Supplementary Figure 8: High-throughput PCR generation of paired gRNA templates for genome editing.
From: Harnessing type I CRISPR–Cas systems for genome engineering in human cells
a, Oligo-templated PCR strategy to generate amplicons for paired gRNA expression from a human U6 (hU6) promoter in mammalian cells. The Rev inner oligonucleotide encodes both gRNA sequences and is modified for new target sites, whereas the remaining primers are invariant. b, Editing efficiencies at Target 7 (from Fig. 2) after co-nucleofecting HEK293 cells with the polycistronic plasmid encoding FokI-EcoCascade and either a paired gRNA expression plasmid or paired gRNA expression amplicon, n=1 nucleofection.
