Fig. 3
From: CRISPR-Switch regulates sgRNA activity by Cre recombination for sequential editing of two loci
CRISPR-Switch-Pulse. a, b Schematics of CRISPR pulse construct. The Cas9 cassette provides expression in a non-recombined state (a, left) while sgRNA transcription is terminated by polythymidine signal (poly-T). Upon recombination the Cas9 cassette is removed and sgRNA is transcribed (a, right) leading to a short time window of both sgRNA and Cas9 present in a cell (b). U6 – U6 promoter. Triangles represent recombination sites. c EGFP deletion efficiency of CRISPR-Switch-Pulse with sgEGFP1 and constitutively expressed Cas9 with sgEGFP1 was compared in CreERT2, EGFP-positive mES cells. Error bars represent standard deviation (n = 3). d Switch-Pulse and constitutive Cas9 lentivirus with sgEGFP1 guide RNA targeting both EGFP and EBFP2 was introduced to NIH3T3 cells encoding CreERT2, EGFP, and EBFP2. Efficiency of single and double deletion was assessed with flow cytometry in FITC and BV421 channels. Plus refers to s.d.
