Fig. 1. Schematic of the pDEST-GADT7 and pDEST-GBKT7 expression cassettes. The Gateway cassette [composed of the recombination sites attR1 and attR2, the chloramphenicol resistance gene (Cm^R) and the ccdB gene] was inserted into the SmaI site of the yeast two-hybrid vectors pGADT7 and pGBKT7. LR recombination with the target gene replaces the region between the attR1 and attR2 recombination sites to create in-frame fusions with either the GAL4 activation domain (GAL4 AD) fused to the HA-tag in the case of pDEST-GADT7 or the GAL4 binding domain (GAL4 BD) fused to the c-Myc tag for pDEST-GBKT7. The promoter from ADH1 (PADH1) and the ADH1 terminator (T_ADH1) were used to control the expression of the gene fusion in yeast, and the T7 promoter (P_T7) was used for in vitro expression. Nucleic acid sequence and predicted amino acid sequence of the junctions between the original yeast two-hybrid vectors and the gateway cassette are given below each vector.