Next-generation sequencing of targeted loci modified by SMART editing

HEK293T/17 cells were genetically edited by SMART editing to assess the effectiveness of SMART editing. There are 4 target genes: CXCR4, Lmnb1, Nrl and NRXN3. Cultured cells were first synchronized at the G2/M phase. Then, cells were transfected with Cas9 RNP along with SMART or traditional templates. After culture for two to three days, cells were harvested, and genomic DNA was extracted. The targeted loci were amplified by PCR and sequenced on an Illumina MiSeq.