Yeast 1-hybrid assay

The SlPaO promoter region (Solyc11g066440) of 1,434 bp was amplified using a primer pair containing EcoRI restriction sites (Supplemental Table ^S1), cloned into pHis2 vector (Clontech) previously linearised using EcoRI. The bait plasmid (pPaO-pHis2) was used to transform Saccharomyces cerevisiae Y187 strain (Clontech). The Solyc12g03648/HEB gene (981 bp) cloned into pBlueScript II SK(+) vector was purchased from Biomatik Corporation (Cambridge, Canada), excised with EcoRI and XhoI and ligated into pGADT7 (Clontech) EcoRI/XhoI digested. The prey plasmid (HEB-pGADT7) was introduced into Saccharomyces cerevisiae AH109 strain (Clontech) and transformants mated with Y187 strain containing the bait plasmid as described by Resentini et al.¹⁰⁴. Diploids were selected on medium lacking Trp and Leu. Growth diploid colonies were scraped on selective media lacking Trp, Leu and His and supplemented with 0, 1, 2 or 5 mM 3-AT (Sigma-Aldrich). Yeast 1-Hybrid was also performed using as bait and prey plasmids pHis2 and pGADT7 respectively as controls.