the tertiary gene pool of wheat, tall wheatgrass Thinopyrum ponticum (2n = 10x = 70) is an excellent source of resistance genes against numerous wheat diseases. The creation of wheat-Th. ponticum partial amphiploids is an intermediate step for transferring the useful genes from Th. ponticum to w...

Seasonal and Cyclical Changes in Genetic Composition of the Marine Intertidal Rock Pool Copepod Tigriopus brevicornis.

PubMed

Van Wormhoudt, Alain

2015-06-01

Tigriopus brevicornis is a marine rock pool copepod widely distributed along Atlantic coasts. Due to the absence of a known dispersal mechanism by free swimming stages, exchanges between populations over long distances are questionable. In order to analyse the evolution of an isolated supralittoral rock pool population, sampling of the copepod was performed monthly during 1 year and compared to samplings over 5 years in the same rock pool, as well as from other rock pools. Using ITS1 analysis, cyclical changes in genetic composition were detected. Our results give clear indications concerning the segregation of the rock pool population and a lack of gene flow among outside populations. A network analysis shows the presence of several shared dominant haplotypes and also singletons differing by one mutation point. F(st) analyses indicate that the main changes occur in autumn and winter. The few analogies of ITS1 sequences with nearby populations may indicate that new migrants must re-colonise the pools from surrounding rock crevices in the intertidal habitat where they may have found a refuge after bad weather conditions.

Making a Splash.

ERIC Educational Resources Information Center

Maland, Jim

1998-01-01

Presents a 10-step process that allows swimming-pool owners to objectively scrutinize their existing facility's needs, construction, and operation and maintenance budgets before renovating a structurally deficient or costly pool. Four examples of problems involving pool-renovation decision making are highlighted. (GR)

Analysis of electric power industry restructuring

NASA Astrophysics Data System (ADS)

Al-Agtash, Salem Yahya

1998-10-01

This thesis evaluates alternative structures of the electric power industry in a competitive environment. One structure is based on the principle of creating a mandatory power pool to foster competition and manage system economics. The structure is PoolCo (pool coordination). A second structure is based on the principle of allowing independent multilateral trading and decentralized market coordination. The structure is DecCo (decentralized coordination). The criteria I use to evaluate these two structures are: economic efficiency, system reliability and freedom of choice. Economic efficiency evaluation considers strategic behavior of individual generators as well as behavioral variations of different classes of consumers. A supply-function equilibria model is characterized for deriving bidding strategies of competing generators under PoolCo. It is shown that asymmetric equilibria can exist within the capacities of generators. An augmented Lagrangian approach is introduced to solve iteratively for global optimal operations schedules. Under DecCo, the process involves solving iteratively for system operations schedules. The schedules reflect generators strategic behavior and brokers' interactions for arranging profitable trades, allocating losses and managing network congestion. In the determination of PoolCo and DecCo operations schedules, overall costs of power generation (start-up and shut-down costs and availability of hydro electric power) as well as losses and costs of transmission network are considered. For system reliability evaluation, I examine the effect of PoolCo and DecCo operating conditions on the system security. Random component failure perturbations are generated to simulate the actual system behavior. This is done using Monte Carlo simulation. Freedom of choice evaluation accounts for schemes' beneficial opportunities and capabilities to respond to consumers expressed preferences. An IEEE 24-bus test system is used to illustrate the concepts developed for economic efficiency evaluation. The system was tested over two years time period. The results indicate 2.6684 and 2.7269 percent of efficiency loss on average for PoolCo and DecCo, respectively. These values, however, do not represent forecasts of efficiency losses of PoolCo- and DecCo-based competitive industries. Rather, they are illustrations of the efficiency losses for the given IEEE test system and based on the modeling assumptions underlying framework development.

"Every Gene Is Everywhere but the Environment Selects": Global Geolocalization of Gene Sharing in Environmental Samples through Network Analysis.

PubMed

Fondi, Marco; Karkman, Antti; Tamminen, Manu V; Bosi, Emanuele; Virta, Marko; Fani, Renato; Alm, Eric; McInerney, James O

2016-05-13

The spatial distribution of microbes on our planet is famously formulated in the Baas Becking hypothesis as "everything is everywhere but the environment selects." While this hypothesis does not strictly rule out patterns caused by geographical effects on ecology and historical founder effects, it does propose that the remarkable dispersal potential of microbes leads to distributions generally shaped by environmental factors rather than geographical distance. By constructing sequence similarity networks from uncultured environmental samples, we show that microbial gene pool distributions are not influenced nearly as much by geography as ecology, thus extending the Bass Becking hypothesis from whole organisms to microbial genes. We find that gene pools are shaped by their broad ecological niche (such as sea water, fresh water, host, and airborne). We find that freshwater habitats act as a gene exchange bridge between otherwise disconnected habitats. Finally, certain antibiotic resistance genes deviate from the general trend of habitat specificity by exhibiting a high degree of cross-habitat mobility. The strong cross-habitat mobility of antibiotic resistance genes is a cause for concern and provides a paradigmatic example of the rate by which genes colonize new habitats when new selective forces emerge. © The Author(s) 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

TRAM (Transcriptome Mapper): database-driven creation and analysis of transcriptome maps from multiple sources

PubMed Central

2011-01-01

Background Several tools have been developed to perform global gene expression profile data analysis, to search for specific chromosomal regions whose features meet defined criteria as well as to study neighbouring gene expression. However, most of these tools are tailored for a specific use in a particular context (e.g. they are species-specific, or limited to a particular data format) and they typically accept only gene lists as input. Results TRAM (Transcriptome Mapper) is a new general tool that allows the simple generation and analysis of quantitative transcriptome maps, starting from any source listing gene expression values for a given gene set (e.g. expression microarrays), implemented as a relational database. It includes a parser able to assign univocal and updated gene symbols to gene identifiers from different data sources. Moreover, TRAM is able to perform intra-sample and inter-sample data normalization, including an original variant of quantile normalization (scaled quantile), useful to normalize data from platforms with highly different numbers of investigated genes. When in 'Map' mode, the software generates a quantitative representation of the transcriptome of a sample (or of a pool of samples) and identifies if segments of defined lengths are over/under-expressed compared to the desired threshold. When in 'Cluster' mode, the software searches for a set of over/under-expressed consecutive genes. Statistical significance for all results is calculated with respect to genes localized on the same chromosome or to all genome genes. Transcriptome maps, showing differential expression between two sample groups, relative to two different biological conditions, may be easily generated. We present the results of a biological model test, based on a meta-analysis comparison between a sample pool of human CD34+ hematopoietic progenitor cells and a sample pool of megakaryocytic cells. Biologically relevant chromosomal segments and gene clusters with differential expression during the differentiation toward megakaryocyte were identified. Conclusions TRAM is designed to create, and statistically analyze, quantitative transcriptome maps, based on gene expression data from multiple sources. The release includes FileMaker Pro database management runtime application and it is freely available at http://apollo11.isto.unibo.it/software/, along with preconfigured implementations for mapping of human, mouse and zebrafish transcriptomes. PMID:21333005

Structure of Escherichia coli dGTP Triphosphohydrolase: A Hexameric Enzyme with DNA Effector Molecules

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

Singh, Deepa; Gawel, Damian; Itsko, Mark

The Escherichia coli dgt gene encodes a dGTP triphosphohydrolase whose detailed role still remains to be determined. Deletion of dgt creates a mutator phenotype, indicating that the dGTPase has a fidelity role, possibly by affecting the cellular dNTP pool. In the present paper, we have investigated the structure of the Dgt protein at 3.1-Å resolution. One of the obtained structures revealed a protein hexamer that contained two molecules of single-stranded DNA. The presence of DNA caused significant conformational changes in the enzyme, including in the catalytic site of the enzyme. Dgt preparations lacking DNA were able to bind single-stranded DNAmore » with high affinity (K d ~ 50 nM). DNA binding positively affected the activity of the enzyme: dGTPase activity displayed sigmoidal (cooperative) behavior without DNA but hyperbolic (Michaelis-Menten) kinetics in its presence, consistent with a specific lowering of the apparent K m for dGTP. A mutant Dgt enzyme was also created containing residue changes in the DNA binding cleft. This mutant enzyme, whereas still active, was incapable of DNA binding and could no longer be stimulated by addition of DNA. We also created an E. coli strain containing the mutant dgt gene on the chromosome replacing the wild-type gene. The mutant also displayed a mutator phenotype. Finally, our results provide insight into the allosteric regulation of the enzyme and support a physiologically important role of DNA binding.« less

Structure of Escherichia coli dGTP Triphosphohydrolase: A Hexameric Enzyme with DNA Effector Molecules

DOE PAGES

Singh, Deepa; Gawel, Damian; Itsko, Mark; ...

2015-02-18

The Escherichia coli dgt gene encodes a dGTP triphosphohydrolase whose detailed role still remains to be determined. Deletion of dgt creates a mutator phenotype, indicating that the dGTPase has a fidelity role, possibly by affecting the cellular dNTP pool. In the present paper, we have investigated the structure of the Dgt protein at 3.1-Å resolution. One of the obtained structures revealed a protein hexamer that contained two molecules of single-stranded DNA. The presence of DNA caused significant conformational changes in the enzyme, including in the catalytic site of the enzyme. Dgt preparations lacking DNA were able to bind single-stranded DNAmore » with high affinity (K d ~ 50 nM). DNA binding positively affected the activity of the enzyme: dGTPase activity displayed sigmoidal (cooperative) behavior without DNA but hyperbolic (Michaelis-Menten) kinetics in its presence, consistent with a specific lowering of the apparent K m for dGTP. A mutant Dgt enzyme was also created containing residue changes in the DNA binding cleft. This mutant enzyme, whereas still active, was incapable of DNA binding and could no longer be stimulated by addition of DNA. We also created an E. coli strain containing the mutant dgt gene on the chromosome replacing the wild-type gene. The mutant also displayed a mutator phenotype. Finally, our results provide insight into the allosteric regulation of the enzyme and support a physiologically important role of DNA binding.« less

The Impact of Sampling Schemes on the Site Frequency Spectrum in Nonequilibrium Subdivided Populations

PubMed Central

Städler, Thomas; Haubold, Bernhard; Merino, Carlos; Stephan, Wolfgang; Pfaffelhuber, Peter

2009-01-01

Using coalescent simulations, we study the impact of three different sampling schemes on patterns of neutral diversity in structured populations. Specifically, we are interested in two summary statistics based on the site frequency spectrum as a function of migration rate, demographic history of the entire substructured population (including timing and magnitude of specieswide expansions), and the sampling scheme. Using simulations implementing both finite-island and two-dimensional stepping-stone spatial structure, we demonstrate strong effects of the sampling scheme on Tajima's D (DT) and Fu and Li's D (DFL) statistics, particularly under specieswide (range) expansions. Pooled samples yield average DT and DFL values that are generally intermediate between those of local and scattered samples. Local samples (and to a lesser extent, pooled samples) are influenced by local, rapid coalescence events in the underlying coalescent process. These processes result in lower proportions of external branch lengths and hence lower proportions of singletons, explaining our finding that the sampling scheme affects DFL more than it does DT. Under specieswide expansion scenarios, these effects of spatial sampling may persist up to very high levels of gene flow (Nm > 25), implying that local samples cannot be regarded as being drawn from a panmictic population. Importantly, many data sets on humans, Drosophila, and plants contain signatures of specieswide expansions and effects of sampling scheme that are predicted by our simulation results. This suggests that validating the assumption of panmixia is crucial if robust demographic inferences are to be made from local or pooled samples. However, future studies should consider adopting a framework that explicitly accounts for the genealogical effects of population subdivision and empirical sampling schemes. PMID:19237689

Monitoring and source tracking of tetracycline resistance genes in lagoons and groundwater adjacent to swine production facilities over a 3-year period

USGS Publications Warehouse

Koike, S.; Krapac, I.G.; Oliver, H.D.; Yannarell, A.C.; Chee-Sanford, J. C.; Aminov, R.I.; Mackie, R.I.

2007-01-01

To monitor the dissemination of resistance genes into the environment, we determined the occurrence of tetracycline resistance (Tcr) genes in groundwater underlying two swine confinement operations. Monitoring well networks (16 wells at site A and 6 wells at site C) were established around the lagoons at each facility. Groundwater (n = 124) and lagoon (n = 12) samples were collected from the two sites at six sampling times from 2000 through 2003. Total DNA was extracted, and PCR was used to detect seven Tcr genes [tet(M), tet(O), tet(Q), tet(W), tet(C), tet(H), and tet(Z)]. The concentration of Tcr genes was quantified by real-time quantitative PCR. To confirm the Tcr gene source in groundwater, comparative analysis of tet(W) gene sequences was performed on groundwater and lagoon samples. All seven Tcr genes were continually detected in groundwater during the 3-year monitoring period at both sites. At site A, elevated detection frequency and concentration of Tcr genes were observed in the wells located down-gradient of the lagoon. Comparative analysis of tet(W) sequences revealed that the impacted groundwater contained gene sequences almost identical (99.8% identity) to those in the lagoon, but these genes were not found in background libraries. Novel sequence clusters and unique indigenous resistance gene pools were also found in the groundwater. Thus, antibiotic resistance genes in groundwater are affected by swine manure, but they are also part of the indigenous gene pool. Copyright ?? 2007, American Society for Microbiology. All Rights Reserved.

Monitoring and Source Tracking of Tetracycline Resistance Genes in Lagoons and Groundwater Adjacent to Swine Production Facilities over a 3-Year Period▿

PubMed Central

Koike, S.; Krapac, I. G.; Oliver, H. D.; Yannarell, A. C.; Chee-Sanford, J. C.; Aminov, R. I.; Mackie, R. I.

2007-01-01

To monitor the dissemination of resistance genes into the environment, we determined the occurrence of tetracycline resistance (Tcr) genes in groundwater underlying two swine confinement operations. Monitoring well networks (16 wells at site A and 6 wells at site C) were established around the lagoons at each facility. Groundwater (n = 124) and lagoon (n = 12) samples were collected from the two sites at six sampling times from 2000 through 2003. Total DNA was extracted, and PCR was used to detect seven Tcr genes [tet(M), tet(O), tet(Q), tet(W), tet(C), tet(H), and tet(Z)]. The concentration of Tcr genes was quantified by real-time quantitative PCR. To confirm the Tcr gene source in groundwater, comparative analysis of tet(W) gene sequences was performed on groundwater and lagoon samples. All seven Tcr genes were continually detected in groundwater during the 3-year monitoring period at both sites. At site A, elevated detection frequency and concentration of Tcr genes were observed in the wells located down-gradient of the lagoon. Comparative analysis of tet(W) sequences revealed that the impacted groundwater contained gene sequences almost identical (99.8% identity) to those in the lagoon, but these genes were not found in background libraries. Novel sequence clusters and unique indigenous resistance gene pools were also found in the groundwater. Thus, antibiotic resistance genes in groundwater are affected by swine manure, but they are also part of the indigenous gene pool. PMID:17545324

Toward an animal model of cystic fibrosis: Targeted interruption of exon 10 of the cystic fibrosis transmembrane regulator gene in embryonic stem cells

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

Koller, B.H.; Hyungsuk Kim; Latour, A.M.

1991-12-01

A gene-targeting construct was made containing 7.8 kilobases of DNA spanning exon 10 of the mouse cystic fibrosis transmembrane regulator (CFTR) gene in which part of the exon has been replaced by two neomycin-resistance (Neo) genes driven by different promoters. (This replacement introduces a chain-termination codon at amino acid position 489 in the CFTR sequence). A herpes simplex thymidine kinase gene was on each end of the construct, which was electroporated into embryonic stem (ES) cells. Colonies resistant to G418, or to G418 plus ganciclovir, were selected and screened by Southern blotting or by PCR amplification. Five pools of G418-resistantmore » cells gave PCR products diagnostic of targeting. Four independent clones of ES cells with a disrupted CFTR gene have been isolated from these pools. The frequency of targeting was 1/2500 G418-resistant colonies. This low frequency is not the consequence of marginal expression of the Neo genes in the targeted cells. The CFTR targeting events were clustered among our experiments in a manner suggesting that some unidentified factor(s), possible passage number, influences the recovery of CFTR-targeted cells.« less

Mating system and gene flow in the red seaweed Gracilaria gracilis: effect of haploid-diploid life history and intertidal rocky shore landscape on fine-scale genetic structure.

PubMed

Engel, C R; Destombe, C; Valero, M

2004-04-01

The impact of haploid-diploidy and the intertidal landscape on a fine-scale genetic structure was explored in a red seaweed Gracilaria gracilis. The pattern of genetic structure was compared in haploid and diploid stages at a microgeographic scale (< 5 km): a total of 280 haploid and 296 diploid individuals located in six discrete, scattered rock pools were genotyped using seven microsatellite loci. Contrary to the theoretical expectation of predominantly endogamous mating systems in haploid-diploid organisms, G. gracilis showed a clearly allogamous mating system. Although within-population allele frequencies were similar between haploids and diploids, genetic differentiation among haploids was more than twice that of diploids, suggesting that there may be a lag between migration and (local) breeding due to the long generation times in G. gracilis. Weak, but significant, population differentiation was detected in both haploids and diploids and varied with landscape features, and not with geographic distance. Using an assignment test, we establish that effective migration rates varied according to height on the shore. In this intertidal species, biased spore dispersal may occur during the transport of spores and gametes at low tide when small streams flow from high- to lower-shore pools. The longevity of both haploid and diploid free-living stages and the long generation times typical of G. gracilis populations may promote the observed pattern of high genetic diversity within populations relative to that among populations.

Myosin Heavy Chain Gene Expression in Developing Neonatal Skeletal Muscle: Involvement of the Nerve, Gravity, and Thyroid State

NASA Technical Reports Server (NTRS)

Baldwin, K. M.; Adams, G.; Haddad, F.; Zeng, M.; Qin, A.; Qin, L.; McCue, S.; Bodell, P.

1999-01-01

The myosin heavy chain (MHC) gene family encodes at least six MHC proteins (herein designated as neonatal, embryonic, slow type I (beta), and fast IIa, IIx, and IIb) that are expressed in skeletal muscle in a muscle-specific and developmentally-regulated fashion. At birth, both antigravity (e.g. soleus) and locomotor (e.g., plantaris) skeletal muscles are undifferentiated relative to the adult MHC phenotype such that the neonatal and embryonic MHC isoforms account for 80 - 90% of the MHC pool in a fast locomotor muscle; whereas, the embryonic and slow, type I isoforms account for approx. 90% of the pool in a typical antigravity muscle. The goal of this study was to investigate the role of an intact nerve, gravity and thyroid hormone (T3), as well as certain interactions of these interventions, on MHC gene expression in developing neonatal skeletal muscles of rodents.

Genome-wide analysis of ivermectin response by Onchocerca volvulus reveals that genetic drift and soft selective sweeps contribute to loss of drug sensitivity

PubMed Central

Nana-Djeunga, Hugues C.; Kengne-Ouafo, Jonas A.; Pion, Sébastien D. S.; Bopda, Jean; Kamgno, Joseph; Wanji, Samuel; Che, Hua; Kuesel, Annette C.; Walker, Martin; Basáñez, Maria-Gloria; Boakye, Daniel A.; Osei-Atweneboana, Mike Y.; Boussinesq, Michel; Prichard, Roger K.; Grant, Warwick N.

2017-01-01

Background Treatment of onchocerciasis using mass ivermectin administration has reduced morbidity and transmission throughout Africa and Central/South America. Mass drug administration is likely to exert selection pressure on parasites, and phenotypic and genetic changes in several Onchocerca volvulus populations from Cameroon and Ghana—exposed to more than a decade of regular ivermectin treatment—have raised concern that sub-optimal responses to ivermectin's anti-fecundity effect are becoming more frequent and may spread. Methodology/Principal findings Pooled next generation sequencing (Pool-seq) was used to characterise genetic diversity within and between 108 adult female worms differing in ivermectin treatment history and response. Genome-wide analyses revealed genetic variation that significantly differentiated good responder (GR) and sub-optimal responder (SOR) parasites. These variants were not randomly distributed but clustered in ~31 quantitative trait loci (QTLs), with little overlap in putative QTL position and gene content between the two countries. Published candidate ivermectin SOR genes were largely absent in these regions; QTLs differentiating GR and SOR worms were enriched for genes in molecular pathways associated with neurotransmission, development, and stress responses. Finally, single worm genotyping demonstrated that geographic isolation and genetic change over time (in the presence of drug exposure) had a significantly greater role in shaping genetic diversity than the evolution of SOR. Conclusions/Significance This study is one of the first genome-wide association analyses in a parasitic nematode, and provides insight into the genomics of ivermectin response and population structure of O. volvulus. We argue that ivermectin response is a polygenically-determined quantitative trait (QT) whereby identical or related molecular pathways but not necessarily individual genes are likely to determine the extent of ivermectin response in different parasite populations. Furthermore, we propose that genetic drift rather than genetic selection of SOR is the underlying driver of population differentiation, which has significant implications for the emergence and potential spread of SOR within and between these parasite populations. PMID:28746337

Complete mitochondrial genome from South American catfish Pseudoplatystoma reticulatum (Eigenmann & Eigenmann) and its impact in Siluriformes phylogenetic tree.

PubMed

Villela, Luciana Cristine Vasques; Alves, Anderson Luis; Varela, Eduardo Sousa; Yamagishi, Michel Eduardo Beleza; Giachetto, Poliana Fernanda; da Silva, Naiara Milagres Augusto; Ponzetto, Josi Margarete; Paiva, Samuel Rezende; Caetano, Alexandre Rodrigues

2017-02-01

The cachara (Pseudoplatystoma reticulatum) is a Neotropical freshwater catfish from family Pimelodidae (Siluriformes) native to Brazil. The species is of relative economic importance for local aquaculture production and basic biological information is under development to help boost efforts to domesticate and raise the species in commercial systems. The complete cachara mitochondrial genome was obtained by assembling Illumina RNA-seq data from pooled samples. The full mitogenome was found to be 16,576 bp in length, showing the same basic structure, order, and genetic organization observed in other Pimelodidae, with 13 protein-coding genes, 2 rNA genes, 22 trNAs, and a control region. Observed base composition was 24.63% T, 28.47% C, 31.45% A, and 15.44% G. With the exception of NAD6 and eight tRNAs, all of the observed mitochondrial genes were found to be coded on the H strand. A total of 107 SNPs were identified in P. reticulatum mtDNA, 67 of which were located in coding regions. Of these SNPs, 10 result in amino acid changes. Analysis of the obtained sequence with 94 publicly available full Siluriformes mitogenomes resulted in a phylogenetic tree that generally agreed with available phylogenetic proposals for the order. The first report of the complete Pseudoplatystoma reticulatum mitochondrial genome sequence revealed general gene organization, structure, content, and order similar to most vertebrates. Specific sequence and content features were observed and may have functional attributes which are now available for further investigation.

Synthesis of customized petroleum-replica fuel molecules by targeted modification of free fatty acid pools in Escherichia coli

PubMed Central

Howard, Thomas P.; Middelhaufe, Sabine; Moore, Karen; Edner, Christoph; Kolak, Dagmara M.; Taylor, George N.; Parker, David A.; Lee, Rob; Smirnoff, Nicholas; Aves, Stephen J.; Love, John

2013-01-01

Biofuels are the most immediate, practical solution for mitigating dependence on fossil hydrocarbons, but current biofuels (alcohols and biodiesels) require significant downstream processing and are not fully compatible with modern, mass-market internal combustion engines. Rather, the ideal biofuels are structurally and chemically identical to the fossil fuels they seek to replace (i.e., aliphatic n- and iso-alkanes and -alkenes of various chain lengths). Here we report on production of such petroleum-replica hydrocarbons in Escherichia coli. The activity of the fatty acid (FA) reductase complex from Photorhabdus luminescens was coupled with aldehyde decarbonylase from Nostoc punctiforme to use free FAs as substrates for alkane biosynthesis. This combination of genes enabled rational alterations to hydrocarbon chain length (Cn) and the production of branched alkanes through upstream genetic and exogenous manipulations of the FA pool. Genetic components for targeted manipulation of the FA pool included expression of a thioesterase from Cinnamomum camphora (camphor) to alter alkane Cn and expression of the branched-chain α-keto acid dehydrogenase complex and β-keto acyl-acyl carrier protein synthase III from Bacillus subtilis to synthesize branched (iso-) alkanes. Rather than simply reconstituting existing metabolic routes to alkane production found in nature, these results demonstrate the ability to design and implement artificial molecular pathways for the production of renewable, industrially relevant fuel molecules. PMID:23610415

High-Throughput Parallel Sequencing to Measure Fitness of Leptospira interrogans Transposon Insertion Mutants during Acute Infection

PubMed Central

Matsunaga, James; Haake, David A.

2016-01-01

Pathogenic species of Leptospira are the causative agents of leptospirosis, a zoonotic disease that causes mortality and morbidity worldwide. The understanding of the virulence mechanisms of Leptospira spp is still at an early stage due to the limited number of genetic tools available for this microorganism. The development of random transposon mutagenesis in pathogenic strains a decade ago has contributed to the identification of several virulence factors. In this study, we used the transposon sequencing (Tn-Seq) technique, which combines transposon mutagenesis with massive parallel sequencing, to study the in vivo fitness of a pool of Leptospira interrogans mutants. We infected hamsters with a pool of 42 mutants (input pool), which included control mutants with insertions in four genes previously analyzed by virulence testing (loa22, ligB, flaA1, and lic20111) and 23 mutants with disrupted signal transduction genes. We quantified the mutants in different tissues (blood, kidney and liver) at 4 days post-challenge by high-throughput sequencing and compared the frequencies of mutants recovered from tissues to their frequencies in the input pool. Control mutants that were less fit in the Tn-Seq experiment were attenuated for virulence when tested separately in the hamster model of lethal leptospirosis. Control mutants with unaltered fitness were as virulent as the wild-type strain. We identified two mutants with the transposon inserted in the same putative adenylate/guanylate cyclase gene (lic12327) that had reduced in vivo fitness in blood, kidney and liver. Both lic12327 mutants were attenuated for virulence when tested individually in hamsters. Growth of the control mutants and lic12327 mutants in culture medium were similar to that of the wild-type strain. These results demonstrate the feasibility of screening large pools of L. interrogans transposon mutants for those with altered fitness, and potentially attenuated virulence, by transposon sequencing. PMID:27824878

High-resolution mapping and sequence analysis of 597 cDNA clones transcribed from the 1 Mb region in human chromosome 4q16.3 containing Huntington disease gene

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

Hadano, S.; Ishida, Y.; Tomiyasu, H.

1994-09-01

To complete a transcription map of the 1 Mb region in human chromosome 4p16.3 containing the Huntington disease (HD) gene, the isolation of cDNA clones are being performed throughout. Our method relies on a direct screening of the cDNA libraries probed with single copy microclones from 3 YAC clones spanning 1 Mbp of the HD gene region. AC-DNAs were isolated by a preparative pulsed-field gel electrophoresis, amplified by both a single unique primer (SUP)-PCR and a linker ligation PCR, and 6 microclone-DNA libraries were generated. Then, 8,640 microclones from these libraries were independently amplified by PCR, and arrayed onto themore » membranes. 800-900 microclones that were not cross-hybridized with total human and yeast genomic DNA, TAC vector DNA, and ribosomal cDNA on a dot hybridization (putatively carrying single copy sequences) were pooled to make 9 probe pools. A total of {approximately}1.8x10{sup 7} plaques from the human brain cDNA libraries was screened with 9 pool-probes, and then 672 positive cDNA clones were obtained. So far, 597 cDNA clones were defined and arrayed onto a map of the 1 Mbp of the HD gene region by hybridization with HD region-specific cosmid contigs and YAC clones. Further characterization including a DNA sequencing and Northern blot analysis is currently underway.« less

A DNA pooling based system to detect Escherichia coli virulence factors in fecal and wastewater samples

PubMed Central

Luz María Chacón, J; Lizeth Taylor, C; Carmen Valiente, A; Irene Alvarado, P; Ximena Cortés, B

2012-01-01

The availability of a useful tool for simple and timely detection of the most important virulent varieties of Escherichia coli is indispensable. To this end, bacterial DNA pools which had previously been categorized were obtained from isolated colonies as well as selected in terms of utilized phenotype; the pools were assessed by two PCR Multiplex for the detection of virulent E. coli eaeA, bfpA, stx1, stx2, ipaH, ST, LT, and aatA genes, with the 16S gene used as DNA control. The system was validated with 66 fecal samples and 44 wastewater samples. At least one positive isolate was detected by a virulent gene among the 20 that were screened. The analysis of fecal samples from children younger than 6 years of age detected frequencies of 25% LT positive strains, 8.3% eae, 8.3% bfpA, 16.7% ipaH, as well as 12.5 % aatA and ST. On the other hand, wastewater samples revealed frequencies of 25.7% eaeA positive, 30.3% stx1, 15.1% LT and 19.7% aatA. This study is an initial step toward carrying out epidemiological field research that will reveal the presence of these bacterial varieties. PMID:24031959

Why barcode? High-throughput multiplex sequencing of mitochondrial genomes for molecular systematics.

PubMed

Timmermans, M J T N; Dodsworth, S; Culverwell, C L; Bocak, L; Ahrens, D; Littlewood, D T J; Pons, J; Vogler, A P

2010-11-01

Mitochondrial genome sequences are important markers for phylogenetics but taxon sampling remains sporadic because of the great effort and cost required to acquire full-length sequences. Here, we demonstrate a simple, cost-effective way to sequence the full complement of protein coding mitochondrial genes from pooled samples using the 454/Roche platform. Multiplexing was achieved without the need for expensive indexing tags ('barcodes'). The method was trialled with a set of long-range polymerase chain reaction (PCR) fragments from 30 species of Coleoptera (beetles) sequenced in a 1/16th sector of a sequencing plate. Long contigs were produced from the pooled sequences with sequencing depths ranging from ∼10 to 100× per contig. Species identity of individual contigs was established via three 'bait' sequences matching disparate parts of the mitochondrial genome obtained by conventional PCR and Sanger sequencing. This proved that assembly of contigs from the sequencing pool was correct. Our study produced sequences for 21 nearly complete and seven partial sets of protein coding mitochondrial genes. Combined with existing sequences for 25 taxa, an improved estimate of basal relationships in Coleoptera was obtained. The procedure could be employed routinely for mitochondrial genome sequencing at the species level, to provide improved species 'barcodes' that currently use the cox1 gene only.

Biodiversity in intertidal rock pools: informing engineering criteria for artificial habitat enhancement in the built environment.

PubMed

Firth, Louise B; Schofield, Meredith; White, Freya J; Skov, Martin W; Hawkins, Stephen J

2014-12-01

Coastal defence structures are proliferating to counter rising and stormier seas. With increasing concern about the ecological value of built environments, efforts are being made to create novel habitat to increase biodiversity. Rock pools are infrequent on artificial structures. We compared biodiversity patterns between rock pools and emergent rock and assessed the role of pool depth and substratum incline in determining patterns of biodiversity. Rock pools were more taxon rich than emergent substrata. Patterns varied with depth and incline with algal groups being more positively associated with shallow than deeper habitats. Substratum incline had little influence on colonising epibiota, with the exception of canopy algae in deeper habitats where vertical surfaces supported greater taxon richness than horizontal surfaces. The creation of artificial rock pools in built environments will have a positive effect on biodiversity. Building pools of varying depths and inclines and shore heights will provide a range of habitats, increase environmental heterogeneity, therefore creating more possible ecological niches, promoting local biodiversity. Copyright © 2014 Elsevier Ltd. All rights reserved.

POOL server: machine learning application for functional site prediction in proteins.

PubMed

Somarowthu, Srinivas; Ondrechen, Mary Jo

2012-08-01

We present an automated web server for partial order optimum likelihood (POOL), a machine learning application that combines computed electrostatic and geometric information for high-performance prediction of catalytic residues from 3D structures. Input features consist of THEMATICS electrostatics data and pocket information from ConCavity. THEMATICS measures deviation from typical, sigmoidal titration behavior to identify functionally important residues and ConCavity identifies binding pockets by analyzing the surface geometry of protein structures. Both THEMATICS and ConCavity (structure only) do not require the query protein to have any sequence or structure similarity to other proteins. Hence, POOL is applicable to proteins with novel folds and engineered proteins. As an additional option for cases where sequence homologues are available, users can include evolutionary information from INTREPID for enhanced accuracy in site prediction. The web site is free and open to all users with no login requirements at http://www.pool.neu.edu. m.ondrechen@neu.edu Supplementary data are available at Bioinformatics online.

Prevalence of tick-borne encephalitis virus in ixodid ticks collected from the republic of Korea during 2011-2012.

PubMed

Yun, Seok-Min; Song, Bong Gu; Choi, Wooyoung; Park, Won Il; Kim, Sung Yun; Roh, Jong Yul; Ryou, Jungsang; Ju, Young Ran; Park, Chan; Shin, E-Hyun

2012-12-01

In this study, we demonstrated that TBEV-infected ticks have been distributed in the ROK, combined with our previous results. These results suggest that TBEV may exist in the ROK, and H. longicornis, H. flava, and I. nipponensis may be potential vectors of TBEV. In addition, these results emphasize the need for further epidemiological research of TBEV. We examined for the presence of RNA of TBEV by reverse transcriptase-nested polymerase chain reaction (RT-nested PCR) using ixodid ticks captured in 25 localities of 10 provinces. Ticks were collected by the flagging and dragging method or using sentinel BG traps at forests, grass thickets, and grassland. A total of 13,053 ticks belonging to two genera and four species were collected and pooled (1292 pools), according to collection site, species of tick, and developmental stage. Among 1292 pools, the envelope (E) protein gene of TBEV was detected using RT-nested PCR in 10 pools (3 pools of the 1,331 adult ticks and 7 pools of the 11,169 nymph ticks) collected from Gangwon-do province, Jeonrabuk-do province, and Jeju Island. The minimum infection rates for TBEV of Haemaphysalis longicornis, Haemaphysalis flava, and Ixodes nipponensis were 0.06%, 0.17%, and 2.38%, respectively. Phylogenetic analysis based on the partial E protein gene was performed to identify relationships between the TBEV strains. This showed that 10 Korean strains clustered with the Western subtype. In this study, we investigated the prevalence of tick-borne encephalitis virus (TBEV) in ixodid ticks from various regions of the Republic of Korea (ROK) during 2011-2012 to identify whether TBEV is circulating and to determine the endemic regions of TBEV.

Genetic and epigenetic mechanisms of gene regulation during lens development

PubMed Central

Cvekl, Ales; Duncan, Melinda K.

2007-01-01

Recent studies demonstrated a number of links between chromatin structure, gene expression, extracellular signaling and cellular differentiation during lens development. Lens progenitor cells originate from a pool of common progenitor cells, the pre-placodal region (PPR) which is formed due to a complex exchange of extracellular signals between the neural plate, naïve ectoderm and mesendoderm. A specific commitment to the lens program over alternate choices such as the formation of olfactory epithelium or the anterior pituitary is manifested by the formation of a thickened surface ectoderm, the lens placode. Mouse lens progenitor cells are characterized by the expression of a complement of lens lineage-specific transcription factors including Pax6, Six3 and Sox2, controlled by FGF and BMP signaling, followed later by c-Maf, Mab21like1, Prox1 and FoxE3. Proliferation of lens progenitors together with their morphogenetic movements results in the formation of the lens vesicle. This transient structure, comprised of lens precursor cells, is polarized with its anterior cells retaining their epithelial morphology and proliferative capacity, whereas the posterior lens precursor cells initiate terminal differentiation forming the primary lens fibers. Lens differentiation is marked by expression and accumulation of crystallins and other structural proteins. The transcriptional control of crystallin genes is characterized by the reiterative use of transcription factors required for the establishment of lens precursors in combination with more ubiquitously expressed factors (e.g. AP-1, AP-2α, CREB and USF) and recruitment of histone acetyltransferases (HATs) CBP and p300, and chromatin remodeling complexes SWI/SNF and ISWI. These studies have poised the study of lens development at the forefront of efforts to understand the connections between development, cell signaling, gene transcription and chromatin remodeling. PMID:17905638

Genetic structure and viability selection in the golden eagle (Aquila chrysaetos), a vagile raptor with a Holarctic distribution

USGS Publications Warehouse

Doyle, Jacqueline M.; Katzner, Todd E.; Roemer, Gary; Cain, James W.; Millsap, Brian; McIntyre, Carol; Sonsthagen, Sarah A.; Fernandez, Nadia B.; Wheeler, Maria; Bulut, Zafer; Bloom, Peter; DeWoody, J. Andrew

2016-01-01

Molecular markers can reveal interesting aspects of organismal ecology and evolution, especially when surveyed in rare or elusive species. Herein, we provide a preliminary assessment of golden eagle (Aquila chrysaetos) population structure in North America using novel single nucleotide polymorphisms (SNPs). These SNPs included one molecular sexing marker, two mitochondrial markers, 85 putatively neutral markers that were derived from noncoding regions within large intergenic intervals, and 74 putatively nonneutral markers found in or very near protein-coding genes. We genotyped 523 eagle samples at these 162 SNPs and quantified genotyping error rates and variability at each marker. Our samples corresponded to 344 individual golden eagles as assessed by unique multilocus genotypes. Observed heterozygosity of known adults was significantly higher than of chicks, as was the number of heterozygous loci, indicating that mean zygosity measured across all 159 autosomal markers was an indicator of fitness as it is associated with eagle survival to adulthood. Finally, we used chick samples of known provenance to test for population differentiation across portions of North America and found pronounced structure among geographic sampling sites. These data indicate that cryptic genetic population structure is likely widespread in the golden eagle gene pool, and that extensive field sampling and genotyping will be required to more clearly delineate management units within North America and elsewhere.

Horizontal gene transfer and mobile genetic elements in marine systems.

PubMed

Sobecky, Patricia A; Hazen, Tracy H

2009-01-01

The pool of mobile genetic elements (MGE) in microbial communities consists of viruses, plasmids, and associated elements (insertion sequences, transposons, and integrons) that are either self-transmissible or use mobile plasmids and viruses as vehicles for their dissemination. This mobilome facilitates the horizontal transfer of genes that promote the evolution and adaptation of microbial communities. Efforts to characterize MGEs from microbial populations resident in a variety of ecological habitats have revealed a surprisingly novel and seemingly untapped biodiversity. To better understand the impact of horizontal gene transfer (HGT), as well as the agents that promote HGT in marine ecosystems and to determine whether or not environmental parameters can effect the composition and structure of the mobilome in marine microbial communities, information on the distribution, diversity, and ecological traits of the marine mobilome is presented. In this chapter we discuss recent insights gained from different methodological approaches used to characterize the biodiversity and ecology of MGE in marine environments and their contributions to HGT. In addition, we present case studies that highlight specific HGT examples in coastal, open-ocean, and deep-sea marine ecosystems.

Fragment structure from vapor explosions during the impact of molten metal droplets into a liquid pool

NASA Astrophysics Data System (ADS)

Kouraytem, Nadia; Li, Er Qiang; Vakarelski, Ivan Uriev; Thoroddsen, Sigurdur

2015-11-01

High-speed video imaging is used in order to look at the impact of a molten metal drop falling into a liquid pool. The interaction regimes are three: film boiling, nucleate boiling or vapor explosion. Following the vapor explosion, the metal fragments and different textures are observed. It was seen that, using a tin alloy, a porous structure results whereas using a distinctive eutectic metal, Field's metal, micro beads are formed. Different parameters such as the metal type, molten metal temperature, pool surface tension and pool boiling temperature have been altered in order to assess the role they play on the explosion dynamics and the molten metal's by product.

Cambrian-Ordovician Knox production in Ohio: Three case studies of structural-stratigraphic traps

USGS Publications Warehouse

Riley, R.A.; Wicks, J.; Thomas, Joan

2002-01-01

The Knox Dolomite (Cambrian-Ordovician) in Ohio consists of a mixed carbonate-siliciclastic sequence deposited in a tidal-flat to shallow-marine environment along a broad continental shelf. Knox hydrocarbon production occurs in porous sandstone and dolomite reservoirs in the Copper Ridge dolomite, Rose Run sandstone, and Beekmantown dolomite. In Ohio, historical Knox exploration and development have been focused on paleogeomorphic traps within the prolific Morrow Consolidated field, and more recently, within and adjacent to the Rose Run subcrop. Although these paleogeomorphic traps have yielded significant Knox production, structural and stratigraphic traps are being largely ignored. Three Knox-producing pools demonstrate structural and stratigraphic traps: the Birmingham-Erie pool in southern Erie and southwestern Lorain counties, the South Canaan pool in northern Wayne County, and the East Randolph pool in south-central Portage County. Enhanced porosity and permeability from fractures, as evident in the East Randolph pool, are also an underexplored mechanism for Knox hydrocarbon accumulation. An estimated 800 bcf of gas from undiscovered Knox resources makes the Knox one of the most attractive plays in the Appalachian basin.

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

Culbert, A.A.; Wallis, G.A.; Kadler, K.E.

The brittleness of bone in people with lethal (type II) osteogenesis imperfecta, a heritable disorder caused by mutations in the type I collagen genes, arises from the deposition of abnormal collagen in the bone matrix. The inability of the abnormal collagen to participate in mineralization may be caused by its failure to interact with other bone proteins. Here, we have designed a strategy to isolate the genes important for mineralization of collagen during bone formation. Cells isolated from 16-day embryonic chick calvaria and seeded post-confluence in culture deposited a mineralized matrix over a period of 2 weeks. Chick skin fibroblastsmore » seeded and cultured under the same conditions did not mineralize. Using RT-PCR, we prepared short cDNAs ({approximately}300 bp) corresponding to the 3{prime} ends of mRNA from fibroblasts and separately from the mineralizing calvarial cells. Subtractive cDNA hybridization generated a pool of cDNAs that were specific to mineralizing calvarial cells but not to fibroblasts. Screening of 100,000 plaques of a chick bone ZAP Express cDNA library with this pool of mineralizing-specific cDNAs identified ten clones which comprised full-length cDNAs for the bone proteins osteopontin (eight of the ten positives), bone sialoprotein II (one of the ten positives), and cystatin (one of the ten positives). cDNAs for type I collagen, fibronectin, alkaline phosphatase, house-keeping genes, and other genes expressed in fibroblasts were not identified in this preliminary screen. The pool of short cDNAs is likely to comprise cDNAs for further bone-specific genes and will be used to screen the entire bone cDNA library of 4.2 million clones. 30 refs., 4 figs.« less

High Temperature Inhibits Ascorbate Recycling and Light Stimulation of the Ascorbate Pool in Tomato despite Increased Expression of Biosynthesis Genes

PubMed Central

Massot, Capucine; Bancel, Doriane; Lopez Lauri, Félicie; Truffault, Vincent; Baldet, Pierre; Stevens, Rebecca; Gautier, Hélène

2013-01-01

Understanding how the fruit microclimate affects ascorbate (AsA) biosynthesis, oxidation and recycling is a great challenge in improving fruit nutritional quality. For this purpose, tomatoes at breaker stage were harvested and placed in controlled environment conditions at different temperatures (12, 17, 23, 27 and 31°C) and irradiance regimes (darkness or 150 µmol m-2 s-1). Fruit pericarp tissue was used to assay ascorbate, glutathione, enzymes related to oxidative stress and the AsA/glutathione cycle and follow the expression of genes coding for 5 enzymes of the AsA biosynthesis pathway (GME, VTC2, GPP, L-GalDH, GLDH). The AsA pool size in pericarp tissue was significantly higher under light at temperatures below 27°C. In addition, light promoted glutathione accumulation at low and high temperatures. At 12°C, increased AsA content was correlated with the enhanced expression of all genes of the biosynthesis pathway studied, combined with higher DHAR and MDHAR activities and increased enzymatic activities related to oxidative stress (CAT and APX). In contrast, at 31°C, MDHAR and GR activities were significantly reduced under light indicating that enzymes of the AsA/glutathione cycle may limit AsA recycling and pool size in fruit pericarp, despite enhanced expression of genes coding for AsA biosynthesis enzymes. In conclusion, this study confirms the important role of fruit microclimate in the regulation of fruit pericarp AsA content, as under oxidative conditions (12°C, light) total fruit pericarp AsA content increased up to 71%. Moreover, it reveals that light and temperature interact to regulate both AsA biosynthesis gene expression in tomato fruits and AsA oxidation and recycling. PMID:24367665

High temperature inhibits ascorbate recycling and light stimulation of the ascorbate pool in tomato despite increased expression of biosynthesis genes.

PubMed

Massot, Capucine; Bancel, Doriane; Lopez Lauri, Félicie; Truffault, Vincent; Baldet, Pierre; Stevens, Rebecca; Gautier, Hélène

2013-01-01

Understanding how the fruit microclimate affects ascorbate (AsA) biosynthesis, oxidation and recycling is a great challenge in improving fruit nutritional quality. For this purpose, tomatoes at breaker stage were harvested and placed in controlled environment conditions at different temperatures (12, 17, 23, 27 and 31 °C) and irradiance regimes (darkness or 150 µmol m(-2) s(-1)). Fruit pericarp tissue was used to assay ascorbate, glutathione, enzymes related to oxidative stress and the AsA/glutathione cycle and follow the expression of genes coding for 5 enzymes of the AsA biosynthesis pathway (GME, VTC2, GPP, L-GalDH, GLDH). The AsA pool size in pericarp tissue was significantly higher under light at temperatures below 27 °C. In addition, light promoted glutathione accumulation at low and high temperatures. At 12 °C, increased AsA content was correlated with the enhanced expression of all genes of the biosynthesis pathway studied, combined with higher DHAR and MDHAR activities and increased enzymatic activities related to oxidative stress (CAT and APX). In contrast, at 31 °C, MDHAR and GR activities were significantly reduced under light indicating that enzymes of the AsA/glutathione cycle may limit AsA recycling and pool size in fruit pericarp, despite enhanced expression of genes coding for AsA biosynthesis enzymes. In conclusion, this study confirms the important role of fruit microclimate in the regulation of fruit pericarp AsA content, as under oxidative conditions (12 °C, light) total fruit pericarp AsA content increased up to 71%. Moreover, it reveals that light and temperature interact to regulate both AsA biosynthesis gene expression in tomato fruits and AsA oxidation and recycling.

