title: Detection of genomic variation by selection of a 9 mb DNA region and high throughput sequencing.
abstract: Detection of the rare polymorphisms and causative mutations of genetic diseases in a targeted genomic area has become a major goal in order to understand genomic and phenotypic variability. We have interrogated repeat-masked regions of 8.9 Mb on human chromosomes 21 (7.8 Mb) and 7 (1.1 Mb) from an individual from the International HapMap Project (NA12872). We have optimized a method of genomic selection for high throughput sequencing. Microarray-based selection and sequencing resulted in 260-fold enrichment, with 41% of reads mapping to the target region. 83% of SNPs in the targeted region had at least 4-fold sequence coverage and 54% at least 15-fold. When assaying HapMap SNPs in NA12872, our sequence genotypes are 91.3% concordant in regions with coverage > or = 4-fold, and 97.9% concordant in regions with coverage > or = 15-fold. About 81% of the SNPs recovered with both thresholds are listed in dbSNP. We observed that regions with low sequence coverage occur in close proximity to low-complexity DNA. Validation experiments using Sanger sequencing were performed for 46 SNPs with 15-20 fold coverage, with a confirmation rate of 96%, suggesting that DNA selection provides an accurate and cost-effective method for identifying rare genomic variants.
authors.affiliation: Department of Genetic Medicine and Development, University of Geneva Medical School, Geneva, Switzerland. Sergey.Nikolaev@unige.ch
authors.forename: Sergey I
authors.initials: SI
authors.lastname: Nikolaev
authors.forename: Christian
authors.initials: C
authors.lastname: Iseli
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Sharp
authors.forename: Daniel
authors.initials: D
authors.lastname: Robyr
authors.forename: Jacques
authors.initials: J
authors.lastname: Rougemont
authors.forename: Corinne
authors.initials: C
authors.lastname: Gehrig
authors.forename: Laurent
authors.initials: L
authors.lastname: Farinelli
authors.forename: Stylianos E
authors.initials: SE
authors.lastname: Antonarakis
mesh_terms: D002874:Chromosome Mapping; D002891:Chromosomes, Human, Pair 21; D004247:DNA / Q000235:genetics*; D014644:Genetic Variation; D015894:Genome, Human; D006801:Humans; D020641:Polymorphism, Single Nucleotide
chemical_list: D004247:DNA
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