Diversity of 534 pairs of duplicated genes in allotetraploid Gossypium hirsutum

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MethylScreen: DNA methylation density monitoring using quantitative PCR

Aberrant gene silencing of genes through cytosine methylation has been demonstrated during the development of many types of cancers including prostate cancer. Several genes including GSTP1 have been shown to be methylated in prostate cancer leading to the suggestion and demonstration that methylation status of such genes could be used as cancer diagnosis markers alone or in support of histology. We developed a bisulfite-free alternative, MethylScreen technology, an assay for DNA methylation detection utilizing combined restriction from both methylation-sensitive restriction enzymes (MSRE) and methylation-dependent restriction enzymes (MDRE). MethylScreen was used to analyze the 5′ region of GSTP1 in cell lines, in vitro methylated DNA populations, and flash-frozen tissue samples in an effort to characterize the output and analytical performance characteristics of the assay. The output from the quantitative PCR assay suggested that it could not only detect fully methylated molecules in a mixed population below the 1% level, but it could also quantify the abundance of intermediately methylated molecules. Interestingly, the interpreted output from the four quantitative PCRs closely resembled the molecular population as described by clone-based bisulfite genomic sequencing

Unequal Accumulation Of Nucleotide Diversity In The Two Co-Resident Genomes Of Allopolyploid Cotton

Rates of molecular evolution are highly variable within and among genes and among lineages. The forces responsible for this variation include a suite of internal genomic mechanisms (e.g., recombinational environment, repair efficiency) and external population level phenomena (e.g., selection, effective population size). A promising model for disentangling intrinsic and extrinsic forces are allopolyploid plants, which combine two (or more) set of homoeologous genes in a single nucleus and in the same ecological context. Thus, the null hypothesis may be tested that homoeologs will accumulate nucleotide diversity at equivalent rates in a set of populations within a species. Here we test this hypothesis using Gossypium hirsutum (cotton), a natural allopolyploid derived from the merger, 1-2 million years ago (MYA) of progenitor diploid genomes (A and D), which diverged from a common ancestor ~5-10 million years ago. Homoeologous chromosomes in the allopolyploid (AT and DtT remain highly similar to those of their diploid orthologs, with high synteny and colinearity. Using sequence capture and 454 sequencing, we screened 40 diverse G. hirsutum accessions, targeting exonic regions in ~400 homoeologous gene pairs. Analyzed sequences included approximately twice the target space, due to recovery of introns and flanking UTRs. Nucleotide diversity levels in G. hirsutum are low, consistent with earlier indications. Notwithstanding this low level of overall diversity, preliminary analyses suggest that diversity is higher in the At genome than the Dt genome, genome-wide. This enhanced diversity may reflect differences related to the two-fold difference in progenitor genome size. Our results demonstrate that genic environment plays a key role in the genesis of genetic novelty, and how allopolyploidy can create novel allelic combinations with the potential for adaptive traits

Unequal Accumulation Of Nucleotide Diversity In The Two Co-Resident Genomes Of Allopolyploid Cotton

Rates of molecular evolution are highly variable within and among genes and among lineages. The forces responsible for this variation include a suite of internal genomic mechanisms (e.g., recombinational environment, repair efficiency) and external population level phenomena (e.g., selection, effective population size). A promising model for disentangling intrinsic and extrinsic forces are allopolyploid plants, which combine two (or more) set of homoeologous genes in a single nucleus and in the same ecological context. Thus, the null hypothesis may be tested that homoeologs will accumulate nucleotide diversity at equivalent rates in a set of populations within a species. Here we test this hypothesis using Gossypium hirsutum (cotton), a natural allopolyploid derived from the merger, 1-2 million years ago (MYA) of progenitor diploid genomes (A and D), which diverged from a common ancestor ~5-10 million years ago. Homoeologous chromosomes in the allopolyploid (AT and DtT remain highly similar to those of their diploid orthologs, with high synteny and colinearity. Using sequence capture and 454 sequencing, we screened 40 diverse G. hirsutum accessions, targeting exonic regions in ~400 homoeologous gene pairs. Analyzed sequences included approximately twice the target space, due to recovery of introns and flanking UTRs. Nucleotide diversity levels in G. hirsutum are low, consistent with earlier indications. Notwithstanding this low level of overall diversity, preliminary analyses suggest that diversity is higher in the At genome than the Dt genome, genome-wide. This enhanced diversity may reflect differences related to the two-fold difference in progenitor genome size. Our results demonstrate that genic environment plays a key role in the genesis of genetic novelty, and how allopolyploidy can create novel allelic combinations with the potential for adaptive traits