Quantitative RT-PCR

Indicated populations were sorted into RLT Plus lysis buffer (Qiagen) and stored at −80°C, then processed using Allprep DNA/RNA micro kit (Qiagen) per manufacturer’s protocol. For qPCR analyses, RNA was reverse transcribed using SuperScript III cDNA synthesis kit (ThermoFisher) and amplified using Power SYBR Green PCR master mix (ThermoFisher). Aldh1a2 primers: 5′-GACTTGTAGCAGCTGTCTTCACT-3′, forward, and 5′-TCACCCATTTCTCTCCCATTTCC-3′, reverse. Rsp17 gene was used as a housekeeping gene and amplified with the following primers: ATTGAGGTGGATCCCGACAC, forward; TGCCAACTGTAGGCTGAGTG, reverse.