香石竹斑驳病毒（CarMVsh）外壳蛋白基因克隆及原核表达

香石竹斑驳病毒上海分离株是从上海地区栽培的香石竹上分离并鉴定的，以提纯的病毒为材料，ＳＤＳ－酚法纯化的基因组ＲＮＡ作为模板，ＲＴ－ＰＣＲ合成并扩增外壳蛋白基因ｃＤＮＡ，ｃＤＮＡ克隆于ｐＧＥＭ－Ｔ ｅａｓｙ ｖｅｃｔｏｒ，转化为Ｅ．ｃｏｌｉＪＭ１０９。阳笥克隆ｐＴＣａＣＰ经序列分析，证明带有全长ＣＰ基因。

Cloning and Expression of Coat Protein Gene of Carnation Mottle Virus Shanghai Isolate in E. coli

An Shanghai isolate of Carnation mottle virus (CarMVsh) was isolated from carnations in Shanghai and identified by its particle shape and size, susceptibility to indicator plants and the reactions to specific antisera. Genomic RNA extracted from virus preparation was employed as template to clone cDNA of coat protein gene by RT PCR. PCR products were purified with agarose gel and cloned into pGEM T easy vector to construct the recombinant plasmid pTCaCP for nucleotide sequencing. The homology of the nucleotide sequence is 96.46% to CarMVsh reported previously by Guilley et al . The plasmids of pTrc99A and pBAD/His B were used to construct CP gene expression vector. The CP gene was expressed in E. coli Top10 strain transformed by the recombinant pBHCaCP vector, but failed with the recombinant pTrCaCP vector. The product was identified by SDS PAGE and Western Blot with CarMVsh antiserum, and showed to be CarMVsh CP with M r 38 000.