Hifair 1st strand cdna synthesis supermix

Total RNA was isolated from cells and tissues using TRIzol reagent (Takara, Shiga, Japan) according to the manufacturer’s instructions. The concentrations of RNA were measured using a NanoDrop 2000 (Thermo Fisher Scientific, Waltham, MA, USA). One microgram of RNA per sample was reverse transcribed using Hifair™ 1st Strand cDNA Synthesis SuperMix (Yeasen, Shanghai, China). qRT-PCR was performed using the Hieff™ qPCR SYRB Green Master Mix (Yeasen). The following primers were used for qRT-PCR: GAPDH, forward 5′-GGGAAGGTGAAGGTCGGAGT-3′ and reverse 5′-GGGGTCATTGATGGCAACA-3′; HOXA13, forward 5′-GAACGGCCAAATGTACTGCC-3′ and reverse 5′-GTATAAGGCACGCGCTTCTTTC-3′; VEGFA, forward 5′-ATCGAGTACATCTTCAAGCCAT-3′ and reverse 5′-GTGAGGTTTGATCCGCATAATC-3′; EGFR, forward 5′-ACCCATATGTACCATCGATGTC-3′ and reverse 5′-GAATTCGATGATCAACTCACGG-3′; FGF2, forward 5′-CATCAAGCTACAACTTCAAGCA-3′ and reverse 5′-CCGTAACACATTTAGAAGCCAG-3′; FGFR2, forward 5′-CTAAAGGCAACCTCCGAGAATA-3′ and reverse 5′-ACATTTTTGGGAAGCCAAGTAC-3′. Relative expression was normalized to GAPDH expression. Each qRT-PCR reaction was performed in triplicate, and the relative mRNA levels were calculated using the 2^−ΔΔCt method.