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为探讨甘蔗花叶病毒(Sugarcane mosaic virus,SCMV)CI基因的功能,对CI蛋白进行了免疫定位。利用RT-PCR方法,扩增甘蔗花叶病毒北京株系(SCMV Beijing isolate,SCMV-BJ)CI基因部分片段,将其连接到表达载体pET-28a上,重组质粒pETCIF1导入大肠杆菌BL21(DE3)pLysS中。SDS-PAGE分析表明,经IPTG诱导,CI基因获得了高效表达。将融合蛋白纯化后免疫兔子获得特异性较高的抗血清,利用该抗血清,对健康玉米和受病毒侵染的玉米茎、叶脉和叶片进行免疫金标记,在茎和叶脉韧皮部筛管的细胞壁处有金颗粒。
To investigate the function of CI gene of Sugarcane mosaic virus (SCMV), the CI protein was immunolocalized. A partial fragment of the CI gene of SCMV Beijing isolate (SCMV-BJ) was amplified by RT-PCR and ligated into the expression vector pET-28a. The recombinant plasmid pETCIF1 was introduced into E. coli BL21 (DE3) pLysS. SDS-PAGE analysis showed that the CI gene was highly expressed by IPTG induction. The fusion protein was purified and immunized into rabbits to obtain antiserum with higher specificity. The antisera was used to immunize the corn and virus infected corn stalks, veins and leaves, Department of gold particles.