Skip to main content
. Author manuscript; available in PMC: 2017 Jul 1.
Published in final edited form as: Nat Protoc. 2016 Dec 8;12(1):88–103. doi: 10.1038/nprot.2016.152

Figure 1. Enhanced genome editing with Dox-inducible Cas9.

Figure 1

a. Overview. A piggy-Bac transposon encapsulating the reverse tet activator, a tet-activator responsive promoter driving humanized Cas9, and a puromycin resistance cassette were integrated into the genome of wild-type human iPSCs. Treatment with Dox and co-transfection with gRNA and donor DNA oligonucleotide efficiently yields mutant iPSC clones. The transposon is efficiently removed by transfection with an excision-only piggyBac transposase, either as a separate step or concurrently with the gRNA and donor oligo. b. PCR genotyping of PGP1 cells confirming hCas9-PB stable integration.