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. 2018 Aug 16;210(2):607–619. doi: 10.1534/genetics.118.301470

Figure 2.

Figure 2

YDA interacts with EIN3. (A) HIS-YDA fusing protein exhibited specific affinity for GST-EIN3 but not GST in vitro. Input is considered as a positive control. (B) GFP-YDA showed specific affinity with FLAG-EIN3 in vivo. FLAG-EIN3 was associated with membranes and can be detected with anti-GFP antibodies. Wild type was used as a negative control. Input is considered as a positive control. (C) The cDNA of YODA and EIN3 were respectively inserted into pGBKT7 and pGADT7 to obtain the bait plasmid and the prey plasmids. Saccharomyces cerevisiae strain Y2H gold was cotransformed with pGBKT7-YODA and pGADT7-EIN3 vectors, and transformed Y2H Gold cells containing both vectors were selected using SD/-Leu-Trp(DDO), SD/-Leu-Trp-His-Ade(QDO), and SD/-Leu-Trp-His-Ade with X-α-gal and Aba(QDO/A/X). pGBKT7 and pGADT7 were bait and prey vectors without inserts. pGBKT7-53 and pGADT7-T encode two fusion proteins that are known to interact (Clontech). pGBKT7-Lam and pGADT7-T were used as negative controls. Serial dilutions of cotransformed yeast cells were used to measure the strength of the interaction.