Comparative genome-wide polymorphic microsatellite markers in Antarctic penguins through next generation sequencing.

Microsatellites are valuable molecular markers for evolutionary and ecological studies. Next generation sequencing is responsible for the increasing number of microsatellites for non-model species. Penguins of the Pygoscelis genus are comprised of three species: Adélie (P. adeliae), Chinstrap (P. antarcticus) and Gentoo penguin (P. papua), all distributed around Antarctica and the sub-Antarctic. The species have been affected differently by climate change, and the use of microsatellite markers will be crucial to monitor population dynamics. We characterized a large set of genome-wide microsatellites and evaluated polymorphisms in all three species. SOLiD reads were generated from the libraries of each species, identifying a large amount of microsatellite loci: 33,677, 35,265 and 42,057 for P. adeliae, P. antarcticus and P. papua, respectively. A large number of dinucleotide (66,139), trinucleotide (29,490) and tetranucleotide (11,849) microsatellites are described. Microsatellite abundance, diversity and orthology were characterized in penguin genomes. We evaluated polymorphisms in 170 tetranucleotide loci, obtaining 34 polymorphic loci in at least one species and 15 polymorphic loci in all three species, which allow to perform comparative studies. Polymorphic markers presented here enable a number of ecological, population, individual identification, parentage and evolutionary studies of Pygoscelis, with potential use in other penguin species.

Genome diversification within a clonal population of pandemic Vibrio parahaemolyticus seems to depend on the life circumstances of each individual bacteria.

BACKGROUND: New strains of Vibrio parahaemolyticus that cause diarrhea in humans by seafood ingestion periodically emerge through continuous evolution in the ocean. Influx and expansion in the Southern Chilean ocean of a highly clonal V. parahaemolyticus (serotype O3:K6) population from South East Asia caused one of the largest seafood-related diarrhea outbreaks in the world. Here, genomics analyses of isolates from this rapidly expanding clonal population offered an opportunity to observe the molecular evolutionary changes often obscured in more diverse populations. RESULTS: Whole genome sequence comparison of eight independent isolates of this population from mussels or clinical cases (from different years) was performed. Differences of 1366 to 217,729 bp genome length and 13 to 164 bp single nucleotide variants (SNVs) were found. Most genomic differences corresponded to the presence of regions unique to only one or two isolates, and were probably acquired by horizontal gene transfer (HGT). Some DNA gain was chromosomal but most was in plasmids. One isolate had a large region (8,644 bp) missing, which was probably caused by excision of a prophage. Genome innovation by the presence of unique DNA, attributable to HGT from related bacteria, varied greatly among the isolates, with values of 1,366 (ten times the number of highest number of SNVs) to 217,729 (a thousand times more than the number of highest number of SNVs). CONCLUSIONS: The evolutionary forces (SNVs, HGT) acting on each isolate of the same population were found to differ to an extent that probably depended on the ecological scenario and life circumstances of each bacterium.

Yellow lupin (Lupinus luteus L.) transcriptome sequencing: molecular marker development and comparative studies.

BACKGROUND: Yellow lupin (Lupinus luteus L.) is a minor legume crop characterized by its high seed protein content. Although grown in several temperate countries, its orphan condition has limited the generation of genomic tools to aid breeding efforts to improve yield and nutritional quality. In this study, we report the construction of 454-expresed sequence tag (EST) libraries, carried out comparative studies between L. luteus and model legume species, developed a comprehensive set of EST-simple sequence repeat (SSR) markers, and validated their utility on diversity studies and transferability to related species. RESULTS: Two runs of 454 pyrosequencing yielded 205 Mb and 530 Mb of sequence data for L1 (young leaves, buds and flowers) and L2 (immature seeds) EST- libraries. A combined assembly (L1L2) yielded 71,655 contigs with an average contig length of 632 nucleotides. L1L2 contigs were clustered into 55,309 isotigs. 38,200 isotigs translated into proteins and 8,741 of them were full length. Around 57% of L. luteus sequences had significant similarity with at least one sequence of Medicago, Lotus, Arabidopsis, or Glycine, and 40.17% showed positive matches with all of these species. L. luteus isotigs were also screened for the presence of SSR sequences. A total of 2,572 isotigs contained at least one EST-SSR, with a frequency of one SSR per 17.75 kbp. Empirical evaluation of the EST-SSR candidate markers resulted in 222 polymorphic EST-SSRs. Two hundred and fifty four (65.7%) and 113 (30%) SSR primer pairs were able to amplify fragments from L. hispanicus and L. mutabilis DNA, respectively. Fifty polymorphic EST-SSRs were used to genotype a sample of 64 L. luteus accessions. Neighbor-joining distance analysis detected the existence of several clusters among L. luteus accessions, strongly suggesting the existence of population subdivisions. However, no clear clustering patterns followed the accession's origin. CONCLUSION: L. luteus deep transcriptome sequencing will facilitate the further development of genomic tools and lupin germplasm. Massive sequencing of cDNA libraries will continue to produce raw materials for gene discovery, identification of polymorphisms (SNPs, EST-SSRs, INDELs, etc.) for marker development, anchoring sequences for genome comparisons and putative gene candidates for QTL detection.

