| 摘要: |
| 【目的】利用农杆菌介导法将抗 CMV 和 ToMV 的RNAi表达载体 pBi35STC12 转入红番3号加工番茄，获得抗 CMV 和 ToMV 的加工番茄植株，为加工番茄病毒病的防治奠定基础。【方法】采用 RT-PCR 扩增，选取新疆加工番茄 CMV 分离物 NS04 1a复制酶第1 051-1 350 bp 序列和 ToMV 分离物 SCS-2 130/180 ku复制酶第1 920-2 200 bp序列，作为干扰片段 CR12 和 To12。通过T克隆载体将CR12和To12连接成拼接片段TC12，构建成含反向重复结构拼接片段的RNAi表达载体 pBi35STC12。通过农杆菌介导法，用其转化红番3号加工番茄，经过筛选、PCR检测得到转基因植株。【结果】成功构建了抗 CMV 和 ToMV 的RNAi表达载体pBi35STC12，并用其转化番茄植株，从转化的1 500个愈伤组织中获得33株再生植株，利用 PCR 对再生植株进行检测，显示从33株再生植株中获得了19株转基因植株，转化率为1.26%。【结论】构建了抗CMV和ToMV的RNAi表达载体 pBi35STC12，用其转化红番3号加工番茄，获得了转基因植株。 |
| 关键词: CMV ToMV RNAi载体 加工番茄 遗传转化 |
| DOI： |
| 分类号: |
| 基金项目:国际科技合作与交流专项(20072072)；国家自然科学基金项目(31260420) |
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| Construction of RNAi vector with resistance to Cucumber mosaic virus and Tomato mosaic virus and genetic transformation of processing tomato |
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QIAO Ya-hong,TIAN Gui-ying,ZHENG Yin-ying,et al
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| Abstract: |
| 【Objective】The purpose of the study was to obtain transgenic processing tomato plant with the resistance to Cucumber mosaic virus (CMV) and Tomato mosaic virus (ToMV) by Agrobacterium mediated transformation,and improve the prevention and control of virus disease.【Method】The 1 051-1 350 bp sequence of 1a replicase cloned from CMV isolate NS04 and the 1 920-2 200 bp sequence of 130/180 ku replicase cloned from ToMV isolate SCS2 of Xinjiang processing tomato amplified by RT-PCR were used as interference fragments CR12 and To12.The two fragments CR12 and To12 were spliced into one fragment through pGEM-T easy.The recombinant plant expression vector pBi35STC12 containing inverted repeat fragments was constructed and introduced into processing tomato Hongfan 3 by Agrobacterium mediated transformation.Then the transgenic plants were identified by PCR.【Result】The RNAi vector pBi35STC12 was successfully constructed and transformed to the processing tomato.33 regenerated plants were obtained from 1 500 callus of processing tomato Hongfan 3.19 of them were identified to be transgenic plants by PCR and the transformational rate was 1.26%.【Conclusion】The RNAi vector pBi35STC12 was constructed and successfully transformed to the processing tomato Hongfan 3. |
| Key words: Cucumber mosaic virus Tomato mosaic virus RNAi vector processing tomato genetic transformation |
