TGF-β1 Expression Quantification in Rats

TRIzol reagent (Invitrogen; Thermo Fisher Scientific, Inc., Waltham, MA, USA) was used to extract total RNA from rats. cDNA was synthesized from 1 µg of RNA using one-step reverse transcription kit (no. 639505; Takara Bio, Inc., Otsu, Japan). The mRNA levels of each index were measured using a fluorescence quantitative PCR kit (FSQ-101; Toyobo Life Science, Osaka, Japan). GAPDH was taken as an internal reference. TGF-β1 gene localization: NC_000019.10. Primers were designed and synthesized by Shanghai Sangon Biotechnology Co., Ltd. (Shanghai, China). Sequence: upstream, 5′-GGCCAGATCCTGTCCAAGC-3′ and downstream, 5′-GTGGGTTTCCACCATTAGCAC-3′; internal reference GAPDH: upstream, 5′-TGGCCTTCCGTGTTCCTAC-3′ and downstream, 5′-GAGTTGCTGTTGAAGTCGCA-3′. The relative expression level of each index was calculated by 2^−ΔCq [ΔCq = Cq (target gene) - Cq (GAPDH)] (14 (link)–16 (link)).