(A) Multi-tissue RT-PCR analyses of mouse Adad1 and Adad2. La, ladder; He, heart; Li, liver; Sp, spleen; Lu, lung; Ki, kidney; Br, brain; St, stomach; In, intestine; Te, testis; Ov, ovary; Ut, uterus; Ep, epididymis. The expression of Hprt was analyzed in parallel as a loading control. (B) Postnatal testis RT-PCR analyses of mouse Adad1 and Adad2. The expression of Adad1 and Adad2 was analyzed in mouse testes from postnatal day 5 to 42. (C – D) Schematic representation of Adad2 null mouse generation by CRISPR/Cas9. Single guide RNAs (sgRNAs) #1 and #2, which target the first and the tenth exons, respectively, were used to delete the entire coding region of Adad2. Fw and Rv #1 primers were used to detect the wildtype allele, whereas Fw and Rv #2 primers were used to detect the null allele by genomic PCR. (E) Immunodetection of ADAD2 in wildtype and homozygous null testis lysates using mAb#19–8. BASIGIN was analyzed as a loading control. (F) Fertility tests of Adad2 null male mice. (G) Testis relative to body weight in wildtype and Adad2 null males. ns, no significant difference. (H) Observation of sperm extracted from cauda epididymis from wildtype and Adad2 null males.