CAJ | 학술논문

为获得具有活性的猪繁殖与呼吸障碍综合征病毒( PRRSV)非结构蛋白1α( nsp1α),以BJ-4 PRRSV nsp1α真核表达质粒pcDNA3.1-nsp1α为模板扩增nsp1α,克隆到原核表达载体pET -28a,1 mmol/L异丙基硫代半乳糖苷(IPTG)诱导含有重组质粒的大肠杆菌BL21(DE3)。 SDS–PAGE结果显示,nsp1α以包涵体形式高效表达,重组蛋白大小约为20 ku；Western blot结果显示,表达的重组蛋白能与小鼠抗His标签单克隆抗体反应；包涵体蛋白复性后,ELISA方法检测结果显示,重组蛋白能与1∶6400稀释的PRRSV阳性血清反应。表明成功表达了nsp1α蛋白,且表达的重组蛋白具有较好的免疫学活性。
위획득구유활성적저번식여호흡장애종합정병독( PRRSV)비결구단백1α( nsp1α),이BJ-4 PRRSV nsp1α진핵표체질립pcDNA3.1-nsp1α위모판확증nsp1α,극륭도원핵표체재체pET -28a,1 mmol/L이병기류대반유당감(IPTG)유도함유중조질립적대장간균BL21(DE3)。 SDS–PAGE결과현시,nsp1α이포함체형식고효표체,중조단백대소약위20 ku；Western blot결과현시,표체적중조단백능여소서항His표첨단극륭항체반응；포함체단백복성후,ELISA방법검측결과현시,중조단백능여1∶6400희석적PRRSV양성혈청반응。표명성공표체료nsp1α단백,차표체적중조단백구유교호적면역학활성。
In order to obtain activated nonstructural protein 1α( nsp1α) of porcine reproductive and res-piratory syndrome virus (PRRSV),the nsp1α gene was amplified from the expression plasmid pcDNA3. 1-nsp1α of PRRSV BJ-4 strain and was subcloned into prokaryotic expression vector pET-28a. Then the pET-28a-nsp1α was transformed into E. coli BL21 (DE3). The E. coli BL21 (DE3) was induced by 1 mmol/L IPTG. SDS-PAGE result showed that the nsp1αwas mainly expressed in form of inclusion body and the molecular weight was 20 ku. Western blot result showed that nsp1αcould recognized with His-tag mouse Mcab. ELISA result showed that nspld could react with the PRRSV porcine positive serum which was diluted to 1∶6 400 . In this research the PRRSV nsp1α was expressed successfully and had good im- munological activity.