Yeast two-hybrid analysis

Yeast two-hybrid interaction analyses were performed in the PJ694α yeast strain with the MatchMaker two-hybrid system using the vectors pGBT9 and pGADT7 (Clontech) that express protein fusions to the GAL4 BD or AD domains, respectively. The pGADT7 constructs containing either POLA3 or POLA5 fragments were described by (9); and the pGBT9-CLF and pGBT9 EMF2 were previously described by (38). Selection was performed on synthetic complete (SC) minimal medium without Leu, Trp, His and Ade supplemented with different concentrations of 3-amino-1,2,4-triazole (3-AT).