whole exome-seq of homo sapiens: adult female blood
For the generation of standard exome capture libraries, we used the Agilent SureSelect Target Enrichment protocol for Illumina paired-end sequencing library (ver. B.3, June 2015) together with 200ng input FFPE DNA. In all cases, the SureSelectHuman All Exon V5 probe set was used. The quantification of DNA and the DNA quality is measured by PicoGreen and Nanodrop. Fragmentation of 1ug of genomic DNA was performed using adaptive focused acoustic technology.(AFA; Covaris) The fragmented DNA is repaired, an A is ligated to the 3 end, agilnet adapters are then ligated to the fragments. Once ligation had been assessed, the adapter ligated product is PCR amplified. The final purified product is then quantified using qPCR according to the qPCR Quantification Protocol Guide and qualified using the Caliper LabChipHigh Sensitivity DNA. (PerkinElmer).For exome capture, 250 ng of DNA library was mixed with hybridization buffers, blocking mixes, RNase block and 5 l of SureSelect all exon capture library, according to the standard Agilent SureSelect Target Enrichment protocol. Hybridization to the capture baits was conducted at 65C using heated thermal cycler lid option at 105C for 24 hours on PCR machine. The captured DNA was then amplified. The final purified product is then quantified using qPCR according to the qPCR Quantification Protocol Guide and qualified using the TapeStationDNAscreentape(Agilent). And then we sequenced using the HiSeq 2500 platform (Illumina,San Diego, USA)
0802AHP-0040_HAC10