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作物学报 ›› 2012, Vol. 38 ›› Issue (09): 1583-1591.doi: 10.3724/SP.J.1006.2012.01583

• 作物遗传育种·种质资源·分子遗传学 • 上一篇    下一篇

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

薛丽琰1,**，罗兵1,3,**，朱利泉1,*，杨永军1，张贺翠1，常登龙1，陈松1，彭一波1，杨红1，曾静1，杨昆1，高启国2，李成琼2，任雪松2，王小佳2   

  1. 1西南大学植物生理生化实验室, 重庆 400716; 2重庆市蔬菜学重点实验室, 重庆 400716; 3常熟理工学院生物与食品工程学院, 江苏常熟 215500
  • 收稿日期:2011-12-16 修回日期:2012-04-20 出版日期:2012-09-12 网络出版日期:2012-07-03
  • 通讯作者: 朱利泉, E-mail: zhuliquan@swu.edu.cn, Tel: 023-68250794
  • 基金资助:

    本研究由国家自然科学基金项目(30971849)和重庆市自然科学基金重点项目(cstc2012jjB80010)资助。

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

XUE Li-Yan1,**,LUO Bing1,3,**,ZHU Li-Quan1,*,YANG Yong-Jun1,ZHANG He-Cui1,CHANG Deng-Long1,CHEN Song1,PENG Yi-Bo1,YANG Hong1,ZENG Jing1,YANG Kun1,GAO Qi-Guo2,LI Cheng-Qiong2,REN Xue-Song2,WANG Xiao-Jia2   

  1. 1Plant Physiology and Biochemistry Laboratory of Southwest University, Chongqing 400716, China; 2 Chongqing Key Laboratory of Olericulture, Chongqing 400716, China; 3 College of Biology and Food Engineering, Changshu Institute of Technology, Changshu 215500, China
  • Received:2011-12-16 Revised:2012-04-20 Published:2012-09-12 Published online:2012-07-03
  • Contact: 朱利泉, E-mail: zhuliquan@swu.edu.cn, Tel: 023-68250794

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摘要： 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。

关键词: 甘蓝, 自交不亲和, S位点富含半胱氨酸蛋白(SCR), 酵母双杂交

Abstract: 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.

Key words: Brassica oleracea, Self-incompatibility, SCR, Yeast two-hybrid system

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