Generation of SirT1 Conditional Knockout Mice

A previously described SirT1 targeting construct, KOII [23 (link)], was used to generate mice harboring a conditional targeted mutation in the SirT1 gene (SirT1^co/co mice) (see Additional file 1). The breeding of SirT1^co/co mice and CMV-Cre transgenic mice results in mice harboring a germline-transmitted deletion of exon 4 of the SirT1 gene (SirT1^+/ko mice). Both SirT1^co/co mice and SirT1^+/co mice were used to establish breeding colonies for generating SirT1^co/co and SirT1^ko/ko mice, respectively. Both SirT1^co/co mice and SirT1^ko/ko mice were in a mixed 129SvJ/C57B6 background. Mice were housed in a special-pathogen-free facility and all procedures were approved by the University of Washington Animal Care and Use Committee. A PCR-based genotyping method was established to identify the wild-type, co, and ko loci of the SirT1 gene using three primers: 5' co primer, 5'-GGTTGACTTAGGTCTTGTCTG; 5' ko primer, 5'-AGGCGGATTTCTGAGTTCGA; 3' primer, 5'-CGTCCCTTGTAATGTTTCCC. Murine embryonic fibroblasts (MEFs) were isolated from the embryos between embryonic day 12.5 and embryonic day 14.5. The body weight was measured once a week.