Fig. 1. Generation of DNA repair gene knock-outs in human intestinal stem cell cultures.

Targeting strategy for the generation of MLH1 (A) and NTHL1 (B) knockout organoids using CRISPR-Cas9 genome editing. sgRNA, single guide RNA. (C) qRT-PCR for MLH1 in normal and MLH1^KO organoids. Expression was normalized to GAPDH. Mean and SD (error bars) of n = 3 independent experiments are indicated. (D) Same as in (C), but for NTHL1. (E) Western blot analysis of MLH1 expression in normal and MLH1^KO organoids (representative from n = 3). Tubulin was used as a loading control. The asterisk indicates a background band. (F) Same as in (E), but for NTHL1. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) was used as a loading control.