Figure 1

Generation of CIDEA reporter mice. (a) Schematic structure of the Cidea-P2A-Luc2-T2A-tdT (Cidea^Luc2-tdT) construct. Reporter cassette encoding Porcine teschovirus-1 2A sequence (P2A), luciferase 2 (Luc2), Thosea asigna virus 2A sequence (T2A), tandem-dimer Tomato (tdT), polyadenylation signal (pA), and a positive selection marker (Neo, neomycin-resistant gene) replaced the stop codon of the Cidea gene. Diphtheria toxin A (DTA) gene was a negative selection marker. Genotyping primer sets for distinguishing wild type (WT) and CIDEA reporter mice were presented. (b) Quantitative PCR analysis of relative Cidea expression level in each tissue of mice (n = 6). (c) Immunoblot analysis of BAT of WT, heterozygous (HET), and homozygous (HOMO) CIDEA reporter mice (n = 8). (d–f) Immunoblot (d,e) and qPCR analysis (f) of BAT and iWAT of WT and HOMO CIDEA reporter mice (TG) maintained in thermoneutral (30 °C for 4 weeks), cold (4 °C for a week), or room temperature control condition (22 °C) (n = 4). Data were analyzed by an unpaired, two-tailed t test in (c) and two-way analysis of variance (ANOVA) with Bonferroni post hoc tests in (d–f) (mean ± SEM; *P < 0.05, **P < 0.01, and ***P < 0.001).