黄瓜发根农杆菌转化体系的建立  

Establishment of Transformation System of Agrobacterium rhizogenes in Cucumber

作　　者：樊颖伦[1] 徐风琳 周慧珍 刘欣欣 杨欣玥 吕山花[1] Fan Yinglun;Xu Fenglin;Zhou Huizhen;Liu Xinxin;Yang Xinyue;LüShanhua(College of Agronomy,Liaocheng University,Liaocheng,252000)

机构地区：[1]聊城大学农学院,聊城252000

出　　处：《分子植物育种》2025年第1期113-118,共6页Molecular Plant Breeding

基　　金：国家自然科学基金项目(31271751)资助。

摘　　要：黄瓜是根癌农杆菌介导遗传转化获得转基因植株较为困难的物种之一,尽管黄瓜的基因组测序已经完成,但大多数基因的功能尚未确定。发根农杆菌介导的毛状根转化能高效地诱导植物产生转基因根,为研究相关基因在根中的生物学功能提供了一种方便、快捷的途径。本研究建立了由发根农杆菌介导的高效、快速的适用于在黄瓜活体上诱导产生转基因根的转化方法。利用携带红色荧光蛋白基因DsRed2的发根农杆菌K599穿刺接种不同基因型黄瓜幼苗的下胚轴,在高湿状态下培养2周左右。在3种不同基因型黄瓜幼苗的下胚轴穿刺部位生长出毛状根,通过红色荧光蛋白观察及PCR验证,表明89%~94%的穿刺植株携带有转基因毛状根,新生毛状根中转基因根的比例约为55%。由此建立了在活体黄瓜下胚轴诱导产生转基因根的方法,红色荧光蛋白基因DsRed2可以作为报告基因来筛选转基因根与非转基因根,剪去非转基因根和主根后进行移栽种植,即为携带有转基因根而地上茎为非转基因的野生型黄瓜植株,该方法为大规模分析黄瓜根中相关基因的功能、地上-地下信号转导等研究奠定了基础。Cucumber(Cucumis sativus L.)is one of the species which is difficult to obtain transgenic plants by Agrobacterium tumefaciens-mediated transformation.Although the cucumber genome has been sequenced,the function of most genes has not been characterized.Agrobacterium rhizogenes-mediated hairy root transformation can efficiently induce plants to produce transgenic roots,providing a convenient and rapid approach for studying the biological functions of related genes in roots.In this study,an efficient and rapid transformation method mediated by Agrobacterium rhizogenes for inducing transgenic roots on living cucumber plants was established.A.rhizogenes K599 carrying red fluorescent protein(RFP)gene,DsRed2,was inoculated into hypocotyl of cucumber seedlings.The seedings inoculated were cultured about 2 weeks under the control of high humidity.Hairy roots were induced at the puncture sites of the hypocotyls in three different cucumber genotypes.Observation using red fluorescent protein and PCR verification indicated that approximately 89%to 94%of the punctured plants carried transgenic hairy roots,and the proportion of transgenic roots among the newly formed hairy roots was around 55%.The method of inducing transgenic roots from hypocotyls of cucumber in vivo was established,and the RFP gene,DsRed2,can be used as reporter gene to distinguish transgenic roots from non-transgenic roots.The cucumber plants with transgenic roots and wild-tpye shoot generated,which were transplanted after cutting off the non-transgenic roots and primary root,were suitable for the large-scale functional studies in root-related genes and root-shoot signal communication.

关 键 词：黄瓜 发根农杆菌 转基因根 红色荧光蛋白 

分 类 号：S642.2[农业科学—蔬菜学]