Quantitative Real-Time PCR for Gene Expression

A Tetro cDNA synthesis kit (catalogue no. BIO-65043, Bioline, London, UK) was used to synthesise cDNA using random primers, while following the manufacturer’s instructions. Two genes from each condition (YPD-28 °C, YPD-16 °C, and YP + glycerol-16 °C) showing significant transcriptional changes in RNAseq dataset were picked for further validation of the expression by real-time PCR. Real-time PCR was performed in a Roche thermocycler on the cDNA of hybrids and parental strains using the iTaq Universal SYBR Green Supermix (catalogue no. 1725121, BioRad, Deeside, UK). Reverse transcription and real time PCR was done, as described previously [14 (link),15 (link)]. Table S8 shows the primers used for real time.