Fig. 4 Gene disruption via chip. Gene disruption via chip. Plasmids encoding both sgRNA targeting AAVS1 locus or NUAK2 and Cas9 protein were delivered into MCF7 and HeLa cells, respectively. After 7 days of cell culture, genomic DNA was extracted. PCR product sequencing for specific targeting regions was performed. (A) PCR product sequencing data for the sgAAVS1 targeting region. The 20-bp target sequence is shown in red; the PAM sequence is shown in blue. Representative sequences for indels are shown. Short black lines denote different deletions. Black arrow denotes an insertion. (B) Surveyor mutation detection assay for sgAAVS1- and Cas9 protein–mediated indels via chip. Arrows indicate the expected positions of DNA bands cleaved by Surveyor Nuclease S. The symbol * indicates the cleavage lane of DNA bands after cells went through the same chip three times. (C) Illustration of sgNUAK2 targeting region at the first exon. The 20-bp target sequence is shown in red; the PAM sequence is shown in blue. (D) PCR product sequencing data for the sgNUAK2 targeting region. Representative sequences for deletions are shown. Short black lines denote different deletions. (E) Surveyor mutation detection assay for sgNUAK2- and Cas9 protein–mediated indels via chip. Arrows indicate the expected positions of DNA bands cleaved by Surveyor Nuclease S. Xin Han et al. Sci Adv 2015;1:e1500454 Copyright © 2015, The Authors