Figure 2 | Scientific Reports

Figure 2

From: NF90–NF45 is essential for β cell compensation under obesity-inducing metabolic stress through suppression of p53 signaling pathway

Figure 2

Generation of NF45 flox/flox mice. (A) Targeting strategy for the generation of NF45 floxed mice using CRISPR/Cas9 system with two single guide RNAs (sgRNAs) targeted to sequences flanking NF45 exon 1 (sgRNA-1 and 2) and a long single-stranded oligonucleotide DNA (lssODNA) containing two loxP sites flanked by short arms with homology to the desired insertion sites. (B) Genotyping of founder mice (#1–#13) injected with sgRNAs-1 and -2, lssODNA and Cas9 mRNA. PCR products corresponding to floxed NF45 locus or knockout of the NF45 locus were indicated by an arrow. M, λHindIII marker (C) DNA sequencing of the CRISPR/Cas9-targeted NF45 locus of founder mice #1 and #4. SgRNAs and loxP sites were showed as blue and red, respectively. (D and E) Genotyping of F1 pups derived from founder #1 and #4 by PCR using primer pairs targeted loxP sites shown in A. M, 100 bp DNA ladder. (F) Western blot analysis of the levels of NF45 in liver, spleen and pancreas from wild-type and NF45 flox/flox (flx/flx) mice. P.C, positive control. Protein extraction from murine testis was used as positive control.

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