Figure 2.

Generation of Tmprss12^del/del mice using CRISPR/Cas9. (A) Structure of Tmprss12 gene and CRISPR/Cas9 targeting scheme in exon 2 and 5. Asterisk (^*) indicates the 8 bp deletion. Black boxes show the coding region. (B) Wave pattern sequence of Tmprss12 in WT and Tmprss12^del/del mice. (C) Genotyping of Tmprss12^del/del mice using two primer sets that amplify the WT allele or the large deletion allele. (D) Amino acid translation in Tmprss12^del/del mice. The large deletion in Tmprss12^del/del mice causes a premature stop codon at amino acid 70. (E) Protein expression of TMPRSS12 in testis and cauda epididymal spermatozoa from WT, Tmprss12^d8/d8, and Tmprss12^del/del mice. GAPDH was used as a loading control. TGC: testicular germ cells. (F) Immunofluorescence analysis of spermatozoa from Tmprss12 ^del/del mice labeled with antibodies against TMPRSS12 (red). Fluorescence observed in the WT sperm head disappears in spermatozoa from Tmprss12^del/del mice. Nuclei were stained with Hoechst 33342 (blue). Scale bars: 25 μm.