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应用RT-PCR技术克隆了水稻瘤矮病毒(Rice gall dwarf virus,RGDV)广东分离物基因组的第10片段,并测定了全序列。结果表明,RGDV广东分离物S10(登录号EF532325)全长1 198 bp,含有一个ORF,编码一条由320氨基酸组成、推测分子量约36 kDa的多肽。与泰国分离物相应组分相比,基因结构基本一致,核苷酸和氨基酸序列同源性分别为96.2%和98.8%;S10编码多肽与水稻矮缩病毒(Rice dwarf virus,RDV)S9编码蛋白及伤瘤病毒(Wound tumor virus,WTV)S11编码蛋白也分别具有29%和33%的相似性。本研究还将S10 cDNA克隆至原核表达载体pET28b(+)上,通过IPTG诱导在大肠杆菌BL21(DE3)中得到了高效表达,并利用His. Bind树脂纯化得到电泳纯级制品。本工作为进一步研究S10编码蛋白的结构与功能奠定了一定的基础。
The tenth fragment of genome of rice gall dwarf virus (RGDV) Guangdong isolate was cloned by RT-PCR and sequenced. The results showed that RGDV Guangdong isolate S10 (accession number EF532325) was 1 198 bp in length and contained an ORF encoding a polypeptide of 320 amino acids with a predicted molecular weight of about 36 kDa. Compared with the corresponding components of the Thai isolate, the gene structure was basically the same, and the nucleotide and amino acid sequence homologies were 96.2% and 98.8% respectively. The encoded protein of S10 and rice dwarf virus (RDV) And Wound tumor virus (WTV) S11 encoded protein also have 29% and 33% similarity, respectively. In the present study, S10 cDNA was cloned into the prokaryotic expression vector pET28b (+) and expressed in E.coli BL21 (DE3) induced by IPTG. The purified product was purified with His. Bind resin. This work laid a solid foundation for further study on the structure and function of S10-encoded protein.