Microbiological Analysis in Three Diverse Natural Geothermal Bathing Pools in Iceland

PubMed Central

Thorolfsdottir, Berglind Osk Th.; Marteinsson, Viggo Thor

2013-01-01

Natural thermal bathing pools contain geothermal water that is very popular to bathe in but the water is not sterilized, irradiated or treated in any way. Increasing tourism in Iceland will lead to increasing numbers of bath guests, which can in turn affect the microbial flora in the pools and therefore user safety. Today, there is no legislation that applies to natural geothermal pools in Iceland, as the water is not used for consumption and the pools are not defined as public swimming pools. In this study, we conducted a microbiological analysis on three popular but different natural pools in Iceland, located at Lýsuhóll, Hveravellir and Landmannalaugar. Total bacterial counts were performed by flow cytometry, and with plate count at 22 °C, 37 °C and 50 °C. The presence of viable coliforms, Enterococcus spp. and pseudomonads were investigated by growth experiments on selective media. All samples were screened for noroviruses by real time PCR. The results indicate higher fecal contamination in the geothermal pools where the geothermal water flow was low and bathing guest count was high during the day. The number of cultivated Pseudomonas spp. was high (13,000–40,000 cfu/100 mL) in the natural pools, and several strains were isolated and classified as opportunistic pathogens. Norovirus was not detected in the three pools. DNA was extracted from one-liter samples in each pool and analyzed by partial 16S rRNA gene sequencing. Microbial diversity analysis revealed different microbial communities between the pools and they were primarily composed of alpha-, beta- and gammaproteobacteria. PMID:23493033

Genetic variation in Mediterranean Helichrysum italicum (Asteraceae; Gnaphalieae): do disjunct populations of subsp. microphyllum have a common origin?

PubMed

Galbany-Casals, M; Blanco-Moreno, J M; Garcia-Jacas, N; Breitwieser, I; Smissen, R D

2011-07-01

The yellow-flowered everlasting daisy Helichrysum italicum (Asteraceae, Gnaphalieae) is widely distributed in the Mediterranean basin, where it grows in continuous and widespread populations in diverse open habitats. Helichrysum italicum subsp. microphyllum has a disjunct distribution in the Balearic Islands (Majorca and Dragonera), Corsica, Sardinia, Crete and Cyprus. Numerous morphological intermediates between subsp. italicum and subsp. microphyllum are known from Corsica, where the two subspecies co-occur. The aims of the study were to investigate if subsp. microphyllum has a common origin, constituting an independent gene pool from subsp. italicum, or if the morphological differences between subsp. microphyllum and subsp. italicum have arisen independently in different locations from a common wider gene pool. Our analyses of AFLP, cpDNA sequences and morphological characters show that there is geographic structure to the genetic variation within H. italicum, with eastern and western Mediterranean groups, which do not correspond with the division into subsp. microphyllum and subsp. italicum as currently circumscribed. Local selection on quantitative trait loci provides sufficient explanation for the morphological divergence observed and is consistent with genetic data. Within the western Mediterranean group of the species we found considerable polymorphism in chloroplast DNA sequences among and within some populations. Comparison with chloroplast DNA sequences from other Helichrysum species showed that some chloroplast haplotypes are shared across species. © 2010 German Botanical Society and The Royal Botanical Society of the Netherlands.

Possible roles of uncultured archaea in carbon cycling in methane-seep sediments

NASA Astrophysics Data System (ADS)

Yoshinaga, Marcos Y.; Lazar, Cassandre S.; Elvert, Marcus; Lin, Yu-Shih; Zhu, Chun; Heuer, Verena B.; Teske, Andreas; Hinrichs, Kai-Uwe

2015-09-01

Studies on microbial carbon cycling uniformly confirm that anaerobic methane-oxidizing archaea (ANME) and sulfate-reducing bacteria represent the dominant and most active fraction of the sedimentary microbial community in methane-seep sediments. However, little is known about other frequently observed and abundant microbial taxa, their role in carbon cycling and association with the anaerobic oxidation of methane (AOM). Here, we provide a comprehensive characterization of stable carbon isotopes (δ13C) from several intact polar lipid (IPL) classes and metabolite pools in a downcore profile at a cold seep within the oxygen minimum zone off Pakistan. We aimed to evaluate microbial carbon metabolism using IPLs in relation to redox conditions, metabolites and 16S rRNA gene libraries. The 13C-depleted signature of carbon pools and microbial metabolites in pore waters (e.g., dissolved inorganic carbon, lactate and acetate) demonstrated high accumulation of AOM-associated biomass and subsequent turnover thereof. ANMEs accounted for a small fraction of the archaeal 16S rRNA gene survey, whereas sequences of other uncultured benthic archaea dominated the clone libraries, particularly the Marine Benthic Group D. On the basis of lipid diversity and carbon isotope information, we suggest that structurally diverse phospho- and glycolipids, including the recently identified unsaturated tetraethers that are particularly abundant in this setting, are likely derived from archaea other than ANMEs. Through the evaluation of δ13C values of individual IPL, our results indicate heterotrophy as a possible metabolic pathway of archaea in these AOM-dominated sediments.

17 CFR 229.1113 - (Item 1113) Structure of the transaction.

Code of Federal Regulations, 2011 CFR

2011-04-01

... securities, and within each class, with respect to cash flows, credit enhancement or other support and any... narrative discussion of the allocation and priority structure of pool cash flows, present the flow of funds... any requirements directing cash flows from the pool assets (such as to reserve accounts, cash...

17 CFR 229.1113 - (Item 1113) Structure of the transaction.

Code of Federal Regulations, 2012 CFR

2012-04-01

... securities, and within each class, with respect to cash flows, credit enhancement or other support and any... narrative discussion of the allocation and priority structure of pool cash flows, present the flow of funds... any requirements directing cash flows from the pool assets (such as to reserve accounts, cash...

17 CFR 229.1113 - (Item 1113) Structure of the transaction.

Code of Federal Regulations, 2013 CFR

2013-04-01

... securities, and within each class, with respect to cash flows, credit enhancement or other support and any... narrative discussion of the allocation and priority structure of pool cash flows, present the flow of funds... any requirements directing cash flows from the pool assets (such as to reserve accounts, cash...

17 CFR 229.1113 - (Item 1113) Structure of the transaction.

Code of Federal Regulations, 2014 CFR

2014-04-01

... securities, and within each class, with respect to cash flows, credit enhancement or other support and any... narrative discussion of the allocation and priority structure of pool cash flows, present the flow of funds... any requirements directing cash flows from the pool assets (such as to reserve accounts, cash...

Further evidence of an Amerindian contribution to the Polynesian gene pool on Easter Island.

PubMed

Thorsby, E; Flåm, S T; Woldseth, B; Dupuy, B M; Sanchez-Mazas, A; Fernandez-Vina, M A

2009-06-01

Available evidence suggests a Polynesian origin of the Easter Island population. We recently found that some native Easter Islanders also carried some common American Indian (Amerindian) human leukocyte antigen (HLA) alleles, which probably were introduced before Europeans discovered the island in 1722. In this study, we report molecular genetic investigations of 21 other selected native Easter Islanders. Analysis of mitochondrial DNA and Y chromosome markers showed no traces of an Amerindian contribution. However, high-resolution genomic HLA typing showed that two individuals carried some other common Amerindian HLA alleles, different from those found in our previous investigations. The new data support our previous evidence of an Amerindian contribution to the gene pool on Easter Island.

Method for nonlinear optimization for gas tagging and other systems

DOEpatents

Chen, Ting; Gross, Kenny C.; Wegerich, Stephan

1998-01-01

A method and system for providing nuclear fuel rods with a configuration of isotopic gas tags. The method includes selecting a true location of a first gas tag node, selecting initial locations for the remaining n-1 nodes using target gas tag compositions, generating a set of random gene pools with L nodes, applying a Hopfield network for computing on energy, or cost, for each of the L gene pools and using selected constraints to establish minimum energy states to identify optimal gas tag nodes with each energy compared to a convergence threshold and then upon identifying the gas tag node continuing this procedure until establishing the next gas tag node until all remaining n nodes have been established.

Method for nonlinear optimization for gas tagging and other systems

DOEpatents

Chen, T.; Gross, K.C.; Wegerich, S.

1998-01-06

A method and system are disclosed for providing nuclear fuel rods with a configuration of isotopic gas tags. The method includes selecting a true location of a first gas tag node, selecting initial locations for the remaining n-1 nodes using target gas tag compositions, generating a set of random gene pools with L nodes, applying a Hopfield network for computing on energy, or cost, for each of the L gene pools and using selected constraints to establish minimum energy states to identify optimal gas tag nodes with each energy compared to a convergence threshold and then upon identifying the gas tag node continuing this procedure until establishing the next gas tag node until all remaining n nodes have been established. 6 figs.

A family of LRR sequences in the vicinity of the Co-2 locus for anthracnose resistance in Phaseolus vulgaris and its potential use in marker-assisted selection.

PubMed

Geffroy, V; Creusot, F; Falquet, J; Sévignac, M; Adam-Blondon, A F; Bannerot, H; Gepts, P; Dron, M

1998-03-01

Molecular markers offer new opportunities for breeding for disease resistance. Resistance gene pyramiding in a single cultivar, as a strategy for durable resistance, can be facilitated by marker-assisted selection (MAS). A RAPD marker, ROH20(450), linked to the Mesoamerican Co-2 anthracnose resistance gene, was previously transformed into a SCAR marker, SCH20. In the present paper we have further characterized the relevance of the SCH20 SCAR marker in different genetic backgrounds. Since this SCAR marker was found to be useful mainly in the Andean gene pool, we identified a new PCR-based marker (SCAreoli) for indirect scoring of the presence of the Co-2 gene. The SCAreoli SCAR marker is polymorphic in the Mesoamerican as well as in the Andean gene pool and should be useful in MAS. We also report that PvH20, the cloned sequence corresponding to the 450-bp RAPD marker ROH20(450), contains six imperfect leucine-rich repeats, and reveals a family of related sequences in the vicinity of the Co-2 locus. These results are discussed in the context of the recent cloning of some plant resistance genes.

Construction and Evaluation of Normalized cDNA Libraries Enriched with Full-Length Sequences for Rapid Discovery of New Genes from Sisal (Agave sisalana Perr.) Different Developmental Stages

PubMed Central

Zhou, Wen-Zhao; Zhang, Yan-Mei; Lu, Jun-Ying; Li, Jun-Feng

2012-01-01

To provide a resource of sisal-specific expressed sequence data and facilitate this powerful approach in new gene research, the preparation of normalized cDNA libraries enriched with full-length sequences is necessary. Four libraries were produced with RNA pooled from Agave sisalana multiple tissues to increase efficiency of normalization and maximize the number of independent genes by SMART™ method and the duplex-specific nuclease (DSN). This procedure kept the proportion of full-length cDNAs in the subtracted/normalized libraries and dramatically enhanced the discovery of new genes. Sequencing of 3875 cDNA clones of libraries revealed 3320 unigenes with an average insert length about 1.2 kb, indicating that the non-redundancy of libraries was about 85.7%. These unigene functions were predicted by comparing their sequences to functional domain databases and extensively annotated with Gene Ontology (GO) terms. Comparative analysis of sisal unigenes and other plant genomes revealed that four putative MADS-box genes and knotted-like homeobox (knox) gene were obtained from a total of 1162 full-length transcripts. Furthermore, real-time PCR showed that the characteristics of their transcripts mainly depended on the tight expression regulation of a number of genes during the leaf and flower development. Analysis of individual library sequence data indicated that the pooled-tissue approach was highly effective in discovering new genes and preparing libraries for efficient deep sequencing. PMID:23202944

Glycosylation-related genes in NS0 cells are insensitive to moderately elevated ammonium concentrations

PubMed Central

Brodsky, Arthur Nathan; Caldwell, Mary; Bae, Sooneon; Harcum, Sarah W.

2014-01-01

NS0 and Chinese hamster ovary (CHO) cell lines are used to produce recombinant proteins for human therapeutics; however, ammonium accumulation can negatively impact cell growth, recombinant protein production, and protein glycosylation. To improve product quality and decrease costs, the relationship between ammonium and protein glycosylation needs to be elucidated. While ammonium has been shown to adversely affect glycosylation-related gene expression in CHO cells, NS0 studies have not been performed. Therefore, this study sought to determine if glycosylation in NS0 cells were ammonium-sensitive at the gene expression level. Using a DNA microarray that contained mouse glycosylation-related and housekeeping genes, the of these genes was analysed in response to various culture conditions – elevated ammonium, elevated salt, and elevated ammonium with proline. Surprisingly, no significant differences in gene expression levels were observed between the control and these conditions. Further, the elevated ammonium cultures were analysed using real-time quantitative reverse transcriptase PCR (qRT-PCR) for key glycosylation genes, and the qRT-PCR results corroborated the DNA microarray results, demonstrating that NS0 cells are ammonium-insensitive at the gene expression level. Since NS0 are known to have elevated nucleotide sugar pools under ammonium stress, and none of the genes directly responsible for these metabolic pools were changed, consequently cellular control at the translational or substrate-level must be responsible for the universally observed decreased glycosylation quality under elevated ammonium. PMID:25062658

Flow structure through pool-riffle sequences and a conceptual model for their sustainability in gravel-bed rivers

Treesearch

D. Caamano; P. Goodwin; J. M. Buffington

2010-01-01

Detailed field measurements and simulations of three-dimensional flow structure were used to develop a conceptual model to explain the sustainability of self-formed pool-riffle sequences in gravel-bed rivers. The analysis was conducted at the Red River Wildlife Management Area in Idaho, USA, and enabled characterization of the flow structure through two consecutive...

Prospects and challenges for fungal metatranscriptomics of complex communities

Treesearch

Cheryl R. Kuske; Cedar N. Hesse; Jean F. Challacombe; Daniel Cullen; Joshua R. Herr; Rebecca C. Mueller; Adrian Tsang; Rytas Vilgalys

2015-01-01

The ability to extract and purify messenger RNA directly from plants, decomposing organic matter and soil, followed by highthroughput sequencing of the pool of expressed genes, has spawned the emerging research area of metatranscriptomics. Each metatranscriptome provides a snapshot of the composition and relative abundance of actively transcribed genes, and thus...

Pre-breeding for diversification of primary gene pool and genetic enhancement of grain legumes

PubMed Central

Sharma, Shivali; Upadhyaya, H. D.; Varshney, R. K.; Gowda, C. L. L.

2013-01-01

The narrow genetic base of cultivars coupled with low utilization of genetic resources are the major factors limiting grain legume production and productivity globally. Exploitation of new and diverse sources of variation is needed for the genetic enhancement of grain legumes. Wild relatives with enhanced levels of resistance/tolerance to multiple stresses provide important sources of genetic diversity for crop improvement. However, their exploitation for cultivar improvement is limited by cross-incompatibility barriers and linkage drags. Pre-breeding provides a unique opportunity, through the introgression of desirable genes from wild germplasm into genetic backgrounds readily used by the breeders with minimum linkage drag, to overcome this. Pre-breeding activities using promising landraces, wild relatives, and popular cultivars have been initiated at International Crops Research Institute for the Semi-Arid Tropics (ICRISAT) to develop new gene pools in chickpea, pigeonpea, and groundnut with a high frequency of useful genes, wider adaptability, and a broad genetic base. The availability of molecular markers will greatly assist in reducing linkage drags and increasing the efficiency of introgression in pre-breeding programs. PMID:23970889

Extreme-phenotype genome-wide association study (XP-GWAS): a method for identifying trait-associated variants by sequencing pools of individuals selected from a diversity panel.

PubMed

Yang, Jinliang; Jiang, Haiying; Yeh, Cheng-Ting; Yu, Jianming; Jeddeloh, Jeffrey A; Nettleton, Dan; Schnable, Patrick S

2015-11-01

Although approaches for performing genome-wide association studies (GWAS) are well developed, conventional GWAS requires high-density genotyping of large numbers of individuals from a diversity panel. Here we report a method for performing GWAS that does not require genotyping of large numbers of individuals. Instead XP-GWAS (extreme-phenotype GWAS) relies on genotyping pools of individuals from a diversity panel that have extreme phenotypes. This analysis measures allele frequencies in the extreme pools, enabling discovery of associations between genetic variants and traits of interest. This method was evaluated in maize (Zea mays) using the well-characterized kernel row number trait, which was selected to enable comparisons between the results of XP-GWAS and conventional GWAS. An exome-sequencing strategy was used to focus sequencing resources on genes and their flanking regions. A total of 0.94 million variants were identified and served as evaluation markers; comparisons among pools showed that 145 of these variants were statistically associated with the kernel row number phenotype. These trait-associated variants were significantly enriched in regions identified by conventional GWAS. XP-GWAS was able to resolve several linked QTL and detect trait-associated variants within a single gene under a QTL peak. XP-GWAS is expected to be particularly valuable for detecting genes or alleles responsible for quantitative variation in species for which extensive genotyping resources are not available, such as wild progenitors of crops, orphan crops, and other poorly characterized species such as those of ecological interest. © 2015 The Authors The Plant Journal published by Society for Experimental Biology and John Wiley & Sons Ltd.

Spatial Genetic Structure of the Abundant and Widespread Peatmoss Sphagnum magellanicum Brid.

PubMed Central

Kyrkjeeide, Magni Olsen; Hassel, Kristian; Flatberg, Kjell Ivar; Shaw, A. Jonathan; Yousefi, Narjes; Stenøien, Hans K.

2016-01-01

Spore-producing organisms have small dispersal units enabling them to become widespread across continents. However, barriers to gene flow and cryptic speciation may exist. The common, haploid peatmoss Sphagnum magellanicum occurs in both the Northern and Southern hemisphere, and is commonly used as a model in studies of peatland ecology and peatmoss physiology. Even though it will likely act as a rich source in functional genomics studies in years to come, surprisingly little is known about levels of genetic variability and structuring in this species. Here, we assess for the first time how genetic variation in S. magellanicum is spatially structured across its full distribution range (Northern Hemisphere and South America). The morphologically similar species S. alaskense was included for comparison. In total, 195 plants were genotyped at 15 microsatellite loci. Sequences from two plastid loci (trnG and trnL) were obtained from 30 samples. Our results show that S. alaskense and almost all plants of S. magellanicum in the northern Pacific area are diploids and share the same gene pool. Haploid plants occur in South America, Europe, eastern North America, western North America, and southern Asia, and five genetically differentiated groups with different distribution ranges were found. Our results indicate that S. magellanicum consists of several distinct genetic groups, seemingly with little or no gene flow among them. Noteworthy, the geographical separation of diploids and haploids is strikingly similar to patterns found within other haploid Sphagnum species spanning the Northern Hemisphere. Our results confirm a genetic division between the Beringian and the Atlantic that seems to be a general pattern in Sphagnum taxa. The pattern of strong genetic population structuring throughout the distribution range of morphologically similar plants need to be considered in future functional genomic studies of S. magellanicum. PMID:26859563

Spatial Genetic Structure of the Abundant and Widespread Peatmoss Sphagnum magellanicum Brid.

PubMed

Kyrkjeeide, Magni Olsen; Hassel, Kristian; Flatberg, Kjell Ivar; Shaw, A Jonathan; Yousefi, Narjes; Stenøien, Hans K

2016-01-01

Spore-producing organisms have small dispersal units enabling them to become widespread across continents. However, barriers to gene flow and cryptic speciation may exist. The common, haploid peatmoss Sphagnum magellanicum occurs in both the Northern and Southern hemisphere, and is commonly used as a model in studies of peatland ecology and peatmoss physiology. Even though it will likely act as a rich source in functional genomics studies in years to come, surprisingly little is known about levels of genetic variability and structuring in this species. Here, we assess for the first time how genetic variation in S. magellanicum is spatially structured across its full distribution range (Northern Hemisphere and South America). The morphologically similar species S. alaskense was included for comparison. In total, 195 plants were genotyped at 15 microsatellite loci. Sequences from two plastid loci (trnG and trnL) were obtained from 30 samples. Our results show that S. alaskense and almost all plants of S. magellanicum in the northern Pacific area are diploids and share the same gene pool. Haploid plants occur in South America, Europe, eastern North America, western North America, and southern Asia, and five genetically differentiated groups with different distribution ranges were found. Our results indicate that S. magellanicum consists of several distinct genetic groups, seemingly with little or no gene flow among them. Noteworthy, the geographical separation of diploids and haploids is strikingly similar to patterns found within other haploid Sphagnum species spanning the Northern Hemisphere. Our results confirm a genetic division between the Beringian and the Atlantic that seems to be a general pattern in Sphagnum taxa. The pattern of strong genetic population structuring throughout the distribution range of morphologically similar plants need to be considered in future functional genomic studies of S. magellanicum.

Exploiting tRNAs to Boost Virulence

PubMed Central

Albers, Suki; Czech, Andreas

2016-01-01

Transfer RNAs (tRNAs) are powerful small RNA entities that are used to translate nucleotide language of genes into the amino acid language of proteins. Their near-uniform length and tertiary structure as well as their high nucleotide similarity and post-transcriptional modifications have made it difficult to characterize individual species quantitatively. However, due to the central role of the tRNA pool in protein biosynthesis as well as newly emerging roles played by tRNAs, their quantitative assessment yields important information, particularly relevant for virus research. Viruses which depend on the host protein expression machinery have evolved various strategies to optimize tRNA usage—either by adapting to the host codon usage or encoding their own tRNAs. Additionally, several viruses bear tRNA-like elements (TLE) in the 5′- and 3′-UTR of their mRNAs. There are different hypotheses concerning the manner in which such structures boost viral protein expression. Furthermore, retroviruses use special tRNAs for packaging and initiating reverse transcription of their genetic material. Since there is a strong specificity of different viruses towards certain tRNAs, different strategies for recruitment are employed. Interestingly, modifications on tRNAs strongly impact their functionality in viruses. Here, we review those intersection points between virus and tRNA research and describe methods for assessing the tRNA pool in terms of concentration, aminoacylation and modification. PMID:26797637

Phylogeography of the Lutzomyia gomezi (Diptera: Phlebotominae) on the Panama Isthmus

PubMed Central

2014-01-01

Background Lutzomyia gomezi (Nitzulescu, 1931) is one of the main Leishmania (Vianna) panamensis vectors in Panama, and despite its medical significance, there are no population genetic studies regarding this species. In this study, we used the sequences of the mitochondrial gene cytochrome b/start of NADH1 and the nuclear elongation gene α-1 in order to analyze genetic variation and phylogeographic structure of the Lu. gomezi populations. Methods A total of 86 Lu. gomezi individuals were captured in 38 locations where cutaneous leishmaniasis occurred. DNA was extracted with phenol/chloroform methods and amplification of genes was performed using PCR primers for mitochondrial and nuclear markers. Results We found a total of 37 and 26 haplotypes of mitochondrial and nuclear genes, high haplotype diversity (h) for all three populations were detected with both molecular markers. Nucleotide diversity (π) was estimated to be high for all three populations with the mitochondrial marker, which was opposite to the estimate with the nuclear marker. In the AMOVA Φst recorded moderate (mitochondrial) and small (nuclear) population structure with statistical significance among populations. The analysis of the fixation index (Fst) used to measure the differentiation of populations showed that with the exception of the population located in the region of Bocas del Toro, the other populations presented with minor genetic differentiation. The median-Joining network of the mitochondrial marker reveled three clusters and recorded four haplotypes exclusively of localities sampled from Western Panama, demonstrating strong divergence. We found demographic population expansion with Fu´s Fs neutrality test. In the analysis mismatch distribution was observed as a bimodal curve. Conclusion Lu. gomezi is a species with higher genetic pool or variability and mild population structure, due to possible capacity migration and local adaptation to environmental changes or colonization potential. Thus, knowledge of the genetic population and evolutionary history is useful to understand the implications of different population genetic structures for cutaneous leishmaniasis epidemiology. PMID:24398187

The relative abundance of predicted genes associated with ammonia-oxidation, nitrate reduction, and biomass decomposition in mineral soil are altered by intensive timber harvest.

NASA Astrophysics Data System (ADS)

Mushinski, R. M.; Zhou, Y.; Gentry, T. J.; Boutton, T. W.

2017-12-01

Forest ecosystems in the southern United States are substantially altered by anthropogenic disturbances such as timber harvest and land conversion, with effects being observed in carbon and nutrient pools as well as biogeochemical processes. Furthermore, the desire to develop renewable energy sources in the form of biomass extraction from logging residues may result in alterations in soil community structure and function. While the impact of forest management on soil physicochemical properties of the region has been studied, its' long-term effect on soil bacterial community composition and metagenomic potential is relatively unknown, especially at deeper soil depths. This study investigates how intensive organic matter removal intensities associated with timber harvest influence decadal-scale alterations in bacterial community structure and functional potential in the upper 1-m of the soil profile, 18 years post-harvest in a Pinus taeda L. forest of eastern Texas. Amplicon sequencing of the 16S rRNA gene was used in conjunction with soil chemical analyses to evaluate treatment-induced differences in community composition and potential environmental drivers of associated change. Furthermore, functional potential was assessed by using amplicon data to make metagenomic predictions. Results indicate that increasing organic matter removal intensity leads to altered community composition and the relative abundance of dominant OTUs annotated to Burkholderia and Aciditerrimonas. The relative abundance of predicted genes associated with dissimilatory nitrate reduction and denitrification were highest in the most intensively harvested treatment while genes involved in nitrification were significantly lower in the most intensively harvested treatment. Furthermore, genes associated with glycosyltransferases were significantly reduced with increasing harvest intensity while polysaccharide lyases increased. These results imply that intensive organic matter removal may create long-term alterations in bacterial community structure with concurrent alterations to mineral soil carbon and nutrient cycling which may have future consequences on forest regeneration and subsequent stand productivity.

The Genetic Structure of Marijuana and Hemp.

PubMed

Sawler, Jason; Stout, Jake M; Gardner, Kyle M; Hudson, Darryl; Vidmar, John; Butler, Laura; Page, Jonathan E; Myles, Sean

2015-01-01

Despite its cultivation as a source of food, fibre and medicine, and its global status as the most used illicit drug, the genus Cannabis has an inconclusive taxonomic organization and evolutionary history. Drug types of Cannabis (marijuana), which contain high amounts of the psychoactive cannabinoid Δ9-tetrahydrocannabinol (THC), are used for medical purposes and as a recreational drug. Hemp types are grown for the production of seed and fibre, and contain low amounts of THC. Two species or gene pools (C. sativa and C. indica) are widely used in describing the pedigree or appearance of cultivated Cannabis plants. Using 14,031 single-nucleotide polymorphisms (SNPs) genotyped in 81 marijuana and 43 hemp samples, we show that marijuana and hemp are significantly differentiated at a genome-wide level, demonstrating that the distinction between these populations is not limited to genes underlying THC production. We find a moderate correlation between the genetic structure of marijuana strains and their reported C. sativa and C. indica ancestry and show that marijuana strain names often do not reflect a meaningful genetic identity. We also provide evidence that hemp is genetically more similar to C. indica type marijuana than to C. sativa strains.

The Genetic Structure of Marijuana and Hemp

PubMed Central

Sawler, Jason; Stout, Jake M.; Gardner, Kyle M.; Hudson, Darryl; Vidmar, John; Butler, Laura; Page, Jonathan E.; Myles, Sean

2015-01-01

Despite its cultivation as a source of food, fibre and medicine, and its global status as the most used illicit drug, the genus Cannabis has an inconclusive taxonomic organization and evolutionary history. Drug types of Cannabis (marijuana), which contain high amounts of the psychoactive cannabinoid Δ 9-tetrahydrocannabinol (THC), are used for medical purposes and as a recreational drug. Hemp types are grown for the production of seed and fibre, and contain low amounts of THC. Two species or gene pools (C. sativa and C. indica) are widely used in describing the pedigree or appearance of cultivated Cannabis plants. Using 14,031 single-nucleotide polymorphisms (SNPs) genotyped in 81 marijuana and 43 hemp samples, we show that marijuana and hemp are significantly differentiated at a genome-wide level, demonstrating that the distinction between these populations is not limited to genes underlying THC production. We find a moderate correlation between the genetic structure of marijuana strains and their reported C. sativa and C. indica ancestry and show that marijuana strain names often do not reflect a meaningful genetic identity. We also provide evidence that hemp is genetically more similar to C. indica type marijuana than to C. sativa strains. PMID:26308334

Expression of growth hormone and its transcription factor, Pit-1, in early bovine development.

PubMed

Joudrey, E M; Lechniak, D; Petrik, J; King, W A

2003-03-01

During bovine embryogenesis, bovine growth hormone (bGH) contributes to proliferation, differentiation, and modulation of embryo metabolism. Pituitary-specific transcription factor-1 (Pit-1) is a transcription factor that binds to promoters of GH, prolactin (PRL), and thyroid-stimulating hormone-beta (TSHbeta) encoding genes. A polymorphism in the fifth exon of the bGH gene resulting in a leucine (Leu) to valine (Val) substitution provides an Alu I restriction site when the Leu allele is present. To determine the onset of embryonic expression of the bGH gene, oocytes derived from ovaries homozygous for Leu alleles were fertilized in vitro with spermatozoa obtained from a Val homozygote. For each developmental stage examined, three separate pools of embryos composed of approximately 100 cell samples underwent RNA isolation, reverse transcription to cDNA, and amplification by nested PCR (nPCR). Bovine GH gene transcripts were identified at 2- to 4-cell (n = 162), 8- to 16-cell (n = 73), morulae (n = 51), and blastocyst (n = 15) stages. Likewise, transcripts for Pit-1 were detected at 2-cell (n = 125), 4-cell (n = 114), 8-cell (n = 56), 12-to-32-cell (n = 32), morulae (n = 68), and blastocyst (n = 14) stages. After digestion with Alu1, bGH cDNA was genotyped by restriction fragment length polymorphism (RFLP) analysis. Bovine GH mRNA was present in all pools of stages examined. Both Leu and Val alleles (maternal and paternal) were only detected in pools of embryos that had reached 8- to 16-cell stage. Results suggest that transcription of the bGH gene begins at the 8- to 16-cell stage in bovine embryos, possibly under control of the transcription factor, Pit-1, and that RFLP analysis of the bGH gene can be used to determine parental origin of transcripts in early embryonic development. Copyright 2003 Wiley-Liss, Inc.

Association studies of 23 positional/functional candidate genes on chromosome 10 in late-onset Alzheimer's disease.

PubMed

Morgan, A R; Turic, D; Jehu, L; Hamilton, G; Hollingworth, P; Moskvina, V; Jones, L; Lovestone, S; Brayne, C; Rubinsztein, D C; Lawlor, B; Gill, M; O'Donovan, M C; Owen, M J; Williams, J

2007-09-05

Late-onset Alzheimer's disease (LOAD) is a common neurodegenerative disorder, with a complex etiology. APOE is the only confirmed susceptibility gene for LOAD. Others remain yet to be found. Evidence from linkage studies suggests that a gene (or genes) conferring susceptibility for LOAD resides on chromosome 10. We studied 23 positional/functional candidate genes from our linkage region on chromosome 10 (APBB1IP, ALOX5, AD037, SLC18A3, DKK1, ZWINT, ANK3, UBE2D1, CDC2, SIRT1, JDP1, NET7, SUPV3L1, NEN3, SAR1, SGPL1, SEC24C, CAMK2G, PP3CB, SNCG, CH25H, PLCE1, ANXV111) in the MRC genetic resource for LOAD. These candidates were screened for sequence polymorphisms in a sample of 14 LOAD subjects and detected polymorphisms tested for association with LOAD in a three-stage design involving two stages of genotyping pooled DNA samples followed by a third stage in which markers showing evidence for association in the first stages were subjected to individual genotyping. One hundred and twenty polymorphisms were identified and tested in stage 1 (4 case + 4 control pools totaling 366 case and 366 control individuals). Single nucleotide polymorphisms (SNPs) showing evidence of association with LOAD were then studied in stage 2 (8 case + 4 control pools totaling 1,001 case and 1,001 control individuals). Five SNPs, in four genes, showed evidence for association (P < 0.1) at stage 2 and were individually genotyped in the complete dataset, comprising 1,160 LOAD cases and 1,389 normal controls. Two SNPs in SGPL1 demonstrated marginal evidence of association, with uncorrected P values of 0.042 and 0.056, suggesting that variation in SGPL1 may confer susceptibility to LOAD. Copyright 2007 Wiley-Liss, Inc.

Pharmacokinetics and pharmacogenetics of the MEK1/2 inhibitor, selumetinib, in Asian and Western healthy subjects: a pooled analysis.

PubMed

Dymond, Angela W; Elks, Cathy; Martin, Paul; Carlile, David J; Mariani, Gabriella; Lovick, Susan; Huang, Yifan; Lorch, Ulrike; Brown, Helen; So, Karen

2017-06-01

Emerging data on selumetinib, a MEK1/2 inhibitor in clinical development, suggest a possible difference in pharmacokinetics (PK) between Japanese and Western patients. This pooled analysis sought to assess the effect of ethnicity on selumetinib exposure in healthy Western and Asian subjects, and to identify any association between genetic variants in the UGT1A1, CYP2C19 and ABCG2 genes and observed differences in selumetinib PK. A pooled analysis of data from ten Phase I studies, one in Asian subjects (encompassing Japanese, non-Japanese Asian and Indian Asian subjects) and nine in Western subjects, was conducted. Key findings were derived from the collective exposure data across doses of 25, 35, 50 and 75 mg selumetinib; primary variables were dose-normalized AUC and C max . PK data from 308 subjects (10 studies) were available for the pooled analysis; genetic data from 87 subjects (3 studies) were available for the pharmacogenetic analysis. Dose-normalized AUC and C max were 35% (95% CI: 25-47%) and 39% (95% CI: 24-56%) higher in the pooled Asian group, respectively, compared with Western subjects. PK exposure parameters were similar between the Japanese, non-Japanese Asian and Indian groups. There was no evidence that the polymorphisms assessed in the genes UGT1A1, CYP2C19 and ABCG2 account for observed PK differences. Selumetinib exposure was higher in healthy Asian subjects compared with Western subjects, and these data provide valuable insight for clinicians to consider when treating patients of Asian ethnicity with selumetinib.

[Legal and management issues of public health requirements for swimming pools in Regione Toscana].

PubMed

Linguanti, Saverio; Totaro, Michele; Frendo, Lorenzo; Giorgi, Serena; Porretta, Andrea; Valentini, Paola; Baggiani, Angelo

2018-01-01

In this paper we describe the laws and rules applying to swimming pools. Authorization activity title for recreational swimming pools is regulated according to articles 80 and 86 of the TULPS R.D.n°773/1931. In Regione Toscana periodic management of the hygienic requirements for swimming pools is regulated by Regional Decree 54R/2015. It requires the evaluation of physical-chemical and microbiological indicators. The law applies to structural and organizational requirements of swimming pools as well as defines responsabilities of managers.

Uneven colonization of the lymphoid periphery by T cells that undergo early TCR{alpha} rearrangements.

PubMed

Hendricks, Deborah W; Fink, Pamela J

2009-04-01

A sparse population of thymocytes undergoes TCRalpha gene rearrangement early in development, before the double-positive stage. The potential of these cells to contribute to the peripheral T cell pool is unknown. To examine the peripheral T cell compartment expressing a repertoire biased to early TCR gene rearrangements, we developed a mouse model in which TCRalpha rearrangements are restricted to the double-negative stage of thymocyte development. These mice carry floxed RAG2 alleles and a Cre transgene driven by the CD4 promoter. As expected, conventional T cell development is compromised in such Cre(+) RAG2(fl/fl) mice, and the TCRalphabeta(+) T cells that develop are limited in their TCRalpha repertoire, preferentially using early rearranging Valpha genes. In the gut, the Thy-1(+)TCRalphabeta(+) intraepithelial lymphocyte (IEL) compartment is surprisingly intact, whereas the Thy-1(-)TCRalphabeta(+) subset is almost completely absent. Thus, T cells expressing a TCRalpha repertoire that is the product of early gene rearrangements can preferentially populate distinct IEL compartments. Despite this capacity, Cre(+) RAG2(fl/fl) T cell progenitors cannot compete with wild-type T cell progenitors in mixed bone marrow chimeras, suggesting that in normal mice, there is only a small contribution to the peripheral T cell pool by cells that have undergone early TCRalpha rearrangements. In the absence of wild-type competitors, aggressive homeostatic proliferation in the IEL compartment can promote a relatively normal Thy-1(+) TCRalphabeta(+) T cell pool from the limited population derived from Cre(+) RAG2(fl/fl) progenitors.

Uneven colonization of the lymphoid periphery by T cells that undergo early TCRα rearrangements1

PubMed Central

Hendricks, Deborah W.; Fink, Pamela J.

2009-01-01

A sparse population of thymocytes undergoes TCRα gene rearrangement early in development, before the double positive stage. The potential of these cells to contribute to the peripheral T cell pool is unknown. To examine the peripheral T cell compartment expressing a repertoire biased to early TCR gene rearrangements, we developed a mouse model in which TCRα rearrangements are restricted to the double negative stage of thymocyte development. These mice carry floxed RAG2 alleles and a Cre transgene driven by the CD4 promoter. As expected, conventional T cell development is compromised in such Cre(+) RAG2fl/fl mice, and the TCRαβ+ T cells that develop are limited in their TCRα repertoire, preferentially utilizing early-rearranging Vα genes. In the gut, the Thy-1+TCRαβ+ intraepithelial lymphocyte (IEL) compartment is surprisingly intact, while the Thy-1−TCRαβ+ subset is almost completely absent. Thus, T cells expressing a TCRα repertoire that is the product of early gene rearrangements can preferentially populate distinct IEL compartments. Despite this capacity, Cre(+) RAG2fl/fl T cell progenitors cannot compete with wild-type (WT) T cell progenitors in mixed bone marrow chimeras, suggesting that in normal mice, there is only a small contribution to the peripheral T cell pool by cells that have undergone early TCRα rearrangements. In the absence of WT competitors, aggressive homeostatic proliferation in the IEL compartment can promote a relatively normal Thy-1+ TCRαβ+ T cell pool from the limited population derived from Cre(+) RAG2fl/fl progenitors. PMID:19299725

Induction of cardiomyocyte-like cells in infarct hearts by gene transfer of Gata4, Mef2c, and Tbx5.

PubMed

Inagawa, Kohei; Miyamoto, Kazutaka; Yamakawa, Hiroyuki; Muraoka, Naoto; Sadahiro, Taketaro; Umei, Tomohiko; Wada, Rie; Katsumata, Yoshinori; Kaneda, Ruri; Nakade, Koji; Kurihara, Chitose; Obata, Yuichi; Miyake, Koichi; Fukuda, Keiichi; Ieda, Masaki

2012-10-12

After myocardial infarction (MI), massive cell death in the myocardium initiates fibrosis and scar formation, leading to heart failure. We recently found that a combination of 3 cardiac transcription factors, Gata4, Mef2c, and Tbx5 (GMT), reprograms fibroblasts directly into functional cardiomyocytes in vitro. To investigate whether viral gene transfer of GMT into infarcted hearts induces cardiomyocyte generation. Coronary artery ligation was used to generate MI in the mouse. In vitro transduction of GMT retrovirus converted cardiac fibroblasts from the infarct region into cardiomyocyte-like cells with cardiac-specific gene expression and sarcomeric structures. Injection of the green fluorescent protein (GFP) retrovirus into mouse hearts, immediately after MI, infected only proliferating noncardiomyocytes, mainly fibroblasts, in the infarct region. The GFP expression diminished after 2 weeks in immunocompetent mice but remained stable for 3 months in immunosuppressed mice, in which cardiac induction did not occur. In contrast, injection of GMT retrovirus into α-myosin heavy chain (αMHC)-GFP transgenic mouse hearts induced the expression of αMHC-GFP, a marker of cardiomyocytes, in 3% of virus-infected cells after 1 week. A pooled GMT injection into the immunosuppressed mouse hearts induced cardiac marker expression in retrovirus-infected cells within 2 weeks, although few cells showed striated muscle structures. To transduce GMT efficiently in vivo, we generated a polycistronic retrovirus expressing GMT separated by 2A "self-cleaving" peptides (3F2A). The 3F2A-induced cardiomyocyte-like cells in fibrotic tissue expressed sarcomeric α-actinin and cardiac troponin T and had clear cross striations. Quantitative RT-PCR also demonstrated that FACS-sorted 3F2A-transduced cells expressed cardiac-specific genes. GMT gene transfer induced cardiomyocyte-like cells in infarcted hearts.

Genotyping-by-Sequencing Analysis for Determining Population Structure of Finger Millet Germplasm of Diverse Origins.

PubMed

Kumar, Anil; Sharma, Divya; Tiwari, Apoorv; Jaiswal, J P; Singh, N K; Sood, Salej

2016-07-01

Finger millet [ (L.) Gaertn.] is grown mainly by subsistence farmers in arid and semiarid regions of the world. To broaden its genetic base and to boost its production, it is of paramount importance to characterize and genotype the diverse gene pool of this important food and nutritional security crop. However, as a result of nonavailability of the genome sequence of finger millet, the progress could not be made in realizing the molecular basis of unique qualities of the crop. In the present investigation, attempts have been made to characterize the genetically diverse collection of 113 finger millet accessions through whole-genome genotyping-by-sequencing (GBS), which resulted in a genome-wide set of 23,000 single-nucleotide polymorphisms (SNPs) segregating across the entire collection and several thousand SNPs segregating within every accession. A model-based population structure analysis reveals the presence of three subpopulations among the finger millet accessions, which are in parallel with the results of phylogenetic analysis. The observed population structure is consistent with the hypothesis that finger millet was domesticated first in Africa, and from there it was introduced to India some 3000 yr ago. A total of 1128 gene ontology (GO) terms were assigned to SNP-carrying genes for three main categories: biological process, cellular component, and molecular function. Facilitated access to high-throughput genotyping and sequencing technologies are likely to improve the breeding process in developing countries, and as such, this data will be very useful to breeders who are working for the genetic improvement of finger millet. Copyright © 2016 Crop Science Society of America.

Influence of modes of metal transfer on grain structure and direction of grain growth in low nickel austenitic stainless steel weld metals

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

Mukherjee, Manidipto; Saha, Saptarshi; Pal, Tapan Kumar, E-mail: tkpal.ju@gmail.com

2015-04-15

The present study elaborately discussed the effect of different modes of metal transfer (i.e., short circuit mode, spray mode and pulse mode) on grain structure and direction of grain growth in low nickel austenitic stainless steel weld metals. Electron backscattered diffraction (EBSD) analysis was used to study the grain growth direction and grain structure in weld metals. The changes in grain structure and grain growth direction were found to be essentially varied with the weld pool shape and acting forces induced by modes of metal transfer at a constant welding speed. Short circuit mode of metal transfer owing to highermore » Marangoni force (M{sub a}) and low electromagnetic force (R{sub m}) promotes the lower weld pool volume (Γ) and higher weld pool maximum radius (r{sub m}). Short circuit mode also shows curved and tapered columnar grain structures and the grain growth preferentially occurred in <001> direction. In contrast, spray mode of metal transfer increases the Γ and reduces the r{sub m} values due to very high R{sub m} and typically reveals straight and broad columnar grain structures with preferential growth direction in <111>. In the pulse mode of metal transfer relatively high M{sub a} and R{sub m} simultaneously increase the weld pool width and the primary penetration which might encourage relatively complex grain growth directions in the weld pool and cause a shift of major intensity from <001> to <111> direction. It can also be concluded that the fusion zone grain structure and direction of grain growth are solely dependent on modes of metal transfer and remain constant for a particular mode of metal transfer irrespective of filler wire used. - Highlights: • Welded joints of LNiASS were prepared by varying modes of metal transfer. • Weld pool shape, grain structure and grain growth direction were studied. • Short circuit mode shows curved and tapered grain growth in <001> direction. • Spray mode shows straight and broad columnar grain growth in <111> direction. • Pulse mode shows complex grain growth with a shift in growth direction.« less

Molecular genetic studies of natives on Easter Island: evidence of an early European and Amerindian contribution to the Polynesian gene pool.

PubMed

Lie, B A; Dupuy, B M; Spurkland, A; Fernández-Viña, M A; Hagelberg, E; Thorsby, E

2007-01-01

Most archaeological and linguistic evidence suggest a Polynesian origin of the population of Easter Island (Rapanui), and this view has been supported by the identification of Polynesian mitochondrial DNA (mtDNA) polymorphisms in prehistoric skeletal remains. However, some evidence of an early South American contact also exists (the sweet potato, bottle gourd etc.), but genetic studies have so far failed to show an early Amerindian contribution to the gene pool on Easter Island. To address this issue, we analyzed mtDNA and Y chromosome markers and performed high-resolution human leukocyte antigen (HLA) genotyping of DNA harvested from previously collected sera of 48 reputedly nonadmixed native Easter Islanders. All individuals carried mtDNA types and HLA alleles previously found in Polynesia, and most men carried Y chromosome markers of Polynesian origin, providing further evidence of a Polynesian origin of the population of Easter Island. A few individuals carried HLA alleles and/or Y chromosome markers of European origin. More interestingly, some individuals carried the HLA alleles A*0212 and B*3905, which are of typical Amerindian origin. The genealogy of some of the individuals carrying these non-Polynesian HLA alleles and their haplotypic backgrounds suggest an introduction into Easter Island in the early 1800s, or earlier. Thus, there may have been an early European and Amerindian contribution to the Polynesian gene pool of Easter Island.

Pooled Resequencing of 122 Ulcerative Colitis Genes in a Large Dutch Cohort Suggests Population-Specific Associations of Rare Variants in MUC2.

PubMed

Visschedijk, Marijn C; Alberts, Rudi; Mucha, Soren; Deelen, Patrick; de Jong, Dirk J; Pierik, Marieke; Spekhorst, Lieke M; Imhann, Floris; van der Meulen-de Jong, Andrea E; van der Woude, C Janneke; van Bodegraven, Adriaan A; Oldenburg, Bas; Löwenberg, Mark; Dijkstra, Gerard; Ellinghaus, David; Schreiber, Stefan; Wijmenga, Cisca; Rivas, Manuel A; Franke, Andre; van Diemen, Cleo C; Weersma, Rinse K

2016-01-01

Genome-wide association studies have revealed several common genetic risk variants for ulcerative colitis (UC). However, little is known about the contribution of rare, large effect genetic variants to UC susceptibility. In this study, we performed a deep targeted re-sequencing of 122 genes in Dutch UC patients in order to investigate the contribution of rare variants to the genetic susceptibility to UC. The selection of genes consists of 111 established human UC susceptibility genes and 11 genes that lead to spontaneous colitis when knocked-out in mice. In addition, we sequenced the promoter regions of 45 genes where known variants exert cis-eQTL-effects. Targeted pooled re-sequencing was performed on DNA of 790 Dutch UC cases. The Genome of the Netherlands project provided sequence data of 500 healthy controls. After quality control and prioritization based on allele frequency and pathogenicity probability, follow-up genotyping of 171 rare variants was performed on 1021 Dutch UC cases and 1166 Dutch controls. Single-variant association and gene-based analyses identified an association of rare variants in the MUC2 gene with UC. The associated variants in the Dutch population could not be replicated in a German replication cohort (1026 UC cases, 3532 controls). In conclusion, this study has identified a putative role for MUC2 on UC susceptibility in the Dutch population and suggests a population-specific contribution of rare variants to UC.

