烟草花叶病毒及其弱毒株基因组的cDNA克隆和序列分析

Genomic cloning and nucleotide sequence analysis of the cDNAs of Tobacco mosaic virus and its mild strains

利用RT PCR技术获得了覆盖整个烟草花叶病毒 (Tobaccomosaicvirus,TMV)强毒株TMV W、2个弱毒株TMV 0 17和TMV 15 2基因组 (RNA)的cDNA克隆。序列测定结果表明 ,强、弱毒株全长均为 6 395个核苷酸 ,具有 4个开放阅读框 (openreadingframe,ORF) ,分别编码 12 6、183、30、17.6kDa蛋白。TMV W和普通株系TMV U1的基因组同源率达到 98% ,TMV W为普通株系。TMV 0 17、TMV 15 2基因组发生变异的部位主要在 12 6 /183kDaORF中。和TMV W相比 ,TMV 0 17仅在 12 6kDa蛋白ORF中有 18个碱基发生变异 ,其中 10个碱基引起氨基酸的改变 ;而TMV 15 2在 183kDaORF中有 16个碱基发生变异 ,其中有 11个引起氨基酸的改变。TMV 0 17和TMV 15 2有 11个共同变异的碱基 ,其中有 8个引起氨基酸的变异。TMV 15 2在 30kDaORF中有 1个碱基发生变异而引起氨基酸的改变。其它区域 ,三者完全相同。

The genomic RNAs of one virulent strain and two mild strains of Tobacco mosaic virus (TMV) were cloned and sequenced. Results showed that the genomic RNAs of all the three strains were 6 395 nt in length, with 4 open reading frames (ORFs) which encoded proteins of 126, 183, 30 and 17.6 kDa respectively. The nucleotide sequence homology between TMV W (virulent strain) and TMV U1 was 98%, indicating that TMV W was also a common strain. Compared with TMV W, the nucleotide sequence variations of the 2 mild strains (TMV 017 and TMV 152) located mainly in the ORFs of 126 or 183 kDa. In TMV 017, variations only occurred in the ORF of 126 kDa, where 18 nucleotide mutations caused the alteration of 10 amino acids. There were 16 nucleotide mutations in the 183 kDa ORF of TMV 152, 11 of which caused change of predicted proteins. There were 11 common nucleotide mutations in 126 kDa ORFs of TMV 017 and TMV 152, 8 of which caused the alteration of amino acid. In addition, there was 1 nucleotide mutation that caused the alteration of 1 amino acid in 30 kDa ORF of TMV 152 when compared to that of TMV W. The other regions were identical among the three strains.