Nimblegen v2

Whole-exome sequencing was performed on genomic DNA from 51 patients and 47 controls (the initial screening group). Whole-exome capture libraries were constructed from approximately 3μg of purified genomic DNA following fragmentation, end repair, phosphorylation, and ligation. Ligated samples were hybridized to the exon capture arrays using Roche NimbleGen V2 (44.1 Mbp). Captured DNA was annealed at 95°C and single-stranded DNA amplified and sequenced on an Illumina HiSeq2000.
Exome sequencing data were analyzed using the Exome Analysis Pipeline (DNAnexus, http://www.dnanexus.com). Variant detection and genotyping were performed on exomes and the flanking 500 bp downstream of the 5’- or 3’-UTRs. A BAM (binary alignment map) file for each sample was generated from the alignment of the Illumina sequence reads to the human genome (hg19). SNV calls were evaluated and each variant annotated.

Cloned parasites were initially typed with 10 microsatellites to screen for potential recombinant progeny as described^{36 (link)} using primers in Supplementary Table 2. DNA labeling and hybridization to a custom-made SNP-typing microarray were conducted essentially as described using 1 μg of gDNA^{27 (link)}. Scanned images were gridded and processed using NimbleGen v2.5 (Roche NimbleGen Inc.) to extract signal data. SNP genotypes were called when the probe sets at each locus indicated complementary SNP genotypes on both DNA strands. Parental clones were also hybridized in duplicate, and genotypes were scored at each locus as inheriting one of the parental genotypes or as N/A if a non-parental genotype was called. A total of 60 progeny with unique microsatellite patterns were genotyped with the SNP array.