水稻瘤矮病毒广东分离物基因组第10组分的序列分析及原核表达

 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1198bp,含有一个ORF,编码一条由320氨基酸组成、推测分子量约36kDa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S10cDNA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。

Sequencing and prokaryotic expression of segment 10 of Rice gall dwarf virus Guangdong isolate

The full length cDNA of S10 sequence of Rice gall dwarf virus (RGDV) Guangdong isolate was cloned into pMD18-T vector and the complete nucleotide sequence was determined. The results showed that S10 had a length of 1198 nt encoding a polypeptide of 320 amino acids with a Mr of 36 kDa. Sequence comparison showed that the S10 of RGDV Guangdong isolate had the same genome organization with the Thailand isolate, and shared 96.2% nucleotide and 98.8% amino acid sequence identities. The predicted polypeptide of RGDV $10 also shared 29% and 33% identities respectively with the proteins encoded by Rice dwarf virus (RDV) S9 and Wound tumor virus (WTV) S11. The RGDV S10 cDNA was cloned to pET28b (+) and highly expressed in Escherichia coli BL21 (DE3), and the expressed recombinant protein was purified by His. Bind column. These works will promote the structural and functional research of RGDV S10 protein.