慢病毒介导的猫四连接素蛋白过表达系统的构建  

Construction of a lentiviral expression system for feline Tetranectin

作　　者：杨泽昊 胡佳艺 赵源杰 刘萌萌 陈宇 YANG Zehao;HU Jiayi;ZHAO Yuanjie;LIU Mengmeng;CHEN Yu(School of Tropical Agriculture and Forestry,Hainan university,Danzhou,Hainan 571737,China)

机构地区：[1]海南大学热带农林学院,海南儋州571737

出　　处：《热带生物学报》2025年第1期58-63,共6页Journal of Tropical Biology

基　　金：海南大学高层次人才科研启动费项目(XJ2400005267)。

摘　　要：四连接素(Tetranectin)是一种具有结合钙离子和肝素能力的纤溶酶原结合蛋白。通过构建稳定表达猫四连接素蛋白的细胞系,为后续探究其与猫心血管疾病的关系提供实验材料。本研究利用分子克隆技术构建过表达猫四连接素基因的重组慢病毒质粒,并通过慢病毒感染获得过表达猫四连接素蛋白的HEK293T细胞系,蛋白免疫印迹法验证蛋白表达。慢病毒质粒测序结果表明,实测猫四连接素基因CDS序列与NCBI数据库预测序列一致。经嘌呤霉素筛选过表达细胞系构建成功,猫四连接素重组蛋白以可溶形式分泌于细胞上清中,分子量约为25kDa并带有flag标签。Tetranectin is a plasminogen-binding protein with the ability to bind calcium ions and heparin.To establish a stable cell line expressing feline tetranectin and provide experimental materials for subsequent investigations into its association with feline cardiovascular diseases,we constructed a recombinant lentiviral plasmid overexpressing the feline tetranectin gene.A HEK293T cell line overexpressing feline tetranectin was obtained through lentiviral infection,and protein expression was verified by Western blotting.Sequencing of the lentiviral plasmid confirmed that the coding sequence of the feline tetranectin gene was consistent with the predicted sequence in the NCBI database.The overexpression cell line was successfully established after puromycin selection.The recombinant feline tetranectin protein was secreted into the cell supernatant in a soluble form with a molecular weight of approximately 25 kDa and a FLAG tag.

关 键 词：猫四连接素蛋白 CLEC3B 慢病毒包装系统 真核表达系统 

分 类 号：S852.23[农业科学—基础兽医学]