Generation of CRISPR/Cas9-mediated bicistronic knock-in <i>ins1-cre</i> driver mice
In the present study, we generated novel <i>cre</i> driver mice for gene manipulation in pancreatic β cells. Using the CRISPR/Cas9 system, stop codon sequences of <i>Ins1</i> were targeted for insertion of <i>cre</i>, including <i>2A</i> sequences. A founder of C57BL/6J-<i>Ins1<sup>em1 (cre) Utr</sup></i> strain was produced from an oocyte injected with <i>pX330</i> containing the sequences encoding gRNA and Cas9 and a DNA donor plasmid carrying <i>2A-cre</i>. (R26GRR x C57BL/6J-<i>Ins1<sup>em1 (cre) Utr</sup></i>) F<sub>1</sub> mice were histologically characterized for cre-loxP recombination in the embryonic and adult stages; cre-loxP recombination was observed in all pancreatic islets examined in which almost all insulin-positive cells showed tdsRed fluorescence, suggesting β cell-specific recombination. Furthermore, there were no significant differences in results of glucose tolerance test among genotypes (homo/hetero/wild). Taken together, these observations indicated that C57BL/6J-<i>Ins1<sup>em1 (cre) Utr</sup></i> is useful for studies of glucose metabolism and the strategy of bicistronic <i>cre</i> knock-in using the CRISPR/Cas9 system could be useful for production of <i>cre</i> driver mice.
実験動物彙報 65(3), 319-327, 2016