Empirical Validation of Pooled Whole Genome Population Re-Sequencing in Drosophila melanogaster
The sequencing of pooled non-barcoded individuals is an inexpensive and efficient means of assessing genome-wide population allele frequencies, yet its accuracy has not been thoroughly tested. We assessed the accuracy of this approach on whole, complex eukaryotic genomes by resequencing pools of largely isogenic, individually sequenced Drosophila melanogaster strains. We called SNPs in the pooled data and estimated false positive and false negative rates using the SNPs called in individual strain as a reference. We also estimated allele frequency of the SNPs using “pooled” data and compared them with “true” frequencies taken from the estimates in the individual strains. We demonstrate that pooled sequencing provides a faithful estimate of population allele frequency with the error well approximated by binomial sampling, and is a reliable means of novel SNP discovery with low false positive rates. However, a sufficient number of strains should be used in the pooling because variation in the amount of DNA derived from individual strains is a substantial source of noise when the number of pooled strains is low. Our results and analysis confirm that pooled sequencing is a very powerful and cost-effective technique for assessing of patterns of sequence variation in populations on genome-wide scales, and is applicable to any dataset where sequencing individuals or individual cells is impossible, difficult, time consuming, or expensive.
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2012-07-26
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dig-ibe-es-10261-999502021-12-28T16:27:33Z Empirical Validation of Pooled Whole Genome Population Re-Sequencing in Drosophila melanogaster Zhu, Yuan Bergland, Alan O. González Pérez, Josefa Petrov, Dmitri A. Ministerio de Ciencia e Innovación (España) European Commission National Institutes of Health (US) Agency for Science, Technology and Research A*STAR (Singapore) The sequencing of pooled non-barcoded individuals is an inexpensive and efficient means of assessing genome-wide population allele frequencies, yet its accuracy has not been thoroughly tested. We assessed the accuracy of this approach on whole, complex eukaryotic genomes by resequencing pools of largely isogenic, individually sequenced Drosophila melanogaster strains. We called SNPs in the pooled data and estimated false positive and false negative rates using the SNPs called in individual strain as a reference. We also estimated allele frequency of the SNPs using “pooled” data and compared them with “true” frequencies taken from the estimates in the individual strains. We demonstrate that pooled sequencing provides a faithful estimate of population allele frequency with the error well approximated by binomial sampling, and is a reliable means of novel SNP discovery with low false positive rates. However, a sufficient number of strains should be used in the pooling because variation in the amount of DNA derived from individual strains is a substantial source of noise when the number of pooled strains is low. Our results and analysis confirm that pooled sequencing is a very powerful and cost-effective technique for assessing of patterns of sequence variation in populations on genome-wide scales, and is applicable to any dataset where sequencing individuals or individual cells is impossible, difficult, time consuming, or expensive. YZ was supported by the A*STAR National Science Scholarship PhD. AOB was supported by the NIH NRSA fellowship (F32 GM097837-01). JG was supported by a Spanish Ministry of Science and Innovation (RYC-2010-07306) and a European Commission Marie Curie CIG Grant (PCIG09-GA-2011-293860). DAP was supported by NIH grants 1R01GM089926 and P50HG002568. Peer reviewed 2014-07-16T07:52:21Z 2014-07-16T07:52:21Z 2012-07-26 artículo http://purl.org/coar/resource_type/c_6501 PLoS ONE 7(7): e41901 (2012) 1932-6203 http://hdl.handle.net/10261/99950 10.1371/journal.pone.0041901 http://dx.doi.org/10.13039/501100004837 http://dx.doi.org/10.13039/501100000780 http://dx.doi.org/10.13039/100000002 http://dx.doi.org/10.13039/501100001348 22848651 en Publisher's version http://dx.doi.org/10.1371/journal.pone.0041901 Sí open Public Library of Science |
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The sequencing of pooled non-barcoded individuals is an inexpensive and efficient means of assessing genome-wide population allele frequencies, yet its accuracy has not been thoroughly tested. We assessed the accuracy of this approach on whole, complex eukaryotic genomes by resequencing pools of largely isogenic, individually sequenced Drosophila melanogaster strains. We called SNPs in the pooled data and estimated false positive and false negative rates using the SNPs called in individual strain as a reference. We also estimated allele frequency of the SNPs using “pooled” data and compared them with “true” frequencies taken from the estimates in the individual strains. We demonstrate that pooled sequencing provides a faithful estimate of population allele frequency with the error well approximated by binomial sampling, and is a reliable means of novel SNP discovery with low false positive rates. However, a sufficient number of strains should be used in the pooling because variation in the amount of DNA derived from individual strains is a substantial source of noise when the number of pooled strains is low. Our results and analysis confirm that pooled sequencing is a very powerful and cost-effective technique for assessing of patterns of sequence variation in populations on genome-wide scales, and is applicable to any dataset where sequencing individuals or individual cells is impossible, difficult, time consuming, or expensive. |
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Ministerio de Ciencia e Innovación (España) |
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Ministerio de Ciencia e Innovación (España) Zhu, Yuan Bergland, Alan O. González Pérez, Josefa Petrov, Dmitri A. |
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artículo |
author |
Zhu, Yuan Bergland, Alan O. González Pérez, Josefa Petrov, Dmitri A. |
spellingShingle |
Zhu, Yuan Bergland, Alan O. González Pérez, Josefa Petrov, Dmitri A. Empirical Validation of Pooled Whole Genome Population Re-Sequencing in Drosophila melanogaster |
author_sort |
Zhu, Yuan |
title |
Empirical Validation of Pooled Whole Genome Population Re-Sequencing in Drosophila melanogaster |
title_short |
Empirical Validation of Pooled Whole Genome Population Re-Sequencing in Drosophila melanogaster |
title_full |
Empirical Validation of Pooled Whole Genome Population Re-Sequencing in Drosophila melanogaster |
title_fullStr |
Empirical Validation of Pooled Whole Genome Population Re-Sequencing in Drosophila melanogaster |
title_full_unstemmed |
Empirical Validation of Pooled Whole Genome Population Re-Sequencing in Drosophila melanogaster |
title_sort |
empirical validation of pooled whole genome population re-sequencing in drosophila melanogaster |
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Public Library of Science |
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2012-07-26 |
url |
http://hdl.handle.net/10261/99950 http://dx.doi.org/10.13039/501100004837 http://dx.doi.org/10.13039/501100000780 http://dx.doi.org/10.13039/100000002 http://dx.doi.org/10.13039/501100001348 |
work_keys_str_mv |
AT zhuyuan empiricalvalidationofpooledwholegenomepopulationresequencingindrosophilamelanogaster AT berglandalano empiricalvalidationofpooledwholegenomepopulationresequencingindrosophilamelanogaster AT gonzalezperezjosefa empiricalvalidationofpooledwholegenomepopulationresequencingindrosophilamelanogaster AT petrovdmitria empiricalvalidationofpooledwholegenomepopulationresequencingindrosophilamelanogaster |
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1777668574076207104 |