title: Generation of CRISPR/Cas9-mediated bicistronic knock-in ins1-cre driver mice.
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.
authors.affiliation: Laborarory Animal Resource Center, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8575, Japan.
authors.forename: Yoshikazu
authors.initials: Y
authors.lastname: Hasegawa
authors.forename: Yoshikazu
authors.initials: Y
authors.lastname: Hoshino
authors.forename: Abdelaziz E
authors.initials: AE
authors.lastname: Ibrahim
authors.forename: Kanako
authors.initials: K
authors.lastname: Kato
authors.forename: Yoko
authors.initials: Y
authors.lastname: Daitoku
authors.forename: Yoko
authors.initials: Y
authors.lastname: Tanimoto
authors.forename: Yoshihisa
authors.initials: Y
authors.lastname: Ikeda
authors.forename: Hisashi
authors.initials: H
authors.lastname: Oishi
authors.forename: Satoru
authors.initials: S
authors.lastname: Takahashi
authors.forename: Atsushi
authors.initials: A
authors.lastname: Yoshiki
authors.forename: Ken-Ichi
authors.initials: K
authors.lastname: Yagami
authors.forename: Hiroyoshi
authors.initials: H
authors.lastname: Iseki
authors.forename: Seiya
authors.initials: S
authors.lastname: Mizuno
authors.forename: Fumihiro
authors.initials: F
authors.lastname: Sugiyama
mesh_terms: D000818:Animals; D064113:CRISPR-Cas Systems; D018388:Codon, Terminator / Q000235:genetics; D050719:Crk-Associated Substrate Protein / Q000008:administration & dosage; D000072669:Gene Editing / Q000379:methods*; D005947:Glucose / Q000378:metabolism; D007267:Injections; D007328:Insulin / Q000235:genetics*; D050417:Insulin-Secreting Cells; D019426:Integrases / Q000008:administration & dosage / Q000235:genetics*; D008810:Mice, Inbred C57BL; D008817:Mice, Mutant Strains / Q000235:genetics; D016254:Mutagenesis, Insertional; D009865:Oocytes; D012313:RNA / Q000008:administration & dosage; D011995:Recombination, Genetic
chemical_list: C493842:Bcar1 protein, mouse; D018388:Codon, Terminator; D050719:Crk-Associated Substrate Protein; C514817:Ins1 protein, mouse; D007328:Insulin; D012313:RNA; C045073:Cre recombinase; D019426:Integrases; D005947:Glucose