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Exp Anim. 2016 Jul 29;65(3):319-27. doi: 10.1538/expanim.16-0016. Epub 2016 Apr 7.

Generation of CRISPR/Cas9-mediated bicistronic knock-in ins1-cre driver mice.

Author information

1
Laborarory Animal Resource Center, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8575, Japan.

Abstract

In the present study, we generated novel cre driver mice for gene manipulation in pancreatic β cells. Using the CRISPR/Cas9 system, stop codon sequences of Ins1 were targeted for insertion of cre, including 2A sequences. A founder of C57BL/6J-Ins1(em1 (cre) Utr) strain was produced from an oocyte injected with pX330 containing the sequences encoding gRNA and Cas9 and a DNA donor plasmid carrying 2A-cre. (R26GRR x C57BL/6J-Ins1(em1 (cre) Utr)) F1 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-Ins1(em1 (cre) Utr) is useful for studies of glucose metabolism and the strategy of bicistronic cre knock-in using the CRISPR/Cas9 system could be useful for production of cre driver mice.

PMID:
27053096
PMCID:
PMC4976246
DOI:
10.1538/expanim.16-0016
[Indexed for MEDLINE]
Free PMC Article

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