GSM8140768: MDSL-Cas9, BRUNELLO, T19, AZA, Rp2; Homo sapiens; OTHER
Large scale DNA Extractions were performed using the Qiagen Blood & Cell Culture DNA Maxi Kit (Qiagen, #13362). sgRNA cassettes were amplified by PCR through multiple 50uL reactions, each containing 5ug gDNA, using staggered P7 primers and indexed P5 primers, and 2X NEBNext Q5 high fidelity master mix (NEB, #M0541L)as previously described. PCR products were pooled, purified using AMPure XP beads. 1.0pM of the pooled CRISPR libraries were sequenced on the NextSeq 500 platform using the NextSeq 550 high-output kit with a read length of 1 x 75bp and a 20% PhiX spike-in. CRISPR-Cas9 dropout screen