Association of nicotinic acid with a poly(amidoamine) dendrimer studied by molecular dynamics simulations.

The interaction of poly(amidoamine)-G3 (PAMAM-G3) dendrimer with nicotinic acid (NA) was investigated by using molecular dynamics (MD) simulations. First, sample free energy profiles of NA crossing PAMAM-G3 at pH 6 and 3 were computed using the adaptive biasing force (ABF) method. We found that PAMAM-G3 provides a more appropriate environment for NA inclusion when internal tertiary amine groups are unprotonated (at pH 6). However, when internal tertiary amine groups are protonated (at pH 3), the PAMAM cavities are less hydrophobic; therefore the drug-dendrimer interactions become similar to drug-solvent interactions. Traditional MD simulations were also performed to investigate the structural stability of the PAMAM-NA complexes near the free energy minima at pH 6. We found that association of NA and PAMAM adopts a preferred binding mode around the surface of PAMAM, where hydrogen bond (HB) interactions with the amino and amide NH groups of the nearby monomers are established. These interactions are very stable whether additional van der Waals interactions between pyridine ring of NA and methylene groups of the more external monomers of PAMAM are established.

Comparative genome-wide polymorphic microsatellite markers in Antarctic penguins through next generation sequencing

Abstract Microsatellites are valuable molecular markers for evolutionary and ecological studies. Next generation sequencing is responsible for the increasing number of microsatellites for non-model species. Penguins of the Pygoscelis genus are comprised of three species: Adélie (P. adeliae), Chinstrap (P. antarcticus) and Gentoo penguin (P. papua), all distributed around Antarctica and the sub-Antarctic. The species have been affected differently by climate change, and the use of microsatellite markers will be crucial to monitor population dynamics. We characterized a large set of genome-wide microsatellites and evaluated polymorphisms in all three species. SOLiD reads were generated from the libraries of each species, identifying a large amount of microsatellite loci: 33,677, 35,265 and 42,057 for P. adeliae, P. antarcticus and P. papua, respectively. A large number of dinucleotide (66,139), trinucleotide (29,490) and tetranucleotide (11,849) microsatellites are described. Microsatellite abundance, diversity and orthology were characterized in penguin genomes. We evaluated polymorphisms in 170 tetranucleotide loci, obtaining 34 polymorphic loci in at least one species and 15 polymorphic loci in all three species, which allow to perform comparative studies. Polymorphic markers presented here enable a number of ecological, population, individual identification, parentage and evolutionary studies of Pygoscelis, with potential use in other penguin species.

Characterization of novel genic SSR markers in Linum usitatissimum (L. ) and their transferability across eleven Linum species

Little is known about the evolutionary relationships among Linum species, basically because of the lack of transferable molecular markers. Currently, expressed sequence tags available in public databases provide an opportunity for the rapid and inexpensive development of simple sequence repeat (SSR) markers in wild flax species. In this regard, fifty expressed sequence tag-derived microsatellite markers (EST-SSRs) were evaluated for polymorphism and transferability in 50 Linum usitatissimum cultivars/accessions and 11 Linum species. Among them 23 EST-SSRs were polymorphic in L. usitatissimum, while 2-4 alleles were detected (average 2.26 per locus). The polymorphism information content value ranged from 0.08 to 0.55 (average 0.38). Forty one genic markers (95.3 percent) produced strong amplicons in at least two of the 11 Linum species. The percentage of cross amplification ranged from 34.1 percent to 92.7 percent in L. tauricum and L. bienne, respectively. Moreover, the rate of transferability was associated positively with the botanical section. Our results suggest that the high degree of EST-SSRs transferability to Linum species can be a useful enhancement of the current database of SSR markers for future genetic and evolutionary studies.