水稻黑条矮缩病毒基因组组分10编码的外壳蛋白基因的克隆及表达

从我国山东发病的玉米材料中提取水稻黑条矮缩病毒 ,抽提病毒RNA ,经RT PCR ,克隆了编码外层外壳蛋白的基因组组分 10 (S10 )的cDNA ,并进行了序列测定 .与已报道的日本株和湖北等地的S10进行了序列同源性比较 .结果表明 ,与日本株的同源性为 92 % ,与湖北等地的同源性在 97%～ 98%之间 .将该序列构建到pGEX 3X表达载体中 ,经IPTG诱导 ,表达了分子质量约为 76ku的GST融合蛋白 .经亲和层析纯化和Western印迹分析 ,证实了该基因以可溶性的GST融合蛋白形式在原核中表达 .

Cloning and Expression of Genome Segment 10 Gene Encoding Capsid Protein of Rice Black Streaked Dwarf Virus

The rice black streaked dwarf virus (RBSDV) and its RNA were isolated from infected maize plants from Shandong in China. The genome segment 10 (S10) gene encoding outer capsid protein was obtained by RT PCR and its sequence was compared with that in strains from Japan, Hubei and other areas in China. The homology of S10 between Shandong and Japanese strain was 92%, much lower than that with Chinese strains ranging from 97% to 98%. The expression vector pGEX S10 was constructed with S10 and pGEX 3X. When the E. coli strain BL21 harboring the pGEX S10 was induced by IPTG for 6 hours, SDS PAGE analysis showed that a 76 ku protein was expressed. This 76 ku protein was purified by Glutathione Sepharose 4B and western blot analysis. It is confirmed that the RBSDV S10 gene was expressed as GST fusion protein in E. coli .