Exome capture of normal sample HS1846
Genomic DNA was extracted following the protocol supplied in the Qiagen AllPrep DNA/RNA Mini Kit (Cat#80204), and quantified using a Quant-iT dsDNA HS assay kit and a Qubit fluorometer (Invitrogen). Approximately 500ng DNA was sheared for 75 seconds at duty cycle “20%” and intensity of “5” using a Covaris E210, and run on an 8% PAGE gel. A 200 to 250bp DNA size fraction was excised and eluted from the gel slice, and was ligated to Illumina paired-end adapters following the standard protocol. The adapter ligated DNA was amplified for 10 cycles using the PE primer set (Illumina) and purified as a pre-exome capture library. The DNA was assessed using an Agilent DNA 1000 Series II assay, and 500ng DNA was hybridized to the 38Mb Human exon probe using the All Exon Kit (Cat#G3362) following the Agilent SureSelect Paired-End Target Enrichment System Protocol (Version 1.0, September 2009). The captured DNA was purified using a Qiagen MinElute column, and amplified for 12 cycles using PE primer set. The PCR products were run on an 8% PAGE gel, the desired size range (320 to 370bp) was excised and purified, and was then assessed using an Agilent DNA 1000 series II assay and diluted to 10nM. The final library DNA concentration was confirmed using a Quant-iT dsDNA HS assay kit and Qubit fluorometer. Clusters were generated on the Illumina cluster station and paired-end reads generated using an Illumina Genome Analyzer (GAIIx) following the manufacturer's instructions.