Targetted sequencing of H. sapiens: Sample s_C_006135_P001_d
Genomic DNA from tumor- and patient-matched normal samples were extracted using the Qiagen DNeasy Tissue kit and the EZ1 Advanced XL system (Qiagen, Valencia, CA), respectively. Genomic DNA was sheared using the Covaris E200 instrument (Covaris, Woburn, MA). Sequencing libraries were prepared using the KAPA HTP protocol (Kapa Biosystems, Wilmington, MA) and the Biomek FX system (Beckman Coulter, Brea, CA) through several enzymatic steps, including end repair, A-base addition, ligation of Illumina sequence adaptors followed by PCR amplification, and clean-up. Forty-eight adaptors containing 6-mer unique barcodes corresponding to the Illumina TruSeq barcoding system were used in the library preparation. Tumor and normal libraries were pooled at a 3:1 ratio. A mix of blocker oligos synthesized at IDT (Integrated DNA Technologies, Coralville, IA) and compatible with the TruSeq barcodes was also added to the pool to help prevent nonspecific binding and improve target capture specificity. Custom DNA probes targeting exons and selected introns of the MSK-IMPACT genes were synthesized using the NimbleGen SeqCap EZ library custom oligo system and were biotinylated to allow for sequence enrichment by capture using streptavidin-conjugated beads. Pooled libraries containing captured DNA fragments were subsequently sequenced on the Illumina HiSeq sequencers. Complete details are in ref (https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5808190/)
alignment_software
BWA ver 0.7.12-r1039