甘蓝SCR识别与结合SRK胞外域核心编码区DNA序列的酵母双杂交检测

SCR是芸薹属自交不亲和性的雄性决定因子。为研究SCR与SRK之间相互作用的核心区段，通过构建结球甘蓝的包含系列不同长度的SCR的cDNA序列的pGBKT7融合载体，利用酵母双杂交系统检测SCR与SRK之间的相互作用。结果显示，构建的载体均未出现自激活现象，所获得SCR全长与其相对应SRK胞外域具有相互作用，且相互作用核心编码区位于SCR编码基因的第97~186 bp处。实验结果还显示，此单倍型的SCR信号肽剪切位点及相邻的几个氨基酸残基对相互作用实验结果有干扰作用。以上结论为包括甘蓝在内的芸薹属自交不亲和机制的研究提供了新的实验依据。

Identifications of DNA Sequences Encoding Key Region of SCR Interacting with SRK Extracellular Domain by Using Yeast Two-Hybrid System

The S-locus cystein-rich protein(SCR)is the male-determining factor of self-incompatibility in Brassica.In this study, the SCR fragments with different lengths amplified from Brassica oleracea L.were ligated with pGBKT7 to construct recombinant bait plasmids,which were then transformed into yeast Y2HGold cells for detecting their interaction with the S-locus receptor kinase extracellular domain(eSRK)in Brassica oleracea by using the yeast two-hybrid system.The results showed that recombinant vectors were not activated autonomously.The full length SCR could interact with eSRK,and the core region in the SCR was located between 97 and 186 bp.Moreover,the result also indicated that the splicing site of signal peptides of this haplotype SCR and its several adjacent amino acid residues could affect the interaction.These conclusions add some novel insights into the mechanism research of self-incompatibility in Brassica.