Fig. 4. Constitutive unphosphorylated PER2 and hypophosphorylated CLOCK in the mPER2 PCD knock-in mice.
a A diagram depicting domains of mPER2 protein and the knock-in (KI) mutations made in the mPER2 PCD domain. b Comparison of the DNA sequencing results in the wild-type (WT) and the Per2m/m mice. c Immunoprecipitation assays and Western blot results showing the levels and phosphorylation profiles of mPER2 and other proteins in the liver extracts of the indicated mouse strains. mPER2 antibody was used in the immunoprecipitation assays. Three independent experiments were performed to validate the results. d Western blot results showing the comparison of mPER2 phosphorylation profiles of the liver extracts collected at CT20 of the WT and Per1−/−; Per2m/m mice. The protein samples were treated with lambda phosphatase. Three independent experiments were performed to validate the results. e Western blot results comparing the mPER2 phosphorylation profiles of the liver extracts collected at CT20 of the WT, Per1−/−; Per2m/m and Per1−/−; Per2m/m; Per3−/− mice. Three independent experiments were performed to validate the results. f Western blot results showing the levels and phosphorylation profiles of mouse clock proteins at the indicated time points in constant darkness. Three independent experiments were performed to validate the results. g The quantification of mPER2 levels of Western blot results in the WT and Per1−/−; Per2m/m mice. Data are represented as mean ± SD, n = 3. h Western blot results showing the levels and phosphorylation profiles of the nuclear mPER2 protein at the indicated time points in constant darkness. See also Supplementary Fig. 3. i The quantification of the nuclear mPER2 levels of Western blot results in the WT and Per1−/−; Per2m/m mice. Data are represented as mean ± SD, n = 3.