| On the top panel, the schematic draw depicts localisation of each sgRNA on the MIR105/767 locus, as well as the position of primers used for PCR
amplification prior to the Surveyor nuclease treatment. Sizes of initial (uncleaved) PCR products are also given. Transfections with each of the Cas9/sgRNA
vectors (sgRNA1 to -8) were performed in HT1080 cells, and DNA was extracted from transfectants 3 days later. DNA samples were submitted to PCR
amplification with indicated primers, and then exposed to Surveyor nuclease. Resulting DNA products were analyzed by gel electrophoresis. Presence of
DNA fragments with a size lower than the original PCR product indicates efficient sgRNA-directed cleavage. The ratio between the intensity of the lower
band and that of the starting PCR band provides a measurement of cleavage efficiency (% indel). |
