Fig. 2

Identification of positive targeting mice for the CRISPR/Cas9 genomic engineering. a If DNA sequencing of mouse tail PCR products produced double peaks (frame shift in one allele), the products were subjected to TA cloning, and the further sequencing of independent clones would provide the detailed genomic information of the mosaic mice. b The R172P mutation and gRNA site were framed with a primer pair (607 bp) in the genome for PCR identification. c The electropherogram (bottom panel) shows the PCR identification of Cas9 engineered mice. d The direct sequencing result of PCR products shows the continuously overlapping peaks (double peaks) caused by different allelic substitutions. e The LB agar plate shows the E. coli bacterial clones during TA cloning. f DNA sequencing of TA clones identified 10 different genomic mutations after Cas9targeting. g DNA sequencing of TA clones confirmed that the donor carried 6 synonymous mutations in the gRNA region and a G- > C mutation in the mouse genome, which produced the R172P mutation in the TRP53 of tumour suppressor