Fig. 2.

Dual sgRNA-directed large deletion of TYR in zygotes. a Determination of 105-kb deletions of TYR gene in rabbit embryos by PCR. A clear band demonstrates the large-fragment deletion of TYR in #5 embryos; primers 1F and 1R were used for large-fragment determination. M DNA ladder. b Mutation detection for each sgRNA in zygotes. Primers TYR-2F/2R were used for mutation determination for sgRNA1 and sgRNA2, and TYR-3F/3R were used for sgRNA3 and sgRNA 4. M DNA marker. c T-cloning sequencing of deletion of TYR in #5 blastocyst. PAM sites are underlined and highlighted in red; target sequences are green; deletions (−) and insertions (+) are shown. WT wild-type control. d T-cloning sequencing of the target site for each sgRNA in injected embryos. PAM sites are underlined and highlighted in red; target sequences are green; deletions (−) and insertions (+) are shown. WT wild-type control