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Bioinformatics Advance Access originally published online on June 3, 2009
Bioinformatics 2009 25(16):2074-2075; doi:10.1093/bioinformatics/btp344
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© 2009 The Author(s)
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

Detecting SNPs and estimating allele frequencies in clonal bacterial populations by sequencing pooled DNA

Kathryn E. Holt 1,*, Yik Y. Teo 2, Heng Li 1, Satheesh Nair 3, Gordon Dougan 1, John Wain 3 and Julian Parkhill 1

1 Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge CB10 1SA, 2 Wellcome Trust Centre for Human Genetics, University of Oxford, Roosevelt Drive, Oxford OX3 7BN and 3 Laboratory of Gastrointestinal Pathogens, Centre for Infections, Health Protection Agency, 61 Colindale Avenue, London NW9 5HT, UK

*To whom correspondence should be addressed.


  Abstract

Summary: Here, we present a method for estimating the frequencies of SNP alleles present within pooled samples of DNA using high-throughput short-read sequencing. The method was tested on real data from six strains of the highly monomorphic pathogen Salmonella Paratyphi A, sequenced individually and in a pool. A variety of read mapping and quality-weighting procedures were tested to determine the optimal parameters, which afforded ≥80% sensitivity of SNP detection and strong correlation with true SNP frequency at poolwide read depth of 40x, declining only slightly at read depths 20–40x.

Availability: The method was implemented in Perl and relies on the opensource software Maq for read mapping and SNP calling. The Perl script is freely available from ftp://ftp.sanger.ac.uk/pub/pathogens/pools/.

Contact: kh2{at}sanger.ac.uk

Supplementary information: Supplementary data are available at Bioinformatics online.

Associate Editor: Joaquin Dopazo

Received on March 19, 2009; revised on May 25, 2009; accepted on May 29, 2009

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