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中国农学通报

中国农学通报 ›› 2005, Vol. 21 ›› Issue (8): 28-28.

所属专题： 生物技术； 农业工程； 园艺； 烟草种植与生产

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番茄双抗烟草花叶病毒和黄瓜花叶病毒的基因工程研究

廖俊杰,李进进,许继勇   

  • 出版日期:2005-08-05 发布日期:2005-08-05

Studies on Transgenic Tomato Plants with Resistance to Tobacco Mosaic Virus and Cucumber Mosaic Virus

Liao Junjie , Li Jinjin, Xu Jiyong   

  • Online:2005-08-05 Published:2005-08-05

摘要： 在烟草花叶病毒（TMV）外壳蛋白（CP）基因的5′和3′端分别合成寡聚核苷酸T5和T3，并分别引入ClaⅠ和BamHⅠ位点，采用PCR技术扩增得到TMV CP基因，用ClaⅠ / BamHⅠ消化后重组到pBluescript KS+中，并将该基因与黄瓜花叶病毒（CMV）CP基因重组在同一个表达载体pKYLX7上获得双价CP基因表达载体pKDTC，并转入根癌农杆菌LBA4404中，通过“叶盘法”转化番茄“丽春”、“京韩一号”，获得了番茄再生植株。通过点杂交、PCR检测和Southern杂交，证实“丽春”的L-2、L-5、“京韩一号”的JH-6号为转TMV CP和CMV CP基因的工程植株。工程植株在室温中表现正常，与对照植株相比，转基因植株表现明显的抗TMV、CMV性能。

Abstract: The TMV CP gene was synthesized with Polymerase Chain Reaction (PCR) y using the cDNA of its genomic RNA as a template. A synthetic ClaⅠsite was included in T5 primer while a BamHⅠsite in T3 primer, the PCR product was cloned into the ClaⅠ/BamHⅠ site of pBluescript KS+. Then bivalent plant expression vector, pKDTC (TMV-CP+CMV-CP), with double resistance to TMV and CMV, was constructed, in which each CP gene was controlled by a 35S promotor of CaMV. Transgenic tomato plants were regenerated via Agrobacterium transformation system, and analyzed by dot blot, PCR and Southern blot, these results showed that both genes were integrated into the transgenic plant lines L-2, L-5 of Lichun and JH-6 of Jinghan No 1. The transgenic plants grew normally and obviously resisted to the infection of corresponding viruses in greenhouse.