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Development of Transcriptomic Markers for Population Analysis Using Restriction Site Associated RNA Sequencing (RARseq).

Alabady MS, Rogers WL, Malmberg RL - PLoS ONE (2015)

Bottom Line: The average numbers of RNA SNPs and alleles per loci are 1.89 and 2.17, respectively.Our results suggest that the RARseq protocol allows good depth of coverage per loci for detecting RNA SNPs and polymorphic loci for population genomics and mapping analyses.In non-model systems where complete genomes sequences are not always available, RARseq data can be analyzed in reference to the transcriptome.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant Biology, University of Georgia, Athens, GA, 30604, United States of America.

ABSTRACT
We describe restriction site associated RNA sequencing (RARseq), an RNAseq-based genotype by sequencing (GBS) method. It includes the construction of RNAseq libraries from double stranded cDNA digested with selected restriction enzymes. To test this, we constructed six single- and six-dual-digested RARseq libraries from six F2 pitcher plant individuals and sequenced them on a half of a Miseq run. On average, the de novo approach of population genome analysis detected 544 and 570 RNA SNPs, whereas the reference transcriptome-based approach revealed an average of 1907 and 1876 RNA SNPs per individual, from single- and dual-digested RARseq data, respectively. The average numbers of RNA SNPs and alleles per loci are 1.89 and 2.17, respectively. Our results suggest that the RARseq protocol allows good depth of coverage per loci for detecting RNA SNPs and polymorphic loci for population genomics and mapping analyses. In non-model systems where complete genomes sequences are not always available, RARseq data can be analyzed in reference to the transcriptome. In addition to enriching for functional markers, this method may prove particularly useful in organisms where the genomes are not favorable for DNA GBS.

No MeSH data available.


The correlation between haplotype and gene diversity inferred from RARseq data.Both haplotype and gene diversities were calculated from the de novo analysis (top plots) and the reference-based analysis (bottom plots) of MseI and MesI-Styl RARtags.
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pone.0134855.g003: The correlation between haplotype and gene diversity inferred from RARseq data.Both haplotype and gene diversities were calculated from the de novo analysis (top plots) and the reference-based analysis (bottom plots) of MseI and MesI-Styl RARtags.

Mentions: The total number of catalog loci (maker loci with sequence coverage in all individuals) with at least one SNP ranges from 345 (ref-based MseI-Styl) to 549 (de novo MseI). More SNP/allelic loci were detected using the de novo approach compared to the reference approach. The average number of SNPs and alleles per locus are very similar in the four datasets (Table 4.) The number of inferred haplotypes from de novo and reference-based approaches differed dramatically. For instance, out of 549 SNP/allelic loci, only 28 haplotypes were inferred in the de novo MseI dataset, whereas 351 haplotypes were inferred from 377 SNP/allelic loci in the reference-based MseI dataset (Table 4.) This large difference may be attributed to the pair-wise locus comparison in the de novo analysis, whereas loci are compared to one reference in the reference-based approach. Both haplotype and gene diversity measures were very similar across all datasets (Table 4.) The plot in Fig 3 shows the correlation between mean gene and haplotype diversities in the inferred haplotypes. As evident from the R2 and P values, there is a weak significant correlation between both gene diversity and haplotype diversity. This could indicate low level of linkage disequilibrium within haplotypes, possibly related to the distribution of expressed genes corresponding to the cDNAs.


Development of Transcriptomic Markers for Population Analysis Using Restriction Site Associated RNA Sequencing (RARseq).

Alabady MS, Rogers WL, Malmberg RL - PLoS ONE (2015)

The correlation between haplotype and gene diversity inferred from RARseq data.Both haplotype and gene diversities were calculated from the de novo analysis (top plots) and the reference-based analysis (bottom plots) of MseI and MesI-Styl RARtags.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4524703&req=5

pone.0134855.g003: The correlation between haplotype and gene diversity inferred from RARseq data.Both haplotype and gene diversities were calculated from the de novo analysis (top plots) and the reference-based analysis (bottom plots) of MseI and MesI-Styl RARtags.
Mentions: The total number of catalog loci (maker loci with sequence coverage in all individuals) with at least one SNP ranges from 345 (ref-based MseI-Styl) to 549 (de novo MseI). More SNP/allelic loci were detected using the de novo approach compared to the reference approach. The average number of SNPs and alleles per locus are very similar in the four datasets (Table 4.) The number of inferred haplotypes from de novo and reference-based approaches differed dramatically. For instance, out of 549 SNP/allelic loci, only 28 haplotypes were inferred in the de novo MseI dataset, whereas 351 haplotypes were inferred from 377 SNP/allelic loci in the reference-based MseI dataset (Table 4.) This large difference may be attributed to the pair-wise locus comparison in the de novo analysis, whereas loci are compared to one reference in the reference-based approach. Both haplotype and gene diversity measures were very similar across all datasets (Table 4.) The plot in Fig 3 shows the correlation between mean gene and haplotype diversities in the inferred haplotypes. As evident from the R2 and P values, there is a weak significant correlation between both gene diversity and haplotype diversity. This could indicate low level of linkage disequilibrium within haplotypes, possibly related to the distribution of expressed genes corresponding to the cDNAs.

Bottom Line: The average numbers of RNA SNPs and alleles per loci are 1.89 and 2.17, respectively.Our results suggest that the RARseq protocol allows good depth of coverage per loci for detecting RNA SNPs and polymorphic loci for population genomics and mapping analyses.In non-model systems where complete genomes sequences are not always available, RARseq data can be analyzed in reference to the transcriptome.

View Article: PubMed Central - PubMed

Affiliation: Department of Plant Biology, University of Georgia, Athens, GA, 30604, United States of America.

ABSTRACT
We describe restriction site associated RNA sequencing (RARseq), an RNAseq-based genotype by sequencing (GBS) method. It includes the construction of RNAseq libraries from double stranded cDNA digested with selected restriction enzymes. To test this, we constructed six single- and six-dual-digested RARseq libraries from six F2 pitcher plant individuals and sequenced them on a half of a Miseq run. On average, the de novo approach of population genome analysis detected 544 and 570 RNA SNPs, whereas the reference transcriptome-based approach revealed an average of 1907 and 1876 RNA SNPs per individual, from single- and dual-digested RARseq data, respectively. The average numbers of RNA SNPs and alleles per loci are 1.89 and 2.17, respectively. Our results suggest that the RARseq protocol allows good depth of coverage per loci for detecting RNA SNPs and polymorphic loci for population genomics and mapping analyses. In non-model systems where complete genomes sequences are not always available, RARseq data can be analyzed in reference to the transcriptome. In addition to enriching for functional markers, this method may prove particularly useful in organisms where the genomes are not favorable for DNA GBS.

No MeSH data available.