Avian influenza virus (AIV) is a highly variable pathogen. The M2 gene of AIV is a transmembrane protein. It has highly conservative antigenic epitopes and is a potential antigen for cross-protection. In this study, t...Avian influenza virus (AIV) is a highly variable pathogen. The M2 gene of AIV is a transmembrane protein. It has highly conservative antigenic epitopes and is a potential antigen for cross-protection. In this study, the recombinants containing the full-length M2 and the transmembrane segment deleted M2 were constructed respectively. When measuring the expression of the two constructs, we found that the full-length M2 failed to express in the prokaryotic expression system, whereas transmembrane segment deleted M2 was highly expressed as fusion protein in a soluble form. The fusion protein reacted with SPF chicken serum specific for AIV A/goose / Guangdong /1996(H5N1). The protein was purified by GST purification system and the purified protein was used to prepare anti-M2 serum in mice. Immunofluorescence test demonstrated the binding of the antiserum with AIV (H5N1) infected MDCK cells. The results suggest that the recombinant transmembrane segment deleted M2 could provide good antigenicity.展开更多
目的:构建头孢菌素C酰化酶(Cephalosporin C Acylase)基因的两种原核表达载体,确定高效原核表达体系。方法:人工合成头孢菌素C酰化酶S12基因,分别克隆到含有Lac启动子的载体pBC KS和T7-lac启动子的载体pET-28a上,命名为pBC-S12和pET-28-...目的:构建头孢菌素C酰化酶(Cephalosporin C Acylase)基因的两种原核表达载体,确定高效原核表达体系。方法:人工合成头孢菌素C酰化酶S12基因,分别克隆到含有Lac启动子的载体pBC KS和T7-lac启动子的载体pET-28a上,命名为pBC-S12和pET-28-S12,重组质粒分别转化感受态大肠杆菌DH5α和BL21(DE3),表达纯化目的蛋白S12,测定酶活。结果:成功地构建了头孢菌素C酰化酶S12的原核表达菌株,组成型表达和诱导型表达体系的表达量低,目的蛋白具有酶活,诱导型表达产生的酶比组成型表达产生的酶具有更强活性,可作为大肠杆菌表达头孢菌素C酰化酶的高效表达体系。展开更多
文摘Avian influenza virus (AIV) is a highly variable pathogen. The M2 gene of AIV is a transmembrane protein. It has highly conservative antigenic epitopes and is a potential antigen for cross-protection. In this study, the recombinants containing the full-length M2 and the transmembrane segment deleted M2 were constructed respectively. When measuring the expression of the two constructs, we found that the full-length M2 failed to express in the prokaryotic expression system, whereas transmembrane segment deleted M2 was highly expressed as fusion protein in a soluble form. The fusion protein reacted with SPF chicken serum specific for AIV A/goose / Guangdong /1996(H5N1). The protein was purified by GST purification system and the purified protein was used to prepare anti-M2 serum in mice. Immunofluorescence test demonstrated the binding of the antiserum with AIV (H5N1) infected MDCK cells. The results suggest that the recombinant transmembrane segment deleted M2 could provide good antigenicity.
文摘目的:构建头孢菌素C酰化酶(Cephalosporin C Acylase)基因的两种原核表达载体,确定高效原核表达体系。方法:人工合成头孢菌素C酰化酶S12基因,分别克隆到含有Lac启动子的载体pBC KS和T7-lac启动子的载体pET-28a上,命名为pBC-S12和pET-28-S12,重组质粒分别转化感受态大肠杆菌DH5α和BL21(DE3),表达纯化目的蛋白S12,测定酶活。结果:成功地构建了头孢菌素C酰化酶S12的原核表达菌株,组成型表达和诱导型表达体系的表达量低,目的蛋白具有酶活,诱导型表达产生的酶比组成型表达产生的酶具有更强活性,可作为大肠杆菌表达头孢菌素C酰化酶的高效表达体系。