Cross-species transcriptomic approach reveals genes in hamster implantation sites.

PubMed

Lei, Wei; Herington, Jennifer; Galindo, Cristi L; Ding, Tianbing; Brown, Naoko; Reese, Jeff; Paria, Bibhash C

2014-12-01

The mouse model has greatly contributed to understanding molecular mechanisms involved in the regulation of progesterone (P4) plus estrogen (E)-dependent blastocyst implantation process. However, little is known about contributory molecular mechanisms of the P4-only-dependent blastocyst implantation process that occurs in species such as hamsters, guineapigs, rabbits, pigs, rhesus monkeys, and perhaps humans. We used the hamster as a model of P4-only-dependent blastocyst implantation and carried out cross-species microarray (CSM) analyses to reveal differentially expressed genes at the blastocyst implantation site (BIS), in order to advance the understanding of molecular mechanisms of implantation. Upregulation of 112 genes and downregulation of 77 genes at the BIS were identified using a mouse microarray platform, while use of the human microarray revealed 62 up- and 38 down-regulated genes at the BIS. Excitingly, a sizable number of genes (30 up- and 11 down-regulated genes) were identified as a shared pool by both CSMs. Real-time RT-PCR and in situ hybridization validated the expression patterns of several up- and down-regulated genes identified by both CSMs at the hamster and mouse BIS to demonstrate the merit of CSM findings across species, in addition to revealing genes specific to hamsters. Functional annotation analysis found that genes involved in the spliceosome, proteasome, and ubiquination pathways are enriched at the hamster BIS, while genes associated with tight junction, SAPK/JNK signaling, and PPARα/RXRα signalings are repressed at the BIS. Overall, this study provides a pool of genes and evidence of their participation in up- and down-regulated cellular functions/pathways at the hamster BIS. © 2014 Society for Reproduction and Fertility.

Afterlife of a Drop Impacting a Liquid Pool

NASA Astrophysics Data System (ADS)

Saha, Abhishek; Wei, Yanju; Tang, Xiaoyu; Law, Chung K.

2017-11-01

Drop impact on liquid pool is ubiquitous in industrial processes, such as inkjet printing and spray coating. While merging of drop with the impacted liquid surface is essential to facilitate the printing and coating processes, it is the afterlife of this merged drop and associated mixing which control the quality of the printed or coated surface. In this talk we will report an experimental study on the structural evolution of the merged droplet inside the liquid pool. First, we will analyze the depth of the crater created on the pool surface by the impacted drop for a range of impact inertia, and we will derive a scaling relation and the associated characteristic time-scale. Next, we will focus on the toroidal vortex formed by the moving drop inside the liquid pool and assess the characteristic time and length scales of the penetration process. The geometry of the vortex structure which qualitatively indicates the degree of mixedness will also be discussed. Finally, we will present the results from experiments with various viscosities to demonstrate the role of viscous dissipation on the geometry and structure formed by the drop. This work is supported by the Army Research Office and the Xerox Corporation.

Fish assemblage structure in an Oklahoma Ozark stream before and after rainbow trout introduction

USGS Publications Warehouse

Walsh, M.G.; Winkelman, D.L.

2005-01-01

Rainbow trout Oncorhynchus mykiss have been widely stocked throughout the United States as a popular sport fish. Our study was initiated to evaluate potential effects of rainbow trout introduction on native fishes to inform future decisions about trout stocking in northeastern Oklahoma streams. We sampled fish assemblages in pools, glides, and riffles in Brush Creek, Delaware County, Oklahoma, from February 2000 to September 2002, and experimentally stocked rainbow trout into the stream from November 2000 to March 2001 and November 2001 to March 2002. We used a combination of multivariate analyses to evaluate seasonal and habitat effects on native fish assemblages and to compare assemblage structure between prestocking, the first year of stocking, and the second year of stocking. Mesohabitat type significantly affected assemblage structure among years, whereas we did not detect an effect of season. We did not detect differences in assemblage structure among years in glide or riffle habitats. Native fish assemblage structure in pool habitats before rainbow trout introduction differed from assemblage structure in both the first and second year of stocking. Declines in seven species, including two native game fish (smallmouth bass Micropterus dolomieu and bluegill Lepomis machrochirus), contributed to assemblage dissimilarity in pool habitats between prestocking conditions and the second year of stocking. Our results indicate that stocking rainbow trout may cause local disruption in assemblage structure in pool habitats. ?? 2004 by the American Fisheries Society.

Musing on the Memes of Open and Distance Education

ERIC Educational Resources Information Center

Latchem, Colin

2014-01-01

Just as genes propagate themselves in the gene pool by leaping from body to body, so memes (ideas, behaviours, and actions) transmit cultural ideas or practices from one mind to another through writing, speech, or other imitable phenomena. This paper considers the memes that influence the evolution of open and distance education. If the…

Vertical microbial community variability of carbonate-based cones may provide insight into ancient conical stromatolite formation

NASA Astrophysics Data System (ADS)

Bradley, James; Daille, Leslie; Trivedi, Christopher; Bojanowski, Caitlin; Nunn, Heather; Stamps, Blake; Johnson, Hope; Stevenson, Bradley; Berelson, Will; Corsetti, Frank; Spear, John

2016-04-01

Stromatolite morphogenesis is poorly understood, and the process by which microbial mats become mineralized is a primary question in microbialite formation. Ancient conical stromatolites are primarily carbonate-based whereas the few modern analogues in hot springs are either non-mineralized or mineralized by silica. A team from the 2015 International GeoBiology Course investigated carbonate-rich microbial cones from near Little Hot Creek (LHC), Long Valley Caldera, California, to investigate how conical stromatolites might form in a hot spring carbonate system. The cones rise up from a layered microbial mat on the east side of a 45° C pool with very low flow that is super-saturated with respect to CaCO3. Cone structures are 8-30 mm in height, are rigid and do not deform when removed from the pool. Morphological characterization through environmental scanning electronic microscopy revealed that the cone structure is maintained by a matrix of intertwining microbial filaments around carbonate grains. This matrix gives rise to cone-filaments that are arranged vertically or horizontally, and provides further stability to the cone. Preliminary 16S rRNA gene analysis indicated variability of community composition between different vertical levels of the cone. The cone tip had comparatively greater abundance of filamentous cyanobacteria including Leptolingbya, Phormidium and Isosphaera and fewer heterotrophs (e.g. Chloroflexi) compared to the cone bottom. This supports the hypothesis that cone formation may depend on the differential abundance of the microbial community and their potential functional roles. Metagenomic analyses of the cones revealed potential genes related to chemotaxis and motility. Specifically, a genomic bin identified as a member of the genus Isosphaera contained an hmp chemotaxis operon implicated in gliding motility in the cyanobacterium Nostoc punctiforme. Isosphaera is a Planctomycete shown to have phototactic capabilities, and may play a role in conjunction with cyanobacteria in the vertical formation of the cones. This analysis of actively growing cones indicates a complex interplay of geochemistry and microbiology that form structures which can serve as models for processes that occurred in the past and are preserved in the rock record.

Impact and quantification of the sources of error in DNA pooling designs.

PubMed

Jawaid, A; Sham, P

2009-01-01

The analysis of genome wide variation offers the possibility of unravelling the genes involved in the pathogenesis of disease. Genome wide association studies are also particularly useful for identifying and validating targets for therapeutic intervention as well as for detecting markers for drug efficacy and side effects. The cost of such large-scale genetic association studies may be reduced substantially by the analysis of pooled DNA from multiple individuals. However, experimental errors inherent in pooling studies lead to a potential increase in the false positive rate and a loss in power compared to individual genotyping. Here we quantify various sources of experimental error using empirical data from typical pooling experiments and corresponding individual genotyping counts using two statistical methods. We provide analytical formulas for calculating these different errors in the absence of complete information, such as replicate pool formation, and for adjusting for the errors in the statistical analysis. We demonstrate that DNA pooling has the potential of estimating allele frequencies accurately, and adjusting the pooled allele frequency estimates for differential allelic amplification considerably improves accuracy. Estimates of the components of error show that differential allelic amplification is the most important contributor to the error variance in absolute allele frequency estimation, followed by allele frequency measurement and pool formation errors. Our results emphasise the importance of minimising experimental errors and obtaining correct error estimates in genetic association studies.

Investigation of Bartonella infection in ixodid ticks from California.

PubMed

Chang, Chao-chin; Hayashidani, Hideki; Pusterla, Nicola; Kasten, Rickie W; Madigan, John E; Chomel, Bruno B

2002-07-01

A total of 1253 ixodid ticks (254 tick pools) collected between the end of 1995 and the spring of 1997 from six California counties (El Dorado, Los Angeles, Orange, Santa Cruz, Shasta and Sonoma) were examined for the presence of Bartonella DNA by PCR of the citrate synthase gene. Of 1,119 adult Ixodes pacificus ticks tested, 26 (11.6%) of 224 pools, each containing five ticks, were positive (minimum percentage of ticks harboring detectable Bartonella DNA, 2.3%). Bartonella PCR-positive ticks were identified in five counties but none of the ticks from Los Angeles County was positive. Among 47 nymphal I. pacificus ticks collected in Sonoma County, one (10%) positive pool out of 10 pools was identified (minimum percentage of ticks harboring detectable Bartonella DNA, 2.1%). Among the 54 Dermacentor occidentalis grouped in 12 pools from Orange County, one pool (8.3%) was PCR positive for Bartonella and similarly one pool (14.3%) was positive among the 30 Dermacentor variabilis ticks grouped in seven pools. None of the three D. occidentalis from El Dorado County were positive. None of the nine tick pools positive for Ehrlichia phagocytophila were positive for Bartonella. Following our previous findings of Bartonella PCR-positive adult I. pacificus ticks in central coastal California, this is the first preliminary report of the presence of Bartonella DNA in I. pacificus nymphs and in Dermacentor sp. ticks. Distribution of Bartonella among ixodid ticks appears widespread in California.

AvrPm2 encodes an RNase-like avirulence effector which is conserved in the two different specialized forms of wheat and rye powdery mildew fungus.

PubMed

Praz, Coraline R; Bourras, Salim; Zeng, Fansong; Sánchez-Martín, Javier; Menardo, Fabrizio; Xue, Minfeng; Yang, Lijun; Roffler, Stefan; Böni, Rainer; Herren, Gerard; McNally, Kaitlin E; Ben-David, Roi; Parlange, Francis; Oberhaensli, Simone; Flückiger, Simon; Schäfer, Luisa K; Wicker, Thomas; Yu, Dazhao; Keller, Beat

2017-02-01

There is a large diversity of genetically defined resistance genes in bread wheat against the powdery mildew pathogen Blumeria graminis (B. g.) f. sp. tritici. Many confer race-specific resistance to this pathogen, but until now only the mildew avirulence gene AvrPm3 a2/f2 that is recognized by Pm3a/f was known molecularly. We performed map-based cloning and genome-wide association studies to isolate a candidate for the mildew avirulence gene AvrPm2. We then used transient expression assays in Nicotiana benthamiana to demonstrate specific and strong recognition of AvrPm2 by Pm2. The virulent AvrPm2 allele arose from a conserved 12 kb deletion, while there is no protein sequence diversity in the gene pool of avirulent B. g. tritici isolates. We found one polymorphic AvrPm2 allele in B. g. triticale and one orthologue in B. g. secalis and both are recognized by Pm2. AvrPm2 belongs to a small gene family encoding structurally conserved RNase-like effectors, including Avr a13 from B. g. hordei, the cognate Avr of the barley resistance gene Mla13. These results demonstrate the conservation of functional avirulence genes in two cereal powdery mildews specialized on different hosts, thus providing a possible explanation for successful introgression of resistance genes from rye or other grass relatives to wheat. © 2016 The Authors. New Phytologist © 2016 New Phytologist Trust.

The Population Genetics of Cultivation: Domestication of a Traditional Chinese Medicine, Scrophularia ningpoensis Hemsl. (Scrophulariaceae)

PubMed Central

Chen, Chuan; Li, Pan; Wang, Rui-Hong; Schaal, Barbara A.; Fu, Cheng-Xin

2014-01-01

Background Domestic cultivation of medicinal plants is an important strategy for protecting these species from over harvesting. Some species of medicinal plants have been brought into cultivation for more than hundreds years. Concerns about severe loss of genetic diversity and sustainable cultivation can potentially limit future use of these valuable plants. Genetic studies with comprehensive sampling of multiple medicinal species by molecular markers will allow for assessment and management of these species. Here we examine the population genetic consequences of cultivation and domestication in Scrophularia ningpoensis Hemsl. We used chloroplast DNA and genomic AFLP markers to clarify not only the effects of domestication on genetic diversity, but also determine the geographic origins of cultivars and their genetic divergence from native populations. These results will allow both better management of cultivated populations, but also provide insights for crop improvement. Results Twenty-one cpDNA haplotypes of S. ningpoensis were identified. Wild populations contain all haplotypes, whereas only three haplotypes were found in cultivated populations with wild populations having twice the haplotype diversity of cultivated populations. Genetic differentiation between cultivated populations and wild populations was significant. Genomic AFLP markers revealed similar genetic diversity patterns. Furthermore, Structure analysis grouped all wild populations into two gene pools; two of which shared the same gene pool with cultivated S. ningpoensis. The result of Neighbor-Joining analysis was consistent with the structure analysis. In principal coordinate analysis, three cultivated populations from Zhejiang Province grouped together and were separated from other cultivated populations. Conclusions These results suggest that cultivated S. ningpoensis has experienced dramatic loss of genetic diversity under anthropogenic influence. We postulate that strong artificial selection for medicinal quality has resulted in genetic differentiation between cultivated and wild populations. Furthermore, it appears that wild populations in Jiangxi-Hunan area were involved in the origin of cultivated S. ningpoensis. PMID:25157628

Mass effects meet species sorting: transformations of microbial assemblages in epiphreatic subsurface karst water pools.

PubMed

Shabarova, Tanja; Widmer, Franco; Pernthaler, Jakob

2013-09-01

We investigated the transformations of the microbial communities in epiphreatic karst cave pools with different flooding frequencies. Fingerprinting of 16S rRNA genes was combined with microscopic and sequence analysis to examine if source water would transport comparable microbial inocula into the pools at consecutive flood events, and to assess possible effects of residence time on the microbial assemblages during stagnant periods. Variability in the concentrations of dissolved organic carbon and conductivity indicated differences between floods and changes of pool water over time. High numbers of Betaproteobacteria affiliated with Methylophilaceae and Comamonadaceae were introduced into the pools during floodings. While the former persisted in the pools, the latter exhibited considerable microdiversification. These Betaproteobacteria might thus represent core microbial groups in karst water. A decrease in the estimated total diversity of the remaining bacterial taxa was apparent after a few weeks of residence: Some were favoured by stagnant conditions, whereas the majority was rapidly outcompeted. Thus, the microbial communities consisted of different components governed by complementary assembly mechanisms (dispersal versus environmental filtering) upon introduction into the pools. High overlap of temporary and persistent community members between samplings from two winters, moreover, reflected the seasonal recurrence of the studied microbial assemblages. © 2013 John Wiley & Sons Ltd and Society for Applied Microbiology.

Oncogenomic disruptions in arsenic-induced carcinogenesis

PubMed Central

Ng, Kevin W.; Stewart, Greg L.; Dummer, Trevor J.B.; Lam, Wan L.; Martinez, Victor D

2017-01-01

Chronic exposure to arsenic affects more than 200 million people worldwide, and has been associated with many adverse health effects, including cancer in several organs. There is accumulating evidence that arsenic biotransformation, a step in the elimination of arsenic from the human body, can induce changes at a genetic and epigenetic level, leading to carcinogenesis. At the genetic level, arsenic interferes with key cellular processes such as DNA damage-repair and chromosomal structure, leading to genomic instability. At the epigenetic level, arsenic places a high demand on the cellular methyl pool, leading to global hypomethylation and hypermethylation of specific gene promoters. These arsenic-associated DNA alterations result in the deregulation of both oncogenic and tumour-suppressive genes. Furthermore, recent reports have implicated aberrant expression of non-coding RNAs and the consequential disruption of signaling pathways in the context of arsenic-induced carcinogenesis. This article provides an overview of the oncogenomic anomalies associated with arsenic exposure and conveys the importance of non-coding RNAs in the arsenic-induced carcinogenic process. PMID:28179585

A Novel Strategy for Selection and Validation of Reference Genes in Dynamic Multidimensional Experimental Design in Yeast

PubMed Central

Cankorur-Cetinkaya, Ayca; Dereli, Elif; Eraslan, Serpil; Karabekmez, Erkan; Dikicioglu, Duygu; Kirdar, Betul

2012-01-01

Background Understanding the dynamic mechanism behind the transcriptional organization of genes in response to varying environmental conditions requires time-dependent data. The dynamic transcriptional response obtained by real-time RT-qPCR experiments could only be correctly interpreted if suitable reference genes are used in the analysis. The lack of available studies on the identification of candidate reference genes in dynamic gene expression studies necessitates the identification and the verification of a suitable gene set for the analysis of transient gene expression response. Principal Findings In this study, a candidate reference gene set for RT-qPCR analysis of dynamic transcriptional changes in Saccharomyces cerevisiae was determined using 31 different publicly available time series transcriptome datasets. Ten of the twelve candidates (TPI1, FBA1, CCW12, CDC19, ADH1, PGK1, GCN4, PDC1, RPS26A and ARF1) we identified were not previously reported as potential reference genes. Our method also identified the commonly used reference genes ACT1 and TDH3. The most stable reference genes from this pool were determined as TPI1, FBA1, CDC19 and ACT1 in response to a perturbation in the amount of available glucose and as FBA1, TDH3, CCW12 and ACT1 in response to a perturbation in the amount of available ammonium. The use of these newly proposed gene sets outperformed the use of common reference genes in the determination of dynamic transcriptional response of the target genes, HAP4 and MEP2, in response to relaxation from glucose and ammonium limitations, respectively. Conclusions A candidate reference gene set to be used in dynamic real-time RT-qPCR expression profiling in yeast was proposed for the first time in the present study. Suitable pools of stable reference genes to be used under different experimental conditions could be selected from this candidate set in order to successfully determine the expression profiles for the genes of interest. PMID:22675547

Selection of Salmonella enterica Serovar Typhi Genes Involved during Interaction with Human Macrophages by Screening of a Transposon Mutant Library

PubMed Central

Sabbagh, Sébastien C.; Lepage, Christine; McClelland, Michael; Daigle, France

2012-01-01

The human-adapted Salmonella enterica serovar Typhi (S. Typhi) causes a systemic infection known as typhoid fever. This disease relies on the ability of the bacterium to survive within macrophages. In order to identify genes involved during interaction with macrophages, a pool of approximately 105 transposon mutants of S. Typhi was subjected to three serial passages of 24 hours through human macrophages. Mutants recovered from infected macrophages (output) were compared to the initial pool (input) and those significantly underrepresented resulted in the identification of 130 genes encoding for cell membrane components, fimbriae, flagella, regulatory processes, pathogenesis, and many genes of unknown function. Defined deletions in 28 genes or gene clusters were created and mutants were evaluated in competitive and individual infection assays for uptake and intracellular survival during interaction with human macrophages. Overall, 26 mutants had defects in the competitive assay and 14 mutants had defects in the individual assay. Twelve mutants had defects in both assays, including acrA, exbDB, flhCD, fliC, gppA, mlc, pgtE, typA, waaQGP, SPI-4, STY1867-68, and STY2346. The complementation of several mutants by expression of plasmid-borne wild-type genes or gene clusters reversed defects, confirming that the phenotypic impairments within macrophages were gene-specific. In this study, 35 novel phenotypes of either uptake or intracellular survival in macrophages were associated with Salmonella genes. Moreover, these results reveal several genes encoding molecular mechanisms not previously known to be involved in systemic infection by human-adapted typhoidal Salmonella that will need to be elucidated. PMID:22574205

Identification of Differentially Expressed Genes in Blood Cells of Narcolepsy Patients

PubMed Central

Tanaka, Susumu; Honda, Yutaka; Honda, Makoto

2007-01-01

Study Objective: A close association between the human leukocyte antigen (HLA)-DRB1*1501/DQB1*0602 and abnormalities in some inflammatory cytokines have been demonstrated in narcolepsy. Specific alterations in the immune system have been suggested to occur in this disorder. We attempted to identify alterations in gene expression underlying the abnormalities in the blood cells of narcoleptic patients. Designs: Total RNA from 12 narcolepsy-cataplexy patients and from 12 age- and sex-matched healthy controls were pooled. The pooled samples were initially screened for candidate genes for narcolepsy by differential display analysis using annealing control primers (ACP). The second screening of the samples was carried out by semiquantitative PCR using gene-specific primers. Finally, the expression levels of the candidate genes were further confirmed by quantitative real-time PCR using a new set of samples (20 narcolepsy-cataplexy patients and 20 healthy controls). Results: The second screening revealed differential expression of 4 candidate genes. Among them, MX2 was confirmed as a significantly down-regulated gene in the white blood cells of narcoleptic patients by quantitative real-time PCR. Conclusion: We found the MX2 gene to be significantly less expressed in comparison with normal subjects in the white blood cells of narcoleptic patients. This gene is relevant to the immune system. Although differential display analysis using ACP technology has a limitation in that it does not help in determining the functional mechanism underlying sleep/wakefulness dysregulation, it is useful for identifying novel genetic factors related to narcolepsy, such as HLA molecules. Further studies are required to explore the functional relationship between the MX2 gene and narcolepsy pathophysiology. Citation: Tanaka S; Honda Y; Honda M. Identification of differentially expressed genes in blood cells of narcolepsy patients. SLEEP 2007;30(8):974-979. PMID:17702266

Evaluation and Design of Genome-Wide CRISPR/SpCas9 Knockout Screens

PubMed Central

Hart, Traver; Tong, Amy Hin Yan; Chan, Katie; Van Leeuwen, Jolanda; Seetharaman, Ashwin; Aregger, Michael; Chandrashekhar, Megha; Hustedt, Nicole; Seth, Sahil; Noonan, Avery; Habsid, Andrea; Sizova, Olga; Nedyalkova, Lyudmila; Climie, Ryan; Tworzyanski, Leanne; Lawson, Keith; Sartori, Maria Augusta; Alibeh, Sabriyeh; Tieu, David; Masud, Sanna; Mero, Patricia; Weiss, Alexander; Brown, Kevin R.; Usaj, Matej; Billmann, Maximilian; Rahman, Mahfuzur; Costanzo, Michael; Myers, Chad L.; Andrews, Brenda J.; Boone, Charles; Durocher, Daniel; Moffat, Jason

2017-01-01

The adaptation of CRISPR/SpCas9 technology to mammalian cell lines is transforming the study of human functional genomics. Pooled libraries of CRISPR guide RNAs (gRNAs) targeting human protein-coding genes and encoded in viral vectors have been used to systematically create gene knockouts in a variety of human cancer and immortalized cell lines, in an effort to identify whether these knockouts cause cellular fitness defects. Previous work has shown that CRISPR screens are more sensitive and specific than pooled-library shRNA screens in similar assays, but currently there exists significant variability across CRISPR library designs and experimental protocols. In this study, we reanalyze 17 genome-scale knockout screens in human cell lines from three research groups, using three different genome-scale gRNA libraries. Using the Bayesian Analysis of Gene Essentiality algorithm to identify essential genes, we refine and expand our previously defined set of human core essential genes from 360 to 684 genes. We use this expanded set of reference core essential genes, CEG2, plus empirical data from six CRISPR knockout screens to guide the design of a sequence-optimized gRNA library, the Toronto KnockOut version 3.0 (TKOv3) library. We then demonstrate the high effectiveness of the library relative to reference sets of essential and nonessential genes, as well as other screens using similar approaches. The optimized TKOv3 library, combined with the CEG2 reference set, provide an efficient, highly optimized platform for performing and assessing gene knockout screens in human cell lines. PMID:28655737

Biogenic silicon pools in terrestrial biogeosystems and their significance for silicon cycling

NASA Astrophysics Data System (ADS)

Puppe, Daniel; Höhn, Axel; Kaczorek, Danuta; Ehrmann, Otto; Wanner, Manfred; Sommer, Michael

2017-04-01

On a global scale the biogeochemical cycles of silicon (Si) and carbon are connected by weathering processes and fluxes of dissolved Si from terrestrial to aquatic ecosystems. Various pro- and eukaryotic organisms are evolutionarily adapted to synthesize amorphous siliceous structures (biosilicification). In soils these siliceous structures can accumulate and form biogenic Si (BSi) pools, whereat it can be differentiated between phytogenic (BSi synthesized by plants), zoogenic (BSi synthesized by sponges), microbial (BSi synthesized by bacteria and fungi) and protistic (BSi synthesized by unicellular organisms) pools. Accumulation and recycling of BSi in terrestrial biogeosystems influence fluxes of dissolved Si from the continents to the oceans, thus act as a filter in the global Si cycle. As research has primarily been focused on the role of phytogenic Si pools until now there is only little information available on the other BSi pools in soils. In order to fill this knowledge gap we examined different BSi pools in soils of initial and forested terrestrial biogeosystems using modern microscopical methods (laser scanning and scanning electron microscopy). In forested biogeosystems we further analyzed abiotic (e.g. soil pH) and biotic (earthworm biomasses) influencing factors on BSi pool size, while samples of initial biogeosystems were used to analyze spatiotemporal BSi pool dynamics. We found that especially biotic interactions are important factors for protistic BSi pools (represented by testate amoebae) and that phytogenic Si pools are about several 100-times bigger than protistic (testate amoebae) Si pools (0.2-4.7 kg Si ha-1). However, annual biosilicification rates of testate amoebae (up to 80 kg Si ha-1) are comparable to or even can exceed annual silicon uptake by trees. Our studies of initial biogeosystems revealed that BSi pool sizes increased markedly within a relatively short time span (<10 years) of ecosystem development. Differences in quantities, dynamics and resistibility against dissolution of various BSi pools indicated their possibility to influence biogeochemical Si cycling relatively rapid (protistic Si pools) or slow (zoogenic Si pools). In conclusion, our results are crucial for a detailed understanding and a more precise modeling of Si fluxes from terrestrial to aquatic ecosystems.

Cooperative Purchasing of Insurance and Risk Management Services.

ERIC Educational Resources Information Center

Maedke, Thomas F.

1988-01-01

Self Insurance (pooling) offers premium dollar savings, improved coverage, and adequate limits to school districts, governmental entities, and private-sector groups. Outlines a common and safe pool structure. (MLF)

Assembling a Computerized Adaptive Testing Item Pool as a Set of Linear Tests

ERIC Educational Resources Information Center

van der Linden, Wim J.; Ariel, Adelaide; Veldkamp, Bernard P.

2006-01-01

Test-item writing efforts typically results in item pools with an undesirable correlational structure between the content attributes of the items and their statistical information. If such pools are used in computerized adaptive testing (CAT), the algorithm may be forced to select items with less than optimal information, that violate the content…

A genome-wide association study implicates diacylglycerol kinase eta (DGKH) and several other genes in the etiology of bipolar disorder

PubMed Central

Baum, AE; Akula, N; Cabanero, M; Cardona, I; Corona, W; Klemens, B; Schulze, TG; Cichon, S; Rietschel, M; Nöthen, MM; Georgi, A; Schumacher, J; Schwarz, M; Jamra, R Abou; Höfels, S; Propping, P; Satagopan, J; Detera-Wadleigh, SD; Hardy, J; McMahon, FJ

2008-01-01

The genetic basis of bipolar disorder has long been thought to be complex, with the potential involvement of multiple genes, but methods to analyze populations with respect to this complexity have only recently become available. We have carried out a genome-wide association study of bipolar disorder by genotyping over 550,000 SNPs in two independent case-control samples of European origin. The initial association screen was performed using pooled DNA; selected SNPs were confirmed by individual genotyping. While DNA pooling reduces power to detect genetic associations, there is a substantial cost savings and gain in efficiency. A total of 88 SNPs representing 80 different genes met the prior criteria for replication in both samples. Effect sizes were modest: no single SNP of large effect was detected. Of 37 SNPs selected for individual genotyping, the strongest association signal was detected at a marker within the first intron of DGKH (p = 1.5 × 10−8, experiment-wide p<0.01, OR= 1.59). This gene encodes diacylglycerol kinase eta, a key protein in the lithium-sensitive phosphatidyl inositol pathway. This first genome-wide association study of bipolar disorder shows that several genes, each of modest effect, reproducibly influence disease risk. Bipolar disorder may be a polygenic disease. PMID:17486107

The Diagnostic Performance of Stool DNA Testing for Colorectal Cancer: A Systematic Review and Meta-Analysis.

PubMed

Zhai, Rong-Lin; Xu, Fei; Zhang, Pei; Zhang, Wan-Li; Wang, Hui; Wang, Ji-Liang; Cai, Kai-Lin; Long, Yue-Ping; Lu, Xiao-Ming; Tao, Kai-Xiong; Wang, Guo-Bin

2016-02-01

This meta-analysis was designed to evaluate the diagnostic performance of stool DNA testing for colorectal cancer (CRC) and compare the performance between single-gene and multiple-gene tests.MEDLINE, Cochrane, EMBASE databases were searched using keywords colorectal cancers, stool/fecal, sensitivity, specificity, DNA, and screening. Sensitivity analysis, quality assessments, and performance bias were performed for the included studies.Fifty-three studies were included in the analysis with a total sample size of 7524 patients. The studies were heterogeneous with regard to the genes being analyzed for fecal genetic biomarkers of CRC, as well as the laboratory methods being used for each assay. The sensitivity of the different assays ranged from 2% to 100% and the specificity ranged from 81% to 100%. The meta-analysis found that the pooled sensitivities for single- and multigene assays were 48.0% and 77.8%, respectively, while the pooled specificities were 97.0% and 92.7%. Receiver operator curves and diagnostic odds ratios showed no significant difference between both tests with regard to sensitivity or specificity.This meta-analysis revealed that using assays that evaluated multiple genes compared with single-gene assays did not increase the sensitivity or specificity of stool DNA testing in detecting CRC.

Isolation of Novel CreERT2-Driver Lines in Zebrafish Using an Unbiased Gene Trap Approach

PubMed Central

Jungke, Peggy; Hammer, Juliane; Hans, Stefan; Brand, Michael

2015-01-01

Gene manipulation using the Cre/loxP-recombinase system has been successfully employed in zebrafish to study gene functions and lineage relationships. Recently, gene trapping approaches have been applied to produce large collections of transgenic fish expressing conditional alleles in various tissues. However, the limited number of available cell- and tissue-specific Cre/CreERT2-driver lines still constrains widespread application in this model organism. To enlarge the pool of existing CreERT2-driver lines, we performed a genome-wide gene trap screen using a Tol2-based mCherry-T2a-CreERT2 (mCT2aC) gene trap vector. This cassette consists of a splice acceptor and a mCherry-tagged variant of CreERT2 which enables simultaneous labeling of the trapping event, as well as CreERT2 expression from the endogenous promoter. Using this strategy, we generated 27 novel functional CreERT2-driver lines expressing in a cell- and tissue-specific manner during development and adulthood. This study summarizes the analysis of the generated CreERT2-driver lines with respect to functionality, expression, integration, as well as associated phenotypes. Our results significantly enlarge the existing pool of CreERT2-driver lines in zebrafish and combined with Cre–dependent effector lines, the new CreERT2-driver lines will be important tools to manipulate the zebrafish genome. PMID:26083735

Litter and dead wood dynamics in ponderosa pine forests along a 160-year chronosequence.

PubMed

Hall, S A; Burke, I C; Hobbs, N T

2006-12-01

Disturbances such as fire play a key role in controlling ecosystem structure. In fire-prone forests, organic detritus comprises a large pool of carbon and can control the frequency and intensity of fire. The ponderosa pine forests of the Colorado Front Range, USA, where fire has been suppressed for a century, provide an ideal system for studying the long-term dynamics of detrital pools. Our objectives were (1) to quantify the long-term temporal dynamics of detrital pools; and (2) to determine to what extent present stand structure, topography, and soils constrain these dynamics. We collected data on downed dead wood, litter, duff (partially decomposed litter on the forest floor), stand structure, topographic position, and soils for 31 sites along a 160-year chronosequence. We developed a compartment model and parameterized it to describe the temporal trends in the detrital pools. We then developed four sets of statistical models, quantifying the hypothesized relationship between pool size and (1) stand structure, (2) topography, (3) soils variables, and (4) time since fire. We contrasted how much support each hypothesis had in the data using Akaike's Information Criterion (AIC). Time since fire explained 39-80% of the variability in dead wood of different size classes. Pool size increased to a peak as material killed by the fire fell, then decomposed rapidly to a minimum (61-85 years after fire for the different pools). It then increased, presumably as new detritus was produced by the regenerating stand. Litter was most strongly related to canopy cover (r2 = 77%), suggesting that litter fall, rather than decomposition, controls its dynamics. The temporal dynamics of duff were the hardest to predict. Detrital pool sizes were more strongly related to time since fire than to environmental variables. Woody debris peak-to-minimum time was 46-67 years, overlapping the range of historical fire return intervals (1 to > 100 years). Fires may therefore have burned under a wide range of fuel conditions, supporting the hypothesis that this region's fire regime was mixed severity.

Detection of Orientia tsutsugamushi (Rickettsiales: rickettsiaceae) in unengorged chiggers (Acari: Trombiculidae) from Oita Prefecture, Japan, by nested polymerase chain reaction.

PubMed

Pham, X D; Otsuka, Y; Suzuki, H; Takaoka, H

2001-03-01

The current study surveyed the 56-kDa type-specific antigen (TSA) gene DNAs of Orientia tsutsugamushi (Hayashi) in approximately 4.000 unengorged chiggers obtained from the soil or ground surface in an endemic and a nonendemic area of the Tsutsugamushi disease in Oita Prefecture, southwestern Japan, by nested polymerase chain reaction (PCR). Serotypes of O. tsutsugamushi were identified by restriction fragment-length polymorphism (RFLP) analysis. In the endemic area, 242 pools from five species [234 pools of Leptotrombidium scutellare (Nagayo, Miyagawa, Mitamura, Tamiya and Tenjin), two L. pallidum (Nagayo, Miyagawa, Mitamura and Tamiya), four L. kitasatoi (Fukuzumi & Obata), one L. fuji (Kuwata, Berge and Philip), and one Neotrombicula japonica (Tanaka, Kaiwa, Teramura and Kagaya)] were tested, and eight (seven pools of L. scutellare and one N. japonica) were positive for O. tsutsugamushi. Among the seven positive pools of L. scutellare, the distribution of serotypes was as follows: Kuroki (4), Gilliam (1), Karp (1), and Kawasaki (1). The first two serotypes (Kuroki and Gilliam) were identified for the first time in this species. In the nonendemic area, 128 pools from eight species were tested, and 13 were positive for O. tsutsugamushi. The positive rate was as follows: L. pallidum (4/41). L. kitasatoi (1/18), Gahrliepia saduski Womersley (2/10), L. fuji (4/50), L. himizu (Sasa, Kumada, Hayashi, Enomoto, Fukuzumi and Obata) (1/2), and Miyatrombicula kochiensis (Sasa, Kawashima and Egashira) (1/3). The latter three species were shown for the first time to harbor O. tsutsugamushi. All ofthe positive pools were Kuroki, except for two pools (one L. pallidum and one L. fuji), which were Gilliam (this serotype was also detected for the first time in L. pallidum). Further analysis revealed no differences in the nucleotide sequences (125 bp of variable domain 1 of TSA gene) of the same serotypes (i.e., Kuroki and Gilliam) among the positive samples. These data indicate that O. tsutsugamushi was widely distributed in various trombiculid species, even in the nonendemic area. The data are also suggestive of a possible horizontal transmission of O. tsutsugamushi among trombiculid species.

Intra- and extracellular lipid composition and associated gene expression patterns during pollen development in Brassica napus.

PubMed

Piffanelli, P; Ross, J H; Murphy, D J

1997-03-01

Pollen development in angiosperms is regulated by the interaction of products contributed by both the gametophytic (haploid) and sporophytic (diploid) genomes. In entomophilous species, lipids are major products of both sporophytic and gametophytic metabolism during pollen development. Mature pollen grains of Brassica napus are shown to contain three major acyl lipid pools as follows: (i) the extracellular tryphine mainly consisting of medium-chain neutral esters; (ii) the intracellular membranes, particularly endoplasmic reticulum, mainly containing phospholipids; and (iii) the intracellular storage lipids, which are mostly triacylglycerols. This paper reports on the kinetics of accumulation of these lipid classes during pollen maturation and the expression patterns of several lipid biosynthetic genes and their protein products that are differentially regulated in developing microspores/ pollen grains (gametophyte) and tapetal cells (sporophyte) of B. napus. Detailed analysis of three members of the stearoyl-ACP desaturase (sad) gene family by Northern blotting, in situ hybridization and RT-PCR showed that the same individual genes were expressed both in gametophytic and sporophytic tissues, although under different temporal regulation. In the tapetum, maximal expression of two marker genes for lipid biosynthesis (sad and ear) occurred at a bud length of 2-3 mm, and the corresponding gene products SAD and EAR were detected by Western blotting in 3-4 mm buds, coinciding with the maximal rates of tapetal lipid accumulation. These lipids are released following tapetal cell disintegration and are relocated to form the major structural component of the extracellular tryphine layer that coats the mature pollen grain. In contrast, in developing microspores/pollen grains, maximal expression of the lipid marker genes sad, ear, acp and cyb5 was at the 3-5 mm bud stages, with the SAD and EAR gene products detected in 4-7 mm buds. This pattern of expression coincided with accumulation of the intracellular storage and membrane lipid components of pollen. These results suggest that, although the same genes may be expressed in the sporophytic tapetal cells and in gametophytic tissues, they are regulated differentially leading to the production of the various contrasting lipidic structures that are assembled together to give rise to a viable, fertile pollen grain.

Targeted Maximum Likelihood Estimation for Dynamic and Static Longitudinal Marginal Structural Working Models

PubMed Central

Schwab, Joshua; Gruber, Susan; Blaser, Nello; Schomaker, Michael; van der Laan, Mark

2015-01-01

This paper describes a targeted maximum likelihood estimator (TMLE) for the parameters of longitudinal static and dynamic marginal structural models. We consider a longitudinal data structure consisting of baseline covariates, time-dependent intervention nodes, intermediate time-dependent covariates, and a possibly time-dependent outcome. The intervention nodes at each time point can include a binary treatment as well as a right-censoring indicator. Given a class of dynamic or static interventions, a marginal structural model is used to model the mean of the intervention-specific counterfactual outcome as a function of the intervention, time point, and possibly a subset of baseline covariates. Because the true shape of this function is rarely known, the marginal structural model is used as a working model. The causal quantity of interest is defined as the projection of the true function onto this working model. Iterated conditional expectation double robust estimators for marginal structural model parameters were previously proposed by Robins (2000, 2002) and Bang and Robins (2005). Here we build on this work and present a pooled TMLE for the parameters of marginal structural working models. We compare this pooled estimator to a stratified TMLE (Schnitzer et al. 2014) that is based on estimating the intervention-specific mean separately for each intervention of interest. The performance of the pooled TMLE is compared to the performance of the stratified TMLE and the performance of inverse probability weighted (IPW) estimators using simulations. Concepts are illustrated using an example in which the aim is to estimate the causal effect of delayed switch following immunological failure of first line antiretroviral therapy among HIV-infected patients. Data from the International Epidemiological Databases to Evaluate AIDS, Southern Africa are analyzed to investigate this question using both TML and IPW estimators. Our results demonstrate practical advantages of the pooled TMLE over an IPW estimator for working marginal structural models for survival, as well as cases in which the pooled TMLE is superior to its stratified counterpart. PMID:25909047

Sequencing of a QTL-rich region of the Theobroma cacao genome using pooled BACs and the identification of trait specific candidate genes

PubMed Central

2011-01-01

Background BAC-based physical maps provide for sequencing across an entire genome or a selected sub-genomic region of biological interest. Such a region can be approached with next-generation whole-genome sequencing and assembly as if it were an independent small genome. Using the minimum tiling path as a guide, specific BAC clones representing the prioritized genomic interval are selected, pooled, and used to prepare a sequencing library. Results This pooled BAC approach was taken to sequence and assemble a QTL-rich region, of ~3 Mbp and represented by twenty-seven BACs, on linkage group 5 of the Theobroma cacao cv. Matina 1-6 genome. Using various mixtures of read coverages from paired-end and linear 454 libraries, multiple assemblies of varied quality were generated. Quality was assessed by comparing the assembly of 454 reads with a subset of ten BACs individually sequenced and assembled using Sanger reads. A mixture of reads optimal for assembly was identified. We found, furthermore, that a quality assembly suitable for serving as a reference genome template could be obtained even with a reduced depth of sequencing coverage. Annotation of the resulting assembly revealed several genes potentially responsible for three T. cacao traits: black pod disease resistance, bean shape index, and pod weight. Conclusions Our results, as with other pooled BAC sequencing reports, suggest that pooling portions of a minimum tiling path derived from a BAC-based physical map is an effective method to target sub-genomic regions for sequencing. While we focused on a single QTL region, other QTL regions of importance could be similarly sequenced allowing for biological discovery to take place before a high quality whole-genome assembly is completed. PMID:21794110

Sequencing of a QTL-rich region of the Theobroma cacao genome using pooled BACs and the identification of trait specific candidate genes.

PubMed

Feltus, Frank A; Saski, Christopher A; Mockaitis, Keithanne; Haiminen, Niina; Parida, Laxmi; Smith, Zachary; Ford, James; Staton, Margaret E; Ficklin, Stephen P; Blackmon, Barbara P; Cheng, Chun-Huai; Schnell, Raymond J; Kuhn, David N; Motamayor, Juan-Carlos

2011-07-27

BAC-based physical maps provide for sequencing across an entire genome or a selected sub-genomic region of biological interest. Such a region can be approached with next-generation whole-genome sequencing and assembly as if it were an independent small genome. Using the minimum tiling path as a guide, specific BAC clones representing the prioritized genomic interval are selected, pooled, and used to prepare a sequencing library. This pooled BAC approach was taken to sequence and assemble a QTL-rich region, of ~3 Mbp and represented by twenty-seven BACs, on linkage group 5 of the Theobroma cacao cv. Matina 1-6 genome. Using various mixtures of read coverages from paired-end and linear 454 libraries, multiple assemblies of varied quality were generated. Quality was assessed by comparing the assembly of 454 reads with a subset of ten BACs individually sequenced and assembled using Sanger reads. A mixture of reads optimal for assembly was identified. We found, furthermore, that a quality assembly suitable for serving as a reference genome template could be obtained even with a reduced depth of sequencing coverage. Annotation of the resulting assembly revealed several genes potentially responsible for three T. cacao traits: black pod disease resistance, bean shape index, and pod weight. Our results, as with other pooled BAC sequencing reports, suggest that pooling portions of a minimum tiling path derived from a BAC-based physical map is an effective method to target sub-genomic regions for sequencing. While we focused on a single QTL region, other QTL regions of importance could be similarly sequenced allowing for biological discovery to take place before a high quality whole-genome assembly is completed.

Novel quantitative real-time LCR for the sensitive detection of SNP frequencies in pooled DNA: method development, evaluation and application.

PubMed

Psifidi, Androniki; Dovas, Chrysostomos; Banos, Georgios

2011-01-19

Single nucleotide polymorphisms (SNP) have proven to be powerful genetic markers for genetic applications in medicine, life science and agriculture. A variety of methods exist for SNP detection but few can quantify SNP frequencies when the mutated DNA molecules correspond to a small fraction of the wild-type DNA. Furthermore, there is no generally accepted gold standard for SNP quantification, and, in general, currently applied methods give inconsistent results in selected cohorts. In the present study we sought to develop a novel method for accurate detection and quantification of SNP in DNA pooled samples. The development and evaluation of a novel Ligase Chain Reaction (LCR) protocol that uses a DNA-specific fluorescent dye to allow quantitative real-time analysis is described. Different reaction components and thermocycling parameters affecting the efficiency and specificity of LCR were examined. Several protocols, including gap-LCR modifications, were evaluated using plasmid standard and genomic DNA pools. A protocol of choice was identified and applied for the quantification of a polymorphism at codon 136 of the ovine PRNP gene that is associated with susceptibility to a transmissible spongiform encephalopathy in sheep. The real-time LCR protocol developed in the present study showed high sensitivity, accuracy, reproducibility and a wide dynamic range of SNP quantification in different DNA pools. The limits of detection and quantification of SNP frequencies were 0.085% and 0.35%, respectively. The proposed real-time LCR protocol is applicable when sensitive detection and accurate quantification of low copy number mutations in DNA pools is needed. Examples include oncogenes and tumour suppressor genes, infectious diseases, pathogenic bacteria, fungal species, viral mutants, drug resistance resulting from point mutations, and genetically modified organisms in food.

Novel Quantitative Real-Time LCR for the Sensitive Detection of SNP Frequencies in Pooled DNA: Method Development, Evaluation and Application

PubMed Central

Psifidi, Androniki; Dovas, Chrysostomos; Banos, Georgios

2011-01-01

Background Single nucleotide polymorphisms (SNP) have proven to be powerful genetic markers for genetic applications in medicine, life science and agriculture. A variety of methods exist for SNP detection but few can quantify SNP frequencies when the mutated DNA molecules correspond to a small fraction of the wild-type DNA. Furthermore, there is no generally accepted gold standard for SNP quantification, and, in general, currently applied methods give inconsistent results in selected cohorts. In the present study we sought to develop a novel method for accurate detection and quantification of SNP in DNA pooled samples. Methods The development and evaluation of a novel Ligase Chain Reaction (LCR) protocol that uses a DNA-specific fluorescent dye to allow quantitative real-time analysis is described. Different reaction components and thermocycling parameters affecting the efficiency and specificity of LCR were examined. Several protocols, including gap-LCR modifications, were evaluated using plasmid standard and genomic DNA pools. A protocol of choice was identified and applied for the quantification of a polymorphism at codon 136 of the ovine PRNP gene that is associated with susceptibility to a transmissible spongiform encephalopathy in sheep. Conclusions The real-time LCR protocol developed in the present study showed high sensitivity, accuracy, reproducibility and a wide dynamic range of SNP quantification in different DNA pools. The limits of detection and quantification of SNP frequencies were 0.085% and 0.35%, respectively. Significance The proposed real-time LCR protocol is applicable when sensitive detection and accurate quantification of low copy number mutations in DNA pools is needed. Examples include oncogenes and tumour suppressor genes, infectious diseases, pathogenic bacteria, fungal species, viral mutants, drug resistance resulting from point mutations, and genetically modified organisms in food. PMID:21283808

Development of a Self-Report Physical Function Instrument for Disability Assessment: Item Pool Construction and Factor Analysis

PubMed Central

McDonough, Christine M.; Jette, Alan M.; Ni, Pengsheng; Bogusz, Kara; Marfeo, Elizabeth E; Brandt, Diane E; Chan, Leighton; Meterko, Mark; Haley, Stephen M.; Rasch, Elizabeth K.

