Matchmaker Gold yeast two-hybrid system

The Matchmaker Gold yeast two-hybrid system was used to detect the interactions between AccMKK6, Accp38b, and Acckayak, as described in a previous study [33]. The typical MAPK member Accp38b (GenBank accession no. {"type":"entrez-nucleotide","attrs":{"text":"GU321334","term_id":"315258190","term_text":"GU321334"}}GU321334) was isolated as previously described [34] and ligated into the pGBKT7 vector. The ORFs of AccMKK6 and Acckayak were cloned and fused to the pGADT7 vector. The plasmids Accp38b-pGBKT7 and AccMKK6-pGADT7, Accp38b-pGBKT7, and Acckayak-pGADT7 were co-transformed into the Y2H Gold yeast strain, respectively, and plated on DDO (SD/-Leu/-Trp) medium. Then, the positive clones were grown on selective SD (QDO/X, SD/-Ade/-His/-Leu/-Trp with X-α-gal and Aureobasidin A) medium.