霍乱毒素B亚单位植物细胞表达载体的构建及其在人参细胞中的表达  

Construction of Plant Expression Vector for Cholera Toxin B Subunit and ItsExpression in Ginseng Cells

作　　者：任琦[1,2] 宋之明[3] 刘丹[2] 孙洋[4] 于海鹏[2] 李娟[2] 富锐丽[2] 刘英[2] 盛军[1,2] 

机构地区：[1]吉林大学生命科学学院,长春130012 [2]长春生物制品研究所,长春130062 [3]吉林大学第一医院骨科,长春130021 [4]吉林大学白求恩医学院免疫学教研室,长春130021

出　　处：《中国生物制品学杂志》2009年第7期663-666,共4页Chinese Journal of Biologicals

摘　　要：目的构建霍乱毒素B亚单位(CTB)植物细胞表达载体,并在人参细胞中进行表达。方法根据人参偏爱密码子,采用引物延伸PCR法合成CTB基因,连入pBI121质粒,构建植物细胞表达载体,转化人参细胞后,采用PCR、RT-PCR和Western blot进行鉴定。结果测序结果表明,PCR法合成的目的基因序列与设计完全一致,构建的植物细胞表达载体经双酶切鉴定显示,所含基因片段大小与预期相符。提取转基因人参细胞基因组DNA和mRNA,分别进行PCR和RT-PCR鉴定,均可见约400 bp的特异性片段。Western blot分析可见相对分子质量约12 000的特异性条带。结论已成功构建了含霍乱毒素B亚单位的植物细胞表达载体,并在人参细胞中获得表达。Objective To construct a plant expression vector for cholera toxin B subunit （CTB） and express in ginseng cells. Methods CTB gene was synthesized by primer extension PCR using the primers designed according to ginseng cell-preferred codon and inserted into plasmid pBI121. The constructed recombinant plasmid was transformed to ginseng cells, and the transformants were identified by PCR, RT-PCR and Western blot. Results The sequence of amplified target gene was completely identical to that designed. The restriction analysis of constructed plant expression vector proved that the length of inserted gene fragment was consistent with that expected. Either PCR result of genomic DNA or RT-PCR result of mRNA of the transformed ginseng cells showed a specific gene fragment at length of about 400 bp. Western blot showed a specific band with relative molecular mass of about 12 000. Conclusion A plant expression vector for CTB was successfully constructed and expressed in ginseng cells.

关 键 词：霍乱毒素B亚单位(CTB) 植物细胞表达载体 人参细胞 转基因 

分 类 号：Q786[生物学—分子生物学]