(A) The cell lysate containing FtsZ-FLAG or MreB-FLAG was incubated with the lysate containing YgfX-HIS (+) or with that of the wild-type cells (−) before mixed with Ni-NTA column. Eluted fractions were separated by SDS-PAGE and FLAG-tagged proteins were detected using western blot with anti-FLAG antibody conjugated with horseradish peroxidase (Invitrogen, CA).
(B) Yeast two hybrid assay analyzing interaction between YgfX and FtsZ. The full length and various truncated mutants of FtsZ was fused to the activation domain (AD) of pGAD-C1, while YgfX was fused to the binding domain (BD) of pGBD-C1. The interaction was assessed by monitoring the growth on selective media (SD -trp, -leu,-his supplemented with 25 mM 3-aminotriazole).