Cry2Aa2 和PamPAP 双价表达载体的构建及其对辣椒的遗传转化

园艺学报 ›› 2012, Vol. 39 ›› Issue (7): 1285-.

Cry2Aa2 和PamPAP 双价表达载体的构建及其对辣椒的遗传转化

高玉尧, 陈长明, 陈国菊, 曹必好, 雷建军

华南农业大学园艺学院，广州 510642；国家航天育种工程研究中心，广州 510642

出版日期:2012-07-25 发布日期:2012-07-25

Construction of Binary Expression Vector Containing Cry2Aa2 and PamPAP and Its Transfer to Pepper

GAO Yu-Yao, CHEN Chang-Ming, CHEN Guo-Ju, CAO Bi-Hao, LEI Jian-Jun

College of Horticulture，South China Agricultural University，Guangzhou 510642，China；National Engineering Research Center for Space Breeding，Guangzhou 510642，China

Online:2012-07-25 Published:2012-07-25

摘要/Abstract

摘要： 为了获得既抗虫又抗病毒病，又对人体和环境安全的转基因辣椒（Capsicum annuum L．）材料，将启动子CaMV35S、终止子Nos-ter 及缺失无毒型商陆抗病毒蛋白基因PamPAP 一起插入到植物表达载体pCAMBIA1301-Cry2Aa2-PMI 的多克隆位点上，构建双价表达载体，采用农杆菌介导法将其转入辣椒中，并用PMI/甘露糖筛选体系对转化植株筛选。经PCR 扩增和Southern 杂交检测，结果表明Cry2Aa2和PamPAP 已经整合到辣椒基因组中。进一步经RT-PCR 分析，证实Cry2Aa2 和PamPAP 在T₀代转基因辣椒植株中共表达。经抗虫性、抗病毒病表型鉴定：T₀代转基因辣椒植株对黄瓜花叶病毒（CMV）和斜纹夜蛾幼虫的抗性均显著高于非转基因植株。

关键词: 辣椒, PMI/甘露糖筛选体系, Cry2Aa2, PamPAP, 遗传转化, 抗病毒, 抗虫害

Abstract: The aim of this study was to obtain transgenic pepper plants with double resistances to virus and insect that could be used in developing new pepper variety. The non-virulent defective PAP gene （PamPAP），CaMV35S promoter and Nos terminal were inserted into MCS of pCAMBIA1301- Cry2Aa2-PMI. Cry2Aa2 and PamPAP were transferred into pepper（Capsicum annuum L.）mediated by Agrobacterium tumefaciens with PMI/Man selection system．The results of PCR and Southern blot indicated that Cry2Aa2 and PamPAP had been integrated into the pepper genome. RT-PCR analysis further showed that the Cry2Aa2 and PamPAP had been expressed simultaneously. In addition，resistances to virus and insect were assayed，and the results showed that the positive transgenic pepper plants had higher degree of resistances to CMV and Prodenia litura larva than the non-transgenic plants. This suggested that transgenic pepper plants with double resistances to virus and insect could be obtained by transferring Cry2Aa2 and PamPAP.

Key words: pepper, PMI/Man system, Cry2Aa2, PamPAP, genetic transformation, resistance to virus, resistance to insect

中图分类号:

S 641.3

高玉尧, 陈长明, 陈国菊, 曹必好, 雷建军. Cry2Aa2 和PamPAP 双价表达载体的构建及其对辣椒的遗传转化[J]. 园艺学报, 2012, 39(7): 1285-.

GAO Yu-Yao, CHEN Chang-Ming, CHEN Guo-Ju, CAO Bi-Hao, LEI Jian-Jun. Construction of Binary Expression Vector Containing Cry2Aa2 and PamPAP and Its Transfer to Pepper[J]. ACTA HORTICULTURAE SINICA, 2012, 39(7): 1285-.

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