Generation and Characterization of Prx4 and Srx Knockout Mice

All schemes of mouse breeding and experimental protocols were reviewed and approved by the University of Kentucky Institutional Animal Care and Use Committee (UK protocol 2016–2306). All procedures on mice were conducted following the Policy on Humane Care and Use of Laboratory Animals, and the Guidelines of the Animal Care and Laboratory Animal Welfare (NIH). Mice, regardless of strain or genetic background, were all housed in standard cages in temperature-controlled environments under a 12-h light/12-h dark cycle with ad libitum access to standard chow and water unless otherwise indicated. Original Prx4^−/− mice in the C57BL/6 background were originally established by Iuchi et al. [31 (link)]. Prx4^−/− mice in pure FVB/N background were further established by cross breeding the C57BL/6 mice with FVB/N mice for multiple generations (>10). To obtain Prx4^−/− female mice, Prx4 genomic DNA was cloned. In the first intron of Prx4, the phosphoglycerate kinase-driven neomycin cassette flanked by loxP sequences was inserted followed by an extra loxP sequence and introduced upstream of the first exon. Prx4^−/y male mice were generated by breeding Prx4^flox/+ female mice with male mice who express Cre recombinase as described before [31 (link)]. Srx^−/− C57BL/6 mice [32 (link)], were first bred with FVB/N mice and after multiple generations (≥10) of backcross breeding, the offspring Srx^−/− FVB mice were used to breed with Prx4^−/− mice to generate Prx4/Srx double knockout (DKO) mice. The genotype of KO and DKO mice was confirmed by PCR-based mouse genotyping as described previously.