title: GSM5745791: CRISPR screen unsorted population replicate 2; Homo sapiens; OTHER
library_construction_protocol: Genomic DNA was isolated from unsorted cells using the MasterPure Complete DNA Purification kit (Lucigen) and from sorted cells by phenol-chlorofom extraction Sequencing libraries were generated using two sequential rounds of PCR using Herculase II Fusion DNA polymerase (Agilent). In the first round of PCR, 6.6 µg of genomic DNA from unsorted cells was used in each of forty 100 µl reactions, and all recovered genomic DNA from sorted cells was split into two 100 µl reactions. Following 18 cycles of PCR1 amplification, 5 µl from the pooled reaction product was used in the second round of PCR (13 – 14 cycles), using primers containing barcodes and Illumina sequencing adaptors OTHER: CRISPR screen custom DNA library