摘要
设计特异引物,以猪圆环病毒2型(PCV-2)杭州株HZ0201的基因组DNA为模板,PCR扩增出ORF3基因,构建了pGEX-4T-1-ORF3原核表达载体和pEGFP-C2-ORF3真核表达载体。ORF3基因全长315 bp,编码105个氨基酸。SDS-PAGE、Western blot分析及真核PK15细胞转染结果显示:ORF3蛋白在大肠杆菌中以包涵体形式存在,分子量大小约为37.7 ku;ORF3重组蛋白在真核PK15细胞的细胞核和细胞浆都有表达,尤其在细胞核中表达量较高,且对细胞有一定的毒性。
The specific primers were designed and synthesized according to the published sequences of ORF3 gene of porcine circovirus type 2(PCV-2).The complete DNA fragment encoding ORF3 protein was obtained from viral DNA of PCV-2 HZ0201 by PCR and subcloned into prokaryotic expression vector pGEX-4T-1 and eukaryotic expression vector pEGFP-C2.ORF3 gene encoding 105 amino acids was 315 bp in length.The analysis of SDS-PAGE,Western blot and transfection assays showed that the ORF3 recombinant protein with the molecular weight of 37.7 ku mainly existed as inclusion body in E.coli;the recombinant protein was expressed in cellular nucleus and cytoplasma of PK-15 cells and showed some toxicity to PK-15 cells.
出处
《畜牧与兽医》
北大核心
2008年第9期25-30,共6页
Animal Husbandry & Veterinary Medicine
关键词
猪圆环病毒2型
ORF3
原核表达
真核表达
porcine circovirus type 2
ORF3
prokaryotic expression
eukaryotic expression
