title: GSM7360013: WSU-DLCL2 DMSO CRISPR screen; Homo sapiens; OTHER
library_construction_protocol: Genomic DNA of cells containing 800 coverage was harvested (Qiagen # 51192) on day 14 after drug treatment. sgRNAs were recovered by PCR amplification (New England Biolabs # M0530L ) in a single-step reaction of 27 cycles. PCR products from all reactions were pooled and purified by gel extraction (Omega # D2500-02). For each sample, >50ng purified PCR fragment was used for library preparation. These PCR products were treated with End Prep Enzyme Mix for end repairing, 5' Phosphorylation and dA-tailing in one reaction, followed by a T-A ligation to add adaptors to both ends. Adaptor ligated DNA was then performed using DNA Clean Beads. A second PCR reaction was carried out with P5 and P7 primers carrying sequences which can anneal with flowcell to perform bridge PCR and index allowing for multiplexing. The final library product for sequencing were then purified by beads and was qualified. The qualified libraries were sequenced pair end 150bp on illumina Hiseq X10 System.