2014-01-01

Objectives To build a comprehensive item pool representing work-relevant physical functioning and to test the factor structure of the item pool. These developmental steps represent initial outcomes of a broader project to develop instruments for the assessment of function within the context of Social Security Administration (SSA) disability programs. Design Comprehensive literature review; gap analysis; item generation with expert panel input; stakeholder interviews; cognitive interviews; cross-sectional survey administration; and exploratory and confirmatory factor analyses to assess item pool structure. Setting In-person and semi-structured interviews; internet and telephone surveys. Participants A sample of 1,017 SSA claimants, and a normative sample of 999 adults from the US general population. Interventions Not Applicable. Main Outcome Measure Model fit statistics Results The final item pool consisted of 139 items. Within the claimant sample 58.7% were white; 31.8% were black; 46.6% were female; and the mean age was 49.7 years. Initial factor analyses revealed a 4-factor solution which included more items and allowed separate characterization of: 1) Changing and Maintaining Body Position, 2) Whole Body Mobility, 3) Upper Body Function and 4) Upper Extremity Fine Motor. The final 4-factor model included 91 items. Confirmatory factor analyses for the 4-factor models for the claimant and the normative samples demonstrated very good fit. Fit statistics for claimant and normative samples respectively were: Comparative Fit Index = 0.93 and 0.98; Tucker-Lewis Index = 0.92 and 0.98; Root Mean Square Error Approximation = 0.05 and 0.04. Conclusions The factor structure of the Physical Function item pool closely resembled the hypothesized content model. The four scales relevant to work activities offer promise for providing reliable information about claimant physical functioning relevant to work disability. PMID:23542402

Design of inventory pools in spare part support operation systems

NASA Astrophysics Data System (ADS)

Mo, Daniel Y.; Tseng, Mitchell M.; Cheung, Raymond K.

2014-06-01

The objective of a spare part support operation is to fulfill the part request order with different service contracts in the agreed response time. With this objective to achieve different service targets for multiple service contracts and the considerations of inventory investment, it is not only important to determine the inventory policy but also to design the structure of inventory pools and the order fulfilment strategies. In this research, we focused on two types of inventory pools: multiple inventory pool (MIP) and consolidated inventory pool (CIP). The idea of MIP is to maintain separated inventory pools based on the types of service contract, while CIP solely maintains a single inventory pool regardless of service contract. Our research aims to design the inventory pool analytically and propose reserve strategies to manage the order fulfilment risks in CIP. Mathematical models and simulation experiments would be applied for analysis and evaluation.

Overexpression of synapsin Ia in the rat calyx of Held accelerates short-term plasticity and decreases synaptic vesicle volume and active zone area

PubMed Central

Vasileva, Mariya; Renden, Robert; Horstmann, Heinz; Gitler, Daniel; Kuner, Thomas

2013-01-01

Synapsins are synaptic vesicle (SV) proteins organizing a component of the reserve pool of vesicles at most central nervous system synapses. Alternative splicing of the three mammalian genes results in multiple isoforms that may differentially contribute to the organization and maintenance of the SV pools. To address this, we first characterized the expression pattern of synapsin isoforms in the rat calyx of Held. At postnatal day 16, synapsins Ia, Ib, IIb and IIIa were present, while IIa—known to sustain repetitive transmission in glutamatergic terminals—was not detectable. To test if the synapsin I isoforms could mediate IIa-like effect, and if this depends on the presence of the E-domain, we overexpressed either synapsin Ia or synapsin Ib in the rat calyx of Held via recombinant adeno-associated virus-mediated gene transfer. Although the size and overall structure of the perturbed calyces remained unchanged, short-term depression and recovery from depression were accelerated upon overexpression of synapsin I isoforms. Using electron microscopic three-dimensional reconstructions we found a redistribution of SV clusters proximal to the active zones (AZ) alongside with a decrease of both AZ area and SV volume. The number of SVs at individual AZs was strongly reduced. Hence, our data indicate that the amount of synapsin Ia expressed in the calyx regulates the rate and extent of short-term synaptic plasticity by affecting vesicle recruitment to the AZ. Finally, our study reveals a novel contribution of synapsin Ia to define the surface area of AZs. PMID:24391547

Detecting introgressive hybridization between free-ranging domestic dogs and wild wolves (Canis lupus) by admixture linkage disequilibrium analysis.

PubMed

Verardi, A; Lucchini, V; Randi, E

2006-09-01

Occasional crossbreeding between free-ranging domestic dogs and wild wolves (Canis lupus) has been detected in some European countries by mitochondrial DNA sequencing and genotyping unlinked microsatellite loci. Maternal and unlinked genomic markers, however, might underestimate the extent of introgressive hybridization, and their impacts on the preservation of wild wolf gene pools. In this study, we genotyped 220 presumed Italian wolves, 85 dogs and 7 known hybrids at 16 microsatellites belonging to four different linkage groups (plus four unlinked microsatellites). Population clustering and individual assignments were performed using a Bayesian procedure implemented in structure 2.1, which models the gametic disequilibrium arising between linked loci during admixtures, aiming to trace hybridization events further back in time and infer the population of origin of chromosomal blocks. Results indicate that (i) linkage disequilibrium was higher in wolves than in dogs; (ii) 11 out of 220 wolves (5.0%) were likely admixed, a proportion that is significantly higher than one admixed genotype in 107 wolves found previously in a study using unlinked markers; (iii) posterior maximum-likelihood estimates of the recombination parameter r revealed that introgression in Italian wolves is not recent, but could have continued for the last 70 (+/- 20) generations, corresponding to approximately 140-210 years. Bayesian clustering showed that, despite some admixture, wolf and dog gene pools remain sharply distinct (the average proportions of membership to wolf and dog clusters were Q(w) = 0.95 and Q(d) = 0.98, respectively), suggesting that hybridization was not frequent, and that introgression in nature is counteracted by behavioural or selective constraints.

One haploid parent contributes 100% of the gene pool for a widespread species in northwest North America.

PubMed

Karlin, E F; Andrus, R E; Boles, S B; Shaw, A J

2011-02-01

The monoicous peatmoss Sphagnum subnitens has a tripartite distribution that includes disjunct population systems in Europe (including the Azores), northwestern North America and New Zealand. Regional genetic diversity was highest in European S. subnitens but in northwestern North America, a single microsatellite-based multilocus haploid genotype was detected across 16 sites ranging from Coos County, Oregon, to Kavalga Island in the Western Aleutians (a distance of some 4115 km). Two multilocus haploid genotypes were detected across 14 sites on South Island, New Zealand. The microsatellite-based regional genetic diversity detected in New Zealand and North American S. subnitens is the lowest reported for any Sphagnum. The low genetic diversity detected in both of these regions most likely resulted from a founder event associated with vegetative propagation and complete selfing, with one founding haploid plant in northwest North America and two in New Zealand. Thus, one plant appears to have contributed 100% of the gene pool for the population systems of S. subnitens occurring in northwest North America, and this is arguably the most genetically uniform group of plants having a widespread distribution yet detected. Although having a distribution spanning 12.5° of latitude and 56° of longitude, there was no evidence of any genetic diversification in S. subnitens in northwest North America. No genetic structure was detected among the three regions, and it appears that European plants of S. subnitens provided the source for New Zealand and northwest North American populations. © 2010 Blackwell Publishing Ltd.

Metabolic traits of an uncultured archaeal lineage--MSBL1--from brine pools of the Red Sea.

PubMed

Mwirichia, Romano; Alam, Intikhab; Rashid, Mamoon; Vinu, Manikandan; Ba-Alawi, Wail; Anthony Kamau, Allan; Kamanda Ngugi, David; Göker, Markus; Klenk, Hans-Peter; Bajic, Vladimir; Stingl, Ulrich

2016-01-13

The candidate Division MSBL1 (Mediterranean Sea Brine Lakes 1) comprises a monophyletic group of uncultured archaea found in different hypersaline environments. Previous studies propose methanogenesis as the main metabolism. Here, we describe a metabolic reconstruction of MSBL1 based on 32 single-cell amplified genomes from Brine Pools of the Red Sea (Atlantis II, Discovery, Nereus, Erba and Kebrit). Phylogeny based on rRNA genes as well as conserved single copy genes delineates the group as a putative novel lineage of archaea. Our analysis shows that MSBL1 may ferment glucose via the Embden-Meyerhof-Parnas pathway. However, in the absence of organic carbon, carbon dioxide may be fixed via the ribulose bisphosphate carboxylase, Wood-Ljungdahl pathway or reductive TCA cycle. Therefore, based on the occurrence of genes for glycolysis, absence of the core genes found in genomes of all sequenced methanogens and the phylogenetic position, we hypothesize that the MSBL1 are not methanogens, but probably sugar-fermenting organisms capable of autotrophic growth. Such a mixotrophic lifestyle would confer survival advantage (or possibly provide a unique narrow niche) when glucose and other fermentable sugars are not available.

Molecular evidence of RNA polymerase II gene reveals the origin of worldwide cultivated barley

PubMed Central

Wang, Yonggang; Ren, Xifeng; Sun, Dongfa; Sun, Genlou

2016-01-01

The origin and domestication of cultivated barley have long been under debate. A population-based resequencing and phylogenetic analysis of the single copy of RPB2 gene was used to address barley domestication, to explore genetic differentiation of barley populations on the worldwide scale, and to understand gene-pool exchanges during the spread and subsequent development of barley cultivation. Our results revealed significant genetic differentiation among three geographically distinct wild barley populations. Differences in haplotype composition among populations from different geographical regions revealed that modern cultivated barley originated from two major wild barley populations: one from the Near East Fertile Crescent and the other from the Tibetan Plateau, supporting polyphyletic origin of cultivated barley. The results of haplotype frequencies supported multiple domestications coupled with widespread introgression events that generated genetic admixture between divergent barley gene pools. Our results not only provide important insight into the domestication and evolution of cultivated barley, but also enhance our understanding of introgression and distinct selection pressures in different environments on shaping the genetic diversity of worldwide barley populations, thus further facilitating the effective use of the wild barley germplasm. PMID:27786300

Genomics of high molecular weight plasmids isolated from an on-farm biopurification system.

PubMed

Martini, María C; Wibberg, Daniel; Lozano, Mauricio; Torres Tejerizo, Gonzalo; Albicoro, Francisco J; Jaenicke, Sebastian; van Elsas, Jan Dirk; Petroni, Alejandro; Garcillán-Barcia, M Pilar; de la Cruz, Fernando; Schlüter, Andreas; Pühler, Alfred; Pistorio, Mariano; Lagares, Antonio; Del Papa, María F

2016-06-20

The use of biopurification systems (BPS) constitutes an efficient strategy to eliminate pesticides from polluted wastewaters from farm activities. BPS environments contain a high microbial density and diversity facilitating the exchange of information among bacteria, mediated by mobile genetic elements (MGEs), which play a key role in bacterial adaptation and evolution in such environments. Here we sequenced and characterized high-molecular-weight plasmids from a bacterial collection of an on-farm BPS. The high-throughput-sequencing of the plasmid pool yielded a total of several Mb sequence information. Assembly of the sequence data resulted in six complete replicons. Using in silico analyses we identified plasmid replication genes whose encoding proteins represent 13 different Pfam families, as well as proteins involved in plasmid conjugation, indicating a large diversity of plasmid replicons and suggesting the occurrence of horizontal gene transfer (HGT) events within the habitat analyzed. In addition, genes conferring resistance to 10 classes of antimicrobial compounds and those encoding enzymes potentially involved in pesticide and aromatic hydrocarbon degradation were found. Global analysis of the plasmid pool suggest that the analyzed BPS represents a key environment for further studies addressing the dissemination of MGEs carrying catabolic genes and pathway assembly regarding degradation capabilities.

[Hygienic risks in swimming pool: knowledge and behaviours of consumers of three structures in Crema, Parma and Naples].

PubMed

Liguori, G; Castaldi, S; Signorelli, C; Auxilia, F; Alfano, V; Saccani, E; Visciano, A; Fanti, M; Spinelli, A; Pasquarella, C

2007-01-01

The swimmers health's protection and the maintenance of good safety standards of structures can be guaranteed under observance of rules and the well management of the structures and activities. An anonymous questionnaire, with 38 items, was used in order to analyse and better understand the knowledge and behaviour of the users of three swimming pools in Crema, Naples and Parma. Socio-demographic features were similar in the three centres. One of the most important result was that the necessity of showers and foot-bath before entering the swimming pool is not well understood (77% and 78% respectively); caps and foot bath are, instead, almost always worn (98% and 97%). Knowledge on infection diseases transmitted by water is very poor, warts and mycosis being the best known. Sport structures are places where health and wellness can be affected. It is important to underline the role of behavioural rules as the use of showers, caps, and proper shoes. Authors recognize the swimming pool regulations as a valid tool for health promotion. It must be clear and easily understood and it has to be linked to a proper education pathway of all users and employees.

Identification of an ancestral resistance gene cluster involved in the coevolution process between Phaseolus vulgaris and its fungal pathogen Colletotrichum lindemuthianum.

PubMed

Geffroy, V; Sicard, D; de Oliveira, J C; Sévignac, M; Cohen, S; Gepts, P; Neema, C; Langin, T; Dron, M

1999-09-01

The recent cloning of plant resistance (R) genes and the sequencing of resistance gene clusters have shed light on the molecular evolution of R genes. However, up to now, no attempt has been made to correlate this molecular evolution with the host-pathogen coevolution process at the population level. Cross-inoculations were carried out between 26 strains of the fungal pathogen Colletotrichum lindemuthianum and 48 Phaseolus vulgaris plants collected in the three centers of diversity of the host species. A high level of diversity for resistance against the pathogen was revealed. Most of the resistance specificities were overcome in sympatric situations, indicating an adaptation of the pathogen to the local host. In contrast, plants were generally resistant to allopatric strains, suggesting that R genes that were efficient against exotic strains but had been overcome locally were maintained in the plant genome. These results indicated that coevolution processes between the two protagonists led to a differentiation for resistance in the three centers of diversity of the host. To improve our understanding of the molecular evolution of these different specificities, a recombinant inbred (RI) population derived from two representative genotypes of the Andean (JaloEEP558) and Mesoamerican (BAT93) gene pools was used to map anthracnose specificities. A gene cluster comprising both Andean (Co-y; Co-z) and Mesoamerican (Co-9) host resistance specificities was identified, suggesting that this locus existed prior to the separation of the two major gene pools of P. vulgaris. Molecular analysis revealed a high level of complexity at this locus. It harbors 11 restriction fragment length polymorphisms when R gene analog (RGA) clones are used. The relationship between the coevolution process and diversification of resistance specificities at resistance gene clusters is discussed.

Acclimation responses of Arabidopsis thaliana to sustained phosphite treatments

PubMed Central

Berkowitz, Oliver

2013-01-01

Phosphite () induces a range of physiological and developmental responses in plants by disturbing the homeostasis of the macronutrient phosphate. Because of its close structural resemblance to phosphate, phosphite impairs the sensing, membrane transport, and subcellular compartmentation of phosphate. In addition, phosphite induces plant defence responses by an as yet unknown mode of action. In this study, the acclimation of Arabidopsis thaliana plants to a sustained phosphite supply in the growth medium was investigated and compared with plants growing under varying phosphate supplies. Unlike phosphate, phosphite did not suppress the formation of lateral roots in several Arabidopsis accessions. In addition, the expression of well-documented phosphate-starvation-induced genes, such as miRNA399d and At4, was not repressed by phosphite accumulation, whilst the induction of PHT1;1 and PAP1 was accentuated. Thus, a mimicking of phosphate by phosphite was not observed for these classical phosphate-starvation responses. Metabolomic analysis of phosphite-treated plants showed changes in several metabolite pools, most prominently those of aspartate, asparagine, glutamate, and serine. These alterations in amino acid pools provide novel insights for the understanding of phosphite-induced pathogen resistance. PMID:23404904

Bight of Benin: a Maternal Perspective of Four Beninese Populations and their Genetic Implications on the American Populations of African Ancestry.

PubMed

Primativo, Giuseppina; Ottoni, Claudio; Biondi, Gianfranco; Serafino, Sara; Martínez-Labarga, Cristina; Larmuseau, Maarten H D; Scardi, Michele; Decorte, Ronny; Rickards, Olga

2017-03-01

The understanding of the first movements of the ancestral populations within the African continent is still unclear, particularly in West Africa, due to several factors that have shaped the African genetic pool across time. To improve the genetic representativeness of the Beninese population and to better understand the patterns of human settlement inside West Africa and the dynamics of peopling of the Democratic Republic of Benin, we analyzed the maternal genetic variation of 193 Beninese individuals belonging to Bariba, Berba, Dendi, and Fon populations. Results support the oral traditions indicating that the western neighbouring populations have been the ancestors of the first Beninese populations, and the extant genetic structure of the Beninese populations is most likely the result of admixture between populations from neighbouring countries and native people. The present findings highlight how the Beninese populations contributed to the gene pool of the extant populations of some American populations of African ancestry. This strengthens the hypothesis that the Bight of Benin was not only an assembly point for the slave trade during the Trans-Atlantic Slave Trade but also an important slave trapping area. © 2017 John Wiley & Sons Ltd/University College London.

DNA Probe Pooling for Rapid Delineation of Chromosomal Breakpoints

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

Lu, Chun-Mei; Kwan, Johnson; Baumgartner, Adolf

2009-01-30

Structural chromosome aberrations are hallmarks of many human genetic diseases. The precise mapping of translocation breakpoints in tumors is important for identification of genes with altered levels of expression, prediction of tumor progression, therapy response, or length of disease-free survival as well as the preparation of probes for detection of tumor cells in peripheral blood. Similarly, in vitro fertilization (IVF) and preimplantation genetic diagnosis (PGD) for carriers of balanced, reciprocal translocations benefit from accurate breakpoint maps in the preparation of patient-specific DNA probes followed by a selection of normal or balanced oocytes or embryos. We expedited the process of breakpointmore » mapping and preparation of case-specific probes by utilizing physically mapped bacterial artificial chromosome (BAC) clones. Historically, breakpoint mapping is based on the definition of the smallest interval between proximal and distal probes. Thus, many of the DNA probes prepared for multi-clone and multi-color mapping experiments do not generate additional information. Our pooling protocol described here with examples from thyroid cancer research and PGD accelerates the delineation of translocation breakpoints without sacrificing resolution. The turnaround time from clone selection to mapping results using tumor or IVF patient samples can be as short as three to four days.« less

Africanization in the United States: replacement of feral European honeybees (Apis mellifera L.) by an African hybrid swarm.

PubMed

Pinto, M Alice; Rubink, William L; Patton, John C; Coulson, Robert N; Johnston, J Spencer

2005-08-01

The expansion of Africanized honeybees from South America to the southwestern United States in <50 years is considered one of the most spectacular biological invasions yet documented. In the American tropics, it has been shown that during their expansion Africanized honeybees have low levels of introgressed alleles from resident European populations. In the United States, it has been speculated, but not shown, that Africanized honeybees would hybridize extensively with European honeybees. Here we report a continuous 11-year study investigating temporal changes in the genetic structure of a feral population from the southern United States undergoing Africanization. Our microsatellite data showed that (1) the process of Africanization involved both maternal and paternal bidirectional gene flow between European and Africanized honeybees and (2) the panmitic European population was replaced by panmitic mixtures of A. m. scutellata and European genes within 5 years after Africanization. The post-Africanization gene pool (1998-2001) was composed of a diverse array of recombinant classes with a substantial European genetic contribution (mean 25-37%). Therefore, the resulting feral honeybee population of south Texas was best viewed as a hybrid swarm.

Afghan Hindu Kush: Where Eurasian Sub-Continent Gene Flows Converge

PubMed Central

Mazières, Stéphane; Myres, Natalie M.; Lin, Alice A.; Temori, Shah Aga; Metspalu, Mait; Metspalu, Ene; Witzel, Michael; King, Roy J.; Underhill, Peter A.; Villems, Richard; Chiaroni, Jacques

2013-01-01

Despite being located at the crossroads of Asia, genetics of the Afghanistan populations have been largely overlooked. It is currently inhabited by five major ethnic populations: Pashtun, Tajik, Hazara, Uzbek and Turkmen. Here we present autosomal from a subset of our samples, mitochondrial and Y- chromosome data from over 500 Afghan samples among these 5 ethnic groups. This Afghan data was supplemented with the same Y-chromosome analyses of samples from Iran, Kyrgyzstan, Mongolia and updated Pakistani samples (HGDP-CEPH). The data presented here was integrated into existing knowledge of pan-Eurasian genetic diversity. The pattern of genetic variation, revealed by structure-like and Principal Component analyses and Analysis of Molecular Variance indicates that the people of Afghanistan are made up of a mosaic of components representing various geographic regions of Eurasian ancestry. The absence of a major Central Asian-specific component indicates that the Hindu Kush, like the gene pool of Central Asian populations in general, is a confluence of gene flows rather than a source of distinctly autochthonous populations that have arisen in situ: a conclusion that is reinforced by the phylogeography of both haploid loci. PMID:24204668

Molecular detection of Rickettsia aeschlimannii in Hyalomma spp. ticks from camels (Camelus dromedarius) in Nigeria, West Africa.

PubMed

Kamani, J; Baneth, G; Apanaskevich, D A; Mumcuoglu, K Y; Harrus, S

2015-06-01

Several species of the spotted fever group rickettsiae have been identified as emerging pathogens throughout the world, including in Africa. In this study, 197 Hyalomma ticks (Ixodida: Ixodidae) collected from 51 camels (Camelus dromedarius) in Kano, northern Nigeria, were screened by amplification and sequencing of the citrate synthase (gltA), outer membrane protein A (ompA) and 17-kDa antigen gene fragments. Rickettsia sp. gltA fragments were detected in 43.3% (42/97) of the tick pools tested. Rickettsial ompA gene fragments (189 bp and 630 bp) were detected in 64.3% (n = 27) and 23.8% (n = 10) of the gltA-positive tick pools by real-time and conventional polymerase chain reaction (PCR), respectively. The amplicons were 99-100% identical to Rickettsia aeschlimannii TR/Orkun-H and R. aeschlimannii strain EgyRickHimp-El-Arish in GenBank. Furthermore, 17-kDa antigen gene fragments of 214 bp and 265 bp were detected in 59.5% (n = 25) and 38.1% (n = 16), respectively, of tick pools, and sequences were identical to one another and 99-100% identical to those of the R. aeschlimannii strain Ibadan A1 in GenBank. None of the Hyalomma impressum ticks collected were positive for Rickettsia sp. DNA. Rickettsia sp. gltA fragments (133 bp) were detected in 18.8% of camel blood samples, but all samples were negative for the other genes targeted. This is the first report to describe the molecular detection of R. aeschlimannii in Hyalomma spp. ticks from camels in Nigeria. © 2015 The Royal Entomological Society.

Coxiella burnetii and Rickettsia conorii: Two zoonotic pathogens in peridomestic rodents and their ectoparasites in Nigeria.

PubMed

Kamani, Joshua; Baneth, Gad; Gutiérrez, Ricardo; Nachum-Biala, Yaarit; Mumcuoglu, Kosta Y; Harrus, Shimon

2018-01-01

Rodents are hosts of numerous pathogenic agents of public health importance globally. Their ability to harbor these pathogens without showing overt clinical signs of disease has epidemiologic consequences. In some rural settings in Nigeria, humans and rodents do not only share feeds and abode, but the latter may end up on the table of the former as a source of protein, thereby increasing the risks of disease transmission. Molecular assays were used to detect and characterize two agents of zoonotic importance, Coxiella burnetii and Rickettsia spp. in 194 peridomestic rodents captured in a peri-urban setting in Nigeria, and 32 pools of ectoparasites removed from them, to determine their possible role in the epidemiology of these diseases in this country. Targeting and characterizing the insertion sequence IS1111, C. burnetii DNA was detected in 4 out of 194 (2.1%) rodents comprising 3 out of 121 (2.5%) Rattus norvegicus and 1 out of 48 (2.1%) Rattus rattus screened in this study. Rickettsia spp. DNA was detected in two Rhipicephalus sanginueus sensu lato pools (i.e. RT1 and RT4) using the citrate synthase (gltA) gene and further characterized by amplification and sequence analysis of six genes to determine their identity. The RT1 sample consistently gave 98-100% identity to Rickettsia conorii str. Malish 7 for the various genes and loci studied. However, the identity of RT4 could not be definitively determined due to variable identities to different Rickettsia spp. according to the gene or loci under consideration. Further isolation study to determine if the RT4 characterized is a new variant or a mixture of sequences of different rickettsiae within the pool will be worthwhile. Copyright © 2017 Elsevier GmbH. All rights reserved.

From genomes to vaccines: Leishmania as a model.

PubMed Central

Almeida, Renata; Norrish, Alan; Levick, Mark; Vetrie, David; Freeman, Tom; Vilo, Jaak; Ivens, Alasdair; Lange, Uta; Stober, Carmel; McCann, Sharon; Blackwell, Jenefer M

2002-01-01

The 35 Mb genome of Leishmania should be sequenced by late 2002. It contains approximately 8500 genes that will probably translate into more than 10 000 proteins. In the laboratory we have been piloting strategies to try to harness the power of the genome-proteome for rapid screening of new vaccine candidate. To this end, microarray analysis of 1094 unique genes identified using an EST analysis of 2091 cDNA clones from spliced leader libraries prepared from different developmental stages of Leishmania has been employed. The plan was to identify amastigote-expressed genes that could be used in high-throughput DNA-vaccine screens to identify potential new vaccine candidates. Despite the lack of transcriptional regulation that polycistronic transcription in Leishmania dictates, the data provide evidence for a high level of post-transcriptional regulation of RNA abundance during the developmental cycle of promastigotes in culture and in lesion-derived amastigotes of Leishmania major. This has provided 147 candidates from the 1094 unique genes that are specifically upregulated in amastigotes and are being used in vaccine studies. Using DNA vaccination, it was demonstrated that pooling strategies can work to identify protective vaccines, but it was found that some potentially protective antigens are masked by other disease-exacerbatory antigens in the pool. A total of 100 new vaccine candidates are currently being tested separately and in pools to extend this analysis, and to facilitate retrospective bioinformatic analysis to develop predictive algorithms for sequences that constitute potentially protective antigens. We are also working with other members of the Leishmania Genome Network to determine whether RNA expression determined by microarray analyses parallels expression at the protein level. We believe we are making good progress in developing strategies that will allow rapid translation of the sequence of Leishmania into potential interventions for disease control in humans. PMID:11839176

Core element characterization of Rhodococcus promoters and development of a promoter-RBS mini-pool with different activity levels for efficient gene expression.

PubMed

Jiao, Song; Yu, Huimin; Shen, Zhongyao

2018-09-25

To satisfy the urgent demand for promoter engineering that can accurately regulate the metabolic circuits and expression of specific genes in the Rhodococcus microbial platform, a promoter-ribosome binding site (RBS) coupled mini-pool with fine-tuning of different activity levels was successfully established. Transcriptome analyses of R. ruber TH revealed several representative promoters with different activity levels, e.g., Pami, Pcs, Pnh, P50sl36, PcbiM, PgroE and Pniami. β-Galactosidase (LacZ) reporter measurement demonstrated that different gene expression levels could be obtained with these natural promoters combined with an optimal RBS of ami. Further use of these promoters to overexpress the nitrile hydratase (NHase) gene with RBSami in R. ruber THdAdN produced different expression levels consistent with the transcription analyses. The -35 and -10 core elements of different promoters were further analyzed, and the conserved sequences were revealed to be TTGNNN and (T/C)GNNA(A/C)AAT. By mutating the core elements of the strong promoters, Pnh and Pami, into the above consensus sequence, two even stronger promoters, PnhM and PamiM, were obtained with 2.2-fold and 7.7-fold improvements in transcription, respectively. Integrating several strategies, including transcriptome promoter screening, -35 and -10 core element identification, core element point-mutation, RBS optimization and diverse reporter verification, a fine-tuning promoter-RBS combination mini-pool with different activity levels in Rhodococcus strains was successfully established. This development is significant for broad applications of the Rhodococcus genus as a microbial platform. Copyright © 2018 Elsevier B.V. All rights reserved.

How to map your industry's profit pool.

PubMed

Gadiesh, O; Gilbert, J L

1998-01-01

Many managers chart strategy without a full understanding of the sources and distribution of profits in their industry. Sometimes they focus their sights on revenues instead of profits, mistakenly assuming that revenue growth will eventually translate into profit growth. In other cases, they simply lack the data or the analytical tools required to isolate and measure variations in profitability. In this Manager's Tool Kit, the authors present a way to think clearly about where the money's being made in any industry. They describe a framework for analyzing how profits are distributed among the activities that form an industry's value chain. Such an analysis can provide a company's managers with a rich understanding of their industry's profit structure--what the authors call its profit pool--enabling them to identify which activities are generating disproportionately large or small shares of profits. Even more important, a profit-pool map opens a window onto the underlying structure of the industry, helping managers see the various forces that are determining the distribution of profits. As such, a profit-pool map provides a solid basis for strategic thinking. Mapping a profit pool involves four steps: defining the boundaries of the pool, estimating the pool's overall size, estimating the size of each value-chain activity in the pool, and checking and reconciling the calculations. The authors briefly describe each step and then apply the process by providing a detailed example of a hypothetical retail bank. They conclude by looking at ways of organizing the data in chart form as a first step toward plotting a profit-pool strategy.

Parametrization consequences of constraining soil organic matter models by total carbon and radiocarbon using long-term field data

NASA Astrophysics Data System (ADS)

Menichetti, Lorenzo; Kätterer, Thomas; Leifeld, Jens

2016-05-01

Soil organic carbon (SOC) dynamics result from different interacting processes and controls on spatial scales from sub-aggregate to pedon to the whole ecosystem. These complex dynamics are translated into models as abundant degrees of freedom. This high number of not directly measurable variables and, on the other hand, very limited data at disposal result in equifinality and parameter uncertainty. Carbon radioisotope measurements are a proxy for SOC age both at annual to decadal (bomb peak based) and centennial to millennial timescales (radio decay based), and thus can be used in addition to total organic C for constraining SOC models. By considering this additional information, uncertainties in model structure and parameters may be reduced. To test this hypothesis we studied SOC dynamics and their defining kinetic parameters in the Zürich Organic Fertilization Experiment (ZOFE) experiment, a > 60-year-old controlled cropland experiment in Switzerland, by utilizing SOC and SO14C time series. To represent different processes we applied five model structures, all stemming from a simple mother model (Introductory Carbon Balance Model - ICBM): (I) two decomposing pools, (II) an inert pool added, (III) three decomposing pools, (IV) two decomposing pools with a substrate control feedback on decomposition, (V) as IV but with also an inert pool. These structures were extended to explicitly represent total SOC and 14C pools. The use of different model structures allowed us to explore model structural uncertainty and the impact of 14C on kinetic parameters. We considered parameter uncertainty by calibrating in a formal Bayesian framework. By varying the relative importance of total SOC and SO14C data in the calibration, we could quantify the effect of the information from these two data streams on estimated model parameters. The weighing of the two data streams was crucial for determining model outcomes, and we suggest including it in future modeling efforts whenever SO14C data are available. The measurements and all model structures indicated a dramatic decline in SOC in the ZOFE experiment after an initial land use change in 1949 from grass- to cropland, followed by a constant but smaller decline. According to all structures, the three treatments (control, mineral fertilizer, farmyard manure) we considered were still far from equilibrium. The estimates of mean residence time (MRT) of the C pools defined by our models were sensitive to the consideration of the SO14C data stream. Model structure had a smaller effect on estimated MRT, which ranged between 5.9 ± 0.1 and 4.2 ± 0.1 years and 78.9 ± 0.1 and 98.9 ± 0.1 years for young and old pools, respectively, for structures without substrate interactions. The simplest model structure performed the best according to information criteria, validating the idea that we still lack data for mechanistic SOC models. Although we could not exclude any of the considered processes possibly involved in SOC decomposition, it was not possible to discriminate their relative importance.

Vascular tone pathway polymorphisms in relation to primary open-angle glaucoma.

PubMed

Kang, J H; Loomis, S J; Yaspan, B L; Bailey, J C; Weinreb, R N; Lee, R K; Lichter, P R; Budenz, D L; Liu, Y; Realini, T; Gaasterland, D; Gaasterland, T; Friedman, D S; McCarty, C A; Moroi, S E; Olson, L; Schuman, J S; Singh, K; Vollrath, D; Wollstein, G; Zack, D J; Brilliant, M; Sit, A J; Christen, W G; Fingert, J; Forman, J P; Buys, E S; Kraft, P; Zhang, K; Allingham, R R; Pericak-Vance, M A; Richards, J E; Hauser, M A; Haines, J L; Wiggs, J L; Pasquale, L R

2014-06-01

Vascular perfusion may be impaired in primary open-angle glaucoma (POAG); thus, we evaluated a panel of markers in vascular tone-regulating genes in relation to POAG. We used Illumina 660W-Quad array genotype data and pooled P-values from 3108 POAG cases and 3430 controls from the combined National Eye Institute Glaucoma Human Genetics Collaboration consortium and Glaucoma Genes and Environment studies. Using information from previous literature and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, we compiled single-nucleotide polymorphisms (SNPs) in 186 vascular tone-regulating genes. We used the 'Pathway Analysis by Randomization Incorporating Structure' analysis software, which performed 1000 permutations to compare the overall pathway and selected genes with comparable randomly generated pathways and genes in their association with POAG. The vascular tone pathway was not associated with POAG overall or POAG subtypes, defined by the type of visual field loss (early paracentral loss (n=224 cases) or only peripheral loss (n=993 cases)) (permuted P≥0.20). In gene-based analyses, eight were associated with POAG overall at permuted P<0.001: PRKAA1, CAV1, ITPR3, EDNRB, GNB2, DNM2, HFE, and MYL9. Notably, six of these eight (the first six listed) code for factors involved in the endothelial nitric oxide synthase activity, and three of these six (CAV1, ITPR3, and EDNRB) were also associated with early paracentral loss at P<0.001, whereas none of the six genes reached P<0.001 for peripheral loss only. Although the assembled vascular tone SNP set was not associated with POAG, genes that code for local factors involved in setting vascular tone were associated with POAG.

Patterns and architecture of genomic islands in marine bacteria

PubMed Central

2012-01-01

Background Genomic Islands (GIs) have key roles since they modulate the structure and size of bacterial genomes displaying a diverse set of laterally transferred genes. Despite their importance, GIs in marine bacterial genomes have not been explored systematically to uncover possible trends and to analyze their putative ecological significance. Results We carried out a comprehensive analysis of GIs in 70 selected marine bacterial genomes detected with IslandViewer to explore the distribution, patterns and functional gene content in these genomic regions. We detected 438 GIs containing a total of 8152 genes. GI number per genome was strongly and positively correlated with the total GI size. In 50% of the genomes analyzed the GIs accounted for approximately 3% of the genome length, with a maximum of 12%. Interestingly, we found transposases particularly enriched within Alphaproteobacteria GIs, and site-specific recombinases in Gammaproteobacteria GIs. We described specific Homologous Recombination GIs (HR-GIs) in several genera of marine Bacteroidetes and in Shewanella strains among others. In these HR-GIs, we recurrently found conserved genes such as the β-subunit of DNA-directed RNA polymerase, regulatory sigma factors, the elongation factor Tu and ribosomal protein genes typically associated with the core genome. Conclusions Our results indicate that horizontal gene transfer mediated by phages, plasmids and other mobile genetic elements, and HR by site-specific recombinases play important roles in the mobility of clusters of genes between taxa and within closely related genomes, modulating the flexible pool of the genome. Our findings suggest that GIs may increase bacterial fitness under environmental changing conditions by acquiring novel foreign genes and/or modifying gene transcription and/or transduction. PMID:22839777

Mining Gene Regulatory Networks by Neural Modeling of Expression Time-Series.

PubMed

Rubiolo, Mariano; Milone, Diego H; Stegmayer, Georgina

2015-01-01

Discovering gene regulatory networks from data is one of the most studied topics in recent years. Neural networks can be successfully used to infer an underlying gene network by modeling expression profiles as times series. This work proposes a novel method based on a pool of neural networks for obtaining a gene regulatory network from a gene expression dataset. They are used for modeling each possible interaction between pairs of genes in the dataset, and a set of mining rules is applied to accurately detect the subjacent relations among genes. The results obtained on artificial and real datasets confirm the method effectiveness for discovering regulatory networks from a proper modeling of the temporal dynamics of gene expression profiles.

Transcriptomic Analysis Using Olive Varieties and Breeding Progenies Identifies Candidate Genes Involved in Plant Architecture.

PubMed

González-Plaza, Juan J; Ortiz-Martín, Inmaculada; Muñoz-Mérida, Antonio; García-López, Carmen; Sánchez-Sevilla, José F; Luque, Francisco; Trelles, Oswaldo; Bejarano, Eduardo R; De La Rosa, Raúl; Valpuesta, Victoriano; Beuzón, Carmen R

2016-01-01

Plant architecture is a critical trait in fruit crops that can significantly influence yield, pruning, planting density and harvesting. Little is known about how plant architecture is genetically determined in olive, were most of the existing varieties are traditional with an architecture poorly suited for modern growing and harvesting systems. In the present study, we have carried out microarray analysis of meristematic tissue to compare expression profiles of olive varieties displaying differences in architecture, as well as seedlings from their cross pooled on the basis of their sharing architecture-related phenotypes. The microarray used, previously developed by our group has already been applied to identify candidates genes involved in regulating juvenile to adult transition in the shoot apex of seedlings. Varieties with distinct architecture phenotypes and individuals from segregating progenies displaying opposite architecture features were used to link phenotype to expression. Here, we identify 2252 differentially expressed genes (DEGs) associated to differences in plant architecture. Microarray results were validated by quantitative RT-PCR carried out on genes with functional annotation likely related to plant architecture. Twelve of these genes were further analyzed in individual seedlings of the corresponding pool. We also examined Arabidopsis mutants in putative orthologs of these targeted candidate genes, finding altered architecture for most of them. This supports a functional conservation between species and potential biological relevance of the candidate genes identified. This study is the first to identify genes associated to plant architecture in olive, and the results obtained could be of great help in future programs aimed at selecting phenotypes adapted to modern cultivation practices in this species.

Identification and verification of differentially expressed genes in the caprine hypothalamic-pituitary-gonadal axis that are associated with litter size.

PubMed

Feng, Tao; Cao, Gui-Ling; Chu, Ming-Xing; Di, Ran; Huang, Dong-Wei; Liu, Qiu-Yue; Pan, Zhang-Yuan; Jin, Mei; Zhang, Ying-Jie; Li, Ning

2015-02-01

Litter size is a favorable economic trait for the goat industry, but remains a complex trait controlled by multiple genes in multiple organs. Several genes have been identified that may affect embryo survival, follicular development, and the health of fetuses during pregnancy. Jining Grey goats demonstrate the largest litter size among goat breeds indigenous to China. In order to better understand the genetic basis of this trait, six suppression subtractive hybridization (SSH) cDNA libraries were constructed using pooled mRNAs from hypothalamuses, pituitaries, and ovaries of sexually mature and adult polytocous Jining Grey goats, as testers, versus the pooled corresponding mRNAs of monotocous Liaoning Cashmere goats, as drivers. A total of 1,458 true-positive clones--including 955 known genes and 481 known and 22 unknown expressed sequence tags--were obtained from the SSH libraries by sequencing and alignment. The known genes were categorized into cellular processes and signaling information storage and processing, and metabolism. Three genes (FTH1, GH, and SAA) were selected to validate the SSH results by quantitative real-time PCR; all three were up-regulated in the corresponding tissues in the tester group indicating that these are candidate genes associated with the large litter size of Jining Grey goats. Several other identified genes may affect embryo survival, follicular development, and health during pregnancy. This study provides insights into the mechanistic basis by which the caprine hypothalamic-pituitary-gonadal axis affects reproductive traits and provides a theoretical basis for goat production and breeding. © 2015 Wiley Periodicals, Inc.

High frequency of phylogenetically diverse reductive dehalogenase-homologous genes in deep subseafloor sedimentary metagenomes

PubMed Central

Kawai, Mikihiko; Futagami, Taiki; Toyoda, Atsushi; Takaki, Yoshihiro; Nishi, Shinro; Hori, Sayaka; Arai, Wataru; Tsubouchi, Taishi; Morono, Yuki; Uchiyama, Ikuo; Ito, Takehiko; Fujiyama, Asao; Inagaki, Fumio; Takami, Hideto

2014-01-01

Marine subsurface sediments on the Pacific margin harbor diverse microbial communities even at depths of several hundreds meters below the seafloor (mbsf) or more. Previous PCR-based molecular analysis showed the presence of diverse reductive dehalogenase gene (rdhA) homologs in marine subsurface sediment, suggesting that anaerobic respiration of organohalides is one of the possible energy-yielding pathways in the organic-rich sedimentary habitat. However, primer-independent molecular characterization of rdhA has remained to be demonstrated. Here, we studied the diversity and frequency of rdhA homologs by metagenomic analysis of five different depth horizons (0.8, 5.1, 18.6, 48.5, and 107.0 mbsf) at Site C9001 off the Shimokita Peninsula of Japan. From all metagenomic pools, remarkably diverse rdhA-homologous sequences, some of which are affiliated with novel clusters, were observed with high frequency. As a comparison, we also examined frequency of dissimilatory sulfite reductase genes (dsrAB), key functional genes for microbial sulfate reduction. The dsrAB were also widely observed in the metagenomic pools whereas the frequency of dsrAB genes was generally smaller than that of rdhA-homologous genes. The phylogenetic composition of rdhA-homologous genes was similar among the five depth horizons. Our metagenomic data revealed that subseafloor rdhA homologs are more diverse than previously identified from PCR-based molecular studies. Spatial distribution of similar rdhA homologs across wide depositional ages indicates that the heterotrophic metabolic processes mediated by the genes can be ecologically important, functioning in the organic-rich subseafloor sedimentary biosphere. PMID:24624126

High frequency of phylogenetically diverse reductive dehalogenase-homologous genes in deep subseafloor sedimentary metagenomes.

PubMed

Kawai, Mikihiko; Futagami, Taiki; Toyoda, Atsushi; Takaki, Yoshihiro; Nishi, Shinro; Hori, Sayaka; Arai, Wataru; Tsubouchi, Taishi; Morono, Yuki; Uchiyama, Ikuo; Ito, Takehiko; Fujiyama, Asao; Inagaki, Fumio; Takami, Hideto

2014-01-01

Marine subsurface sediments on the Pacific margin harbor diverse microbial communities even at depths of several hundreds meters below the seafloor (mbsf) or more. Previous PCR-based molecular analysis showed the presence of diverse reductive dehalogenase gene (rdhA) homologs in marine subsurface sediment, suggesting that anaerobic respiration of organohalides is one of the possible energy-yielding pathways in the organic-rich sedimentary habitat. However, primer-independent molecular characterization of rdhA has remained to be demonstrated. Here, we studied the diversity and frequency of rdhA homologs by metagenomic analysis of five different depth horizons (0.8, 5.1, 18.6, 48.5, and 107.0 mbsf) at Site C9001 off the Shimokita Peninsula of Japan. From all metagenomic pools, remarkably diverse rdhA-homologous sequences, some of which are affiliated with novel clusters, were observed with high frequency. As a comparison, we also examined frequency of dissimilatory sulfite reductase genes (dsrAB), key functional genes for microbial sulfate reduction. The dsrAB were also widely observed in the metagenomic pools whereas the frequency of dsrAB genes was generally smaller than that of rdhA-homologous genes. The phylogenetic composition of rdhA-homologous genes was similar among the five depth horizons. Our metagenomic data revealed that subseafloor rdhA homologs are more diverse than previously identified from PCR-based molecular studies. Spatial distribution of similar rdhA homologs across wide depositional ages indicates that the heterotrophic metabolic processes mediated by the genes can be ecologically important, functioning in the organic-rich subseafloor sedimentary biosphere.

Genome-Wide Mutagenesis in Borrelia burgdorferi.

PubMed

Lin, Tao; Gao, Lihui

2018-01-01

Signature-tagged mutagenesis (STM) is a functional genomics approach to identify bacterial virulence determinants and virulence factors by simultaneously screening multiple mutants in a single host animal, and has been utilized extensively for the study of bacterial pathogenesis, host-pathogen interactions, and spirochete and tick biology. The signature-tagged transposon mutagenesis has been developed to investigate virulence determinants and pathogenesis of Borrelia burgdorferi. Mutants in genes important in virulence are identified by negative selection in which the mutants fail to colonize or disseminate in the animal host and tick vector. STM procedure combined with Luminex Flex ® Map™ technology and next-generation sequencing (e.g., Tn-seq) are the powerful high-throughput tools for the determination of Borrelia burgdorferi virulence determinants. The assessment of multiple tissue sites and two DNA resources at two different time points using Luminex Flex ® Map™ technology provides a robust data set. B. burgdorferi transposon mutant screening indicates that a high proportion of genes are the novel virulence determinants that are required for mouse and tick infection. In this protocol, an effective signature-tagged Himar1-based transposon suicide vector was developed and used to generate a sequence-defined library of nearly 4800 mutants in the infectious B. burgdorferi B31 clone. In STM, signature-tagged suicide vectors are constructed by inserting unique DNA sequences (tags) into the transposable elements. The signature-tagged transposon mutants are generated when transposon suicide vectors are transformed into an infectious B. burgdorferi clone, and the transposable element is transposed into the 5'-TA-3' sequence in the B. burgdorferi genome with the signature tag. The transposon library is created and consists of many sub-libraries, each sub-library has several hundreds of mutants with same tags. A group of mice or ticks are infected with a mixed population of mutants with different tags, after recovered from different tissues of infected mice and ticks, mutants from output pool and input pool are detected using high-throughput, semi-quantitative Luminex ® FLEXMAP™ or next-generation sequencing (Tn-seq) technologies. Thus far, we have created a high-density, sequence-defined transposon library of over 6600 STM mutants for the efficient genome-wide investigation of genes and gene products required for wild-type pathogenesis, host-pathogen interactions, in vitro growth, in vivo survival, physiology, morphology, chemotaxis, motility, structure, metabolism, gene regulation, plasmid maintenance and replication, etc. The insertion sites of 4480 transposon mutants have been determined. About 800 predicted protein-encoding genes in the genome were disrupted in the STM transposon library. The infectivity and some functions of 800 mutants in 500 genes have been determined. Analysis of these transposon mutants has yielded valuable information regarding the genes and gene products important in the pathogenesis and biology of B. burgdorferi and its tick vectors.

