PEDV nsp15基因真核表达载体的构建及蛋白的表达  

Construction of Eukaryotic Expression Vector and Protein Expression of PEDV Nsp15 Gene

作　　者：张宁 郑亮[1,2] 杨超玥 吴志军 张华[1] 曹宏伟[1] 肖莉杰[1] Zhang Ning;Zheng Liang;Yang Chaoyue;Wu Zhijun;Zhang Hua;Cao Hongwei;Xiao Lijie(College of Life Science and Technology,Heilongjiang Bayi Agricultural University,Daqing163319;College of Animal Science and Technology,Heilongjiang Bayi Agricultural University)

机构地区：[1]黑龙江八一农垦大学生命科学技术学院,大庆163319 [2]黑龙江八一农垦大学动物科技学院

出　　处：《黑龙江八一农垦大学学报》2023年第1期72-77,共6页journal of heilongjiang bayi agricultural university

基　　金：黑龙江八一农垦大学博士科研启动基金(XDB2015-18)。

摘　　要：猪流行性腹泻病毒(porcine epidemic diarrhea virus,PEDV)含有一种名为Nsp15的非结构蛋白,为了研究Nsp15蛋白在侵染宿主细胞时产生的作用,成功构建了PEDV Nsp15基因的真核表达载体。通过PCR技术扩增Nsp15基因片段,经双酶切连接到pCMV-Myc载体上,酶切鉴定及测序结果证实PEDV Nsp15基因成功克隆到pCMV-Myc载体中,并将重组质粒命名为p CMV-Myc-Nsp15。将重组质粒pCMV-Myc-Nsp15转染至IPEC-J2细胞中,利用Western Blot和间接免疫荧光技术检测pCMV-Myc-Nsp15的表达以及定位情况。结果显示,pCMV-Myc-Nsp15在IPEC-J2细胞中成功表达,并且在胞质胞核均匀分布。为进一步研究PEDV Nsp15蛋白的功能奠定基础。Porcine epidemic diarrhea virus(PEDV)contains a nonstructural protein called Nsp15.In order to study the role of Nsp15protein in the infection of host cells,the eukaryotic expression vector of PEDV Nsp15 gene was constructed.Nsp15 gene fragment was amplified by PCR,and was linked to pCMV-myc vector by double enzyme digestion.The results of enzyme digestion identification and sequencing confirmed that PEDV Nsp15 gene was cloned into pCMV-Myc vector,and the recombinant plasmid was named pCMV-Myc-Nsp15.The recombinant plasmid pCM-Myc-Nsp15 was transfected into IPEC-J2,and the expression and localization of pCM-Myc-Nsp15 were detected by Western Blot and immunofluorescence.The results showed that pCMV-Myc-Nsp15 was expressed in IPEC-J2 cells and uniformly distributed in the cytoplasm and nucleus.

关 键 词：猪流行性腹泻病毒 真核表达载体 非结构蛋白15 

分 类 号：S858.26[农业科学—临床兽医学] Q786[农业科学—兽医学]