番茄斑萎病毒N基因克隆及其植物表达载体的构建

目的对从云南楚雄番茄染病材料上得到病毒分离物进行鉴定,建立病毒N基因植物表达载体,以便进行抗病育种研究。方法利用RT-PCR技术克隆得到N基因,全长777 bp,将克隆得到的TSWV-N基因连接至质粒p BI121重组植物表达载体p BI121-TSWV-N。结果 TSWV-N基因与NCBI上已登记的番茄斑萎病毒中国云南分离物同源性达到97%~100%。结论所分离病毒确为番茄斑萎病毒,包含有TSWV-N基因的植物表达载体构建成功。

Cloning of N gene of Tomato spotted wilt virus and construction of its plant expression vectors

Objective To detect and identify the virus from infected tomato plants in Chuxiong, Yunnan province, and construct the recombinant plasmid with the gene N for expressing in plant. Methods The nucleotide sequences of nucleocapsid protein (N) gene of Tomato spotted wilt virus (TSWV) were determined by RT-PCR with the length of 777 bp. Then the cloned TSWV-N gene was connected to pBI121-TSWV-N, which was the recombinant plasmid plant expression vector of pBI121. Results Compared with sequences of corresponding viruses from Yunnan in China that were available in the GeneBank, the N gene of TSWV-Chuxiong shared similarity of 97%~100% at nucleotide level, were ligated into the plant expression vectors, PBI121. Conclusion The virus was identified as TSWV, and the recombinant plasmid was constructed successfully by restriction enzyme analysis.