Multiplexed droplet single-cell RNA-sequencing using natural genetic variation.

PubMed

Kang, Hyun Min; Subramaniam, Meena; Targ, Sasha; Nguyen, Michelle; Maliskova, Lenka; McCarthy, Elizabeth; Wan, Eunice; Wong, Simon; Byrnes, Lauren; Lanata, Cristina M; Gate, Rachel E; Mostafavi, Sara; Marson, Alexander; Zaitlen, Noah; Criswell, Lindsey A; Ye, Chun Jimmie

2018-01-01

Droplet single-cell RNA-sequencing (dscRNA-seq) has enabled rapid, massively parallel profiling of transcriptomes. However, assessing differential expression across multiple individuals has been hampered by inefficient sample processing and technical batch effects. Here we describe a computational tool, demuxlet, that harnesses natural genetic variation to determine the sample identity of each droplet containing a single cell (singlet) and detect droplets containing two cells (doublets). These capabilities enable multiplexed dscRNA-seq experiments in which cells from unrelated individuals are pooled and captured at higher throughput than in standard workflows. Using simulated data, we show that 50 single-nucleotide polymorphisms (SNPs) per cell are sufficient to assign 97% of singlets and identify 92% of doublets in pools of up to 64 individuals. Given genotyping data for each of eight pooled samples, demuxlet correctly recovers the sample identity of >99% of singlets and identifies doublets at rates consistent with previous estimates. We apply demuxlet to assess cell-type-specific changes in gene expression in 8 pooled lupus patient samples treated with interferon (IFN)-β and perform eQTL analysis on 23 pooled samples.

Nitrate Reduction Functional Genes and Nitrate Reduction Potentials Persist in Deeper Estuarine Sediments. Why?

PubMed Central

Papaspyrou, Sokratis; Smith, Cindy J.; Dong, Liang F.; Whitby, Corinne; Dumbrell, Alex J.; Nedwell, David B.

2014-01-01

Denitrification and dissimilatory nitrate reduction to ammonium (DNRA) are processes occurring simultaneously under oxygen-limited or anaerobic conditions, where both compete for nitrate and organic carbon. Despite their ecological importance, there has been little investigation of how denitrification and DNRA potentials and related functional genes vary vertically with sediment depth. Nitrate reduction potentials measured in sediment depth profiles along the Colne estuary were in the upper range of nitrate reduction rates reported from other sediments and showed the existence of strong decreasing trends both with increasing depth and along the estuary. Denitrification potential decreased along the estuary, decreasing more rapidly with depth towards the estuary mouth. In contrast, DNRA potential increased along the estuary. Significant decreases in copy numbers of 16S rRNA and nitrate reducing genes were observed along the estuary and from surface to deeper sediments. Both metabolic potentials and functional genes persisted at sediment depths where porewater nitrate was absent. Transport of nitrate by bioturbation, based on macrofauna distributions, could only account for the upper 10 cm depth of sediment. A several fold higher combined freeze-lysable KCl-extractable nitrate pool compared to porewater nitrate was detected. We hypothesised that his could be attributed to intracellular nitrate pools from nitrate accumulating microorganisms like Thioploca or Beggiatoa. However, pyrosequencing analysis did not detect any such organisms, leaving other bacteria, microbenthic algae, or foraminiferans which have also been shown to accumulate nitrate, as possible candidates. The importance and bioavailability of a KCl-extractable nitrate sediment pool remains to be tested. The significant variation in the vertical pattern and abundance of the various nitrate reducing genes phylotypes reasonably suggests differences in their activity throughout the sediment column. This raises interesting questions as to what the alternative metabolic roles for the various nitrate reductases could be, analogous to the alternative metabolic roles found for nitrite reductases. PMID:24728381

Correlation between Waardenburg syndrome phenotype and genotype in a population of individuals with identified PAX3 mutations.

PubMed

DeStefano, A L; Cupples, L A; Arnos, K S; Asher, J H; Baldwin, C T; Blanton, S; Carey, M L; da Silva, E O; Friedman, T B; Greenberg, J; Lalwani, A K; Milunsky, A; Nance, W E; Pandya, A; Ramesar, R S; Read, A P; Tassabejhi, M; Wilcox, E R; Farrer, L A

1998-05-01

Waardenburg syndrome (WS) type 1 is an autosomal dominant disorder characterized by sensorineural hearing loss, pigmentary abnormalities of the eye, hair, and skin, and dystopia canthorum. The phenotype is variable and affected individuals may exhibit only one or a combination of several of the associated features. To assess the relationship between phenotype and gene defect, clinical and genotype data on 48 families (271 WS individuals) collected by members of the Waardenburg Consortium were pooled. Forty-two unique mutations in the PAX3 gene, previously identified in these families, were grouped in five mutation categories: amino acid (AA) substitution in the paired domain, AA substitution in the homeodomain, deletion of the Ser-Thr-Pro-rich region, deletion of the homeodomain and the Ser-Thr-Pro-rich region, and deletion of the entire gene. These mutation classes are based on the structure of the PAX3 gene and were chosen to group mutations predicted to have similar defects in the gene product. Association between mutation class and the presence of hearing loss, eye pigment abnormality, skin hypopigmentation, or white forelock was evaluated using generalized estimating equations, which allowed for incorporation of a correlation structure that accounts for potential similarity among members of the same family. Odds for the presence of eye pigment abnormality, white forelock, and skin hypopigmentation were 2, 8, and 5 times greater, respectively, for individuals with deletions of the homeodomain and the Pro-Ser-Thr-rich region compared to individuals with an AA substitution in the homeodomain. Odds ratios that differ significantly from 1.0 for these traits may indicate that the gene products resulting from different classes of mutations act differently in the expression of WS. Although a suggestive association was detected for hearing loss with an odds ratio of 2.6 for AA substitution in the paired domain compared with AA substitution in the homeodomain, this odds ratio did not differ significantly from 1.0.

Genetic diversity and genomic signatures of selection among cattle breeds from Siberia, eastern and northern Europe.

PubMed

Iso-Touru, T; Tapio, M; Vilkki, J; Kiseleva, T; Ammosov, I; Ivanova, Z; Popov, R; Ozerov, M; Kantanen, J

2016-12-01

Domestication in the near eastern region had a major impact on the gene pool of humpless taurine cattle (Bos taurus). As a result of subsequent natural and artificial selection, hundreds of different breeds have evolved, displaying a broad range of phenotypic traits. Here, 10 Eurasian B. taurus breeds from different biogeographic and production conditions, which exhibit different demographic histories and have been under artificial selection at various intensities, were investigated using the Illumina BovineSNP50 panel to understand their genetic diversity and population structure. In addition, we scanned genomes from eight breeds for signatures of diversifying selection. Our population structure analysis indicated six distinct breed groups, the most divergent being the Yakutian cattle from Siberia. Selection signals were shared (experimental P-value < 0.01) with more than four breeds on chromosomes 6, 7, 13, 16 and 22. The strongest selection signals in the Yakutian cattle were found on chromosomes 7 and 21, where a miRNA gene and genes related to immune system processes are respectively located. In general, genomic regions indicating selection overlapped with known QTL associated with milk production (e.g. on chromosome 19), reproduction (e.g. on chromosome 24) and meat quality (e.g. on chromosome 7). The selection map created in this study shows that native cattle breeds and their genetic resources represent unique material for future breeding. © 2016 Stichting International Foundation for Animal Genetics.

Molecular characterization of a gene POLR2H encoded an essential subunit for RNA polymerase II from the Giant Panda (Ailuropoda Melanoleuca).

PubMed

Du, Yu-Jie; Hou, Yi-Ling; Hou, Wan-Ru

2013-02-01

The Giant Panda is an endangered and valuable gene pool in genetic, its important functional gene POLR2H encodes an essential shared peptide H of RNA polymerases. The genomic DNA and cDNA sequences were cloned successfully for the first time from the Giant Panda (Ailuropoda melanoleuca) adopting touchdown-PCR and reverse transcription polymerase chain reaction (RT-PCR), respectively. The length of the genomic sequence of the Giant Panda is 3,285 bp, including five exons and four introns. The cDNA fragment cloned is 509 bp in length, containing an open reading frame of 453 bp encoding 150 amino acids. Alignment analysis indicated that both the cDNA and its deduced amino acid sequence were highly conserved. Protein structure prediction showed that there was one protein kinase C phosphorylation site, four casein kinase II phosphorylation sites and one amidation site in the POLR2H protein, further shaping advanced protein structure. The cDNA cloned was expressed in Escherichia coli, which indicated that POLR2H fusion with the N-terminally His-tagged form brought about the accumulation of an expected 20.5 kDa polypeptide in line with the predicted protein. On the basis of what has already been achieved in this study, further deep-in research will be conducted, which has great value in theory and practical significance.

The gravity of pollination: integrating at-site features into spatial analysis of contemporary pollen movement.

PubMed

DiLeo, Michelle F; Siu, Jenna C; Rhodes, Matthew K; López-Villalobos, Adriana; Redwine, Angela; Ksiazek, Kelly; Dyer, Rodney J

2014-08-01

Pollen-mediated gene flow is a major driver of spatial genetic structure in plant populations. Both individual plant characteristics and site-specific features of the landscape can modify the perceived attractiveness of plants to their pollinators and thus play an important role in shaping spatial genetic variation. Most studies of landscape-level genetic connectivity in plants have focused on the effects of interindividual distance using spatial and increasingly ecological separation, yet have not incorporated individual plant characteristics or other at-site ecological variables. Using spatially explicit simulations, we first tested the extent to which the inclusion of at-site variables influencing local pollination success improved the statistical characterization of genetic connectivity based upon examination of pollen pool genetic structure. The addition of at-site characteristics provided better models than those that only considered interindividual spatial distance (e.g. IBD). Models parameterized using conditional genetic covariance (e.g. population graphs) also outperformed those assuming panmixia. In a natural population of Cornus florida L. (Cornaceae), we showed that the addition of at-site characteristics (clumping of primary canopy opening above each maternal tree and maternal tree floral output) provided significantly better models describing gene flow than models including only between-site spatial (IBD) and ecological (isolation by resistance) variables. Overall, our results show that including interindividual and local ecological variation greatly aids in characterizing landscape-level measures of contemporary gene flow. © 2014 John Wiley & Sons Ltd.

Evolution and ecology of influenza A viruses.

PubMed Central

Webster, R G; Bean, W J; Gorman, O T; Chambers, T M; Kawaoka, Y

1992-01-01

In this review we examine the hypothesis that aquatic birds are the primordial source of all influenza viruses in other species and study the ecological features that permit the perpetuation of influenza viruses in aquatic avian species. Phylogenetic analysis of the nucleotide sequence of influenza A virus RNA segments coding for the spike proteins (HA, NA, and M2) and the internal proteins (PB2, PB1, PA, NP, M, and NS) from a wide range of hosts, geographical regions, and influenza A virus subtypes support the following conclusions. (i) Two partly overlapping reservoirs of influenza A viruses exist in migrating waterfowl and shorebirds throughout the world. These species harbor influenza viruses of all the known HA and NA subtypes. (ii) Influenza viruses have evolved into a number of host-specific lineages that are exemplified by the NP gene and include equine Prague/56, recent equine strains, classical swine and human strains, H13 gull strains, and all other avian strains. Other genes show similar patterns, but with extensive evidence of genetic reassortment. Geographical as well as host-specific lineages are evident. (iii) All of the influenza A viruses of mammalian sources originated from the avian gene pool, and it is possible that influenza B viruses also arose from the same source. (iv) The different virus lineages are predominantly host specific, but there are periodic exchanges of influenza virus genes or whole viruses between species, giving rise to pandemics of disease in humans, lower animals, and birds. (v) The influenza viruses currently circulating in humans and pigs in North America originated by transmission of all genes from the avian reservoir prior to the 1918 Spanish influenza pandemic; some of the genes have subsequently been replaced by others from the influenza gene pool in birds. (vi) The influenza virus gene pool in aquatic birds of the world is probably perpetuated by low-level transmission within that species throughout the year. (vii) There is evidence that most new human pandemic strains and variants have originated in southern China. (viii) There is speculation that pigs may serve as the intermediate host in genetic exchange between influenza viruses in avian and humans, but experimental evidence is lacking. (ix) Once the ecological properties of influenza viruses are understood, it may be possible to interdict the introduction of new influenza viruses into humans. Images PMID:1579108

Distribution and Differentiation of Wild, Feral, and Cultivated Populations of Perennial Upland Cotton (Gossypium hirsutum L.) in Mesoamerica and the Caribbean

PubMed Central

Coppens d'Eeckenbrugge, Geo; Lacape, Jean-Marc

2014-01-01

Perennial forms of Gossypium hirsutum are classified under seven races. Five Mesoamerican races would have been derived from the wild race ‘yucatanense’ from northern Yucatán. ‘Marie-Galante’, the main race in the Caribbean, would have developed from introgression with G. barbadense. The racial status of coastal populations from the Caribbean has not been clearly defined. We combined Ecological Niche Modeling with an analysis of SSR marker diversity, to elucidate the relationships among cultivated, feral and wild populations of perennial cottons. Out of 954 records of occurrence in Mesoamerica and the Caribbean, 630 were classified into four categories cultivated, feral (disturbed and secondary habitats), wild/feral (protected habitats), and truly wild cotton (TWC) populations. The widely distributed three first categories cannot be differentiated on ecological grounds, indicating they mostly belong to the domesticated pool. In contrast, TWC are restricted to the driest and hottest littoral habitats, in northern Yucatán and in the Caribbean (from Venezuela to Florida), as confirmed by their climatic envelope in the factorial analysis. Extrapolating this TWC climatic model to South America and the Pacific Ocean points towards places where other wild representatives of tetraploid Gossypium species have been encountered. The genetic analysis sample comprised 42 TWC accessions from 12 sites and 68 feral accessions from 18 sites; at nine sites, wild and feral accessions were collected in close vicinity. Principal coordinate analysis, neighbor joining, and STRUCTURE consistently showed a primary divergence between TWC and feral cottons, and a secondary divergence separating ‘Marie-Galante’ from all other feral accessions. This strong genetic structure contrasts strikingly with the absence of geographic differentiation. Our results show that TWC populations of Mesoamerica and the Caribbean constitute a homogenous gene pool. Furthermore, the relatively low genetic divergence between the Mesoamerican and Caribbean domesticated pools supports the hypothesis of domestication of G. hirsutum in northern Yucatán. PMID:25198534

Stress inducible proteinase inhibitor diversity in Capsicum annuum

PubMed Central

2012-01-01

Background Wound-inducible Pin-II Proteinase inhibitors (PIs) are one of the important plant serine PIs which have been studied extensively for their structural and functional diversity and relevance in plant defense against insect pests. To explore the functional specialization of an array of Capsicum annuum (L.) proteinase inhibitor (CanPIs) genes, we studied their expression, processing and tissue-specific distribution under steady-state and induced conditions. Inductions were performed by subjecting C. annuum leaves to various treatments, namely aphid infestation or mechanical wounding followed by treatment with either oral secretion (OS) of Helicoverpa armigera or water. Results The elicitation treatments regulated the accumulation of CanPIs corresponding to 4-, 3-, and 2-inhibitory repeat domains (IRDs). Fourty seven different CanPI genes composed of 28 unique IRDs were identified in total along with those reported earlier. The CanPI gene pool either from uninduced or induced leaves was dominated by 3-IRD PIs and trypsin inhibitory domains. Also a major contribution by 4-IRD CanPI genes possessing trypsin and chymotrypsin inhibitor domains was specifically revealed in wounded leaves treated with OS. Wounding displayed the highest number of unique CanPIs while wounding with OS treatment resulted in the high accumulation of specifically CanPI-4, -7 and −10. Characterization of the PI protein activity through two dimensional gel electrophoresis revealed tissue and induction specific patterns. Consistent with transcript abundance, wound plus OS or water treated C. annuum leaves exhibited significantly higher PI activity and isoform diversity contributed by 3- and 4-IRD CanPIs. CanPI accumulation and activity was weakly elicited by aphid infestation yet resulted in the higher expression of CanPI-26, -41 and −43. Conclusions Plants can differentially perceive various kinds of insect attacks and respond appropriately through activating plant defenses including regulation of PIs at transcriptional and post-translational levels. Based on the differentially elicited CanPI accumulation patterns, it is intriguing to speculate that generating sequence diversity in the form of multi-IRD PIs is a part of elaborative plant defense strategy to obtain a diverse pool of functional units to confine insect attack. PMID:23153298

Recombinant protein production from stable mammalian cell lines and pools.

PubMed

Hacker, David L; Balasubramanian, Sowmya

2016-06-01

We highlight recent developments for the production of recombinant proteins from suspension-adapted mammalian cell lines. We discuss the generation of stable cell lines using transposons and lentivirus vectors (non-targeted transgene integration) and site-specific recombinases (targeted transgene integration). Each of these methods results in the generation of cell lines with protein yields that are generally superior to those achievable through classical plasmid transfection that depends on the integration of the transfected DNA by non-homologous DNA end-joining. This is the main reason why these techniques can also be used for the generation of stable cell pools, heterogenous populations of recombinant cells generated by gene delivery and genetic selection without resorting to single cell cloning. This allows the time line from gene transfer to protein production to be reduced. Copyright © 2016 Elsevier Ltd. All rights reserved.

The Polynesian gene pool: an early contribution by Amerindians to Easter Island.

PubMed

Thorsby, Erik

2012-03-19

It is now generally accepted that Polynesia was first settled by peoples from southeast Asia. An alternative that eastern parts of Polynesia were first inhabited by Amerindians has found little support. There are, however, many indications of a 'prehistoric' (i.e. before Polynesia was discovered by Europeans) contact between Polynesia and the Americas, but genetic evidence of a prehistoric Amerindian contribution to the Polynesian gene pool has been lacking. We recently carried out genomic HLA (human leucocyte antigen) typing as well as typing for mitochondrial DNA (mtDNA) and Y chromosome markers of blood samples collected in 1971 and 2008 from reputedly non-admixed Easter Islanders. All individuals carried HLA alleles and mtDNA types previously found in Polynesia, and most of the males carried Y chromosome markers of Polynesian origin (a few had European Y chromosome markers), further supporting an initial Polynesian population on Easter Island. The HLA investigations revealed, however, that some individuals also carried HLA alleles which have previously almost only been found in Amerindians. We could trace the introduction of these Amerindian alleles to before the Peruvian slave trades, i.e. before the 1860s, and provide suggestive evidence that they were introduced already in prehistoric time. Our results demonstrate an early Amerindian contribution to the Polynesian gene pool on Easter Island, and illustrate the usefulness of typing for immunogenetic markers such as HLA to complement mtDNA and Y chromosome analyses in anthropological investigations.

Comparison of microsatellite variations between Red Junglefowl and a commercial chicken gene pool.

PubMed

Tadano, R; Kinoshita, K; Mizutani, M; Tsudzuki, M

2014-02-01

It is assumed that Red Junglefowl (Gallus gallus) is one of the main ancestors of domestic chickens (Gallus gallus domesticus). Differences in microsatellite polymorphisms between Red Junglefowl and modern commercial chickens, which are used for egg and meat production, have not been fully reported. A total of 361 individuals from 1 Red Junglefowl population that has been maintained as a closed flock, 5 final cross-bred commercial layer populations (white-, tinted-, and brown-egg layers), and 2 final cross-bred commercial broiler populations were genotyped for 40 autosomal microsatellite loci. We compared microsatellite variations in Red Junglefowl with those in a commercial chicken gene pool. The contribution of each population to the genetic diversity was also estimated based on the molecular coancestry. In total, 302 distinct alleles were detected in 1 Red Junglefowl and 7 commercial chicken populations, of which 31 alleles (10.3%) were unique to Red Junglefowl, most of which occurred at a high frequency. The genetic differentiation between Red Junglefowl and commercial chickens (pairwise FST) ranged from 0.32 to 0.47. According to the neighbor-joining tree based on the modified Cavalli-Sforza chord distances and the Bayesian clustering analysis, Red Junglefowl was genetically distant from the commercial chicken gene pool tested. In all of the populations analyzed, Red Junglefowl made the highest contribution to genetic diversity. These results suggest that Red Junglefowl has a distinct distribution of microsatellite alleles and that there is a high level of genetic divergence between Red Junglefowl and commercial chickens.

Association of Endothelial Nitric Oxide Synthase Gene Polymorphisms with Coronary Artery Disease: An Updated Meta-Analysis and Systematic Review

PubMed Central

Parveen, Farah; Kapoor, Aditya; Sinha, Nakul

2014-01-01

Several association studies of endothelial nitric oxide synthase (NOS3) gene polymorphisms with respect to coronary artery disease (CAD) have been published in the past two decades. However, their association with the disease, especially among different ethnic subgroups, still remains controversial. This prompted us to conduct a systematic review and an updated structured meta-analysis, which is the largest so far (89 articles, 132 separate studies, and a sample size of 69,235), examining association of three polymorphic forms of the NOS3 gene (i.e. Glu298Asp, T786-C and 27bp VNTR b/a) with CAD. In a subgroup analysis, we tested their association separately among published studies originating predominantly from European, Middle Eastern, Asian, Asian-Indian and African ancestries. The pooled analysis confirmed the association of all the three selected SNP with CAD in three different genetic models transcending all ancestries worldwide. The Glu298Asp polymorphism showed strongest association (OR range = 1.28–1.52, and P<0.00001 for all comparisons), followed by T786-C (OR range = 1.34–1.42, and P<0.00001 for all comparisons) and 4b/a, (OR range = 1.19–1.41, and P≤0.002 for all comparisons) in our pooled analysis. Subgroup analysis revealed that Glu298Asp (OR range = 1.54–1.87, and P<0.004 for all comparisons) and 4b/a (OR range = 1.71–3.02, and P<0.00001 for all comparisons) have highest degree of association amongst the Middle Easterners. On the other hand, T786-C and its minor allele seem to carry a highest risk for CAD among subjects of Asian ancestry (OR range = 1.61–1.90, and P≤0.01 for all comparisons). PMID:25409023

Comparative Genomics and Immunoinformatics Approach for the Identification of Vaccine Candidates for Enterohemorrhagic Escherichia coli O157:H7

PubMed Central

García-Angulo, Víctor A.; Kalita, Anjana; Kalita, Mridul; Lozano, Luis

2014-01-01

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 strains are major human food-borne pathogens, responsible for bloody diarrhea and hemolytic-uremic syndrome worldwide. Thus far, there is no vaccine for humans against EHEC infections. In this study, a comparative genomics analysis was performed to identify EHEC-specific antigens useful as potential vaccines. The genes present in both EHEC EDL933 and Sakai strains but absent in nonpathogenic E. coli K-12 and HS strains were subjected to an in silico analysis to identify secreted or surface-expressed proteins. We obtained a total of 65 gene-encoding protein candidates, which were subjected to immunoinformatics analysis. Our criteria of selection aided in categorizing the candidates as high, medium, and low priority. Three members of each group were randomly selected and cloned into pVAX-1. Candidates were pooled accordingly to their priority group and tested for immunogenicity against EHEC O157:H7 using a murine model of gastrointestinal infection. The high-priority (HP) pool, containing genes encoding a Lom-like protein (pVAX-31), a putative pilin subunit (pVAX-12), and a fragment of the type III secretion structural protein EscC (pVAX-56.2), was able to induce the production of EHEC IgG and sIgA in sera and feces. HP candidate-immunized mice displayed elevated levels of Th2 cytokines and diminished cecum colonization after wild-type challenge. Individually tested HP vaccine candidates showed that pVAX-12 and pVAX-56.2 significantly induced Th2 cytokines and production of fecal EHEC sIgA, with pVAX-56.2 reducing EHEC cecum colonization. We describe here a bioinformatics approach able to identify novel vaccine candidates potentially useful for preventing EHEC O157:H7 infections. PMID:24595137

Diversity hotspots of the laurel forest on Tenerife, Canary Islands: a phylogeographic study of Laurus and Ixanthus.

PubMed

Betzin, Anja; Thiv, Mike; Koch, Marcus A

2016-09-01

Macaronesian laurel forest is among the worldwide hotspots of threatened biodiversity. With increasing evidence that woodland composition on the Canary Islands changed dramatically during the last few thousand years, the aim of this study was to find evidence for substantial recent population dynamics of two representative species from laurel forest. Amplified fragment length polymorphism (AFLP) was used to evaluate fine-scaled genetic variation of the paradigmatic tree Laurus novocanariensis (Lauraceae) and a long-lived herbaceous gentian from core laurel forest, Ixanthus viscosus (Gentianaceae), on Tenerife. Bioclimatic variables were analysed to study the respective climate niches. A chloroplast DNA screening was performed to evaluate additional genetic variation. Genetic diversity of the laurel tree showed severe geographic partitioning. On Tenerife, fine-scaled Bayesian clustering of genetic variation revealed a western and an eastern gene pool, separated by a zone of high admixture and with a third major gene pool. Compared with genetic clusters found on the other Canary Islands, the East-West differentiation on Tenerife seems to be more recent than differentiation between islands. This is substantiated by the finding of extremly low levels of chloroplast DNA-based polymorphisms. Ixanthus showed no geographic structuring of genetic variation. Genetic data from Tenerife indicate contemporary gene flow and dispersal on a micro/local scale rather than reflecting an old and relic woodland history. In particular for Laurus, it is shown that this species occupies a broad bioclimatic niche. This is not correlated with its respective distribution of genetic variation, therefore indicating its large potential for contemporary rapid and effective colonization. Ixanthus is more specialized to humid conditions and is mostly found in the natural Monteverde húmedo vegetation types, but even for this species indications for long-term persistence in the respective bioclimatically differentiated regions was not find. © The Author 2016. Published by Oxford University Press on behalf of the Annals of Botany Company. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Fecal Carriage of Extended-Spectrum β-Lactamases in Healthy Humans, Poultry, and Wild Birds in León, Nicaragua-A Shared Pool of blaCTX-M Genes and Possible Interspecies Clonal Spread of Extended-Spectrum β-Lactamases-Producing Escherichia coli.

PubMed

Hasan, Badrul; Laurell, Karl; Rakib, Mufti Mahmud; Ahlstedt, Erik; Hernandez, Jorge; Caceres, Mercedes; Järhult, Josef D

2016-12-01

Antibiotic-resistant bacteria are a major concern in the healthcare of today, especially the increasing number of gram-negative bacteria producing β-lactamases such as extended-spectrum β-lactamases (ESBLs). However, little is known about the relationship of ESBL producers in humans and domestic and wild birds, especially in a low-income setting. Therefore, we studied the fecal carriage of ESBL-producing Escherichia coli and Klebsiella pneumoniae in healthy humans, poultry, and wild birds in the vicinity of León, Nicaragua. Three hundred fecal samples were collected during December 2012 from humans (n = 100), poultry (n = 100) and wild birds (n = 100). The samples were examined for ESBL-producing E. coli and K. pneumoniae, revealing the prevalence of 27% in humans, 13% in poultry, and 8% in wild birds. Further characterization of the ESBL-producing isolates was performed through polymerase chain reaction (PCR) (NDM, CTX-M), epidemiological typing (ERIC2-PCR), multilocus sequence typing, and sequencing. ESBL producers harbored bla CTX-M-2 , bla CTX-M-15 , bla CTX-M-22 , and bla CTX-M-3 genotypes. The bla CTX-M-15 constituted the absolute majority of ESBL genes among all samples. ERIC-PCR demonstrated highly related E. coli clones among humans, poultry, and wild birds. Clinically relevant E. coli clone ST648 was found in humans and poultry. There is a shared pool of bla CTX-M genes between humans and domesticated and wild birds in Nicaragua, and the results suggest shared clones of ESBL-producing E. coli. The study adds to the notion that wild birds and poultry can pick up antibiotic-resistant bacteria of human origin and function as a melting pot of resistance. Structured surveillance programs of antimicrobial resistance and a more regulated prescription of antibiotics are warranted in Nicaragua.

Crystal structure of AFV3-109, a highly conserved protein from crenarchaeal viruses

PubMed Central

Keller, Jenny; Leulliot, Nicolas; Cambillau, Christian; Campanacci, Valérie; Porciero, Stéphanie; Prangishvili, David; Forterre, Patrick; Cortez, Diego; Quevillon-Cheruel, Sophie; van Tilbeurgh, Herman

2007-01-01

The extraordinary morphologies of viruses infecting hyperthermophilic archaea clearly distinguish them from bacterial and eukaryotic viruses. Moreover, their genomes code for proteins that to a large extend have no related sequences in the extent databases. However, a small pool of genes is shared by overlapping subsets of these viruses, and the most conserved gene, exemplified by the ORF109 of the Acidianus Filamentous Virus 3, AFV3, is present on genomes of members of three viral familes, the Lipothrixviridae, Rudiviridae, and "Bicaudaviridae", as well as of the unclassified Sulfolobus Turreted Icosahedral Virus, STIV. We present here the crystal structure of the protein (Mr = 13.1 kD, 109 residues) encoded by the AFV3 ORF 109 in two different crystal forms at 1.5 and 1.3 Å resolution. The structure of AFV3-109 is a five stranded β-sheet with loops on one side and three helices on the other. It forms a dimer adopting the shape of a cradle that encompasses the best conserved regions of the sequence. No protein with a related fold could be identified except for the ortholog from STIV1, whose structure was deposited at the Protein Data Bank. We could clearly identify a well bound glycerol inside the cradle, contacting exclusively totally conserved residues. This interaction was confirmed in solution by fluorescence titration. Although the function of AFV3-109 cannot be deduced directly from its structure, structural homology with the STIV1 protein, and the size and charge distribution of the cavity suggested it could interact with nucleic acids. Fluorescence quenching titrations also showed that AFV3-109 interacts with dsDNA. Genomic sequence analysis revealed bacterial homologs of AFV3-109 as a part of a putative previously unidentified prophage sequences in some Firmicutes. PMID:17241456

Interactions between Natural Organic Matter and Native Microbes in the Oak Ridge FRC Groundwater

NASA Astrophysics Data System (ADS)

Wu, X.; Hazen, T.; Fox, P. M.; Nico, P. S.; Li, Q.; Yang, W.; Liu, Y.; Hess, N. J.; Zhang, P.; Qin, Y.; Zhou, J.; Chakraborty, R.

2016-12-01

Natural organic matter (NOM) is central to microbial food webs; however, little is known about the interplay between the physical and chemical characteristics of the carbon in NOM and its turnover by microbial communities. Microbial activity changes NOM's structure and properties, which may further influence the bioavailability of NOM. The change of NOM may reversely affect the microbial community structure as well. To date, our understanding of these interactions is insufficient, and it is critical to identify the role of NOM to carbon turnover, structure of microbial community and to the metabolic potential of that community. In this study, we aimed to study the interactions between NOM and native microbial communities present in groundwater at a background site (FW305 well) at Oak Ridge Field Research Center, TN. The total organic carbon and inorganic carbon in FW305 deep sediment samples were 0.071% and 0.011%, respectively. Water-soluble NOM was extracted from these sediment samples, the extraction efficiencies were 3.2% for organic carbon and 1.6% for inorganic carbon. The extracted NOM was then provided as the sole carbon source to native microbes present in groundwater. Subsamples were harvested several times from these incubations during a 50-day study. 16S rRNA gene amplicon sequencing and Geochip were used to identify the changes of microbial communities and expression of functional genes during transformation of the NOM. Several advanced chemical techniques including FTICR-MS and NEXAFS were used to characterize the C pool (i.e., NOM metabolites and microbial byproducts). Preliminary data clearly showed that microbial community responded to NOM, and shifted as functional groups in NOM transformed. Further detailed metabolite and gene-based analysis to elucidate these changes is currently being conducted.

Transcriptomic Signature of the SHATTERPROOF2 Expression Domain Reveals the Meristematic Nature of Arabidopsis Gynoecial Medial Domain1[OPEN

PubMed Central

Villarino, Gonzalo H.; Hu, Qiwen; Flores-Vergara, Miguel; Sehra, Bhupinder; Brumos, Javier; Stepanova, Anna N.; Sundberg, Eva; Heber, Steffen

2016-01-01

Plant meristems, like animal stem cell niches, maintain a pool of multipotent, undifferentiated cells that divide and differentiate to give rise to organs. In Arabidopsis (Arabidopsis thaliana), the carpel margin meristem is a vital meristematic structure that generates ovules from the medial domain of the gynoecium, the female floral reproductive structure. The molecular mechanisms that specify this meristematic region and regulate its organogenic potential are poorly understood. Here, we present a novel approach to analyze the transcriptional signature of the medial domain of the Arabidopsis gynoecium, highlighting the developmental stages that immediately proceed ovule initiation, the earliest stages of seed development. Using a floral synchronization system and a SHATTERPROOF2 (SHP2) domain-specific reporter, paired with FACS and RNA sequencing, we assayed the transcriptome of the gynoecial medial domain with temporal and spatial precision. This analysis reveals a set of genes that are differentially expressed within the SHP2 expression domain, including genes that have been shown previously to function during the development of medial domain-derived structures, including the ovules, thus validating our approach. Global analyses of the transcriptomic data set indicate a similarity of the pSHP2-expressing cell population to previously characterized meristematic domains, further supporting the meristematic nature of this gynoecial tissue. Our method identifies additional genes including novel isoforms, cis-natural antisense transcripts, and a previously unrecognized member of the REPRODUCTIVE MERISTEM family of transcriptional regulators that are potential novel regulators of medial domain development. This data set provides genome-wide transcriptional insight into the development of the carpel margin meristem in Arabidopsis. PMID:26983993

Seasonal Patterns of the Insect Community Structure in Urban Rain Pools of Temperate Argentina

PubMed Central

Fontanarrosa, M. Soledad; Collantes, Marta B.; Bachmann, Axel O.

2009-01-01

Temporary aquatic environments are widespread in the world, and although there are considerable regional differences in their type and method of formation they have many physical, chemical and biological properties in common. With the aim to increase knowledge of urban temporary pool fauna, the objectives of this work were to assess the seasonal patterns of species composition, richness, and diversity of the aquatic insect community inhabiting rain pools in urban temperate Argentina, and to identify the environmental variables associated to these patterns. Four temporary pools of an urban green space in Buenos Aires City were studied throughout a 1-year period. Eleven flood cycles with very varied hydroperiods and dry periods, mainly associated with rainfall, were identified. Insect species richness in these temporary urban pools, 86 taxa were documented, was found to be within the range reported for wild temporary water bodies of other regions of the world. The present results provide evidence for the existence of a clear link between habitat and community variability. Hydroperiod and seasonality were the main environmental factors involved in structuring the insect communities of the studied water bodies. Urban pools in green spaces have the potential to act to its dwellers like corridors through the urban matrix. Taking into account these characteristics and their accessibility, urban temporary pools can be considered as promising habitats for the study of ecological processes involving the insect community. PMID:19611261

Metagenomic evidence for metabolism of trace atmospheric gases by high-elevation desert Actinobacteria

PubMed Central

Lynch, Ryan C.; Darcy, John L.; Kane, Nolan C.; Nemergut, Diana R.; Schmidt, Steve K.

2014-01-01

Previous surveys of very dry Atacama Desert mineral soils have consistently revealed sparse communities of non-photosynthetic microbes. The functional nature of these microorganisms remains debatable given the harshness of the environment and low levels of biomass and diversity. The aim of this study was to gain an understanding of the phylogenetic community structure and metabolic potential of a low-diversity mineral soil metagenome that was collected from a high-elevation Atacama Desert volcano debris field. We pooled DNA extractions from over 15 g of volcanic material, and using whole genome shotgun sequencing, observed only 75–78 total 16S rRNA gene OTUs3%. The phylogenetic structure of this community is significantly under dispersed, with actinobacterial lineages making up 97.9–98.6% of the 16S rRNA genes, suggesting a high degree of environmental selection. Due to this low diversity and uneven community composition, we assembled and analyzed the metabolic pathways of the most abundant genome, a Pseudonocardia sp. (56–72% of total 16S genes). Our assembly and binning efforts yielded almost 4.9 Mb of Pseudonocardia sp. contigs, which accounts for an estimated 99.3% of its non-repetitive genomic content. This genome contains a limited array of carbohydrate catabolic pathways, but encodes for CO2 fixation via the Calvin cycle. The genome also encodes complete pathways for the catabolism of various trace gases (H2, CO and several organic C1 compounds) and the assimilation of ammonia and nitrate. We compared genomic content among related Pseudonocardia spp. and estimated rates of non-synonymous and synonymous nucleic acid substitutions between protein coding homologs. Collectively, these comparative analyses suggest that the community structure and various functional genes have undergone strong selection in the nutrient poor desert mineral soils and high-elevation atmospheric conditions. PMID:25566214

Genome sequence of Thalassospira xiamenensis type strain M-5.

PubMed

Lai, Qiliang; Shao, Zongze

2012-12-01

Thalassospira xiamenensis M-5(T) was isolated from the surface water of a waste oil pool at the oil storage dock in the city of Xiamen, Fujian Province, China. Here, we present the draft genome of strain M-5(T), which contains 4,705,237 bp with a G+C content of 54.65% and contains 4,343 protein-coding genes and 46 tRNA genes.

Integrative Analysis of High-throughput Cancer Studies with Contrasted Penalization

PubMed Central

Shi, Xingjie; Liu, Jin; Huang, Jian; Zhou, Yong; Shia, BenChang; Ma, Shuangge

2015-01-01

In cancer studies with high-throughput genetic and genomic measurements, integrative analysis provides a way to effectively pool and analyze heterogeneous raw data from multiple independent studies and outperforms “classic” meta-analysis and single-dataset analysis. When marker selection is of interest, the genetic basis of multiple datasets can be described using the homogeneity model or the heterogeneity model. In this study, we consider marker selection under the heterogeneity model, which includes the homogeneity model as a special case and can be more flexible. Penalization methods have been developed in the literature for marker selection. This study advances from the published ones by introducing the contrast penalties, which can accommodate the within- and across-dataset structures of covariates/regression coefficients and, by doing so, further improve marker selection performance. Specifically, we develop a penalization method that accommodates the across-dataset structures by smoothing over regression coefficients. An effective iterative algorithm, which calls an inner coordinate descent iteration, is developed. Simulation shows that the proposed method outperforms the benchmark with more accurate marker identification. The analysis of breast cancer and lung cancer prognosis studies with gene expression measurements shows that the proposed method identifies genes different from those using the benchmark and has better prediction performance. PMID:24395534

Identifying currents in the gene pool for bacterial populations using an integrative approach.

PubMed

Tang, Jing; Hanage, William P; Fraser, Christophe; Corander, Jukka

2009-08-01

The evolution of bacterial populations has recently become considerably better understood due to large-scale sequencing of population samples. It has become clear that DNA sequences from a multitude of genes, as well as a broad sample coverage of a target population, are needed to obtain a relatively unbiased view of its genetic structure and the patterns of ancestry connected to the strains. However, the traditional statistical methods for evolutionary inference, such as phylogenetic analysis, are associated with several difficulties under such an extensive sampling scenario, in particular when a considerable amount of recombination is anticipated to have taken place. To meet the needs of large-scale analyses of population structure for bacteria, we introduce here several statistical tools for the detection and representation of recombination between populations. Also, we introduce a model-based description of the shape of a population in sequence space, in terms of its molecular variability and affinity towards other populations. Extensive real data from the genus Neisseria are utilized to demonstrate the potential of an approach where these population genetic tools are combined with an phylogenetic analysis. The statistical tools introduced here are freely available in BAPS 5.2 software, which can be downloaded from http://web.abo.fi/fak/mnf/mate/jc/software/baps.html.

Intraspecies Genomic Diversity and Natural Population Structure of the Meat-Borne Lactic Acid Bacterium Lactobacillus sakei▿ †

PubMed Central

Chaillou, Stéphane; Daty, Marie; Baraige, Fabienne; Dudez, Anne-Marie; Anglade, Patricia; Jones, Rhys; Alpert, Carl-Alfred; Champomier-Vergès, Marie-Christine; Zagorec, Monique

2009-01-01

Lactobacillus sakei is a food-borne bacterium naturally found in meat and fish products. A study was performed to examine the intraspecies diversity among 73 isolates sourced from laboratory collections in several different countries. Pulsed-field gel electrophoresis analysis demonstrated a 25% variation in genome size between isolates, ranging from 1,815 kb to 2,310 kb. The relatedness between isolates was then determined using a PCR-based method that detects the possession of 60 chromosomal genes belonging to the flexible gene pool. Ten different strain clusters were identified that had noticeable differences in their average genome size reflecting the natural population structure. The results show that many different genotypes may be isolated from similar types of meat products, suggesting a complex ecological habitat in which intraspecies diversity may be required for successful adaptation. Finally, proteomic analysis revealed a slight difference between the migration patterns of highly abundant GapA isoforms of the two prevailing L. sakei subspecies (sakei and carnosus). This analysis was used to affiliate the genotypic clusters with the corresponding subspecies. These findings reveal for the first time the extent of intraspecies genomic diversity in L. sakei. Consequently, identification of molecular subtypes may in the future prove valuable for a better understanding of microbial ecosystems in food products. PMID:19114527

Population structure of elephant foot yams (Amorphophallus paeoniifolius (Dennst.) Nicolson) in Asia

PubMed Central

Misra, Raj Shekhar; Boonkorkaew, Patchareeya; Thanomchit, Kanokwan

2017-01-01

The corms and leaves of elephant foot yams (Amorphophallus paeoniifolius (Dennst.) Nicolson) are important foods in the local diet in many Asian regions. The crop has high productivity and wide agroecological adaptation and exhibits suitability for the agroforestry system. Although the plant is assumed to reproduce via panmixia, a comprehensive study on the genetic background across regions to enhance wider consumer palatability is still lacking. Here, ten informative microsatellites were analyzed in 29 populations across regions in India, Indonesia and Thailand to understand the genetic diversity, population structure and distribution to improve breeding and conservation programs. The genetic diversity was high among and within regions. Some populations exhibited excess heterozygosity and bottlenecking. Pairwise FST indicated very high genetic differentiation across regions (FST = 0.274), and the Asian population was unlikely to be panmictic. Phylogenetic tree construction grouped the populations according to country of origin with the exception of the Medan population from Indonesia. The current gene flow was apparent within the regions but was restricted among the regions. The present study revealed that Indonesia and Thailand populations could be alternative centers of the gene pool, together with India. Consequently, regional action should be incorporated in genetic conservation and breeding efforts to develop new varieties with global acceptance. PMID:28658282

Intraspecies genomic diversity and natural population structure of the meat-borne lactic acid bacterium Lactobacillus sakei.

PubMed

Chaillou, Stéphane; Daty, Marie; Baraige, Fabienne; Dudez, Anne-Marie; Anglade, Patricia; Jones, Rhys; Alpert, Carl-Alfred; Champomier-Vergès, Marie-Christine; Zagorec, Monique

2009-02-01

Lactobacillus sakei is a food-borne bacterium naturally found in meat and fish products. A study was performed to examine the intraspecies diversity among 73 isolates sourced from laboratory collections in several different countries. Pulsed-field gel electrophoresis analysis demonstrated a 25% variation in genome size between isolates, ranging from 1,815 kb to 2,310 kb. The relatedness between isolates was then determined using a PCR-based method that detects the possession of 60 chromosomal genes belonging to the flexible gene pool. Ten different strain clusters were identified that had noticeable differences in their average genome size reflecting the natural population structure. The results show that many different genotypes may be isolated from similar types of meat products, suggesting a complex ecological habitat in which intraspecies diversity may be required for successful adaptation. Finally, proteomic analysis revealed a slight difference between the migration patterns of highly abundant GapA isoforms of the two prevailing L. sakei subspecies (sakei and carnosus). This analysis was used to affiliate the genotypic clusters with the corresponding subspecies. These findings reveal for the first time the extent of intraspecies genomic diversity in L. sakei. Consequently, identification of molecular subtypes may in the future prove valuable for a better understanding of microbial ecosystems in food products.

Saturation of an Intra-Gene Pool Linkage Map: Towards a Unified Consensus Linkage Map for Fine Mapping and Synteny Analysis in Common Bean

PubMed Central

Galeano, Carlos H.; Fernandez, Andrea C.; Franco-Herrera, Natalia; Cichy, Karen A.; McClean, Phillip E.; Vanderleyden, Jos; Blair, Matthew W.

2011-01-01

Map-based cloning and fine mapping to find genes of interest and marker assisted selection (MAS) requires good genetic maps with reproducible markers. In this study, we saturated the linkage map of the intra-gene pool population of common bean DOR364×BAT477 (DB) by evaluating 2,706 molecular markers including SSR, SNP, and gene-based markers. On average the polymorphism rate was 7.7% due to the narrow genetic base between the parents. The DB linkage map consisted of 291 markers with a total map length of 1,788 cM. A consensus map was built using the core mapping populations derived from inter-gene pool crosses: DOR364×G19833 (DG) and BAT93×JALO EEP558 (BJ). The consensus map consisted of a total of 1,010 markers mapped, with a total map length of 2,041 cM across 11 linkage groups. On average, each linkage group on the consensus map contained 91 markers of which 83% were single copy markers. Finally, a synteny analysis was carried out using our highly saturated consensus maps compared with the soybean pseudo-chromosome assembly. A total of 772 marker sequences were compared with the soybean genome. A total of 44 syntenic blocks were identified. The linkage group Pv6 presented the most diverse pattern of synteny with seven syntenic blocks, and Pv9 showed the most consistent relations with soybean with just two syntenic blocks. Additionally, a co-linear analysis using common bean transcript map information against soybean coding sequences (CDS) revealed the relationship with 787 soybean genes. The common bean consensus map has allowed us to map a larger number of markers, to obtain a more complete coverage of the common bean genome. Our results, combined with synteny relationships provide tools to increase marker density in selected genomic regions to identify closely linked polymorphic markers for indirect selection, fine mapping or for positional cloning. PMID:22174773

Teaching the Hardy-Weinberg Law

ERIC Educational Resources Information Center

Dudley, B. A. C.

1972-01-01

Describes an approach to teaching the Hardy-Weinberg Law in high school genetics class. Instructional procedures used help in developing this concept in broad generalization form rather than merely a mathematical model of a gene pool. (PS)

Animal and vegetation patterns in natural and man-made bog pools: implications for restoration

USGS Publications Warehouse

Mazerolle, M.J.; Poulin, M.; Lavoie, C.; Rochefort, L.; Desrochers, A.; Drolet, B.

