Yeast One-Hybrid Assay

In order to identify proteins that bind to the FaQR promoter, the Matchmaker Gold Yeast One-Hybrid Library Screening System (Clontech) was used. The sequence of the FaQR promoter was cloned into the pAbAi vector, and the construct was integrated into the genome of the Y1HGold yeast strain. A mixture of total RNA from strawberry fruit at four stages (G, T, IR, and R) was used to construct the prey cDNA library (TaKaRa). The background AbA^r expression of the Y1HGold FaQR-pAbAi strain was tested according to the system user manual, and then screening for protein-DNA interactions was carried out. Furthermore, the full length of each transcription factor was cloned separately into pGADT7 AD vector to confirm the screening results. The relationship between FaERF#9 and the FaQR promoter also was examined individually. Primers used in this assay are listed in Supplemental Table S6.