Sd leu trp

The target gene bait vectors, pGBKT7-Ats-1, and pGADT7 AD empty vectors were co-transferred to the MATα strain by the LiAc method and coated on SD/–Leu/–Trp plates (Coolaber, Beijing, China). The monoclonal antibody (generally > 2 mm) with good growth on a SD/–Leu/–Trp (Coolaber, Beijing, China) plate was collected and placed into SD/–Leu/–Trp in liquid medium in shakers at 29 °C and 200 rpm for 200 h. The shaken bacterial droplets were aspirated to SD/–Ade/–His/–Leu/–Trp plates (Coolaber, Beijing, China). The growth of colonies on the plates was observed.

The CDS regions of fus3, ste7, ste11, and ste11 were amplified from total cDNA, purified with a Gel Pure Kit columns I (Magen), and attached to linearized pGADT7-AD and pGBKT7-BD with a ClonExpress II One Step Cloning Kit (Vazyme Biotech Co.). The constructed plasmids were sequenced in Sangon Biotech (Shanghai, China), extracted with a TIANprep mini Plasmid Kit (Transgene, Beijing, China), then pairwise co-transfected into the Y2HGold cells by using a Yeastmaker Yeast Transformation System 2 (630439, TaKaRa). All of the selective medium and reagents, including SD/-Leu, SD/-Trp, SD/-Leu/-Trp, SD/-His/-leu/-Trp/-His, and X-α-gal, were purchased from Coolaber (Beijing, China). All primers used in this experiment are listed in Table S5.