2006-01-01

1. Peatlands have suffered great losses following drainage for agriculture, forestry, urbanisation, or peat mining, near inhabited areas. We evaluated the faunal and vegetation patterns after restoration of a peatland formerly mined for peat. We assessed whether bog pools created during restoration are similar to natural bog pools in terms of water chemistry, vegetation structure and composition, as well as amphibian and arthropod occurrence patterns. 2. Both avian species richness and peatland vegetation cover at the site increased following restoration. Within bog pools, however, the vegetation composition differed between natural and man-made pools. The cover of low shrubs, Sphagnum moss, submerged, emergent and floating vegetation in man-made pools was lower than in natural pools, whereas pH was higher than in typical bog pools. Dominant plant species also differed between man-made and natural pools. 3. Amphibian tadpoles, juveniles and adults occurred more often in man-made pools than natural bog pools. Although some arthropods, including Coleoptera bog specialists, readily colonised the pools, their abundance was two to 26 times lower than in natural bog pools. Plant introduction in bog pools, at the stocking densities we applied, had no effect on the occurrence of most groups. 4. We conclude that our restoration efforts were partially successful. Peatland-wide vegetation patterns following restoration mimicked those of natural peatlands, but 4 years were not sufficient for man-made pools to fully emulate the characteristics of natural bog pools.

Differential gene regulation of GHSR signaling pathway in the arcuate nucleus and NPY neurons by fasting, diet-induced obesity, and 17β-estradiol

PubMed Central

Yasrebi, Ali; Hsieh, Anna; Mamounis, Kyle J.; Krumm, Elizabeth A.; Yang, Jennifer A.; Magby, Jason; Hu, Pu; Roepke, Troy A.

2015-01-01

Ghrelin’s receptor, growth hormone secretagogue receptor (GHSR), is highly expressed in the arcuate nucleus (ARC) and in neuropeptide Y (NPY) neurons. Fasting, diet-induced obesity (DIO), and 17β-estradiol (E2) influence ARC Ghsr expression. It is unknown if these effects occur in NPY neurons. Therefore, we examined the expression of Npy, Agrp, and GHSR signaling pathway genes after fasting, DIO, and E2 replacement in ARC and pools of NPY neurons. In males, fasting increased ARC Ghsr and NPY Foxo1 but decreased NPY Ucp2. In males, DIO decreased ARC and NPY Ghsr and Cpt1c. In fed females, E2 increased Agrp, Ghsr, Cpt1c, and Foxo1 in ARC. In NPY pools, E2 decreased Foxo1 in fed females but increased Foxo1 in fasted females. DIO in females suppressed Agrp and augmented Cpt1c in NPY neurons. In summary, genes involved in GHSR signaling are differentially regulated between the ARC and NPY neurons in a sex-dependent manner. PMID:26577678

Functional Analysis With a Barcoder Yeast Gene Overexpression System

PubMed Central

Douglas, Alison C.; Smith, Andrew M.; Sharifpoor, Sara; Yan, Zhun; Durbic, Tanja; Heisler, Lawrence E.; Lee, Anna Y.; Ryan, Owen; Göttert, Hendrikje; Surendra, Anu; van Dyk, Dewald; Giaever, Guri; Boone, Charles; Nislow, Corey; Andrews, Brenda J.

2012-01-01

Systematic analysis of gene overexpression phenotypes provides an insight into gene function, enzyme targets, and biological pathways. Here, we describe a novel functional genomics platform that enables a highly parallel and systematic assessment of overexpression phenotypes in pooled cultures. First, we constructed a genome-level collection of ~5100 yeast barcoder strains, each of which carries a unique barcode, enabling pooled fitness assays with a barcode microarray or sequencing readout. Second, we constructed a yeast open reading frame (ORF) galactose-induced overexpression array by generating a genome-wide set of yeast transformants, each of which carries an individual plasmid-born and sequence-verified ORF derived from the Saccharomyces cerevisiae full-length EXpression-ready (FLEX) collection. We combined these collections genetically using synthetic genetic array methodology, generating ~5100 strains, each of which is barcoded and overexpresses a specific ORF, a set we termed “barFLEX.” Additional synthetic genetic array allows the barFLEX collection to be moved into different genetic backgrounds. As a proof-of-principle, we describe the properties of the barFLEX overexpression collection and its application in synthetic dosage lethality studies under different environmental conditions. PMID:23050238

Polymorphisms in Genes of Relevance for Oestrogen and Oxytocin Pathways and Risk of Barrett's Oesophagus and Oesophageal Adenocarcinoma: A Pooled Analysis from the BEACON Consortium.

PubMed

Lagergren, Katarina; Ek, Weronica E; Levine, David; Chow, Wong-Ho; Bernstein, Leslie; Casson, Alan G; Risch, Harvey A; Shaheen, Nicholas J; Bird, Nigel C; Reid, Brian J; Corley, Douglas A; Hardie, Laura J; Wu, Anna H; Fitzgerald, Rebecca C; Pharoah, Paul; Caldas, Carlos; Romero, Yvonne; Vaughan, Thomas L; MacGregor, Stuart; Whiteman, David; Westberg, Lars; Nyren, Olof; Lagergren, Jesper

2015-01-01

The strong male predominance in oesophageal adenocarcinoma (OAC) and Barrett's oesophagus (BO) continues to puzzle. Hormonal influence, e.g. oestrogen or oxytocin, might contribute. This genetic-epidemiological study pooled 14 studies from three continents, Australia, Europe, and North America. Polymorphisms in 3 key genes coding for the oestrogen pathway (receptor alpha (ESR1), receptor beta (ESR2), and aromatase (CYP19A1)), and 3 key genes of the oxytocin pathway (the oxytocin receptor (OXTR), oxytocin protein (OXT), and cyclic ADP ribose hydrolase glycoprotein (CD38)), were analysed using a gene-based approach, versatile gene-based test association study (VEGAS). Among 1508 OAC patients, 2383 BO patients, and 2170 controls, genetic variants within ESR1 were associated with BO in males (p = 0.0058) and an increased risk of OAC and BO combined in males (p = 0.0023). Genetic variants within OXTR were associated with an increased risk of BO in both sexes combined (p = 0.0035) and in males (p = 0.0012). We followed up these suggestive findings in a further smaller data set, but found no replication. There were no significant associations between the other 4 genes studied and risk of OAC, BO, separately on in combination, in males and females combined or in males only. Genetic variants in the oestrogen receptor alpha and the oxytocin receptor may be associated with an increased risk of BO or OAC, but replication in other large samples are needed.

Lack of association between TPH2 gene polymorphisms with major depressive disorder in multiethnic Malaysian population.

PubMed

Nazree, Nur Elia; Loke, Ai Chin; Zainal, Nor Zuraida; Mohamed, Zahurin

2015-03-01

Numerous association studies of candidate genes studies with major depressive disorder (MDD) have been conducted for many years; however, the evidence of association between genes and the risk of developing MDD still remains inconclusive. In this study, we aimed to investigate the association between the tryptophan hydroxylase 2 (TPH2) gene and MDD in three ethnic groups (Malay, Chinese and Indian) within the Malaysian population. Two hundred and sixty five MDD patients who fulfilled the Diagnostic and Statistical Manual of Mental Disorders-IV criteria for MDD and 332 healthy controls were recruited for the study. All cases and controls were then genotyped for TPH2 polymorphisms rs1386494, rs1386495 and rs7305115. Single locus analysis in pooled and ethnically stratified subjects revealed no association between each of the three variants of the TPH2 gene with susceptibility to MDD. Strong linkage disequilibrium was detected between rs1386495 and rs1386494 in pooled subjects; however, no significant association was found in the haplotype analysis. In this study, we suggest that in both the Chinese and Indian populations, gender distribution differ significantly between cases and controls, showing that women are more at risk of developing MDD compared with men. Therefore, we suggest that the occurrence of MDD in both Chinese and Indians in the Malaysian population may be influenced by gender. Copyright © 2013 Wiley Publishing Asia Pty Ltd.

Fire control method and analytical model for large liquid hydrocarbon pool fires

NASA Technical Reports Server (NTRS)

Fenton, D. L.

1986-01-01

The dominate parameter governing the behavior of a liquid hydrocarbon (JP-5) pool fire is wind speed. The most effective method of controlling wind speed in the vicinity of a large circular (10 m dia.) pool fire is a set of concentric screens located outside the perimeter. Because detailed behavior of the pool fire structure within one pool fire diameter is unknown, an analytical model supported by careful experiments is under development. As a first step toward this development, a regional pool fire model was constructed for the no-wind condition consisting of three zones -- liquid fuel, combustion, and plume -- where the predicted variables are mass burning rate and characteristic temperatures of the combustion and plume zones. This zone pool fire model can be modified to incorporate plume bending by wind, radiation absorption by soot particles, and a different ambient air flow entrainment rate. Results from the zone model are given for a pool diameter of 1.3 m and are found to reproduce values in the literature.

Identification of susceptibility genes and genetic modifiers of human diseases

NASA Astrophysics Data System (ADS)

Abel, Kenneth; Kammerer, Stefan; Hoyal, Carolyn; Reneland, Rikard; Marnellos, George; Nelson, Matthew R.; Braun, Andreas

2005-03-01

The completion of the human genome sequence enables the discovery of genes involved in common human disorders. The successful identification of these genes is dependent on the availability of informative sample sets, validated marker panels, a high-throughput scoring technology, and a strategy for combining these resources. We have developed a universal platform technology based on mass spectrometry (MassARRAY) for analyzing nucleic acids with high precision and accuracy. To fuel this technology, we generated more than 100,000 validated assays for single nucleotide polymorphisms (SNPs) covering virtually all known and predicted human genes. We also established a large DNA sample bank comprised of more than 50,000 consented healthy and diseased individuals. This combination of reagents and technology allows the execution of large-scale genome-wide association studies. Taking advantage of MassARRAY"s capability for quantitative analysis of nucleic acids, allele frequencies are estimated in sample pools containing large numbers of individual DNAs. To compare pools as a first-pass "filtering" step is a tremendous advantage in throughput and cost over individual genotyping. We employed this approach in numerous genome-wide, hypothesis-free searches to identify genes associated with common complex diseases, such as breast cancer, osteoporosis, and osteoarthritis, and genes involved in quantitative traits like high density lipoproteins cholesterol (HDL-c) levels and central fat. Access to additional well-characterized patient samples through collaborations allows us to conduct replication studies that validate true disease genes. These discoveries will expand our understanding of genetic disease predisposition, and our ability for early diagnosis and determination of specific disease subtype or progression stage.

miRNA-mediated 'tug-of-war' model reveals ceRNA propensity of genes in cancers.

PubMed

Swain, Arpit Chandan; Mallick, Bibekanand

2018-06-01

Competing endogenous RNA (ceRNA) are transcripts that cross-regulate each other at the post-transcriptional level by competing for shared microRNA response elements (MREs). These have been implicated in various biological processes impacting cell-fate decisions and diseases including cancer. There are several studies that predict possible ceRNA pairs by adopting various machine-learning and mathematical approaches; however, there is no method that enables us to gauge as well as compare the propensity of the ceRNA of a gene and precisely envisages which among a pair exerts a stronger pull on the shared miRNA pool. In this study, we developed a method that uses the 'tug of war of genes' concept to predict and quantify ceRNA potential of a gene for the shared miRNA pool in cancers based on a score represented by SoCeR (score of competing endogenous RNA). The method was executed on the RNA-Seq transcriptional profiles of genes and miRNA available at TCGA along with CLIP-supported miRNA-target sites to predict ceRNA in 32 cancer types which were validated with already reported cases. The proposed method can be used to determine the sequestering capability of the gene of interest as well as in ranking the probable ceRNA candidates of a gene. Finally, we developed standalone applications (SoCeR tool) to aid researchers in easier implementation of the method in analysing different data sets or diseases. © 2018 The Authors. Published by FEBS Press and John Wiley & Sons Ltd.

Upregulated Genes In Sporadic, Idiopathic Pulmonary Arterial Hypertension

PubMed Central

Edgar, Alasdair J; Chacón, Matilde R; Bishop, Anne E; Yacoub, Magdi H; Polak, Julia M

2006-01-01

Background To elucidate further the pathogenesis of sporadic, idiopathic pulmonary arterial hypertension (IPAH) and identify potential therapeutic avenues, differential gene expression in IPAH was examined by suppression subtractive hybridisation (SSH). Methods Peripheral lung samples were obtained immediately after removal from patients undergoing lung transplant for IPAH without familial disease, and control tissues consisted of similarly sampled pieces of donor lungs not utilised during transplantation. Pools of lung mRNA from IPAH cases containing plexiform lesions and normal donor lungs were used to generate the tester and driver cDNA libraries, respectively. A subtracted IPAH cDNA library was made by SSH. Clones isolated from this subtracted library were examined for up regulated expression in IPAH using dot blot arrays of positive colony PCR products using both pooled cDNA libraries as probes. Clones verified as being upregulated were sequenced. For two genes the increase in expression was verified by northern blotting and data analysed using Student's unpaired two-tailed t-test. Results We present preliminary findings concerning candidate genes upregulated in IPAH. Twenty-seven upregulated genes were identified out of 192 clones examined. Upregulation in individual cases of IPAH was shown by northern blot for tissue inhibitor of metalloproteinase-3 and decorin (P < 0.01) compared with the housekeeping gene glyceraldehydes-3-phosphate dehydrogenase. Conclusion Four of the up regulated genes, magic roundabout, hevin, thrombomodulin and sucrose non-fermenting protein-related kinase-1 are expressed specifically by endothelial cells and one, muscleblind-1, by muscle cells, suggesting that they may be associated with plexiform lesions and hypertrophic arterial wall remodelling, respectively. PMID:16390543

New enzymes from environmental cassette arrays: Functional attributes of a phosphotransferase and an RNA-methyltransferase

PubMed Central

Nield, Blair S.; Willows, Robert D.; Torda, Andrew E.; Gillings, Michael R.; Holmes, Andrew J.; Nevalainen, K.M. Helena; Stokes, H.W.; Mabbutt, Bridget C.

2004-01-01

By targeting gene cassettes by polymerase chain reaction (PCR) directly from environmentally derived DNA, we are able to amplify entire open reading frames (ORFs) independently of prior sequence knowledge. Approximately 10% of the mobile genes recovered by these means can be attributed to known protein families. Here we describe the characterization of two ORFs which show moderate homology to known proteins: (1) an aminoglycoside phosphotransferase displaying 25% sequence identity with APH(7″) from Streptomyces hygroscopicus, and (2) an RNA methyltransferase sharing 25%–28% identity with a group of recently defined bacterial RNA methyltransferases distinct from the SpoU enzyme family. Our novel genes were expressed as recombinant products and assayed for appropriate enzyme activity. The aminoglycoside phosphotransferase displayed ATPase activity, consistent with the presence of characteristic Mg2+-binding residues. Unlike related APH(4) or APH(7″) enzymes, however, this activity was not enhanced by hygromycin B or kanamycin, suggesting the normal substrate to be a different aminoglycoside. The RNA methyltransferase contains sequence motifs of the RNA methyltransferase superfamily, and our recombinant version showed methyltransferase activity with RNA. Our data confirm that gene cassettes present in the environment encode folded enzymes with novel sequence variation and demonstrable catalytic activity. Our PCR approach (cassette PCR) may be used to identify a diverse range of ORFs from any environmental sample, as well as to directly access the gene pool found in mobile gene cassettes commonly associated with integrons. This gene pool can be accessed from both cultured and uncultured microbial samples as a source of new enzymes and proteins. PMID:15152095

New enzymes from environmental cassette arrays: functional attributes of a phosphotransferase and an RNA-methyltransferase.

PubMed

Nield, Blair S; Willows, Robert D; Torda, Andrew E; Gillings, Michael R; Holmes, Andrew J; Nevalainen, K M Helena; Stokes, H W; Mabbutt, Bridget C

2004-06-01

By targeting gene cassettes by polymerase chain reaction (PCR) directly from environmentally derived DNA, we are able to amplify entire open reading frames (ORFs) independently of prior sequence knowledge. Approximately 10% of the mobile genes recovered by these means can be attributed to known protein families. Here we describe the characterization of two ORFs which show moderate homology to known proteins: (1) an aminoglycoside phosphotransferase displaying 25% sequence identity with APH(7") from Streptomyces hygroscopicus, and (2) an RNA methyltransferase sharing 25%-28% identity with a group of recently defined bacterial RNA methyltransferases distinct from the SpoU enzyme family. Our novel genes were expressed as recombinant products and assayed for appropriate enzyme activity. The aminoglycoside phosphotransferase displayed ATPase activity, consistent with the presence of characteristic Mg(2+)-binding residues. Unlike related APH(4) or APH(7") enzymes, however, this activity was not enhanced by hygromycin B or kanamycin, suggesting the normal substrate to be a different aminoglycoside. The RNA methyltransferase contains sequence motifs of the RNA methyltransferase superfamily, and our recombinant version showed methyltransferase activity with RNA. Our data confirm that gene cassettes present in the environment encode folded enzymes with novel sequence variation and demonstrable catalytic activity. Our PCR approach (cassette PCR) may be used to identify a diverse range of ORFs from any environmental sample, as well as to directly access the gene pool found in mobile gene cassettes commonly associated with integrons. This gene pool can be accessed from both cultured and uncultured microbial samples as a source of new enzymes and proteins.

ABC-B transporter genes in Dirofilaria immitis.

PubMed

Bourguinat, Catherine; Che, Hua; Mani, Thangadurai; Keller, Kathy; Prichard, Roger K

2016-08-01

Dirofilaria immitis is a filarial nematode causing infection and heartworm disease in dogs and other canids, cats, and occasionally in humans. Prevention with macrocyclic lactones (ML) is recommended during the mosquito transmission season. Recently, ML resistance has been reported. ABC-B transporter genes are thought to be involved in the mechanism of ML resistance in other nematodes. This study aimed to identify all the ABC-B transporter genes in D. immitis using as a reference the nDi.2.2 D. immitis whole genome, which is not completely annotated. Using bioinformatic tools and PCR amplification on pooled D. immitis genomic DNA and on pooled cDNA, nine ABC transporter genes including one pseudogene were characterized. Bioinformatic and phylogenetic analyses allowed identification of three P-glycoproteins (Pgps) (Dim-pgp-3 Dim-pgp-10, Dim-pgp-11), of two ABC-B half transporter genes (one ortholog of Cel-haf-4 and Cel-haf-9; and one ortholog of Cel-haf-1 and Cel-haf-3), of one ABC half transporter gene (ortholog of Cel-haf-5) that contained an ABC-C motif, and of one additional half transporter that would require functional study for characterization. The number of ABC-B transporter genes identified was lower than in Caenorhabditis elegans and Haemonchus contortus. Further studies are needed to understand their possible role in ML resistance in D. immitis. These ABC transporters constitute a base for ML resistance investigation in D. immitis and advance our understanding of the molecular biology of this parasite. Copyright © 2016. Published by Elsevier Ltd.

Independent and collective roles of surface structures at different length scales on pool boiling heat transfer

PubMed Central

Li, Calvin H.; Rioux, Russell P.

2016-01-01

Spherical Cu nanocavity surfaces are synthesized to examine the individual role of contact angles in connecting lateral Rayleigh-Taylor wavelength to vertical Kevin-Helmholtz wavelength on hydrodynamic instability for the onset of pool boiling Critical Heat Flux (CHF). Solid and porous Cu pillar surfaces are sintered to investigate the individual role of pillar structure pitch at millimeter scale, named as module wavelength, on hydrodynamic instability at CHF. Last, spherical Cu nanocavities are coated on the porous Cu pillars to create a multiscale Cu structure, which is studied to examine the collective role and relative significance of contact angles and module wavelength on hydrodynamic instability at CHF, and the results indicate that module wavelength plays the dominant role on hydrodynamic instability at CHF when the height of surface structures is equal or above ¼ Kelvin-Helmholtz wavelength. Pool boiling Heat Transfer Coefficient (HTC) enhancements on spherical Cu nanocavity surfaces, solid and porous Cu pillar surfaces, and the integrated multiscale structure have been investigated, too. The experimental results reveal that the nanostructures and porous pillar structures can be combined together to achieve even higher enhancement of HTC than that of individual structures. PMID:27841322

Recombination walking: genetic selection of clones from pooled libraries of yeast artificial chromosomes by homologous recombination.

PubMed Central

Miller, A M; Savinelli, E A; Couture, S M; Hannigan, G M; Han, Z; Selden, R F; Treco, D A

1993-01-01

Recombination walking is based on the genetic selection of specific human clones from a yeast artificial chromosome (YAC) library by homologous recombination. The desired clone is selected from a pooled (unordered) YAC library, eliminating labor-intensive steps typically used in organizing and maintaining ordered YAC libraries. Recombination walking represents an efficient approach to library screening and is well suited for chromosome-walking approaches to the isolation of genes associated with common diseases. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:8367472

Isolation of a Novel Insect-Specific Flavivirus from Culiseta melanura in the Northeastern United States

PubMed Central

Misencik, Michael J.; Grubaugh, Nathan D.; Andreadis, Theodore G.; Ebel, Gregory D.

2016-01-01

Abstract The genus Flavivirus includes a number of newly recognized viruses that infect and replicate only within mosquitoes. To determine whether insect-specific flaviviruses (ISFs) may infect Culiseta (Cs.) melanura mosquitoes, we screened pools of field-collected mosquitoes for virus infection by RT-PCR targeting conserved regions of the NS5 gene. NS5 nucleotide sequences amplified from Cs. melanura pools were genetically similar to other ISFs and most closely matched Calbertado virus from Culex tarsalis, sharing 68.7% nucleotide and 76.1% amino acid sequence identity. The complete genome of one virus isolate was sequenced to reveal a primary open reading frame (ORF) encoding a viral polyprotein characteristic of the genus Flavivirus. Phylogenetic analysis showed that this virus represents a distinct evolutionary lineage that belongs to the classical ISF group. The virus was detected solely in Cs. melanura pools, occurred in sampled populations from Connecticut, New York, New Hampshire, and Maine, and infected both adult and larval stages of the mosquito. Maximum likelihood estimate infection rates (MLE-IR) were relatively stable in overwintering Cs. melanura larvae collected monthly from November of 2012 through May of 2013 (MLE-IR = 0.7–2.1/100 mosquitoes) and in host-seeking females collected weekly from June through October of 2013 (MLE-IR = 3.8–11.5/100 mosquitoes). Phylogenetic analysis of viral sequences revealed limited genetic variation that lacked obvious geographic structure among strains in the northeastern United States. This new virus is provisionally named Culiseta flavivirus on the basis of its host association with Cs. melanura. PMID:26807512

Crustacean communities in coastal ephemeral pools in the Araucanía region (38° S, Chile).

PubMed

De Los Ríos-Escalante, P; Acevedo, P

2016-01-01

The fauna communities of ephemeral pools in southern Chile are characterized by heterogeneity of crustacean taxa; nevertheless, no detailed studies exist of their community structure. The aim of the present study was to analyze the crustacean community structure in two groups of ephemeral pools (Puaucho and Nigue pools) in the coastal zone of the Araucanía region. A correlation matrix was made by species abundance against temperature, conductivity, pH and total dissolved solids. In a second step, a null model for species co-occurrence was applied to the total data and to each group. The results for total data revealed a significant direct relation between the abundance of H. costera, C. dubia and Mesocyclops. For the Puaucho pools, the same results were found together with direct associations with total dissolved solids, conductivity and pH. Finally, different results were found for the Nigue pools, with no clear significant associations, either direct or indirect, between the abundance of different crustacean taxa and abiotic parameters. These results were supported by the co-occurrence null model analysis, which revealed the presence of regulator factors for the total data, and for each of the two groups. Ecological topics are discussed with emphasis on meta-community dynamics.

Hydrology of the Floral City Pool of Tsala Apopka Lake, west-central Florida

USGS Publications Warehouse

Bradner, L.A.

1988-01-01

Tsala Apopka Lake, in west-central Florida, has an area of about 19,000 acres and is divided into three water-management pools, with the Floral City Pool, the most upgradient. The Floral City Pool, which has a surface area of approximately 4,750 acres, contains an extensive combination of lakes, wetlands, and connecting canals. The Pool receives inflow from the Withlacoochee River through two canals. Outflow is through one manmade canal and one natural slough. Canal flow is partially controlled by manmade structures. A cumulative deficit of 19.4 inches of rainfall from August 1984 through May 1985 reduced surface-water inflow to the Floral City Pool to about 0.5 cu ft/sec by May 1985. During May 1985, pool levels declined approximately 0.04 ft/day. By the end of May, there was no observable outflow. From June 1985 through September 1985, 39.8 inches of rainfall caused above-average inflow to the Floral City Pool and a pool-level increase of 6.2 ft. The inflow of 340 CFS nearly equaled the outflow of 338 CFS by the end of September. (USGS)

Alcoholism Detection by Data Augmentation and Convolutional Neural Network with Stochastic Pooling.

PubMed

Wang, Shui-Hua; Lv, Yi-Ding; Sui, Yuxiu; Liu, Shuai; Wang, Su-Jing; Zhang, Yu-Dong

2017-11-17

Alcohol use disorder (AUD) is an important brain disease. It alters the brain structure. Recently, scholars tend to use computer vision based techniques to detect AUD. We collected 235 subjects, 114 alcoholic and 121 non-alcoholic. Among the 235 image, 100 images were used as training set, and data augmentation method was used. The rest 135 images were used as test set. Further, we chose the latest powerful technique-convolutional neural network (CNN) based on convolutional layer, rectified linear unit layer, pooling layer, fully connected layer, and softmax layer. We also compared three different pooling techniques: max pooling, average pooling, and stochastic pooling. The results showed that our method achieved a sensitivity of 96.88%, a specificity of 97.18%, and an accuracy of 97.04%. Our method was better than three state-of-the-art approaches. Besides, stochastic pooling performed better than other max pooling and average pooling. We validated CNN with five convolution layers and two fully connected layers performed the best. The GPU yielded a 149× acceleration in training and a 166× acceleration in test, compared to CPU.

CRISPR-UMI: single-cell lineage tracing of pooled CRISPR-Cas9 screens.

PubMed

Michlits, Georg; Hubmann, Maria; Wu, Szu-Hsien; Vainorius, Gintautas; Budusan, Elena; Zhuk, Sergei; Burkard, Thomas R; Novatchkova, Maria; Aichinger, Martin; Lu, Yiqing; Reece-Hoyes, John; Nitsch, Roberto; Schramek, Daniel; Hoepfner, Dominic; Elling, Ulrich

2017-12-01

Pooled CRISPR screens are a powerful tool for assessments of gene function. However, conventional analysis is based exclusively on the relative abundance of integrated single guide RNAs (sgRNAs) between populations, which does not discern distinct phenotypes and editing outcomes generated by identical sgRNAs. Here we present CRISPR-UMI, a single-cell lineage-tracing methodology for pooled screening to account for cell heterogeneity. We generated complex sgRNA libraries with unique molecular identifiers (UMIs) that allowed for screening of clonally expanded, individually tagged cells. A proof-of-principle CRISPR-UMI negative-selection screen provided increased sensitivity and robustness compared with conventional analysis by accounting for underlying cellular and editing-outcome heterogeneity and detection of outlier clones. Furthermore, a CRISPR-UMI positive-selection screen uncovered new roadblocks in reprogramming mouse embryonic fibroblasts as pluripotent stem cells, distinguishing reprogramming frequency and speed (i.e., effect size and probability). CRISPR-UMI boosts the predictive power, sensitivity, and information content of pooled CRISPR screens.

Observed Structure and Characteristics of Cold Pools over Tropical Oceans using Vector Wind Retrievals and WRF simulations

NASA Astrophysics Data System (ADS)

Garg, P.; Nesbitt, S. W.; Lang, T. J.; Chronis, T.; Thayer, J. D.; Hence, D. A.

2017-12-01

Cold pools generated in the wake of convective activity can enhance the surface sensible heat flux, latent heat flux, and also changes in evaporation out of, and fresh water flux into, the ocean. Recent studies have shown that over the open ocean, cold pool outflow boundaries and their intersections can organize and initiate a spectrum of deep convective clouds, which is a key driver of shallow and deep convection over conditionally-unstable tropical oceans. The primary goal of this study is to understand the structure and characteristics of cold pools over the tropical oceans using observations. With the idea that cold pools will have strong wind gradients at their boundaries, we use ASCAT vector wind retrievals. We identify regions of steep gradients in wind vectors as gradient features (GFs), akin to cold pools. Corresponding to these GFs, sensible and latent heat fluxes were calculated using the observed winds and background temperatures from MERRA-2 reanalysis. To evaluate the proposed technique, cold pools were observed using S-PolKa radar from the DYNAMO/AMIE field campaign in the Indian Ocean for the period of 1 October 2011 to 31 March 2012 and were compared with ASCAT GFs. To relate the thermodynamic and kinematic characteristics of observed and simulated cold pools, simulations were carried out on WRF on a 3-km domain explicitly. The areas of cold pools were identified in the models using virtual temperature (Tv), which is a direct measure of air density, while GFs were identified using model simulated winds. Quantitative measures indicate that GFs are highly correspondent with model-simulated cold pools. In global measurements of cold pools from 2007-2015, it is possible to examine the characteristics of GFs across all tropical ocean basins, and relate them to meteorological conditions, as well as the characteristics of the parent precipitation systems. Our results indicate that while there is a general relationship between the amount of precipitation and the number of cold pools, the largest cold pools exist over the Eastern Pacific basin, where the most stratiform rain is produced from oceanic MCSs. It is anticipated that improved understanding of cold pools, which are a primary triggering mechanism of oceanic shallow and deep convection, will improve prediction of this important component of the climate system.

Evaluation of concurrent priority queue algorithms. Technical report

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

Huang, Q.

1991-02-01

The priority queue is a fundamental data structure that is used in a large variety of parallel algorithms, such as multiprocessor scheduling and parallel best-first search of state-space graphs. This thesis addresses the design and experimental evaluation of two novel concurrent priority queues: a parallel Fibonacci heap and a concurrent priority pool, and compares them with the concurrent binary heap. The parallel Fibonacci heap is based on the sequential Fibonacci heap, which is theoretically the most efficient data structure for sequential priority queues. This scheme not only preserves the efficient operation time bounds of its sequential counterpart, but also hasmore » very low contention by distributing locks over the entire data structure. The experimental results show its linearly scalable throughput and speedup up to as many processors as tested (currently 18). A concurrent access scheme for a doubly linked list is described as part of the implementation of the parallel Fibonacci heap. The concurrent priority pool is based on the concurrent B-tree and the concurrent pool. The concurrent priority pool has the highest throughput among the priority queues studied. Like the parallel Fibonacci heap, the concurrent priority pool scales linearly up to as many processors as tested. The priority queues are evaluated in terms of throughput and speedup. Some applications of concurrent priority queues such as the vertex cover problem and the single source shortest path problem are tested.« less

Microorganism genomics, compositions and methods related thereto

DOEpatents

Handelsman, Jo; Goodman, Robert M.; Rondon, Michelle R.

2001-01-01

The present invention provides methods and compositions for accessing, in a generally unbaised manner, a diverse genetic pool for genes involved in biosynthetic pathways. The invention also provides compounds which can be identified by cloning biosynthetic pathways.

The microbe-mediated mechanisms affecting topsoil carbon stock in Tibetan grasslands

DOE PAGES

Yue, Haowei; Wang, Mengmeng; Wang, Shiping; ...

2015-02-17

Warming has been shown to cause soil carbon (C) loss in northern grasslands owing to accelerated microbial decomposition that offsets increased grass productivity. Yet, a multi-decadal survey indicated that the surface soil C stock in Tibetan alpine grasslands remained relatively stable. To investigate this inconsistency, we analyzed the feedback responses of soil microbial communities to simulated warming by soil transplant in Tibetan grasslands. Microbial functional diversity decreased in response to warming, whereas microbial community structure did not correlate with changes in temperature. The relative abundance of catabolic genes associated with nitrogen (N) and C cycling decreased with warming, most notablymore » in genes encoding enzymes associated with more recalcitrant C substrates. By contrast, genes associated with C fixation increased in relative abundance. The relative abundance of genes associated with urease, glutamate dehydrogenase and ammonia monoxygenase ( ureC, gdh and amoA) were significantly correlated with N 2O efflux. These results suggest that unlike arid/semiarid grasslands, Tibetan grasslands maintain negative feedback mechanisms that preserve terrestrial C and N pools. To examine whether these trends were applicable to the whole plateau, we included these measurements in a model and verified that topsoil C stocks remained relatively stable. Thus, by establishing linkages between microbial metabolic potential and soil biogeochemical processes, we conclude that long-term C loss in Tibetan grasslands is ameliorated by a reduction in microbial decomposition of recalcitrant C substrates.« less

Decreased Nucleotide and Expression Diversity and Modified Coexpression Patterns Characterize Domestication in the Common Bean[W][OPEN

PubMed Central

Bellucci, Elisa; Bitocchi, Elena; Ferrarini, Alberto; Benazzo, Andrea; Biagetti, Eleonora; Klie, Sebastian; Minio, Andrea; Rau, Domenico; Rodriguez, Monica; Panziera, Alex; Venturini, Luca; Attene, Giovanna; Albertini, Emidio; Jackson, Scott A.; Nanni, Laura; Fernie, Alisdair R.; Nikoloski, Zoran; Bertorelle, Giorgio; Delledonne, Massimo; Papa, Roberto

2014-01-01

Using RNA sequencing technology and de novo transcriptome assembly, we compared representative sets of wild and domesticated accessions of common bean (Phaseolus vulgaris) from Mesoamerica. RNA was extracted at the first true-leaf stage, and de novo assembly was used to develop a reference transcriptome; the final data set consists of ∼190,000 single nucleotide polymorphisms from 27,243 contigs in expressed genomic regions. A drastic reduction in nucleotide diversity (∼60%) is evident for the domesticated form, compared with the wild form, and almost 50% of the contigs that are polymorphic were brought to fixation by domestication. In parallel, the effects of domestication decreased the diversity of gene expression (18%). While the coexpression networks for the wild and domesticated accessions demonstrate similar seminal network properties, they show distinct community structures that are enriched for different molecular functions. After simulating the demographic dynamics during domestication, we found that 9% of the genes were actively selected during domestication. We also show that selection induced a further reduction in the diversity of gene expression (26%) and was associated with 5-fold enrichment of differentially expressed genes. While there is substantial evidence of positive selection associated with domestication, in a few cases, this selection has increased the nucleotide diversity in the domesticated pool at target loci associated with abiotic stress responses, flowering time, and morphology. PMID:24850850

Admixture Aberration Analysis: Application to Mapping in Admixed Population Using Pooled DNA

NASA Astrophysics Data System (ADS)

Bercovici, Sivan; Geiger, Dan

Admixture mapping is a gene mapping approach used for the identification of genomic regions harboring disease susceptibility genes in the case of recently admixed populations such as African Americans. We present a novel method for admixture mapping, called admixture aberration analysis (AAA), that uses a DNA pool of affected admixed individuals. We demonstrate through simulations that AAA is a powerful and economical mapping method under a range of scenarios, capturing complex human diseases such as hypertension and end stage kidney disease. The method has a low false-positive rate and is robust to deviation from model assumptions. Finally, we apply AAA on 600 prostate cancer-affected African Americans, replicating a known risk locus. Simulation results indicate that the method can yield over 96% reduction in genotyping. Our method is implemented as a Java program called AAAmap and is freely available.

Molecular characterization of primary gene pool of chickpea based on ISSR markers.

PubMed

Choudhary, Pooja; Khanna, Suruchi M; Jain, Pradeep K; Bharadwaj, Chellapilla; Kumar, Jitendra; Lakhera, Pramesh C; Srinivasan, Ramamurthy

2013-04-01

Genetic diversity and relationships within and among members of the primary gene pool of chickpea, including 38 accessions of Cicer arietinum, six of C. reticulatum,, and four of C. echinospermum, were investigated using 31 ISSR markers. The study revealed moderate diversity, detecting 141 fragments, of which 79 (56%) were polymorphic. Averages were 0.125 for polymorphic information content, 0.350 for marker index, and 0.715 for resolving power. The UPGMA dendrogram and the principal coordinate analysis revealed a clear differentiation between wild and cultivated accessions. The clustering pattern did not strictly follow the grouping of accessions by geographic origin but was in good agreement with the pedigree data and the seed type. The study demonstrates that ISSRs provide promising marker tools in revealing genetic diversity and relationships in chickpea and can contribute to efficient identification, conservation, and utilization of germplasm for plant improvement through conventional as well as molecular breeding approaches.

Donor specificity of YjiC glycosyltransferase determines the conjugation of cytosolic NDP-sugar in in vivo glycosylation reactions.

PubMed

Pandey, Ramesh Prasad; Parajuli, Prakash; Gurung, Rit Bahadur; Sohng, Jae Kyung

2016-09-01

Escherichia coli BL21 (DE3) was engineered by blocking glucose-1-phosphate utilizing glucose phosphate isomerase (pgi), glucose-6-phosphate dehydrogenase (zwf) and uridylyltransferase (galU) genes to produce pool of four different rare dTDP-sugars. The cytosolic pool of dTDP-l-rhamnose, dTDP-d-viosamine, dTDP-4-amino 4,6-dideoxy-d-galactose, and dTDP-3-amino 3,6-dideoxy-d-galactose was generated by overexpressing respective dTDP-sugars biosynthesis genes from various microbial sources. A flexible glycosyltransferase YjiC, from Bacillus licheniformis DSM 13 was also overexpressed to transfer sugar moieties to 3-hydroxyl group of 3-hydroxyflavone, a core unit of flavonoids. Among four rare dTDP-sugars generated in cytosol of engineered strains, YjiC solely transferred l-rhamnose from dTDP-l-rhamnose and tuned to rhamnosyltransferase. Copyright © 2016. Published by Elsevier Inc.

Identification of presumed ancestral DNA sequences of phaseolin in Phaseolus vulgaris.

PubMed Central

Kami, J; Velásquez, V B; Debouck, D G; Gepts, P

1995-01-01

Common bean (Phaseolus vulgaris) consists of two major geographic gene pools, one distributed in Mexico, Central America, and Colombia and the other in the southern Andes (southern Peru, Bolivia, and Argentina). Amplification and sequencing of members of the multigene family coding for phaseolin, the major seed storage protein of the common bean, provide evidence for accumulation of tandem direct repeats in both introns and exons during evolution of the multigene family in this species. The presumed ancestral phaseolin sequences, without tandem repeats, were found in recently discovered but nearly extinct wild common bean populations of Ecuador and northern Peru that are intermediate between the two major gene pools of the species based on geographical and molecular arguments. Our results illustrate the usefulness of tandem direct repeats in establishing the polarity of DNA sequence divergence and therefore in proposing phylogenies. Images Fig. 1 Fig. 3 PMID:7862642

Transcriptomic Analysis Using Olive Varieties and Breeding Progenies Identifies Candidate Genes Involved in Plant Architecture

PubMed Central

González-Plaza, Juan J.; Ortiz-Martín, Inmaculada; Muñoz-Mérida, Antonio; García-López, Carmen; Sánchez-Sevilla, José F.; Luque, Francisco; Trelles, Oswaldo; Bejarano, Eduardo R.; De La Rosa, Raúl; Valpuesta, Victoriano; Beuzón, Carmen R.

2016-01-01

Plant architecture is a critical trait in fruit crops that can significantly influence yield, pruning, planting density and harvesting. Little is known about how plant architecture is genetically determined in olive, were most of the existing varieties are traditional with an architecture poorly suited for modern growing and harvesting systems. In the present study, we have carried out microarray analysis of meristematic tissue to compare expression profiles of olive varieties displaying differences in architecture, as well as seedlings from their cross pooled on the basis of their sharing architecture-related phenotypes. The microarray used, previously developed by our group has already been applied to identify candidates genes involved in regulating juvenile to adult transition in the shoot apex of seedlings. Varieties with distinct architecture phenotypes and individuals from segregating progenies displaying opposite architecture features were used to link phenotype to expression. Here, we identify 2252 differentially expressed genes (DEGs) associated to differences in plant architecture. Microarray results were validated by quantitative RT-PCR carried out on genes with functional annotation likely related to plant architecture. Twelve of these genes were further analyzed in individual seedlings of the corresponding pool. We also examined Arabidopsis mutants in putative orthologs of these targeted candidate genes, finding altered architecture for most of them. This supports a functional conservation between species and potential biological relevance of the candidate genes identified. This study is the first to identify genes associated to plant architecture in olive, and the results obtained could be of great help in future programs aimed at selecting phenotypes adapted to modern cultivation practices in this species. PMID:26973682

In situ environment rather than substrate type dictates microbial community structure of biofilms in a cold seep system

PubMed Central

Lee, On On; Wang, Yong; Tian, Renmao; Zhang, Weipeng; Shek, Chun Shum; Bougouffa, Salim; Al-Suwailem, Abdulaziz; Batang, Zenon B.; Xu, Wei; Wang, Guang Chao; Zhang, Xixiang; Lafi, Feras F.; Bajic, Vladmir B.; Qian, Pei-Yuan

2014-01-01

Using microscopic and molecular techniques combined with computational analysis, this study examined the structure and composition of microbial communities in biofilms that formed on different artificial substrates in a brine pool and on a seep vent of a cold seep in the Red Sea to test our hypothesis that initiation of the biofilm formation and spreading mode of microbial structures differs between the cold seep and the other aquatic environments. Biofilms on different substrates at two deployment sites differed morphologically, with the vent biofilms having higher microbial abundance and better structural features than the pool biofilms. Microbes in the pool biofilms were more taxonomically diverse and mainly composed of various sulfate-reducing bacteria whereas the vent biofilms were exclusively dominated by sulfur-oxidizing Thiomicrospira. These results suggest that the redox environments at the deployment sites might have exerted a strong selection on microbes in the biofilms at two sites whereas the types of substrates had limited effects on the biofilm development. PMID:24399144

Molecular detection of Rickettsia species in ticks collected from the southwestern provinces of the Republic of Korea.

PubMed

Noh, Yoontae; Lee, Yeong Seon; Kim, Heung-Chul; Chong, Sung-Tae; Klein, Terry A; Jiang, Ju; Richards, Allen L; Lee, Hae Kyeong; Kim, Su Yeon

2017-01-10

Rickettsiae constitute a group of arthropod-borne, Gram-negative, obligate intracellular bacteria that are the causative agents of diseases ranging from mild to life threatening that impact on medical and veterinary health worldwide. A total of 6,484 ticks were collected by tick drag from June-October 2013 in the southwestern provinces of the Republic of Korea (ROK) (Jeollanam, n = 3,995; Jeollabuk, n = 680; Chungcheongnam, n = 1,478; and Chungcheongbuk, n = 331). Ticks were sorted into 311 pools according to species, collection site, and stage of development. DNA preparations of tick pools were assayed for rickettsiae by 17 kDa antigen gene and ompA nested PCR (nPCR) assays and the resulting amplicons sequenced to determine the identity and prevalence of spotted fever group rickettsiae (SFGR). Haemaphysalis longicornis (4,471; 52 adults, 123 nymphs and 4,296 larvae) were the most commonly collected ticks, followed by Haemaphysalis flava (1,582; 28 adults, 263 nymphs and 1,291 larvae), and Ixodes nipponensis (431; 25 adults, 5 nymphs and 401 larvae). The minimum field infection rate/100 ticks (assuming 1 positive tick/pool) was 0.93% for the 17 kDa antigen gene and 0.82% for the ompA nPCR assays. The partial 17 kDa antigen and ompA gene sequences from positive pools of H. longicornis were similar to: Rickettsia sp. HI550 (99.4-100%), Rickettsia sp. FUJ98 (99.3-100%), Rickettsia sp. HIR/D91 (99.3-100%), and R. japonica (99.7%). One sequence of the partial 17 kDa antigen gene for H. flava was similar to Rickettsia sp. 17kd-005 (99.7%), while seven sequences of the 17 kDa antigen gene obtained from I. nipponensis ticks were similar to R. monacensis IrR/Munich (98.7-100%) and Rickettsia sp. IRS3 (98.9%). SFG rickettsiae were detected in three species of ixodid ticks collected in the southwestern provinces of the ROK during 2013. A number of rickettsiae have been recently reported from ticks in Korea, some of which were identified as medically important. Results from this study and previous reports demonstrate the need to conduct longitudinal investigations to identify tick-borne rickettsiae and better understand their geographical distributions and potential impact on medical and veterinary health, in addition to risk communication and development of rickettsial disease prevention strategies.

Comparative transcriptomics of two environmentally relevant cyanobacteria reveals unexpected transcriptome diversity

PubMed Central

Voigt, Karsten; Sharma, Cynthia M; Mitschke, Jan; Joke Lambrecht, S; Voß, Björn; Hess, Wolfgang R; Steglich, Claudia

2014-01-01

Prochlorococcus is a genus of abundant and ecologically important marine cyanobacteria. Here, we present a comprehensive comparison of the structure and composition of the transcriptomes of two Prochlorococcus strains, which, despite their similarities, have adapted their gene pool to specific environmental constraints. We present genome-wide maps of transcriptional start sites (TSS) for both organisms, which are representatives of the two most diverse clades within the two major ecotypes adapted to high- and low-light conditions, respectively. Our data suggest antisense transcription for three-quarters of all genes, which is substantially more than that observed in other bacteria. We discovered hundreds of TSS within genes, most notably within 16 of the 29 prochlorosin genes, in strain MIT9313. A direct comparison revealed very little conservation in the location of TSS and the nature of non-coding transcripts between both strains. We detected extremely short 5′ untranslated regions with a median length of only 27 and 29 nt for MED4 and MIT9313, respectively, and for 8% of all protein-coding genes the median distance to the start codon is only 10 nt or even shorter. These findings and the absence of an obvious Shine–Dalgarno motif suggest that leaderless translation and ribosomal protein S1-dependent translation constitute alternative mechanisms for translation initiation in Prochlorococcus. We conclude that genome-wide antisense transcription is a major component of the transcriptional output from these relatively small genomes and that a hitherto unrecognized high degree of complexity and variability of gene expression exists in their transcriptional architecture. PMID:24739626

Assessing pooled BAC and whole genome shotgun strategies for assembly of complex genomes.

