*4.5. Reverse-Transcription PCR and Quantitative Real-Time PCR*

The first strand cDNA was synthesized by the RevertAid First Strand cDNA Synthesis Kit (K1621, Thermo, Waltham, MA, USA). Then, the cDNA was adjusted to an equal concentration. Reverse-transcription PCR was performed by EasyTaq PCR SuperMix (AS111, TRANS, Beijing, China) with a PCR program of (1) 94 °C for 3 min; (2) 29 cycles of 94 °C for 30 s, 55 °C for 30 s, and 72 °C for 2 min; and (3) 72 °C for 10 min and 16 °C for storage.

Quantitative real-time PCR was performed using TransStart Tip Green qPCR SuperMix (AQ141, TRANS, Beijing, China) in LightCycler®96 (Roche, Basel, Switzerland) with a PCR program of (1) 94 °C for 30 s; (2) 45 cycles of 94 °C for 5 s, 55 °C for 15 s, and 72 °C for 15 s; and (3) 95 °C for 10 s, 65 °C for 60 s, and 97 °C for 1 s as the melting curve. *Elongation factor 1* (*EF1*) was used as the reference gene, and the primers used in reverse-transcription PCR and quantitative real-time PCR are listed in Supplementary Tables S2 and S3.
