GSM9822534: CITE-seq, cytokine derived iPSC-microglia (iMG), GEX; Homo sapiens; RNA-Seq
iTF-MG are lifted with TrypLE. iMG are lifted by scraping. The CROP-seq library was generated targeting 31 genes that had interesting phenotypes in the original six FACS-based screens. The two sgRNAs with the strongest phenotypes in the FACS-based screens were selected for each gene for inclusion in the CROP-seq library. In total, 31 genes with 2 sgRNAs each were pooled alongside 5 non-targeting sgRNAs that did not influence phenotypes in any of the original FACS-based screens, resulting in a final library size of 65 elements. The crop-seq library was prepared as previously described in Drager et al. Nat Neuro 2022 from arrayed oligonucleotides ordered from Integrated DNA Technologies. Top and bottom oligonucleotides for each sgRNA were annealed and then pooled. This pool was cloned into pMK1334115. The pooled plasmid library was cloned into Stellar Competent cells such that each plasmid was estimated to have been transformed into >500 bacteria. We performed sgRNA enrichment PCR and sequenced this plasmid pool using MiSeq to confirm normal distribution of all sgRNAs.