利用RNAi技术抑制马铃薯类受体激酶基因（StRLK）表达的研究

doi:10.3969/j.issn.1000-7091.2012.04.004

华北农学报 ›› 2012, Vol. 27 ›› Issue (4): 18-22. doi: 10.3969/j.issn.1000-7091.2012.04.004

所属专题： 薯类作物；生物技术；

利用RNAi技术抑制马铃薯类受体激酶基因（StRLK）表达的研究

朱云¹, 樊娜娜², 于耀华¹, 张柏顺², 杨煜², 晁祥建², 郭宝太¹, 金黎平³, 郭晓², 李广存¹

1. 青岛农业大学生命科学院, 山东青岛266109;
2. 山东省农业科学院蔬菜研究所, 国家蔬菜改良中心山东分中心, 山东省设施蔬菜分子生物学重点实验室, 山东济南250100
3. 中国农业科学院蔬菜花卉研究所, 北京100081

出版日期:2012-08-28
作者简介:朱云（1987-），女，安徽马鞍山人，在读硕士，主要从事马铃薯抗病分子育种研究。樊娜娜为同等贡献作者。郭晓（1979-），女，山东新泰人，助理研究员，硕士，主要从事马铃薯抗病育种研究。
基金资助:
现代农业产业技术体系专项资金资助（CARS-10-ES10）;国家科技支撑项目（2012BAD02805）;山东省自然科学基金项目（Y2007D24;ZR2010CQ025）;人力资源和社会保障部留学回国人员科技活动项目择优资助项目

RNA Interference Analysis of Receptor-like Kinase Gene （StRLK） in Potato

ZHU Yun¹, FAN Na-na², YU Yao-hua¹, ZHANG Bai-shun², YANG Yu², CHAO Xiang-jian², GUO Bao-tai¹, JIN Li-ping³, GUO Xiao², LI Guang-eun¹

1. Life Science College of Qingdao Agricultural University, Qingdao 266109, China;
2. Vegetable Research Institute, Shandong Academy of Agricultural Sciences, National Improvement Center for Vegetable, Shandong Branch, Molecular Biology Key Laboratory of Shandong Facility Vegetable, Jinan 250100, China;
3. Institute of Vegetables and Flowers, Chinese Academy of Agricultural Sciences, Beijing 100081, China

Published:2012-08-28

摘要/Abstract

摘要： 植物类受体蛋白激酶（Receptor-like protein kinase，RLK）是蛋白激酶的一个亚家族，参与植物的信号转导，在植物生长发育、抗病等过程中起重要作用。在前期研究中自二倍体马铃薯高抗青枯病基因型ED13中获得了类受体蛋白激酶基因StRLK。利用StRLK基因的特异区段，以pUCCRNAi为中间克隆载体、pCHF1为植物表达载体，构建了该基因的RNA干扰载体pCHF1-StRLK。进一步以ED13的茎段为外植体，利用农杆菌介导法将pCHF1-StRLK导人ED13中，获得了5株再生植株。用CaMV35S启动子特异引物对5株再生植株进行PCR扩增，均得到大小约500bp的特异性条带，初步证明pCHF1-StRLK成功转入ED13。以ED13为对照，利用StRLK基因的特异引物对这5株再生植株进行半定量RT-PCR分析，结果表明，该基因的表达在5个转基因植株中均受到了不同程度的抑制，说明导入的pCHF1-StRLK发挥了干扰活性，且干扰效果与基因的插入位点有关，为进一步研究StRLK基因的功能提供依据。

关键词: 二倍体马铃薯, StRLK基因, 遗传转化, RNA干扰

Abstract: Plant receptor-like protein kinases （RLKs） are the subfamily of protein kinases and participate plant signal transduction. RLKs play an important role in growth,development and disease resistance in plants. In the pre- vious study,we have obtained the potato receptor-like protein kinase gene （StRLK） from diploid potato genotype ED13 with high resistance to Ralstonia solanacearum. The specific fragment of StRLK was used to construct RNAi vector pCHF1-StRLK through the intermediate cloning vector pUCCRNAi and plants expression vector pCHF1. Then the pCHF1-StRLK was transfered into ED13 via agrobacterium-mediated transformation using the stems of ED13 as explants,and finally five transgenic regeneration lines were obtained. The five lines were tested with polymers Chains Reaction （PCR） using the specific primers of CaMV35S promoter,and the results showed that about 500bp PCR products were got in all of them,indicats that all of the five lines are transgenic lines. RT-PCR analysis of StR- LK gene expression in the five transgenic lines were also performed using the StRLK specific primers with ED13 as control. The results showed that the StRLK gene expression were inhibited in different level in the five different lines, demonstrated that pCHF1-StRLK had played interference activity and the interference effection was related to gene insertion site in the potato genome,which would provide some references in the function study of StRLK gene.

Key words: Diploid potato, StRLK gene, Genetic transformation RNA interference

中图分类号:

朱云, 樊娜娜, 于耀华, 张柏顺, 杨煜, 晁祥建, 郭宝太, 金黎平, 郭晓, 李广存. 利用RNAi技术抑制马铃薯类受体激酶基因（StRLK）表达的研究[J]. 华北农学报, 2012, 27(4): 18-22. doi: 10.3969/j.issn.1000-7091.2012.04.004.

ZHU Yun, FAN Na-na, YU Yao-hua, ZHANG Bai-shun, YANG Yu, CHAO Xiang-jian, GUO Bao-tai, JIN Li-ping, GUO Xiao, LI Guang-eun. RNA Interference Analysis of Receptor-like Kinase Gene （StRLK） in Potato[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2012, 27(4): 18-22. doi: 10.3969/j.issn.1000-7091.2012.04.004.

http://www.hbnxb.net/CN/Y2012/V27/I4/18

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利用RNAi技术抑制马铃薯类受体激酶基因（StRLK）表达的研究

RNA Interference Analysis of Receptor-like Kinase Gene （StRLK） in Potato

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利用RNAi技术抑制马铃薯类受体激酶基因（StRLK）表达的研究

RNA Interference Analysis of Receptor-like Kinase Gene （StRLK） in Potato

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