Enrichment of indels with FAME strategy in diverse target genes.

A. HEK293T cells were transfected with expression plasmids for spCas9, sgRNAs for B2M and individual genes-of-interest (MYC, ZMIZ1 and PTEN). 5 days later, transfected cells were stained with FITC-conjugated antibody against human HLA-A,B&C, sorted into pools of negative and positive cells, and expanded in culture. The pooled cells were amplified for genomic DNA extraction. For each T7EI assay, genomic DNA from 4 samples were included, i.e., wild-type HEK293T cells, MHC-I positive pool, unsorted pool, and MHC-I negative pool. For each sample, both undigested (-) and digested (+) PCR products by T7E1 were run for comparison and quantification. B. Comparison of indel frequencies for MHC-1 (+), unsorted, and MHC-1 (-) cell population. Indel frequencies were calculated using band signal intensity as reported by image analysis of the original gel pictures from the three T7EI assays represented in panel A. Student’s t-test was used to compare the indel frequencies between MHC-1 (+) and unsorted cells, and between unsorted and MHC-1 (-) cells. * p<0.001.