PubMed

Haiminen, Niina; Feltus, F Alex; Parida, Laxmi

2011-04-15

We investigate if pooling BAC clones and sequencing the pools can provide for more accurate assembly of genome sequences than the "whole genome shotgun" (WGS) approach. Furthermore, we quantify this accuracy increase. We compare the pooled BAC and WGS approaches using in silico simulations. Standard measures of assembly quality focus on assembly size and fragmentation, which are desirable for large whole genome assemblies. We propose additional measures enabling easy and visual comparison of assembly quality, such as rearrangements and redundant sequence content, relative to the known target sequence. The best assembly quality scores were obtained using 454 coverage of 15× linear and 5× paired (3kb insert size) reads (15L-5P) on Arabidopsis. This regime gave similarly good results on four additional plant genomes of very different GC and repeat contents. BAC pooling improved assembly scores over WGS assembly, coverage and redundancy scores improving the most. BAC pooling works better than WGS, however, both require a physical map to order the scaffolds. Pool sizes up to 12Mbp work well, suggesting this pooling density to be effective in medium-scale re-sequencing applications such as targeted sequencing of QTL intervals for candidate gene discovery. Assuming the current Roche/454 Titanium sequencing limitations, a 12 Mbp region could be re-sequenced with a full plate of linear reads and a half plate of paired-end reads, yielding 15L-5P coverage after read pre-processing. Our simulation suggests that massively over-sequencing may not improve accuracy. Our scoring measures can be used generally to evaluate and compare results of simulated genome assemblies.

Genome Sequence of Thalassospira xiamenensis Type Strain M-5

PubMed Central

Lai, Qiliang

2012-01-01

Thalassospira xiamenensis M-5T was isolated from the surface water of a waste oil pool at the oil storage dock in the city of Xiamen, Fujian Province, China. Here, we present the draft genome of strain M-5T, which contains 4,705,237 bp with a G+C content of 54.65% and contains 4,343 protein-coding genes and 46 tRNA genes. PMID:23209216

Gene Therapy for Fracture Repair

DTIC Science & Technology

2007-05-01

Methods: We have adopted the Agilent rat oligomer chip to analyze our fracture RNA in our microarray analysis. This chip has 20,046 unique gene...signal during fluorescent labeling of the cDNA. This approach is highly advantageous for reducing the RNA input into the system, minimizing the numbers...perform the analysis on these extremely limited samples without pooling the RNA from multiple individuals. We are therefore able to analyze the

High-density SNP genotyping array for hexaploid wheat and its secondary and tertiary gene pool.

PubMed

Winfield, Mark O; Allen, Alexandra M; Burridge, Amanda J; Barker, Gary L A; Benbow, Harriet R; Wilkinson, Paul A; Coghill, Jane; Waterfall, Christy; Davassi, Alessandro; Scopes, Geoff; Pirani, Ali; Webster, Teresa; Brew, Fiona; Bloor, Claire; King, Julie; West, Claire; Griffiths, Simon; King, Ian; Bentley, Alison R; Edwards, Keith J

2016-05-01

In wheat, a lack of genetic diversity between breeding lines has been recognized as a significant block to future yield increases. Species belonging to bread wheat's secondary and tertiary gene pools harbour a much greater level of genetic variability, and are an important source of genes to broaden its genetic base. Introgression of novel genes from progenitors and related species has been widely employed to improve the agronomic characteristics of hexaploid wheat, but this approach has been hampered by a lack of markers that can be used to track introduced chromosome segments. Here, we describe the identification of a large number of single nucleotide polymorphisms that can be used to genotype hexaploid wheat and to identify and track introgressions from a variety of sources. We have validated these markers using an ultra-high-density Axiom(®) genotyping array to characterize a range of diploid, tetraploid and hexaploid wheat accessions and wheat relatives. To facilitate the use of these, both the markers and the associated sequence and genotype information have been made available through an interactive web site. © 2015 The Authors. Plant Biotechnology Journal published by Society for Experimental Biology and The Association of Applied Biologists and John Wiley & Sons Ltd.

Stimulation of autophagy by the p53 target gene Sestrin2.

PubMed

Maiuri, Maria Chiara; Malik, Shoaib Ahmad; Morselli, Eugenia; Kepp, Oliver; Criollo, Alfredo; Mouchel, Pierre-Luc; Carnuccio, Rosa; Kroemer, Guido

2009-05-15

The oncosuppressor protein p53 regulates autophagy in a dual fashion. The pool of cytoplasmic p53 protein represses autophagy in a transcription-independent fashion, while the pool of nuclear p53 stimulates autophagy through the transactivation of specific genes. Here we report the discovery that Sestrin2, a novel p53 target gene, is involved in the induction of autophagy. Depletion of Sestrin2 by RNA interference reduced the level of autophagy in a panel of p53-sufficient human cancer cell lines responding to distinct autophagy inducers. In quantitative terms, Sestrin2 depletion was as efficient in preventing autophagy induction as was the depletion of Dram, another p53 target gene. Knockout of either Sestrin2 or Dram reduced autophagy elicited by nutrient depletion, rapamycin, lithium or thapsigargin. Moreover, autophagy induction by nutrient depletion or pharmacological stimuli led to an increase in Sestrin2 expression levels in p53-proficient cells. In strict contrast, the depletion of Sestrin2 or Dram failed to affect autophagy in p53-deficient cells and did not modulate the inhibition of baseline autophagy by a cytoplasmic p53 mutant that was reintroduced into p53-deficient cells. We conclude that Sestrin2 acts as a positive regulator of autophagy in p53-proficient cells.

Annotating ebony on the fly.

PubMed

Kohn, Michael H; Wittkopp, Patricia J

2007-07-01

The distinctive black phenotype of ebony mutants has made it one of the most widely used phenotypic markers in Drosophila genetics. Without doubt, ebony showcases the fruits of the fly community's labours to annotate gene function. As of this writing, FlyBase lists 142 references, 1277 fly stocks, 15 phenotypes and 44 alleles. In addition to its namesake pigmentation phenotype, ebony mutants affect other traits, including phototaxis and courtship. With phenotypic consequences of ebony variants readily apparent in the laboratory, does natural selection also see them in the wild? In this issue of Molecular Ecology, Pool & Aquadro investigate this question and found signs of natural selection on the ebony gene that appear to have resulted from selection for darker pigmentation at higher elevations in sub-Saharan populations of Drosophila melanogaster. Such findings from population genomic analysis of wild-derived strains should be included in gene annotations to provide a more holistic view of a gene's function. The evolutionary annotation of ebony added by Pool & Aquadro substantiates that pigmentation can be adaptive and implicates elevation as an important selective factor. This is important progress because the selective factors seem to differ between populations and species. In addition, the study raises issues to consider when extrapolating from selection at the molecular level to selection at the phenotypic level.

[Genetic diversity and relationships of northern eurasia populations for polymorphic Alu-insertions].

PubMed

Khitrinskaia, I Iu; Khar'kov, V N; Voevoda, M I; Stepanova, V A

2014-01-01

We for the first time have examined the autosomal gene pool of the Siberia, Central Asian and the Far East populations (27 populations of 12 ethnic groups) using a set of polymorphic Alu insertions in the human genome. The results of the analysis testify (i) to a significant level of genetic diversity in the Northern Eurasian populations and (ii) to a considerable differentiation of gene pool in the population of this region. It has been shown that at the CD4 locus, the frequency of Alu (-) is inversely related to the Mongoloid component of the population, the lowest and highest frequencies of the Alu deletion at locus CD4 were recorded respectively in Eskimo (0.012) and Russian and Ukrainian (0.35). The analysis of gene flow proved Caucasoid populations (Russian, Tajik and Uzbek), as well as those of Turkic ethnic groups from the Southern Siberia (Altaians and Tuvinians) and Khanty and Mansy populations to be the recipients of a considerable gene flow from the outside of the concerned population system, as compared with the East Siberian and the Far East ethnic groups. The results of the correlation analysis received with use polymorphic Alu insertion testify to the greatest correlation of genetic distances with anthropological characteristics of populations.

SNPGenie: estimating evolutionary parameters to detect natural selection using pooled next-generation sequencing data.

PubMed

Nelson, Chase W; Moncla, Louise H; Hughes, Austin L

2015-11-15

New applications of next-generation sequencing technologies use pools of DNA from multiple individuals to estimate population genetic parameters. However, no publicly available tools exist to analyse single-nucleotide polymorphism (SNP) calling results directly for evolutionary parameters important in detecting natural selection, including nucleotide diversity and gene diversity. We have developed SNPGenie to fill this gap. The user submits a FASTA reference sequence(s), a Gene Transfer Format (.GTF) file with CDS information and a SNP report(s) in an increasing selection of formats. The program estimates nucleotide diversity, distance from the reference and gene diversity. Sites are flagged for multiple overlapping reading frames, and are categorized by polymorphism type: nonsynonymous, synonymous, or ambiguous. The results allow single nucleotide, single codon, sliding window, whole gene and whole genome/population analyses that aid in the detection of positive and purifying natural selection in the source population. SNPGenie version 1.2 is a Perl program with no additional dependencies. It is free, open-source, and available for download at https://github.com/hugheslab/snpgenie. nelsoncw@email.sc.edu or austin@biol.sc.edu Supplementary data are available at Bioinformatics online. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Whole exome sequencing in recurrent early pregnancy loss.

PubMed

Qiao, Ying; Wen, Jiadi; Tang, Flamingo; Martell, Sally; Shomer, Naomi; Leung, Peter C K; Stephenson, Mary D; Rajcan-Separovic, Evica

2016-05-01

Exome sequencing can identify genetic causes of idiopathic recurrent pregnancy loss (RPL). We identified compound heterozygous deleterious mutations affecting DYNC2H1 and ALOX15 in two out of four families with RPL. Both genes have a role in early development. Bioinformatics analysis of all genes with rare and putatively pathogenic mutations in miscarriages and couples showed enrichment in pathways relevant to pregnancy loss, including the complement and coagulation cascades pathways. Next generation sequencing (NGS) is increasingly being used to identify known and novel gene mutations in children with developmental delay and in fetuses with ultrasound-detected anomalies. In contrast, NGS is rarely used to study pregnancy loss. Chromosome microarray analysis detects putatively causative DNA copy number variants (CNVs) in ∼2% of miscarriages and CNVs of unknown significance (predominantly parental in origin) in up to 40% of miscarriages. Therefore, a large number of miscarriages still have an unknown cause. Whole exome sequencing (WES) was performed using Illumina HiSeq 2000 platform on seven euploid miscarriages from four families with RPL. Golden Helix SVS v8.1.5 was used for data assessment and inheritance analysis for deleterious DNA variants predicted to severely disrupt protein-coding genes by introducing a frameshift, loss of the stop codon, gain of the stop codon, changes in splicing or the initial codon. Webgestalt (http://bioinfo.vanderbilt.edu/webgestalt/) was used for pathway and disease association enrichment analysis of a gene pool containing putatively pathogenic variants in miscarriages and couples in comparison to control gene pools. Compound heterozygous mutations in DYNC2H1 and ALOX15 were identified in miscarriages from two families with RPL. DYNC2H1 is involved in cilia biogenesis and has been associated with fetal lethality in humans. ALOX15 is expressed in placenta and its dysregulation has been associated with inflammation, placental, dysfunction, abnormal oxidative stress response and angiogenesis. The pool of putatively pathogenic single nucleotide variants (SNVs) and small insertions and deletions (indels) detected in the miscarriages showed enrichment in 'complement and coagulation cascades pathway', and 'ciliary motility disorders'. We conclude that CNVs, individual SNVs and pool of deleterious gene mutations identified by exome sequencing could contribute to RPL. The size of our sample cohort is small. The functional effect of candidate mutations should be evaluated to determine whether the mutations are causative. This is the first study to assess whether SNVs may contribute to the pathogenesis of miscarriage. Furthermore, our findings suggest that collective effect of mutations in relevant biological pathways could be implicated in RPL. The study was funded by Canadian Institutes of Health Research (grant MOP 106467) and Michael Smith Foundation of Health Research Career Scholar salary award to ERS. © The Author 2016. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

Whole exome sequencing in recurrent early pregnancy loss

PubMed Central

Qiao, Ying; Wen, Jiadi; Tang, Flamingo; Martell, Sally; Shomer, Naomi; Leung, Peter C.K.; Stephenson, Mary D.; Rajcan-Separovic, Evica

2016-01-01

STUDY HYPOTHESIS Exome sequencing can identify genetic causes of idiopathic recurrent pregnancy loss (RPL). STUDY FINDING We identified compound heterozygous deleterious mutations affecting DYNC2H1 and ALOX15 in two out of four families with RPL. Both genes have a role in early development. Bioinformatics analysis of all genes with rare and putatively pathogenic mutations in miscarriages and couples showed enrichment in pathways relevant to pregnancy loss, including the complement and coagulation cascades pathways. WHAT IS KNOWN ALREADY Next generation sequencing (NGS) is increasingly being used to identify known and novel gene mutations in children with developmental delay and in fetuses with ultrasound-detected anomalies. In contrast, NGS is rarely used to study pregnancy loss. Chromosome microarray analysis detects putatively causative DNA copy number variants (CNVs) in ∼2% of miscarriages and CNVs of unknown significance (predominantly parental in origin) in up to 40% of miscarriages. Therefore, a large number of miscarriages still have an unknown cause. STUDY DESIGN, SAMPLES/MATERIALS, METHODS Whole exome sequencing (WES) was performed using Illumina HiSeq 2000 platform on seven euploid miscarriages from four families with RPL. Golden Helix SVS v8.1.5 was used for data assessment and inheritance analysis for deleterious DNA variants predicted to severely disrupt protein-coding genes by introducing a frameshift, loss of the stop codon, gain of the stop codon, changes in splicing or the initial codon. Webgestalt (http://bioinfo.vanderbilt.edu/webgestalt/) was used for pathway and disease association enrichment analysis of a gene pool containing putatively pathogenic variants in miscarriages and couples in comparison to control gene pools. MAIN RESULTS AND THE ROLE OF CHANCE Compound heterozygous mutations in DYNC2H1 and ALOX15 were identified in miscarriages from two families with RPL. DYNC2H1 is involved in cilia biogenesis and has been associated with fetal lethality in humans. ALOX15 is expressed in placenta and its dysregulation has been associated with inflammation, placental, dysfunction, abnormal oxidative stress response and angiogenesis. The pool of putatively pathogenic single nucleotide variants (SNVs) and small insertions and deletions (indels) detected in the miscarriages showed enrichment in ‘complement and coagulation cascades pathway’, and ‘ciliary motility disorders’. We conclude that CNVs, individual SNVs and pool of deleterious gene mutations identified by exome sequencing could contribute to RPL. LIMITATIONS, REASONS FOR CAUTION The size of our sample cohort is small. The functional effect of candidate mutations should be evaluated to determine whether the mutations are causative. WIDER IMPLICATIONS OF THE FINDINGS This is the first study to assess whether SNVs may contribute to the pathogenesis of miscarriage. Furthermore, our findings suggest that collective effect of mutations in relevant biological pathways could be implicated in RPL. STUDY FUNDING AND COMPETING INTEREST(S) The study was funded by Canadian Institutes of Health Research (grant MOP 106467) and Michael Smith Foundation of Health Research Career Scholar salary award to ERS. PMID:26826164

Genomics of crop wild relatives: expanding the gene pool for crop improvement.

PubMed

Brozynska, Marta; Furtado, Agnelo; Henry, Robert J

2016-04-01

Plant breeders require access to new genetic diversity to satisfy the demands of a growing human population for more food that can be produced in a variable or changing climate and to deliver the high-quality food with nutritional and health benefits demanded by consumers. The close relatives of domesticated plants, crop wild relatives (CWRs), represent a practical gene pool for use by plant breeders. Genomics of CWR generates data that support the use of CWR to expand the genetic diversity of crop plants. Advances in DNA sequencing technology are enabling the efficient sequencing of CWR and their increased use in crop improvement. As the sequencing of genomes of major crop species is completed, attention has shifted to analysis of the wider gene pool of major crops including CWR. A combination of de novo sequencing and resequencing is required to efficiently explore useful genetic variation in CWR. Analysis of the nuclear genome, transcriptome and maternal (chloroplast and mitochondrial) genome of CWR is facilitating their use in crop improvement. Genome analysis results in discovery of useful alleles in CWR and identification of regions of the genome in which diversity has been lost in domestication bottlenecks. Targeting of high priority CWR for sequencing will maximize the contribution of genome sequencing of CWR. Coordination of global efforts to apply genomics has the potential to accelerate access to and conservation of the biodiversity essential to the sustainability of agriculture and food production. © 2015 Society for Experimental Biology, Association of Applied Biologists and John Wiley & Sons Ltd.

Lake-specific variation in growth, migration timing and survival of juvenile sockeye salmon Oncorhynchus nerka: separating environmental from genetic influences.

PubMed

Reed, T E; Martinek, G; Quinn, T P

2010-08-01

Time series on juvenile life-history traits obtained from sockeye salmon Oncorhynchus nerka were analysed to assess lake-specific environmental influences on juvenile migration timing, size and survival of fish from a common gene pool. Every year for the past two decades, O. nerka have been spawned at a hatchery facility, and the progeny released into two lakes that differ in average summer temperatures, limnological attributes and growth opportunities. Juveniles reared in the warmer, more productive Crosswind Lake were larger and heavier as smolts compared to those from the cooler, less productive Summit Lake and had higher in-lake and subsequent marine survival. Crosswind Lake smolts migrated from the lake to sea slightly earlier in the season but the migration timing distributions overlapped considerably across years. Fry stocking density had a negative effect on smolt length for both lakes, and a negative effect on in-lake survival in Summit Lake. Taken together, the results revealed a strong effect of lake-rearing environment on the expression of life-history variation in O. nerka. The stocking of these lakes each year with juveniles from a single mixed-source population provided a large-scale reverse common-garden experiment, where the same gene pool was exposed to different environments, rather than the different gene pools in the same environment approach typical of evolutionary ecology studies. Other researchers are encouraged to seek and exploit similar serendipitous situations, which might allow environmental and genetic influences on ecologically important traits to be distinguished in natural or semi-natural settings.

The Lcn972 Bacteriocin-Encoding Plasmid pBL1 Impairs Cellobiose Metabolism in Lactococcus lactis▿

PubMed Central

Campelo, Ana B.; Gaspar, Paula; Roces, Clara; Rodríguez, Ana; Kok, Jan; Kuipers, Oscar P.; Neves, Ana Rute; Martínez, Beatriz

2011-01-01

pBL1 is a Lactococcus lactis theta-replicating 10.9-kbp plasmid that encodes the synthetic machinery of the bacteriocin Lcn972. In this work, the transcriptomes of exponentially growing L. lactis strains with and without pBL1 were compared. A discrete response was observed, with a total of 10 genes showing significantly changed expression. Upregulation of the lactococcal oligopeptide uptake (opp) system was observed, which was likely linked to a higher nitrogen demand required for Lcn972 biosynthesis. Strikingly, celB, coding for the membrane porter IIC of the cellobiose phosphoenolpyruvate-dependent phosphotransferase system (PTS), and the upstream gene llmg0186 were downregulated. Growth profiles for L. lactis strains MG1363, MG1363/pBL1, and MG1363 ΔcelB grown in chemically defined medium (CDM) containing cellobiose confirmed slower growth of MG1363/pBL1 and MG1363 ΔcelB, while no differences were observed with growth on glucose. The presence of pBL1 shifted the fermentation products toward a mixed acid profile and promoted substantial changes in intracellular pool sizes for glycolytic intermediates in cells growing on cellobiose as determined by high-pressure liquid chromatography (HPLC) and nuclear magnetic resonance (NMR). Overall, these data support the genetic evidence of a constriction in cellobiose uptake. Notably, several cell wall precursors accumulated, while other UDP-activated sugar pools were lower, which could reflect rerouting of precursors toward the production of structural or storage polysaccharides. Moreover, cells growing slowly on cellobiose and those lacking celB were more tolerant to Lcn972 than cellobiose-adapted cells. Thus, downregulation of celB could help to build up a response against the antimicrobial activity of Lcn972, enhancing self-immunity of the producer cells. PMID:21890668

Population genomics of early events in the ecological differentiation of bacteria

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

Shapiro, Jesse B.; Friedman, Jonatan; Cordero, Otto X.

Genetic exchange is common among bacteria, but its effect on population diversity during ecological differentiation remains controversial. A fundamental question is whether advantageous mutations lead to selection of clonal genomes or, as in sexual eukaryotes, sweep through populations on their own. Here, we show that in two recently diverged populations of ocean bacteria, ecological differentiation has occurred akin to a sexual mechanism: A few genome regions have swept through subpopulations in a habitat-specific manner, accompanied by gradual separation of gene pools as evidenced by increased habitat specificity of the most recent recombinations. These findings reconcile previous, seemingly contradictory empirical observationsmore » of the genetic structure of bacterial populations and point to a more unified process of differentiation in bacteria and sexual eukaryotes than previously thought.« less

The MIGenAS integrated bioinformatics toolkit for web-based sequence analysis

PubMed Central

Rampp, Markus; Soddemann, Thomas; Lederer, Hermann

2006-01-01

We describe a versatile and extensible integrated bioinformatics toolkit for the analysis of biological sequences over the Internet. The web portal offers convenient interactive access to a growing pool of chainable bioinformatics software tools and databases that are centrally installed and maintained by the RZG. Currently, supported tasks comprise sequence similarity searches in public or user-supplied databases, computation and validation of multiple sequence alignments, phylogenetic analysis and protein–structure prediction. Individual tools can be seamlessly chained into pipelines allowing the user to conveniently process complex workflows without the necessity to take care of any format conversions or tedious parsing of intermediate results. The toolkit is part of the Max-Planck Integrated Gene Analysis System (MIGenAS) of the Max Planck Society available at (click ‘Start Toolkit’). PMID:16844980

Genetic linkage analysis using pooled DNA and infrared detection of tailed STRP primer patterns

NASA Astrophysics Data System (ADS)

Oetting, William S.; Wildenberg, Scott C.; King, Richard A.

1996-04-01

The mapping of a disease locus to a specific chromosomal region is an important step in the eventual isolation and analysis of a disease causing gene. Conventional mapping methods analyze large multiplex families and/or smaller nuclear families to find linkage between the disease and a chromosome marker that maps to a known chromosomal region. This analysis is time consuming and tedious, typically requiring the determination of 30,000 genotypes or more. For appropriate populations, we have instead utilized pooled DNA samples for gene mapping which greatly reduces the amount of time necessary for an initial chromosomal screen. This technique assumes a common founder for the disease locus of interest and searches for a region of a chromosome shared between affected individuals. Our analysis involves the PCR amplification of short tandem repeat polymorphisms (STRP) to detect these shared regions. In order to reduce the cost of genotyping, we have designed unlabeled tailed PCR primers which, when combined with a labeled universal primer, provides for an alternative to synthesizing custom labeled primers. The STRP pattern is visualized with an infrared fluorescence based automated DNA sequencer and the patterns quantitated by densitometric analysis of the allele pattern. Differences in the distribution of alleles between pools of affected and unaffected individuals, including a reduction in the number of alleles in the affected pool, indicate the sharing of a region of a chromosome. We have found this method effective for markers 10 - 15 cM away from the disease locus for a recessive genetic disease.

Evidence for an apartheid-like social structure in early Anglo-Saxon England.

PubMed

Thomas, Mark G; Stumpf, Michael P H; Härke, Heinrich

2006-10-22

The role of migration in the Anglo-Saxon transition in England remains controversial. Archaeological and historical evidence is inconclusive, but current estimates of the contribution of migrants to the English population range from less than 10000 to as many as 200000. In contrast, recent studies based on Y-chromosome variation posit a considerably higher contribution to the modern English gene pool (50-100%). Historical evidence suggests that following the Anglo-Saxon transition, people of indigenous ethnicity were at an economic and legal disadvantage compared to those having Anglo-Saxon ethnicity. It is likely that such a disadvantage would lead to differential reproductive success. We examine the effect of differential reproductive success, coupled with limited intermarriage between distinct ethnic groups, on the spread of genetic variants. Computer simulations indicate that a social structure limiting intermarriage between indigenous Britons and an initially small Anglo-Saxon immigrant population provide a plausible explanation of the high degree of Continental male-line ancestry in England.

Evidence for an apartheid-like social structure in early Anglo-Saxon England

PubMed Central

Thomas, Mark G; Stumpf, Michael P.H; Härke, Heinrich

2006-01-01

The role of migration in the Anglo-Saxon transition in England remains controversial. Archaeological and historical evidence is inconclusive, but current estimates of the contribution of migrants to the English population range from less than 10 000 to as many as 200 000. In contrast, recent studies based on Y-chromosome variation posit a considerably higher contribution to the modern English gene pool (50–100%). Historical evidence suggests that following the Anglo-Saxon transition, people of indigenous ethnicity were at an economic and legal disadvantage compared to those having Anglo-Saxon ethnicity. It is likely that such a disadvantage would lead to differential reproductive success. We examine the effect of differential reproductive success, coupled with limited intermarriage between distinct ethnic groups, on the spread of genetic variants. Computer simulations indicate that a social structure limiting intermarriage between indigenous Britons and an initially small Anglo-Saxon immigrant population provide a plausible explanation of the high degree of Continental male-line ancestry in England. PMID:17002951

Chloroplast Genome Sequence of Pigeonpea (Cajanus cajan (L.) Millspaugh) and Cajanus scarabaeoides (L.) Thouars: Genome Organization and Comparison with Other Legumes

PubMed Central

Kaila, Tanvi; Chaduvla, Pavan K.; Saxena, Swati; Bahadur, Kaushlendra; Gahukar, Santosh J.; Chaudhury, Ashok; Sharma, T. R.; Singh, N. K.; Gaikwad, Kishor

2016-01-01

Pigeonpea (Cajanus cajan (L.) Millspaugh), a diploid (2n = 22) legume crop with a genome size of 852 Mbp, serves as an important source of human dietary protein especially in South East Asian and African regions. In this study, the draft chloroplast genomes of Cajanus cajan and Cajanus scarabaeoides (L.) Thouars were generated. Cajanus scarabaeoides is an important species of the Cajanus gene pool and has also been used for developing promising CMS system by different groups. A male sterile genotype harboring the C. scarabaeoides cytoplasm was used for sequencing the plastid genome. The cp genome of C. cajan is 152,242bp long, having a quadripartite structure with LSC of 83,455 bp and SSC of 17,871 bp separated by IRs of 25,398 bp. Similarly, the cp genome of C. scarabaeoides is 152,201bp long, having a quadripartite structure in which IRs of 25,402 bp length separates 83,423 bp of LSC and 17,854 bp of SSC. The pigeonpea cp genome contains 116 unique genes, including 30 tRNA, 4 rRNA, 78 predicted protein coding genes and 5 pseudogenes. A 50 kb inversion was observed in the LSC region of pigeonpea cp genome, consistent with other legumes. Comparison of cp genome with other legumes revealed the contraction of IR boundaries due to the absence of rps19 gene in the IR region. Chloroplast SSRs were mined and a total of 280 and 292 cpSSRs were identified in C. scarabaeoides and C. cajan respectively. RNA editing was observed at 37 sites in both C. scarabaeoides and C. cajan, with maximum occurrence in the ndh genes. The pigeonpea cp genome sequence would be beneficial in providing informative molecular markers which can be utilized for genetic diversity analysis and aid in understanding the plant systematics studies among major grain legumes. PMID:28018385

Chloroplast Genome Sequence of Pigeonpea (Cajanus cajan (L.) Millspaugh) and Cajanus scarabaeoides (L.) Thouars: Genome Organization and Comparison with Other Legumes.

PubMed

Kaila, Tanvi; Chaduvla, Pavan K; Saxena, Swati; Bahadur, Kaushlendra; Gahukar, Santosh J; Chaudhury, Ashok; Sharma, T R; Singh, N K; Gaikwad, Kishor

2016-01-01

Pigeonpea ( Cajanus cajan (L.) Millspaugh), a diploid (2n = 22) legume crop with a genome size of 852 Mbp, serves as an important source of human dietary protein especially in South East Asian and African regions. In this study, the draft chloroplast genomes of Cajanus cajan and Cajanus scarabaeoides (L.) Thouars were generated. Cajanus scarabaeoides is an important species of the Cajanus gene pool and has also been used for developing promising CMS system by different groups. A male sterile genotype harboring the C. scarabaeoides cytoplasm was used for sequencing the plastid genome. The cp genome of C. cajan is 152,242bp long, having a quadripartite structure with LSC of 83,455 bp and SSC of 17,871 bp separated by IRs of 25,398 bp. Similarly, the cp genome of C. scarabaeoides is 152,201bp long, having a quadripartite structure in which IRs of 25,402 bp length separates 83,423 bp of LSC and 17,854 bp of SSC. The pigeonpea cp genome contains 116 unique genes, including 30 tRNA, 4 rRNA, 78 predicted protein coding genes and 5 pseudogenes. A 50 kb inversion was observed in the LSC region of pigeonpea cp genome, consistent with other legumes. Comparison of cp genome with other legumes revealed the contraction of IR boundaries due to the absence of rps19 gene in the IR region. Chloroplast SSRs were mined and a total of 280 and 292 cpSSRs were identified in C. scarabaeoides and C. cajan respectively. RNA editing was observed at 37 sites in both C. scarabaeoides and C. cajan , with maximum occurrence in the ndh genes. The pigeonpea cp genome sequence would be beneficial in providing informative molecular markers which can be utilized for genetic diversity analysis and aid in understanding the plant systematics studies among major grain legumes.

BAC-pool sequencing and analysis confirms growth-associated QTLs in the Asian seabass genome.

PubMed

Shen, Xueyan; Ngoh, Si Yan; Thevasagayam, Natascha May; Prakki, Sai Rama Sridatta; Bhandare, Pranjali; Tan, Andy Wee Kiat; Tan, Gui Quan; Singh, Siddharth; Phua, Norman Chun Han; Vij, Shubha; Orbán, László

2016-11-08

The Asian seabass is an important marine food fish that has been cultured for several decades in Asia Pacific. However, the lack of a high quality reference genome has hampered efforts to improve its selective breeding. A 3D BAC pool set generated in this study was screened using 22 SSR markers located on linkage group 2 which contains a growth-related QTL region. Seventy-two clones corresponding to 22 FPC contigs were sequenced by Illumina MiSeq technology. We co-assembled the MiSeq-derived scaffolds from each FPC contig with error-corrected PacBio reads, resulting in 187 sequences covering 9.7 Mb. Eleven genes annotated within this region were found to be potentially associated with growth and their tissue-specific expression was investigated. Correlation analysis demonstrated that SNPs in ctsb, skp1 and ppp2ca can be potentially used as markers for selecting fast-growing fingerlings. Conserved syntenies between seabass LG2 and five other teleosts were identified. This study i) provided a 10 Mb targeted genome assembly; ii) demonstrated NGS of BAC pools as a potential approach for mining candidates underlying QTLs of this species; iii) detected eleven genes potentially responsible for growth in the QTL region; and iv) identified useful SNP markers for selective breeding programs of Asian seabass.

On the relative importance of pool morphology and woody debris to distributions of shrimp in a Puerto Rican headwater stream

USGS Publications Warehouse

Pyron, M.; Covich, A.P.; Black, R.W.

1999-01-01

In this paper, we report the sizes and distributional orientation of woody debris in a headwater rainforest stream in the Luquillo Experimental Forest (LEF), Puerto Rico. We also provide results of a 4-month study of a wood addition experiment designed to increase cover for benthic macroinvertebrates (freshwater shrimp). We added branch-sized woody debris to 20 pools in three streams. We trapped four species of freshwater shrimp (two species of benthic detritivores and two predatory shrimp species) during each of the 4 months following wood additions. An analysis of pool morphology (maximum depth, surface area and volume) provided a useful predictor of shrimp abundances. In general, numbers of shrimps increased with sizes of stream pools. A repeated measures ANOVA demonstrated no effect of woody debris additions on total numbers of shrimp per pool area. Two detritivore species (Atya lanipes, a filter feeder and Xiphocaris elongata, a shredder) decreased in abundance with increased woody debris and there was no statistical relationship between woody debris additions and predators (Macrobrachium carcinus and M. crenulatum). Small woody debris additions may have altered flow velocities that were important to filter-feeding Atya at the microhabitat scale, although the overall velocities within pools were not altered by wood additions. Lower numbers of Atya and Xiphocaris in two of the three streams may result from the occurrence of two predaceous fishes (American eel and mountain mullet) and more predatory Macrobrachium in these streams. One likely interpretation of the results of this study is that the stream pools in these study reaches had sufficient habitat structure provided by numerous rock crevices (among large rocks and boulders) to provide refuge from predators. Addition of woody debris did not add significantly to the existing structure. These results may not apply to stream channels with sand and gravel substrata where crevices and undercut banks are lacking and where woody debris often plays a major role by providing structure and refuge.

Study on Dynamic Development of Three-dimensional Weld Pool Surface in Stationary GTAW

NASA Astrophysics Data System (ADS)

Huang, Jiankang; He, Jing; He, Xiaoying; Shi, Yu; Fan, Ding

2018-04-01

The weld pool contains abundant information about the welding process. In particular, the type of the weld pool surface shape, i. e., convex or concave, is determined by the weld penetration. To detect it, an innovative laser-vision-based sensing method is employed to observe the weld pool surface of the gas tungsten arc welding (GTAW). A low-power laser dots pattern is projected onto the entire weld pool surface. Its reflection is intercepted by a screen and captured by a camera. Then the dynamic development process of the weld pool surface can be detected. By observing and analyzing, the change of the reflected laser dots reflection pattern, for shape of the weld pool surface shape, was found to closely correlate to the penetration of weld pool in the welding process. A mathematical model was proposed to correlate the incident ray, reflected ray, screen and surface of weld pool based on structured laser specular reflection. The dynamic variation of the weld pool surface and its corresponding dots laser pattern were simulated and analyzed. By combining the experimental data and the mathematical analysis, the results show that the pattern of the reflected laser dots pattern is closely correlated to the development of weld pool, such as the weld penetration. The concavity of the pool surface was found to increase rapidly after the surface shape was changed from convex to concave during the stationary GTAW process.

Nuclear physics: quantitative single-cell approaches to nuclear organization and gene expression.

PubMed

Lionnet, T; Wu, B; Grünwald, D; Singer, R H; Larson, D R

2010-01-01

The internal workings of the nucleus remain a mystery. A list of component parts exists, and in many cases their functional roles are known for events such as transcription, RNA processing, or nuclear export. Some of these components exhibit structural features in the nucleus, regions of concentration or bodies that have given rise to the concept of functional compartmentalization--that there are underlying organizational principles to be described. In contrast, a picture is emerging in which transcription appears to drive the assembly of the functional components required for gene expression, drawing from pools of excess factors. Unifying this seemingly dual nature requires a more rigorous approach, one in which components are tracked in time and space and correlated with onset of specific nuclear functions. In this chapter, we anticipate tools that will address these questions and provide the missing kinetics of nuclear function. These tools are based on analyzing the fluctuations inherent in the weak signals of endogenous nuclear processes and determining values for them. In this way, it will be possible eventually to provide a computational model describing the functional relationships of essential components.

Landscape genomics reveal signatures of local adaptation in barley (Hordeum vulgare L.)

PubMed Central

Abebe, Tiegist D.; Naz, Ali A.; Léon, Jens

2015-01-01

Land plants are sessile organisms that cannot escape the adverse climatic conditions of a given environment. Hence, adaptation is one of the solutions to surviving in a challenging environment. This study was aimed at detecting adaptive loci in barley landraces that are affected by selection. To that end, a diverse population of barley landraces was analyzed using the genotyping by sequencing approach. Climatic data for altitude, rainfall and temperature were collected from 61 weather sites near the origin of selected landraces across Ethiopia. Population structure analysis revealed three groups whereas spatial analysis accounted significant similarities at shorter geographic distances (< 40 Km) among barley landraces. Partitioning the variance between climate variables and geographic distances indicated that climate variables accounted for most of the explainable genetic variation. Markers by climatic variables association analysis resulted in altogether 18 and 62 putative adaptive loci using Bayenv and latent factor mixed model (LFMM), respectively. Subsequent analysis of the associated SNPs revealed putative candidate genes for plant adaptation. This study highlights the presence of putative adaptive loci among barley landraces representing original gene pool of the farming communities. PMID:26483825

Development of a self-report physical function instrument for disability assessment: item pool construction and factor analysis.

PubMed

McDonough, Christine M; Jette, Alan M; Ni, Pengsheng; Bogusz, Kara; Marfeo, Elizabeth E; Brandt, Diane E; Chan, Leighton; Meterko, Mark; Haley, Stephen M; Rasch, Elizabeth K

2013-09-01

To build a comprehensive item pool representing work-relevant physical functioning and to test the factor structure of the item pool. These developmental steps represent initial outcomes of a broader project to develop instruments for the assessment of function within the context of Social Security Administration (SSA) disability programs. Comprehensive literature review; gap analysis; item generation with expert panel input; stakeholder interviews; cognitive interviews; cross-sectional survey administration; and exploratory and confirmatory factor analyses to assess item pool structure. In-person and semistructured interviews and Internet and telephone surveys. Sample of SSA claimants (n=1017) and a normative sample of adults from the U.S. general population (n=999). Not applicable. Model fit statistics. The final item pool consisted of 139 items. Within the claimant sample, 58.7% were white; 31.8% were black; 46.6% were women; and the mean age was 49.7 years. Initial factor analyses revealed a 4-factor solution, which included more items and allowed separate characterization of: (1) changing and maintaining body position, (2) whole body mobility, (3) upper body function, and (4) upper extremity fine motor. The final 4-factor model included 91 items. Confirmatory factor analyses for the 4-factor models for the claimant and the normative samples demonstrated very good fit. Fit statistics for claimant and normative samples, respectively, were: Comparative Fit Index=.93 and .98; Tucker-Lewis Index=.92 and .98; and root mean square error approximation=.05 and .04. The factor structure of the physical function item pool closely resembled the hypothesized content model. The 4 scales relevant to work activities offer promise for providing reliable information about claimant physical functioning relevant to work disability. Copyright © 2013 American Congress of Rehabilitation Medicine. Published by Elsevier Inc. All rights reserved.

Diversity of nifH gene pools in the rhizosphere of two cultivars of sorghum (Sorghum bicolor) treated with contrasting levels of nitrogen fertilizer.

PubMed

Coelho, Marcia Reed Rodrigues; de Vos, Marjon; Carneiro, Newton Portilho; Marriel, Ivanildo Evódio; Paiva, Edilson; Seldin, Lucy

2008-02-01

The diversity of nitrogen-fixing bacteria was assessed in the rhizospheres of two cultivars of sorghum (IS 5322-C and IPA 1011) sown in Cerrado soil amended with two levels of nitrogen fertilizer (12 and 120 kg ha(-1)). The nifH gene was amplified directly from DNA extracted from the rhizospheres, and the PCR products cloned and sequenced. Four clone libraries were generated from the nifH fragments and 245 sequences were obtained. Most of the clones (57%) were closely related to nifH genes of uncultured bacteria. NifH clones affiliated with Azohydromonas spp., Ideonella sp., Rhizobium etli and Bradyrhizobium sp. were found in all libraries. Sequences affiliated with Delftia tsuruhatensis were found in the rhizosphere of both cultivars sown with high levels of nitrogen, while clones affiliated with Methylocystis sp. were detected only in plants sown under low levels of nitrogen. Moreover, clones affiliated with Paenibacillus durus could be found in libraries from the cultivar IS 5322-C sown either in high or low amounts of fertilizer. This study showed that the amount of nitrogen used for fertilization is the overriding determinative factor that influenced the nitrogen-fixing community structures in sorghum rhizospheres cultivated in Cerrado soil.

Prevalence and diversity of H9N2 avian influenza in chickens of Northern Vietnam, 2014.

PubMed

Thuy, Duong Mai; Peacock, Thomas P; Bich, Vu Thi Ngoc; Fabrizio, Thomas; Hoang, Dang Nguyen; Tho, Nguyen Dang; Diep, Nguyen Thi; Nguyen, Minh; Hoa, Le Nguyen Minh; Trang, Hau Thi Thu; Choisy, Marc; Inui, Ken; Newman, Scott; Trung, Nguyen Vu; van Doorn, Rogier; To, Thanh Long; Iqbal, Munir; Bryant, Juliet E

2016-10-01

Despite their classification as low pathogenicity avian influenza viruses (LPAIV), A/H9N2 viruses cause significant losses in poultry in many countries throughout Asia, the Middle East and North Africa. To date, poultry surveillance in Vietnam has focused on detection of influenza H5 viruses, and there is limited understanding of influenza H9 epidemiology and transmission dynamics. We determined prevalence and diversity of influenza A viruses in chickens from live bird markets (LBM) of 7 northern Vietnamese provinces, using pooled oropharyngeal swabs collected from October to December 2014. Screening by real time RT-PCR revealed 1207/4900 (24.6%) of pooled swabs to be influenza A virus positive; overall prevalence estimates after accounting for pooling (5 swabs/pools) were 5.8% (CI 5.4-6.0). Subtyping was performed on 468 pooled swabs with M gene Ct<26. No influenza H7 was detected; 422 (90.1%) were H9 positive; and 22 (4.7%) were H5 positive. There was no evidence was of interaction between H9 and H5 virus detection rates. We sequenced 17 whole genomes of A/H9N2, 2 of A/H5N6, and 11 partial genomes. All H9N2 viruses had internal genes that clustered with genotype 57 and were closely related to Chinese human isolates of A/H7N9 and A/H10N8. Using a nucleotide divergence cutoff of 98%, we identified 9 distinct H9 genotypes. Phylogenetic analysis suggested multiple introductions of H9 viruses to northern Vietnam rather than in-situ transmission. Further investigations of H9 prevalence and diversity in other regions of Vietnam are warranted to assess H9 endemicity elsewhere in the country. Copyright © 2016 The Authors. Published by Elsevier B.V. All rights reserved.

Genetic diversity and substantial population differentiation in Crassostrea hongkongensis revealed by mitochondrial DNA.

PubMed

Li, Lu; Wu, Xiangyun; Yu, Ziniu

2013-09-01

The Hong Kong oyster, Crassostrea hongkongensis, is an important fisheries resource that is cultivated in the coastal waters of the South China Sea. Despite significant advances in understanding biological and taxonomic aspects of this species, no detailed study of its population genetic diversity in regions of extensive cultivation are available. Direct sequencing of the mtDNA cox1 gene region was used to investigate genetic variation within and between eleven C. hongkongensis populations collected from typical habitats. Sixty-two haplotypes were identified; only haplotype 2 (21.74% of total haplotypes) was shared among all the eleven populations, and most of the observed haplotypes were restricted to individual populations. Both AMOVA and FST analyses revealed significant population structure, and the isolation by distance (IBD) was confirmed. The highest local differentiation was observed between the sample pools from Guangxi versus Guangdong and Fujian, which are separated by a geographic barrier, the Leizhou Peninsula. Current knowledge from seed management suggests that seed transfer from Guangxi province has likely reduced the divergence that somewhat naturally exists between these pools. The findings from the present study could be useful for genetic management and may serve as a baseline by which to monitor future changes in genetic diversity, either due to natural or anthropogenic impacts. Copyright © 2013 Elsevier B.V. All rights reserved.

Species sorting during biofilm assembly by artificial substrates deployed in a cold seep system

PubMed Central

Zhang, Wei Peng; Wang, Yong; Tian, Ren Mao; Bougouffa, Salim; Yang, Bo; Cao, Hui Luo; Zhang, Gen; Wong, Yue Him; Xu, Wei; Batang, Zenon; Al-Suwailem, Abdulaziz; Zhang, Xi Xiang; Qian, Pei-Yuan

2014-01-01

Studies focusing on biofilm assembly in deep-sea environments are rarely conducted. To examine the effects of substrate type on microbial community assembly, biofilms were developed on different substrates for different durations at two locations in the Red Sea: in a brine pool and in nearby bottom water (NBW) adjacent to the Thuwal cold seep II. The composition of the microbial communities in 51 biofilms and water samples were revealed by classification of pyrosequenced 16S rRNA gene amplicons. Together with the microscopic characteristics of the biofilms, the results indicate a stronger selection effect by the substrates on the microbial assembly in the brine pool compared with the NBW. Moreover, the selection effect by substrate type was stronger in the early stages compared with the later stages of the biofilm development. These results are consistent with the hypotheses proposed in the framework of species sorting theory, which states that the power of species sorting during microbial community assembly is dictated by habitat conditions, duration and the structure of the source community. Therefore, the results of this study shed light on the control strategy underlying biofilm-associated marine fouling and provide supporting evidence for ecological theories important for understanding the formation of deep-sea biofilms. PMID:25323200

Historical biome distribution and recent human disturbance shape the diversity of arbuscular mycorrhizal fungi.

PubMed

Pärtel, Meelis; Öpik, Maarja; Moora, Mari; Tedersoo, Leho; Szava-Kovats, Robert; Rosendahl, Søren; Rillig, Matthias C; Lekberg, Ylva; Kreft, Holger; Helgason, Thorunn; Eriksson, Ove; Davison, John; de Bello, Francesco; Caruso, Tancredi; Zobel, Martin

2017-10-01

The availability of global microbial diversity data, collected using standardized metabarcoding techniques, makes microorganisms promising models for investigating the role of regional and local factors in driving biodiversity. Here we modelled the global diversity of symbiotic arbuscular mycorrhizal (AM) fungi using currently available data on AM fungal molecular diversity (small subunit (SSU) ribosomal RNA (rRNA) gene sequences) in field samples. To differentiate between regional and local effects, we estimated species pools (sets of potentially suitable taxa) for each site, which are expected to reflect regional processes. We then calculated community completeness, an index showing the fraction of the species pool present, which is expected to reflect local processes. We found significant spatial variation, globally in species pool size, as well as in local and dark diversity (absent members of the species pool). Species pool size was larger close to areas containing tropical grasslands during the last glacial maximum, which are possible centres of diversification. Community completeness was greater in regions of high wilderness (remoteness from human disturbance). Local diversity was correlated with wilderness and current connectivity to mountain grasslands. Applying the species pool concept to symbiotic fungi facilitated a better understanding of how biodiversity can be jointly shaped by large-scale historical processes and recent human disturbance. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

Survey of aquatic macroinvertebrates and amphibians at Wupatki National Monument, Arizona, USA: An evaluation of selected factors affecting species richness in ephemeral pools

USGS Publications Warehouse

Graham, T.B.

