【摘要】 应用反转录-PCR技术合成并扩增了水稻矮缩病毒(RDV)中国福建分离物基因组第七号片段(S7)cDNA,将PCR产物克隆在载体 pBluescript SK(+)上,进行了亚克隆和序列分析。结果表明克隆片段全长1578bp,含有一个1518bp长的开放阅读框架(ORF),编码一个由506个氨基酸组成的理论分子量为56000的蛋白。该片段与RDV日本分离物基因组的相应片段相比,在核苷酸及氨基酸水平上的同源率分别为93.2％和94.1％。该片段编码的56000蛋白与同属的伤瘤病毒(WTV)基因组第七号片段编码的57000蛋白相比,在靠近N端的区域(aa61～aa141)有较高的氨基酸序列同源性。将RDV S7 cDNA克隆到原核表达载体 pBV221上,通过温度诱导在大肠杆菌中得到表达。表达产物占菌体可溶性总蛋白的13.73％。对表达产物进行了 Western blotting分析。本工作为进一步纯化RDV S7编码蛋白、分析其功能和转化水稻打下了良好基础。更多还原

【Abstract】 The cDNA of segment 7 (S7) of Rice Dwarf Virus (RDV) genome of Chinese . Fujian isolate was synthesized and amplified by RT-PCR. The PCR product was cloned into pBluescript SK(+)and sequenced. The cloned segment is 1578bp in length, contains an single open reading frame with relative molecular weight of 56 000. Comparison of nucleotide and amino acid sequence between this segment and that of RDV genome of Japanese isolate showed 93.2% and 94.1% homology, respectively. The RDV S7 coded 56 000 protein has high amino acid identity in the region near N-termini (aa61-aa141) to the 57 000 protein encoded by segment 7 of the Wound Tumor Virus (WTV) genome. The RDV S7 cDNA was cloned and highly expressed in pBV221 in E. coli. The expressed recombinant RDV S7 -coded protein accounted for 13.73% of the total soluble cell proteins and futher analyzed by Western blotting.更多还原