A bioinformatic filter for improved base-call accuracy and polymorphism detection using the Affymetrix GeneChip whole-genome resequencing platform.
DNA resequencing arrays enable rapid acquisition of high-quality sequence data. This technology represents a promising platform for rapid high-resolution genotyping of microorganisms. Traditional array-based resequencing methods have relied on the use of specific PCR-amplified fragments from the query samples as hybridization targets. While this specificity in the target DNA population reduces the potential for artifacts caused by cross-hybridization, the subsampling of the query genome limits the sequence coverage that can be obtained and therefore reduces the technique's resolution as a genotyping method. We have developed and validated an Affymetrix Inc. GeneChip(R) array-based, whole-genome resequencing platform for Francisella tularensis, the causative agent of tularemia. A set of bioinformatic filters that targeted systematic base-calling errors caused by cross-hybridization between the whole-genome sample and the array probes and by deletions in the sample DNA relative to the chip reference sequence were developed. Our approach eliminated 91% of the false-positive single-nucleotide polymorphism calls identified in the SCHU S4 query sample, at the cost of 10.7% of the true positives, yielding a total base-calling accuracy of 99.992%.
Pandya
Gagan A
GA
Pathogen Functional Genomics Resource Center, The Institute for Genomic Research at the J. Craig Venter Institute, Rockville, MD 20850, USA.
Holmes
Michael H
MH
Sunkara
Sirisha
S
Sparks
Andrew
A
Bai
Yun
Y
Verratti
Kathleen
K
Saeed
Kelly
K
Venepally
Pratap
P
Jarrahi
Behnam
B
Fleischmann
Robert D
RD
Peterson
Scott N
SN
D019295:Computational Biology / Q000379:methods*; D005604:Francisella tularensis / Q000235:genetics; D016680:Genome, Bacterial; D023281:Genomics / Q000379:methods*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods* / Q000592:standards; D020641:Polymorphism, Single Nucleotide; D012015:Reference Standards; D015203:Reproducibility of Results