2002-01-01

Ephemeral aquatic habitats in Wupatki National Monument vary from naturally formed pools in arroyos over 5000 years old, to constructed catchment basins with ages estimated at 60-1000+ years old, and borrow pits and stock ponds 30-60 years old. The different ages of these pools provide different histories of colonization by amphibians and aquatic invertebrates, especially temporary pool specialists such as spadefoot toads and branchiopod crustaceans. Ten pools of five different origins and ages were surveyed in August and/or September 1997 for aquatic organisms; a total of 13 surveys were conducted. Twenty-two taxa were found, with the number of species in a pool during any survey ranging from one to 10. Species composition of the communities changed from one sampling date to the next within individual pools. Community structure is an amalgam of species with different dispersal mechanisms that are influenced by different pool characteristics. Age appears to have little effect overall, but may have influenced branchiopod presence/absence. Distance to permanent water, frequency of disturbance, and current pool size were correlated with presence/absence of some species.

Influence of the arc plasma parameters on the weld pool profile in TIG welding

NASA Astrophysics Data System (ADS)

Toropchin, A.; Frolov, V.; Pipa, A. V.; Kozakov, R.; Uhrlandt, D.

2014-11-01

Magneto-hydrodynamic simulations of the arc and fluid simulations of the weld pool can be beneficial in the analysis and further development of arc welding processes and welding machines. However, the appropriate coupling of arc and weld pool simulations needs further improvement. The tungsten inert gas (TIG) welding process is investigated by simulations including the weld pool. Experiments with optical diagnostics are used for the validation. A coupled computational model of the arc and the weld pool is developed using the software ANSYS CFX. The weld pool model considers the forces acting on the motion of the melt inside and on the surface of the pool, such as Marangoni, drag, electromagnetic forces and buoyancy. The experimental work includes analysis of cross-sections of the workpieces, highspeed video images and spectroscopic measurements. Experiments and calculations have been performed for various currents, distances between electrode and workpiece and nozzle diameters. The studies show the significant impact of material properties like surface tension dependence on temperature as well as of the arc structure on the weld pool behaviour and finally the weld seam depth. The experimental weld pool profiles and plasma temperatures are in good agreement with computational results.

Fine Mapping of a Clubroot Resistance Gene in Chinese Cabbage Using SNP Markers Identified from Bulked Segregant RNA Sequencing

PubMed Central

Huang, Zhen; Peng, Gary; Liu, Xunjia; Deora, Abhinandan; Falk, Kevin C.; Gossen, Bruce D.; McDonald, Mary R.; Yu, Fengqun

2017-01-01

Clubroot, caused by Plasmodiophora brassicae, is an important disease of canola (Brassica napus) in western Canada and worldwide. In this study, a clubroot resistance gene (Rcr2) was identified and fine mapped in Chinese cabbage cv. “Jazz” using single-nucleotide polymorphisms (SNP) markers identified from bulked segregant RNA sequencing (BSR-Seq) and molecular markers were developed for use in marker assisted selection. In total, 203.9 million raw reads were generated from one pooled resistant (R) and one pooled susceptible (S) sample, and >173,000 polymorphic SNP sites were identified between the R and S samples. One significant peak was observed between 22 and 26 Mb of chromosome A03, which had been predicted by BSR-Seq to contain the causal gene Rcr2. There were 490 polymorphic SNP sites identified in the region. A segregating population consisting of 675 plants was analyzed with 15 SNP sites in the region using the Kompetitive Allele Specific PCR method, and Rcr2 was fine mapped between two SNP markers, SNP_A03_32 and SNP_A03_67 with 0.1 and 0.3 cM from Rcr2, respectively. Five SNP markers co-segregated with Rcr2 in this region. Variants were identified in 14 of 36 genes annotated in the Rcr2 target region. The numbers of poly variants differed among the genes. Four genes encode TIR-NBS-LRR proteins and two of them Bra019410 and Bra019413, had high numbers of polymorphic variants and so are the most likely candidates of Rcr2. PMID:28894454

Experimental study on the stability and failure of individual step-pool

NASA Astrophysics Data System (ADS)

Zhang, Chendi; Xu, Mengzhen; Hassan, Marwan A.; Chartrand, Shawn M.; Wang, Zhaoyin

2018-06-01

Step-pools are one of the most common bedforms in mountain streams, the stability and failure of which play a significant role for riverbed stability and fluvial processes. Given this importance, flume experiments were performed with a manually constructed step-pool model. The experiments were carried out with a constant flow rate to study features of step-pool stability as well as failure mechanisms. The results demonstrate that motion of the keystone grain (KS) caused 90% of the total failure events. The pool reached its maximum depth and either exhibited relative stability for a period before step failure, which was called the stable phase, or the pool collapsed before its full development. The critical scour depth for the pool increased linearly with discharge until the trend was interrupted by step failure. Variability of the stable phase duration ranged by one order of magnitude, whereas variability of pool scour depth was constrained within 50%. Step adjustment was detected in almost all of the runs with step-pool failure and was one or two orders smaller than the diameter of the step stones. Two discharge regimes for step-pool failure were revealed: one regime captures threshold conditions and frames possible step-pool failure, whereas the second regime captures step-pool failure conditions and is the discharge of an exceptional event. In the transitional stage between the two discharge regimes, pool and step adjustment magnitude displayed relatively large variabilities, which resulted in feedbacks that extended the duration of step-pool stability. Step adjustment, which was a type of structural deformation, increased significantly before step failure. As a result, we consider step deformation as the direct explanation to step-pool failure rather than pool scour, which displayed relative stability during step deformations in our experiments.

Parallel tagged next-generation sequencing on pooled samples - a new approach for population genetics in ecology and conservation.

PubMed

Zavodna, Monika; Grueber, Catherine E; Gemmell, Neil J

2013-01-01

Next-generation sequencing (NGS) on pooled samples has already been broadly applied in human medical diagnostics and plant and animal breeding. However, thus far it has been only sparingly employed in ecology and conservation, where it may serve as a useful diagnostic tool for rapid assessment of species genetic diversity and structure at the population level. Here we undertake a comprehensive evaluation of the accuracy, practicality and limitations of parallel tagged amplicon NGS on pooled population samples for estimating species population diversity and structure. We obtained 16S and Cyt b data from 20 populations of Leiopelma hochstetteri, a frog species of conservation concern in New Zealand, using two approaches - parallel tagged NGS on pooled population samples and individual Sanger sequenced samples. Data from each approach were then used to estimate two standard population genetic parameters, nucleotide diversity (π) and population differentiation (FST), that enable population genetic inference in a species conservation context. We found a positive correlation between our two approaches for population genetic estimates, showing that the pooled population NGS approach is a reliable, rapid and appropriate method for population genetic inference in an ecological and conservation context. Our experimental design also allowed us to identify both the strengths and weaknesses of the pooled population NGS approach and outline some guidelines and suggestions that might be considered when planning future projects.

Messenger RNA profiling of human platelets by microarray hybridization.

PubMed

Bugert, Peter; Dugrillon, Alex; Günaydin, Ayse; Eichler, Hermann; Klüter, Harald

2003-10-01

Platelets are generally believed to be inactive in terms of de novo protein synthesis. On the other hand, the presence of ribosomes and mRNA molecules is well established. Many studies have used reverse transcriptase (RT) -PCR for detection of gene transcripts in platelets. As RT-PCR is a very sensitive method, any leukocyte contamination of platelet preparations can lead to false results. We performed three filtration procedures to minimize leukocyte contamination of pooled buffy-coat platelet concentrates prior to RNA isolation. Furthermore, by applying a genomic PCR approach with 50 amplification cycles we demonstrated that nucleated cells were not detectable. Microarray hybridization was used to analyze 9,850 individual human genes in RNA from purified platelets. In total we identified 1,526 (15.5%) positive genes. The data were confirmed in six individual experiments each performed on a PC pooled from four individual blood donations. Genes specific for nucleated blood cells such as CD4, CD83 and others were negative and verified the purity of PC. Overrepresentation of positive genes was found in the functional categories of glycoproteins/integrins (22.6% vs. 15.5%, p=0.029) and receptors (20.7% vs. 15.5%, p<0.001). Gene transcripts encoding RANTES, GRO-alpha, MIP-1alpha, MIP-1beta, and others were found at high levels of signal intensity and confirmed literature data. This work provides a mRNA profile of human platelets and a complete list of results can be downloaded from the website of our institute www.ma.uni-heidelberg.de/inst/iti/plt_array.xls. The knowledge about gene transcripts may have an impact on the characterization of novel proteins and their functions in platelets.

Systematic Integration of Brain eQTL and GWAS Identifies ZNF323 as a Novel Schizophrenia Risk Gene and Suggests Recent Positive Selection Based on Compensatory Advantage on Pulmonary Function

PubMed Central

Luo, Xiong-Jian; Mattheisen, Manuel; Li, Ming; Huang, Liang; Rietschel, Marcella; Børglum, Anders D.; Als, Thomas D.; van den Oord, Edwin J.; Aberg, Karolina A.; Mors, Ole; Mortensen, Preben Bo; Luo, Zhenwu; Degenhardt, Franziska; Cichon, Sven; Schulze, Thomas G.; Nöthen, Markus M.; Su, Bing; Zhao, Zhongming; Gan, Lin; Yao, Yong-Gang

2015-01-01

Genome-wide association studies have identified multiple risk variants and loci that show robust association with schizophrenia. Nevertheless, it remains unclear how these variants confer risk to schizophrenia. In addition, the driving force that maintains the schizophrenia risk variants in human gene pool is poorly understood. To investigate whether expression-associated genetic variants contribute to schizophrenia susceptibility, we systematically integrated brain expression quantitative trait loci and genome-wide association data of schizophrenia using Sherlock, a Bayesian statistical framework. Our analyses identified ZNF323 as a schizophrenia risk gene (P = 2.22×10–6). Subsequent analyses confirmed the association of the ZNF323 and its expression-associated single nucleotide polymorphism rs1150711 in independent samples (gene-expression: P = 1.40×10–6; single-marker meta-analysis in the combined discovery and replication sample comprising 44123 individuals: P = 6.85×10−10). We found that the ZNF323 was significantly downregulated in hippocampus and frontal cortex of schizophrenia patients (P = .0038 and P = .0233, respectively). Evidence for pleiotropic effects was detected (association of rs1150711 with lung function and gene expression of ZNF323 in lung: P = 6.62×10–5 and P = 9.00×10–5, respectively) with the risk allele (T allele) for schizophrenia acting as protective allele for lung function. Subsequent population genetics analyses suggest that the risk allele (T) of rs1150711 might have undergone recent positive selection in human population. Our findings suggest that the ZNF323 is a schizophrenia susceptibility gene whose expression may influence schizophrenia risk. Our study also illustrates a possible mechanism for maintaining schizophrenia risk variants in the human gene pool. PMID:25759474

POOL WATER TREATMENT AND COOLING SYSTEM DESCRIPTION DOCUMENT

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

V. King

2000-06-19

The Pool Water Treatment and Cooling System is located in the Waste Handling Building (WHB), and is comprised of various process subsystems designed to support waste handling operations. This system maintains the pool water temperature within an acceptable range, maintains water quality standards that support remote underwater operations and prevent corrosion, detects leakage from the pool liner, provides the capability to remove debris from the pool, controls the pool water level, and helps limit radiological exposure to personnel. The pool structure and liner, pool lighting, and the fuel staging racks in the pool are not within the scope of themore » Pool Water Treatment and Cooling System. Pool water temperature control is accomplished by circulating the pool water through heat exchangers. Adequate circulation and mixing of the pool water is provided to prevent localized thermal hotspots in the pool. Treatment of the pool water is accomplished by a water treatment system that circulates the pool water through filters, and ion exchange units. These water treatment units remove radioactive and non-radioactive particulate and dissolved solids from the water, thereby providing the water clarity needed to conduct waste handling operations. The system also controls pool water chemistry to prevent advanced corrosion of the pool liner, pool components, and fuel assemblies. Removal of radioactivity from the pool water contributes to the project ALARA (as low as is reasonably achievable) goals. A leak detection system is provided to detect and alarm leaks through the pool liner. The pool level control system monitors the water level to ensure that the minimum water level required for adequate radiological shielding is maintained. Through interface with a demineralized water system, adequate makeup is provided to compensate for loss of water inventory through evaporation and waste handling operations. Interface with the Site Radiological Monitoring System provides continuous radiological monitoring of the pool water. The Pool Water Treatment and Cooling System interfaces with the Waste Handling Building System, Site-Generated Radiological Waste Handling System, Site Radiological Monitoring System, Waste Handling Building Electrical System, Site Water System, and the Monitored Geologic Repository Operations Monitoring and Control System.« less

Combinatorial Pooling Enables Selective Sequencing of the Barley Gene Space

PubMed Central

Lonardi, Stefano; Duma, Denisa; Alpert, Matthew; Cordero, Francesca; Beccuti, Marco; Bhat, Prasanna R.; Wu, Yonghui; Ciardo, Gianfranco; Alsaihati, Burair; Ma, Yaqin; Wanamaker, Steve; Resnik, Josh; Bozdag, Serdar; Luo, Ming-Cheng; Close, Timothy J.

2013-01-01

For the vast majority of species – including many economically or ecologically important organisms, progress in biological research is hampered due to the lack of a reference genome sequence. Despite recent advances in sequencing technologies, several factors still limit the availability of such a critical resource. At the same time, many research groups and international consortia have already produced BAC libraries and physical maps and now are in a position to proceed with the development of whole-genome sequences organized around a physical map anchored to a genetic map. We propose a BAC-by-BAC sequencing protocol that combines combinatorial pooling design and second-generation sequencing technology to efficiently approach denovo selective genome sequencing. We show that combinatorial pooling is a cost-effective and practical alternative to exhaustive DNA barcoding when preparing sequencing libraries for hundreds or thousands of DNA samples, such as in this case gene-bearing minimum-tiling-path BAC clones. The novelty of the protocol hinges on the computational ability to efficiently compare hundred millions of short reads and assign them to the correct BAC clones (deconvolution) so that the assembly can be carried out clone-by-clone. Experimental results on simulated data for the rice genome show that the deconvolution is very accurate, and the resulting BAC assemblies have high quality. Results on real data for a gene-rich subset of the barley genome confirm that the deconvolution is accurate and the BAC assemblies have good quality. While our method cannot provide the level of completeness that one would achieve with a comprehensive whole-genome sequencing project, we show that it is quite successful in reconstructing the gene sequences within BACs. In the case of plants such as barley, this level of sequence knowledge is sufficient to support critical end-point objectives such as map-based cloning and marker-assisted breeding. PMID:23592960

Combinatorial pooling enables selective sequencing of the barley gene space.

PubMed

Lonardi, Stefano; Duma, Denisa; Alpert, Matthew; Cordero, Francesca; Beccuti, Marco; Bhat, Prasanna R; Wu, Yonghui; Ciardo, Gianfranco; Alsaihati, Burair; Ma, Yaqin; Wanamaker, Steve; Resnik, Josh; Bozdag, Serdar; Luo, Ming-Cheng; Close, Timothy J

2013-04-01

For the vast majority of species - including many economically or ecologically important organisms, progress in biological research is hampered due to the lack of a reference genome sequence. Despite recent advances in sequencing technologies, several factors still limit the availability of such a critical resource. At the same time, many research groups and international consortia have already produced BAC libraries and physical maps and now are in a position to proceed with the development of whole-genome sequences organized around a physical map anchored to a genetic map. We propose a BAC-by-BAC sequencing protocol that combines combinatorial pooling design and second-generation sequencing technology to efficiently approach denovo selective genome sequencing. We show that combinatorial pooling is a cost-effective and practical alternative to exhaustive DNA barcoding when preparing sequencing libraries for hundreds or thousands of DNA samples, such as in this case gene-bearing minimum-tiling-path BAC clones. The novelty of the protocol hinges on the computational ability to efficiently compare hundred millions of short reads and assign them to the correct BAC clones (deconvolution) so that the assembly can be carried out clone-by-clone. Experimental results on simulated data for the rice genome show that the deconvolution is very accurate, and the resulting BAC assemblies have high quality. Results on real data for a gene-rich subset of the barley genome confirm that the deconvolution is accurate and the BAC assemblies have good quality. While our method cannot provide the level of completeness that one would achieve with a comprehensive whole-genome sequencing project, we show that it is quite successful in reconstructing the gene sequences within BACs. In the case of plants such as barley, this level of sequence knowledge is sufficient to support critical end-point objectives such as map-based cloning and marker-assisted breeding.

Mucopolysaccharidosis VI in cats - clarification regarding genetic testing.

PubMed

Lyons, Leslie A; Grahn, Robert A; Genova, Francesca; Beccaglia, Michela; Hopwood, John J; Longeri, Maria

2016-07-02

The release of new DNA-based diagnostic tools has increased tremendously in companion animals. Over 70 different DNA variants are now known for the cat, including DNA variants in disease-associated genes and genes causing aesthetically interesting traits. The impact genetic tests have on animal breeding and health management is significant because of the ability to control the breeding of domestic cats, especially breed cats. If used properly, genetic testing can prevent the production of diseased animals, causing the reduction of the frequency of the causal variant in the population, and, potentially, the eventual eradication of the disease. However, testing of some identified DNA variants may be unwarranted and cause undo strife within the cat breeding community and unnecessary reduction of gene pools and availability of breeding animals. Testing for mucopolysaccharidosis Type VI (MPS VI) in cats, specifically the genetic testing of the L476P (c.1427T>C) and the D520N (c.1558G>A) variants in arylsulfatase B (ARSB), has come under scrutiny. No health problems are associated with the D520N (c.1558G>A) variant, however, breeders that obtain positive results for this variant are speculating as to possible correlation with health concerns. Birman cats already have a markedly reduced gene pool and have a high frequency of the MPS VI D520N variant. Further reduction of the gene pool by eliminating cats that are heterozygous or homozygous for only the MPS VI D520N variant could lead to more inbreeding depression effects on the breed population. Herein is debated the genetic testing of the MPS VI D520N variant in cats. Surveys from different laboratories suggest the L476P (c.1427T>C) disease-associated variant should be monitored in the cat breed populations, particularly breeds with Siamese derivations and outcrosses. However, the D520N has no evidence of association with disease in cats and testing is not recommended in the absence of L476P genotyping. Selection against the D520N is not warranted in cat populations. More rigorous guidelines may be required to support the genetic testing of DNA variants in all animal species.

Technique for quantitative RT-PCR analysis directly from single muscle fibers.

PubMed

Wacker, Michael J; Tehel, Michelle M; Gallagher, Philip M

2008-07-01

The use of single-cell quantitative RT-PCR has greatly aided the study of gene expression in fields such as muscle physiology. For this study, we hypothesized that single muscle fibers from a biopsy can be placed directly into the reverse transcription buffer and that gene expression data can be obtained without having to first extract the RNA. To test this hypothesis, biopsies were taken from the vastus lateralis of five male subjects. Single muscle fibers were isolated and underwent RNA isolation (technique 1) or placed directly into reverse transcription buffer (technique 2). After cDNA conversion, individual fiber cDNA was pooled and quantitative PCR was performed using primer-probes for beta(2)-microglobulin, glyceraldehyde-3-phosphate dehydrogenase, insulin-like growth factor I receptor, and glucose transporter subtype 4. The no RNA extraction method provided similar quantitative PCR data as that of the RNA extraction method. A third technique was also tested in which we used one-quarter of an individual fiber's cDNA for PCR (not pooled) and the average coefficient of variation between fibers was <8% (cycle threshold value) for all genes studied. The no RNA extraction technique was tested on isolated muscle fibers using a gene known to increase after exercise (pyruvate dehydrogenase kinase 4). We observed a 13.9-fold change in expression after resistance exercise, which is consistent with what has been previously observed. These results demonstrate a successful method for gene expression analysis directly from single muscle fibers.

Genetic Linkage of Soil Carbon Pools and Microbial Functions in Subtropical Freshwater Wetlands in Response to Experimental Warming

PubMed Central

Wang, Hang; He, Zhili; Lu, Zhenmei; Zhou, Jizhong; Van Nostrand, Joy D.; Xu, Xinhua

2012-01-01

Rising climate temperatures in the future are predicted to accelerate the microbial decomposition of soil organic matter. A field microcosm experiment was carried out to examine the impact of soil warming in freshwater wetlands on different organic carbon (C) pools and associated microbial functional responses. GeoChip 4.0, a functional gene microarray, was used to determine microbial gene diversity and functional potential for C degradation. Experimental warming significantly increased soil pore water dissolved organic C and phosphorus (P) concentrations, leading to a higher potential for C emission and P export. Such losses of total organic C stored in soil could be traced back to the decomposition of recalcitrant organic C. Warming preferentially stimulated genes for degrading recalcitrant C over labile C. This was especially true for genes encoding cellobiase and mnp for cellulose and lignin degradation, respectively. We confirmed this with warming-enhanced polyphenol oxidase and peroxidase activities for recalcitrant C acquisition and greater increases in recalcitrant C use efficiency than in labile C use efficiency (average percentage increases of 48% versus 28%, respectively). The relative abundance of lignin-degrading genes increased by 15% under warming; meanwhile, soil fungi, as the primary decomposers of lignin, were greater in abundance by 27%. This work suggests that future warming may enhance the potential for accelerated fungal decomposition of lignin-like compounds, leading to greater microbially mediated C losses than previously estimated in freshwater wetlands. PMID:22923398

Detection of a Pool in Semi-Continuous Castings Made of Heat-Treatable Aluminum Alloys

NASA Astrophysics Data System (ADS)

Krushenko, G. G.; Nazarov, V. P.

2017-12-01

Various products (sheets, sections, etc.) manufactured by metal forming (rolled products, forged pieces, etc.) from semi-continuous castings are widely used in the aerospace industry. The so-called pool, which is the conical volume of a liquid metal, exists at the top of the liquid metal. Experience demonstrates that the geometry, the depth, and the shape of the pool substantially affect the structure formation in a casting and its quality. The application of a titanium nitride nanopowder, which is introduced in a melt in the volume of a rod, as a modifier allowed us to find the exact geometry of the pool.

Assessing pooled BAC and whole genome shotgun strategies for assembly of complex genomes

PubMed Central

2011-01-01

Background We investigate if pooling BAC clones and sequencing the pools can provide for more accurate assembly of genome sequences than the "whole genome shotgun" (WGS) approach. Furthermore, we quantify this accuracy increase. We compare the pooled BAC and WGS approaches using in silico simulations. Standard measures of assembly quality focus on assembly size and fragmentation, which are desirable for large whole genome assemblies. We propose additional measures enabling easy and visual comparison of assembly quality, such as rearrangements and redundant sequence content, relative to the known target sequence. Results The best assembly quality scores were obtained using 454 coverage of 15× linear and 5× paired (3kb insert size) reads (15L-5P) on Arabidopsis. This regime gave similarly good results on four additional plant genomes of very different GC and repeat contents. BAC pooling improved assembly scores over WGS assembly, coverage and redundancy scores improving the most. Conclusions BAC pooling works better than WGS, however, both require a physical map to order the scaffolds. Pool sizes up to 12Mbp work well, suggesting this pooling density to be effective in medium-scale re-sequencing applications such as targeted sequencing of QTL intervals for candidate gene discovery. Assuming the current Roche/454 Titanium sequencing limitations, a 12 Mbp region could be re-sequenced with a full plate of linear reads and a half plate of paired-end reads, yielding 15L-5P coverage after read pre-processing. Our simulation suggests that massively over-sequencing may not improve accuracy. Our scoring measures can be used generally to evaluate and compare results of simulated genome assemblies. PMID:21496274

Non-stationary Drainage Flows and Cold Pools in Gentle Terrain

NASA Astrophysics Data System (ADS)

Mahrt, L.

2015-12-01

Previous studies have concentrated on organized topography with well-defined slopes or valleys in an effort to understand the flow dynamics. However, most of the Earth's land surface consists of gentle terrain that is quasi three dimensional. Different scenarios are briefly classified. A network of measurements are analyzed to examine shallow cold pools and drainage flow down the valley which develop for weak ambient wind and relatively clear skies. However, transient modes constantly modulate or intermittently eliminate the cold pool, which makes extraction and analysis of the horizontal structure of the cold pool difficult with traditional analysis methods. Singular value decomposition successfully isolates the effects of large-scale flow from local down-valley cold air drainage within the cold pool in spite of the intermittent nature of this local flow. The traditional concept of a cold pool must be generalized to include cold pool intermittency, complex variation of temperature related to some three-dimensionality and a diffuse cold pool top. Different types of cold pools are classified in terms of the stratification and gradient of potential temperature along the slope. The strength of the cold pool is related to a forcing temperature scale proportional to the net radiative cooling divided by the wind speed above the valley. The scatter is large partly due to nonstationarity of the marginal cold pool in this shallow valley

Methodological assessment of 2b-RAD genotyping technique for population structure inferences in yellowfin tuna (Thunnus albacares).

PubMed

Pecoraro, Carlo; Babbucci, Massimiliano; Villamor, Adriana; Franch, Rafaella; Papetti, Chiara; Leroy, Bruno; Ortega-Garcia, Sofia; Muir, Jeff; Rooker, Jay; Arocha, Freddy; Murua, Hilario; Zudaire, Iker; Chassot, Emmanuel; Bodin, Nathalie; Tinti, Fausto; Bargelloni, Luca; Cariani, Alessia

2016-02-01

Global population genetic structure of yellowfin tuna (Thunnus albacares) is still poorly understood despite its relevance for the tuna fishery industry. Low levels of genetic differentiation among oceans speak in favour of the existence of a single panmictic population worldwide of this highly migratory fish. However, recent studies indicated genetic structuring at a much smaller geographic scales than previously considered, pointing out that YFT population genetic structure has not been properly assessed so far. In this study, we demonstrated for the first time, the utility of 2b-RAD genotyping technique for investigating population genetic diversity and differentiation in high gene-flow species. Running de novo pipeline in Stacks, a total of 6772 high-quality genome-wide SNPs were identified across Atlantic, Indian and Pacific population samples representing all major distribution areas. Preliminary analyses showed shallow but significant population structure among oceans (FST=0.0273; P-value<0.01). Discriminant Analysis of Principal Components endorsed the presence of genetically discrete yellowfin tuna populations among three oceanic pools. Although such evidence needs to be corroborated by increasing sample size, these results showed the efficiency of this genotyping technique in assessing genetic divergence in a marine fish with high dispersal potential. Copyright © 2015 Elsevier B.V. All rights reserved.

Pool spacing, channel morphology, and the restoration of tidal forested wetlands of the Columbia River, U.S.A.

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

Diefenderfer, Heida L.; Montgomery, David R.

2008-10-09

Tidal forested wetlands have sustained substantial areal losses, and restoration practitioners lack a description of many ecosystem structures associated with these late-successional systems in which surface water is a significant controlling factor on the flora and fauna. The roles of large woody debris in terrestrial and riverine ecosystems have been well described compared to functions in tidal areas. This study documents the role of large wood in forcing channel morphology in Picea-sitchensis (Sitka spruce) dominated freshwater tidal wetlands in the floodplain of the Columbia River, U.S.A. near the Pacific coast. The average pool spacing documented in channel surveys of threemore » freshwater tidal forested wetlands near Grays Bay were 2.2 ± 1.3, 2.3 ± 1.2, and 2.5 ± 1.5. There were significantly greater numbers of pools on tidal forested wetland channels than on a nearby restoration site. On the basis of pool spacing and the observed sequences of log jams and pools, the tidal forested wetland channels were classified consistent with a forced step-pool class. Tidal systems, with bidirectional flow, have not previously been classified in this way. The classification provides a useful basis for restoration project design and planning in historically forested tidal freshwater areas, particularly in regard to the use of large wood in restoration actions and the development of pool habitats for aquatic species. Significant modifications by beaver on these sites warrant further investigation to explore the interactions between these animals and restoration actions affecting hydraulics and channel structure in tidal areas.« less

Quality Improvement: Creating a Float Pool Specialty Within a New Graduate Residency.

PubMed

Shinners, Jean; Alejandro, John Aldrich N; Frigillana, Vanessa; Desmond, Juliann; LaVigne, Ronda

2016-01-01

Creating new norms is essential for success as acute care leaders seek to redesign care delivery. Through the structures of the registered nurse (RN) residency and utilizing a quality improvement process, new graduate RNs demonstrated success in creating a centralized float pool resource.

16 CFR 1207.5 - Design.

Code of Federal Regulations, 2014 CFR

2014-01-01

... pool slide shall be such that no structural failures of any component part shall cause failures of any... such fasteners shall not cause a failure of the tread under the ladder loading conditions specified in... without failure or permanent deformation. (d) Handrails. Swimming pool slide ladders shall be equipped...

16 CFR 1207.5 - Design.

Code of Federal Regulations, 2012 CFR

2012-01-01

... pool slide shall be such that no structural failures of any component part shall cause failures of any... such fasteners shall not cause a failure of the tread under the ladder loading conditions specified in... without failure or permanent deformation. (d) Handrails. Swimming pool slide ladders shall be equipped...

Clinicopathological Characteristics of Patients with Non-Small-Cell Lung Cancer Who Harbor EML4-ALK Fusion Gene: A Meta-Analysis

PubMed Central

Zhao, Fengzhi; Xu, Meng; Lei, Honcho; Zhou, Ziqi; Wang, Liang; Li, Ping; Zhao, Jianfu; Hu, Penghui

2015-01-01

Background A novel fusion gene of echinoderm microtubule-associated protein-like 4 (EML4) and anaplastic lymphoma kinase (ALK) has been recently identified in non-small-cell lung cancers (NSCLCs). Patients with the EML4-ALK fusion gene demonstrate unique clinicopathological and physiological characteristics. Here we present a meta-analysis of large-scale studies to evaluate the clinicopathological characteristics of NSCLC patients harboring the EML4-ALK fusion gene. Methods Both English and Chinese databases were systematically used to search the materials of the clinicopathological characteristics of patients with NSCLC harboring the EML4-ALK fusion gene. Pooled relative risk (RR) estimates and the 95% confidence intervals (95% CI) were calculated with the fixed or random effect model. Publication bias and chi-square test were also calculated. Results 27 retrospective studies were included in our meta-analysis. These studies included a total of 6950 patients. The incidence rate of EML4-ALK fusion in NSCLC patients was found to be 6.8% (472/6950). The correlation of the EML4-ALK fusion gene and clinicopathological characteristics of NSCLC patients demonstrated a significant difference in smoking status, histological types, stage, and ethnic characteristics. The positive rate of the EML4-ALK fusion gene expression in females were slightly higher than that in males, but not significantly (P = 0.52). In addition, the EML4-ALK fusion gene was mutually exclusive of the EGFR and KRAS mutation genes (P = 0.00). Conclusion Our pooled analysis revealed that the EML4-ALK fusion gene was observed predominantly in adenocarcinoma, non-smoking and NSCLC patients, especially those diagnosed in the advanced clinical stage of NSCLC. Additionally, the EML4-ALK fusion gene was exclusive of the EGFR and KRAS mutation genes. We surmise that IHC assay is a valuable tool for the prescreening of patients with ALK fusion gene in clinical practice, and FISH assay can be performed as a confirmation method. These insights might be helpful in guiding the appropriate molecular target therapy for NSCLC. PMID:25706305

Clinicopathological characteristics of patients with non-small-cell lung cancer who harbor EML4-ALK fusion gene: a meta-analysis.

PubMed

Zhao, Fengzhi; Xu, Meng; Lei, Honcho; Zhou, Ziqi; Wang, Liang; Li, Ping; Zhao, Jianfu; Hu, Penghui

2015-01-01

A novel fusion gene of echinoderm microtubule-associated protein-like 4 (EML4) and anaplastic lymphoma kinase (ALK) has been recently identified in non-small-cell lung cancers (NSCLCs). Patients with the EML4-ALK fusion gene demonstrate unique clinicopathological and physiological characteristics. Here we present a meta-analysis of large-scale studies to evaluate the clinicopathological characteristics of NSCLC patients harboring the EML4-ALK fusion gene. Both English and Chinese databases were systematically used to search the materials of the clinicopathological characteristics of patients with NSCLC harboring the EML4-ALK fusion gene. Pooled relative risk (RR) estimates and the 95% confidence intervals (95% CI) were calculated with the fixed or random effect model. Publication bias and chi-square test were also calculated. 27 retrospective studies were included in our meta-analysis. These studies included a total of 6950 patients. The incidence rate of EML4-ALK fusion in NSCLC patients was found to be 6.8% (472/6950). The correlation of the EML4-ALK fusion gene and clinicopathological characteristics of NSCLC patients demonstrated a significant difference in smoking status, histological types, stage, and ethnic characteristics. The positive rate of the EML4-ALK fusion gene expression in females were slightly higher than that in males, but not significantly (P = 0.52). In addition, the EML4-ALK fusion gene was mutually exclusive of the EGFR and KRAS mutation genes (P = 0.00). Our pooled analysis revealed that the EML4-ALK fusion gene was observed predominantly in adenocarcinoma, non-smoking and NSCLC patients, especially those diagnosed in the advanced clinical stage of NSCLC. Additionally, the EML4-ALK fusion gene was exclusive of the EGFR and KRAS mutation genes. We surmise that IHC assay is a valuable tool for the prescreening of patients with ALK fusion gene in clinical practice, and FISH assay can be performed as a confirmation method. These insights might be helpful in guiding the appropriate molecular target therapy for NSCLC.

In-vivo-induced antigenic determinants of Fusobacterium nucleatum subsp. nucleatum.

PubMed

Lee, H-R; Rhyu, I-C; Kim, H-D; Jun, H-K; Min, B-M; Lee, S-H; Choi, B-K

2011-04-01

Fusobacterium nucleatum plays a pivotal role in dental plaque biofilm formation and is known to be involved in chronic inflammatory systemic disease. However, limited knowledge of F. nucleatum genes expressed in vivo interferes with our understanding of pathogenesis. In this study, we identified F. nucleatum genes induced in vivo using in-vivo-induced antigen technology (IVIAT). Among 30,000 recombinant clones screened, 87 reacted reproducibly with pooled sera from 10 patients with periodontitis. The clones encoded for 32 different proteins, of which 28 could be assigned to their functions, which were categorized in translation, transcription, transport, energy metabolism, cell envelope, cellular process, fatty acid and phospholipid metabolism, transposition, cofactor biosynthesis, amino acid biosynthesis, and DNA replication. Putative virulence factors detected were ABC transporter, butyrate-acetoacetate CoA-transferase, hemin receptor, hemolysin, hemolysin-related protein, LysR family transcriptional regulator, serine protease, and transposase. Analysis of immune responses to the in-vivo-induced (ivi) antigens in five patients demonstrated that most were reactive to these proteins, confirming results with pooled sera. IVIAT-identified F. nucleatum genes in this study may accelerate the elucidation of F. nucleatum-mediated molecular pathogenesis. © 2011 John Wiley & Sons A/S.

Identification of Genes from the Fungal Pathogen Cryptococcus neoformans Related to Transmigration into the Central Nervous System

PubMed Central

Tseng, Hsiang-Kuang; Liu, Chang-Pan; Price, Michael S.; Jong, Ambrose Y.; Chang, Jui-Chih; Toffaletti, Dena L.; Betancourt-Quiroz, Marisol; Frazzitta, Aubrey E.; Cho, Wen-Long; Perfect, John R.

2012-01-01

Background A mouse brain transmigration assessment (MBTA) was created to investigate the central nervous system (CNS) pathogenesis of cryptococcal meningoencephalitis. Methodology/Principal Findings Two cryptococcal mutants were identified from a pool of 109 pre-selected mutants that were signature-tagged with the nourseothricin acetyltransferase (NAT) resistance cassette. These two mutants displayed abnormal transmigration into the central nervous system. One mutant displaying decreased transmigration contains a null mutation in the putative FNX1 gene, whereas the other mutant possessing a null mutation in the putative RUB1 gene exhibited increased transmigration into the brain. Two macrophage adhesion-defective mutants in the pool, 12F1 and 3C9, showed reduced phagocytosis by macrophages, but displayed no defects in CNS entry suggesting that transit within macrophages (the “Trojan horse” model of CNS entry) is not the primary mechanism for C. neoformans migration into the CNS in this MBTA. Conclusions/Significance This research design provides a new strategy for genetic impact studies on how Cryptococcus passes through the blood-brain barrier (BBB), and the specific isolated mutants in this assay support a transcellular mechanism of CNS entry. PMID:23028773

Differential gene regulation of GHSR signaling pathway in the arcuate nucleus and NPY neurons by fasting, diet-induced obesity, and 17β-estradiol.

PubMed

Yasrebi, Ali; Hsieh, Anna; Mamounis, Kyle J; Krumm, Elizabeth A; Yang, Jennifer A; Magby, Jason; Hu, Pu; Roepke, Troy A

2016-02-15

Ghrelin's receptor, growth hormone secretagogue receptor (GHSR), is highly expressed in the arcuate nucleus (ARC) and in neuropeptide Y (NPY) neurons. Fasting, diet-induced obesity (DIO), and 17β-estradiol (E2) influence ARC Ghsr expression. It is unknown if these effects occur in NPY neurons. Therefore, we examined the expression of Npy, Agrp, and GHSR signaling pathway genes after fasting, DIO, and E2 replacement in ARC and pools of NPY neurons. In males, fasting increased ARC Ghsr and NPY Foxo1 but decreased NPY Ucp2. In males, DIO decreased ARC and NPY Ghsr and Cpt1c. In fed females, E2 increased Agrp, Ghsr, Cpt1c, and Foxo1 in ARC. In NPY pools, E2 decreased Foxo1 in fed females but increased Foxo1 in fasted females. DIO in females suppressed Agrp and augmented Cpt1c in NPY neurons. In summary, genes involved in GHSR signaling are differentially regulated between the ARC and NPY neurons in a sex-dependent manner. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

Pool boiling with high heat flux enabled by a porous artery structure

NASA Astrophysics Data System (ADS)

Bai, Lizhan; Zhang, Lianpei; Lin, Guiping; Peterson, G. P.

2016-06-01

A porous artery structure utilizing the concept of "phase separation and modulation" is proposed to enhance the critical heat flux of pool boiling. A series of experiments were conducted on a range of test articles in which multiple rectangular arteries were machined directly into the top surface of a 10.0 mm diameter copper rod. The arteries were then covered by a 2.0 mm thickness microporous copper plate through silver brazing. The pool wall was fabricated from transparent Pyrex glass to allow a visualization study, and water was used as the working fluid. Experimental results confirmed that the porous artery structure provided individual flow paths for the liquid supply and vapor venting, and avoided the detrimental effects of the liquid/vapor counter flow. As a result, a maximum heat flux of 610 W/cm2 over a heating area of 0.78 cm2 was achieved with no indication of dryout, prior to reaching the heater design temperature limit. Following the experimental tests, the mechanisms responsible for the boiling critical heat flux and performance enhancement of the porous artery structure were analyzed.

Effects of common seagrass restoration methods on ecosystem structure in subtropical seagrass meadows.

PubMed

Bourque, Amanda S; Fourqurean, James W

2014-06-01

Seagrass meadows near population centers are subject to frequent disturbance from vessel groundings. Common seagrass restoration methods include filling excavations and applying fertilizer to encourage seagrass recruitment. We sampled macrophytes, soil structure, and macroinvertebrate infauna at unrestored and recently restored vessel grounding disturbances to evaluate the effects of these restoration methods on seagrass ecosystem structure. After a year of observations comparing filled sites to both undisturbed reference and unrestored disturbed sites, filled sites had low organic matter content, nutrient pools, and primary producer abundance. Adding a nutrient source increased porewater nutrient pools at disturbed sites and in undisturbed meadows, but not at filled sites. Environmental predictors of infaunal community structure across treatments included soil texture and nutrient pools. At the one year time scale, the restoration methods studied did not result in convergence between restored and unrestored sites. Particularly in filled sites, soil conditions may combine to constrain rapid development of the seagrass community and associated infauna. Our study is important for understanding early recovery trajectories following restoration using these methods. Published by Elsevier Ltd.

Aspen phenylpropanoid genes' expression levels correlate with genets' tannin richness and vary both in responses to soil nitrogen and associations with phenolic profiles.

PubMed

Decker, Vicki H G; Bandau, Franziska; Gundale, Michael J; Cole, Christopher T; Albrectsen, Benedicte R

2017-02-01

Condensed tannin (CT) contents of European aspen (Populus tremula L.) vary among genotypes, and increases in nitrogen (N) availability generally reduce plants' tannin production in favor of growth, through poorly understood mechanisms. We hypothesized that intrinsic tannin production rates may co-vary with gene expression responses to soil N and resource allocation within the phenylpropanoid pathway (PPP). Thus, we examined correlations between soil N levels and both expression patterns of eight PPP genes (measured by quantitative-reverse transcription PCR) and foliar phenolic compounds (measured by liquid chromatography-mass spectrometry) in young aspen genets with intrinsically extreme CT levels. Monitored phenolics included salicinoids, lignins, flavones, flavonols, CT precursors and CTs. The PPP genes were consistently expressed more strongly in high-CT trees. Low N supplements reduced expression of genes throughout the PPP in all genets, while high N doses restored expression of genes at the beginning and end of the pathway. These PPP changes were not reflected in pools of tannin precursors, but varying correlations between gene expression and foliar phenolic pools were detected in young and mature leaves, suggesting that processes linking gene expression and the resulting phenolics vary spatially and temporally. Precursor fluxes suggested that CT-related metabolic rate or sink controls are linked to intrinsic carbon allocation strategies associated with N responses. Overall, we found more negative correlations (indicative of allocation trade-offs) between PPP gene expression and phenolic products following N additions in low-CT plants than in high-CT plants. The tannin-related expression dynamics suggest that, in addition to defense, relative tannin levels may also be indicative of intraspecific variations in the way aspen genets respond to soil fertility. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

Microdissection of Shoot Meristem Functional Domains

USDA-ARS?s Scientific Manuscript database

The shoot apical meristem (SAM) maintains a pool of indeterminate cells within the SAM proper, while lateral organs are initiated from the SAM periphery. Laser microdissection–microarray technology was used to compare transcriptional profiles within these SAM domains to identify novel maize genes th...

Genetic diversity in Gossypium genus

USDA-ARS?s Scientific Manuscript database

The overall objectives of this paper are to report on cotton germplasm resources, morphobiological and agronomic diversity of Gossypium genus and review efforts on molecular genetic diversity of cotton gene pools as well as on the challenges and perspectives of exploiting genetic diversity in cotton...

Structural and Functional Characterization of Ribosomal Protein Gene Introns in Sponges

PubMed Central

Perina, Drago; Korolija, Marina; Mikoč, Andreja; Roller, Maša; Pleše, Bruna; Imešek, Mirna; Morrow, Christine; Batel, Renato; Ćetković, Helena

2012-01-01

Ribosomal protein genes (RPGs) are a powerful tool for studying intron evolution. They exist in all three domains of life and are much conserved. Accumulating genomic data suggest that RPG introns in many organisms abound with non-protein-coding-RNAs (ncRNAs). These ancient ncRNAs are small nucleolar RNAs (snoRNAs) essential for ribosome assembly. They are also mobile genetic elements and therefore probably important in diversification and enrichment of transcriptomes through various mechanisms such as intron/exon gain/loss. snoRNAs in basal metazoans are poorly characterized. We examined 449 RPG introns, in total, from four demosponges: Amphimedon queenslandica, Suberites domuncula, Suberites ficus and Suberites pagurorum and showed that RPG introns from A. queenslandica share position conservancy and some structural similarity with “higher” metazoans. Moreover, our study indicates that mobile element insertions play an important role in the evolution of their size. In four sponges 51 snoRNAs were identified. The analysis showed discrepancies between the snoRNA pools of orthologous RPG introns between S. domuncula and A. queenslandica. Furthermore, these two sponges show as much conservancy of RPG intron positions between each other as between themselves and human. Sponges from the Suberites genus show consistency in RPG intron position conservation. However, significant differences in some of the orthologous RPG introns of closely related sponges were observed. This indicates that RPG introns are dynamic even on these shorter evolutionary time scales. PMID:22880015

Structural and functional characterization of ribosomal protein gene introns in sponges.

PubMed

Perina, Drago; Korolija, Marina; Mikoč, Andreja; Roller, Maša; Pleše, Bruna; Imešek, Mirna; Morrow, Christine; Batel, Renato; Ćetković, Helena

2012-01-01

Ribosomal protein genes (RPGs) are a powerful tool for studying intron evolution. They exist in all three domains of life and are much conserved. Accumulating genomic data suggest that RPG introns in many organisms abound with non-protein-coding-RNAs (ncRNAs). These ancient ncRNAs are small nucleolar RNAs (snoRNAs) essential for ribosome assembly. They are also mobile genetic elements and therefore probably important in diversification and enrichment of transcriptomes through various mechanisms such as intron/exon gain/loss. snoRNAs in basal metazoans are poorly characterized. We examined 449 RPG introns, in total, from four demosponges: Amphimedon queenslandica, Suberites domuncula, Suberites ficus and Suberites pagurorum and showed that RPG introns from A. queenslandica share position conservancy and some structural similarity with "higher" metazoans. Moreover, our study indicates that mobile element insertions play an important role in the evolution of their size. In four sponges 51 snoRNAs were identified. The analysis showed discrepancies between the snoRNA pools of orthologous RPG introns between S. domuncula and A. queenslandica. Furthermore, these two sponges show as much conservancy of RPG intron positions between each other as between themselves and human. Sponges from the Suberites genus show consistency in RPG intron position conservation. However, significant differences in some of the orthologous RPG introns of closely related sponges were observed. This indicates that RPG introns are dynamic even on these shorter evolutionary time scales.

Identification of a t(3;4)(p1.3;q1.5) translocation breakpoint in pigs using somatic cell hybrid mapping and high-resolution mate-pair sequencing

PubMed Central

Fève, Katia; Foissac, Sylvain; Pinton, Alain; Mompart, Florence; Esquerré, Diane; Faraut, Thomas; Yerle, Martine

2017-01-01

Reciprocal translocations are the most frequently occurring constitutional structural rearrangements in mammalian genomes. In phenotypically normal pigs, an incidence of 1/200 is estimated for such rearrangements. Even if constitutional translocations do not necessarily induce defects and diseases, they are responsible for significant economic losses in domestic animals due to reproduction failures. Over the last 30 years, advances in molecular and cytogenetic technologies have led to major improvements in the resolution of the characterization of translocation events. Characterization of translocation breakpoints helps to decipher the mechanisms that lead to such rearrangements and the functions of the genes that are involved in the translocation. Here, we describe the fine characterization of a reciprocal translocation t(3;4) (p1.3;q1.5) detected in a pig line. The breakpoint was identified at the base-pair level using a positional cloning and chromosome walking strategy in somatic cell hybrids that were generated from an animal that carries this translocation. We show that this translocation occurs within the ADAMTSL4 gene and results in a loss of expression in homozygous carriers. In addition, by taking this translocation as a model, we used a whole-genome next-generation mate-pair sequencing approach on pooled individuals to evaluate this strategy for high-throughput screening of structural rearrangements. PMID:29121641