2019-nCoV N蛋白在家蚕杆状病毒表达系统中的表达及纯化  

Expression and Purification of 2019-nCoV Nucleocapsid Protein by Silkworm Baculovirus Expression System

作　　者：海毕瑞 杨扬[1] 刘世火 吴峻 徐家萍[1] Hai Birui;Yang Yang;Liu Shihuo;Wu Jun;Xu Jiaping(Anhui Key Laboratory of Resource Insect Biology and Innovative Utilization,School of Life Sciences,Anhui Agricultural University,Hefei 230036,China;Shanghai Institute of Biomedical Technology,Shanghai 200032,China)

机构地区：[1]安徽农业大学生命科学学院资源昆虫生物学与创新利用安徽省重点实验室,合肥230036 [2]上海市生物医药技术研究院,上海200032

出　　处：《蚕业科学》2023年第4期333-339,共7页ACTA SERICOLOGICA SINICA

基　　金：安徽省国际科技合作项目(202104b11020002)。

摘　　要：核衣壳(nucleocapsid,N)蛋白是新型冠状病毒的结构蛋白,与病毒RNA的转录和复制有关。目前,N蛋白已被广泛应用于新型冠状病毒肺炎的检测诊断及治疗。本研究克隆的2019-nCoV-n基因CDS由1257个碱基构成,编码419个氨基酸。将该基因连接到昆虫表达载体pFastBacTMHTB构建重组Bacmid,转染至BmN细胞中获得重组病毒,将重组病毒注射家蚕蛹体内,收集发病家蚕蛹血淋巴,通过镍柱亲和纯化,SDS-PAGE和Western blot检测表明获得一定纯度具有活性的重组2019-nCoV-N蛋白,为后期进一步开发抗体检测试剂盒鉴定了基础。Nucleocapsid protein is a structural protein of novel coronavirus which is related to the transcription and replication of viral RNA.At present,N protein has been widely used in the detection,diagnosis,and treatment of Corona Virus Disease 2019(COVID-19).CDS of 2019-nCoV-n gene cloned in this study consisted of 1257 bases and encoded 419 amino acids.This gene was constructed to the insect expression vector pFastBacTMHTB to obtain recombinant Bacmid,which was then transfected into BmN cells to obtain recombinant virus.The recombinant virus was injected into the silkworm pupa.The hemolymph of the infected silkworm pupa was collected and purified by nickel column affinity.SDSPAGE and Western blot results showed that recombinant 2019-nCoV-N protein with certain purity and activity was obtained,which provided the basis for further development of antibody detection kit in the future.

关 键 词：杆状病毒表达系统 家蚕蛹 2019-nCoV 核衣壳蛋白 蛋白质纯化 

分 类 号：S3[农业科学—农艺学]