Expression of ARC1 in Vitro and Test of Interaction Between ARC1 and SRK from Brassica oleracea L. in Signal Transduction Pathway of Self-Incompatibility

Abstract ARC1 is an arm repeat containing a downstream gene of S-locus receptor kinase (SRK). It interacts with SRK probably in signal transduction pathway of self-incompatibility. In this study, the interaction between SRK and ARC1 proteins was tested using the expression vectors pET43.1a-ARC1 and pET43.1a-SRK E1 . The coding sequences of ARC1 gene were amplified from stigma mRNA of Brassica oleracea L. and inserted into the expression vector pET43.1a to construct the recombinant plasma pET43.1a-ARC1. The recombinant plasmid was transformed to Escherichia coli (BL21) and the expression of the recombinant protein was detected using sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Using the co-immunoprecipitation theory and characteristic of 6× His Tag combined with Ni+ in pET43.1a-ARC1, a new method was proposed for testing the interaction between proteins. With this method the interaction between ARC1 and SRK was analyzed, showing that ARC1 and SRK could interact with each other to combine and form a complex. This study provides the theoretical and technical bases for further analyzing the mechanism of interaction between ARC1 and SRK proteins and for probing into interaction between ARC1 and downstream targets.