2.4 | Real-time quantitative PCR
Total RNA was isolated from aorta using TRIzol reagent (Invitrogen,
Cat# 15596026) and retro-transcribed to cDNA using cDNA PCR kit (Takara
Bio Inc., Cat# RR037Q). Real-time quantitative PCR was performed using
SYBR-Green-based detection (Takara Bio Inc., Cat# RR420L) with primers
commercially synthesized (Sangon Biotech, China). The primer sequences
from 5’ to 3’ are: 1) PPARγ2 forward, TCGCTGATGCACTGCCTATG, PPARγ2
reverse, GAGAGGTCCACAGAGCTGATT; 2) internal control β-actin forward,
AGAGGGAAATCGTGCGTGAC, β-actin reverse, CAATAGTGATGACCTGGCCGT. Real time
quantitative PCR was performed using the following cycling conditions:
denaturation, annealing, and extension at 95°C, 57°C and 72°C for 10 s,
30 s, and 10 s, respectively, for 40 cycles. Relative expression of
PPARγ2 was analysed using the comparative Ct method
(2−ΔΔCt).