摘要
根据烟草花叶病毒Ｕ１株系ＴＭＶ－Ｕ１序列，人工合成引物，通过ＰＣＲ扩增并克隆了烟草花叶病毒蚕豆株系（ＴＭＶ－Ｂ）的外壳蛋白（ＣＰ）基因和３′端非编码区．ＤＮＡ全序列测定结果表明，外壳蛋白基因全长４５９个碱基，编码１５１个氨基酸，３′端非编码区全长２２４个碱基．与ＴＭＶ－Ｕ１株系相比，ＴＭＶ－Ｂ仅在ＣＰ基因上有一个碱基缺失，并导致ＣＰ基因提前终止，使其编码的氨基酸少７个．将ＣＰ基因及３′端非编码区插入ｐＧＥＭＥＸ－１的Ｔ７基因１０中，转入Ｅ．ｃｏｌｉ后诱导表达，聚丙烯酰胺凝胶电泳分析呈现一特异的蛋白带．Ｗｅｓｔｅｒｎ免疫检测证明，该特异带与ＴＭＶ抗血清呈阳性反应．将ＣＰ基因正向插入植物表达载体ＰＢⅠ１２１中，置于花椰菜花叶病毒３５Ｓ启动子控制之下。
The coat protein gene and the 3′ unencoded end of broad bean strain of TMV is cloned and amplified by PCR (Polymerase Chain Reaction).The molecular length of CP gene is 0.7 kb.There is a deletion of basepair which causes a loss of 7 amino acid.The CP gene is inserted into the T 7 gene 10 of pGEMEX 1 and transformed to the competent cells of E.coli.A specific protein presents on the polyacrilomide gel.Western blot indicates a specific reaction of this protein with the antiserum of TMV.Then the CP gene is inserted into the pBⅠ121 under the control of 35S promoter.
出处
《湖南农业大学学报（自然科学版）》
CAS
CSCD
1997年第1期1-8,共8页
Journal of Hunan Agricultural University(Natural Sciences)
基金
本研究系国际科学基金
关键词
遗传工程
烟草花叶病毒
外壳蛋白基因
蚕豆
genetic engineering tobacco mosaic virus sequences analysis / coat protein gene 3′ unencoded end expression
