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以分别含有BSMV-CH基因组3个组分ds-cDNA全长的pMD-α、pMD-β和pMD-γ质粒为模板,PCR扩增BSMV-CH编码各蛋白基因,克隆至酵母双杂交载体,检测各蛋白是否存在自身激活特性,用酵母双杂交分析BSMV-CH编码蛋白之间的相互作用情况,测定β-半乳糖苷酶活性确认蛋白在酵母体内的作用,Western-blot分析蛋白的体外相互作用。结果表明成功克隆到BSMV-CH编码蛋白基因,并正确克隆至酵母双杂交载体,酵母双杂交检测未发现BSMV-CH编码蛋白之间的相互作用,但确定BSMV-CH编码的βC蛋白可以自身相互作用,体外可以单体或二聚体形式存在。
The full-length pMD-α, pMD-β and pMD-γ plasmids containing three components of the BSMV-CH genome were used as templates to amplify the BSMV-CH protein genes and cloned into the yeast two-hybrid vector. Detection of the presence or absence of self-activation of each protein, yeast two-hybrid analysis of BSMV-CH encoded protein interaction between the determination of β-galactosidase activity to confirm the role of the protein in yeast, Western-blot analysis of proteins in vitro effect. The results showed that the BSMV-CH encoding protein gene was cloned successfully and correctly cloned into the yeast two-hybrid vector. The yeast two-hybrid assay did not find any interaction between the BSMV-CH encoding proteins. However, it was confirmed that BSMV-CH encoded βC proteins could interact with each other Role, in vitro monomer or dimer form.