鸡传染性贫血病毒VP1基因的原核表达

Prokaryotic Expression of Chicken Infectious Anemia Virus VP1

根据GenBank上公布的鸡传染性贫血病毒VP1的基因序列设计一系列引物,通过基因扩增得到了鸡传染性贫血病毒(CAV)VP1基因的全长片段和一系列不同N末端截断的VP1基因片段,分别将这些片段插入表达质粒载体pET28a的多克隆位点上,构建成4种重组表达工程菌(E.coli BL21/pET-28-CAVVP1、E.coli BL21/pET-28-CAVVP1Nd29、E.coli BL21/pET-28-CAV VP1Nd56、E.coli BL21/pET-28-CAV VP1Nd137),研究了4种不同长度VP1片段在重组大肠杆菌中的表达,并通过SDS-PAGE电泳筛选最佳表达方式。结果表明:E.coli BL21/pET-28-CAVVP1Nd29能表达分子质量为44ku的可溶性蛋白,E.coli BL21/pET-28-CAV VP1Nd56能表达分子质量为40ku的不可溶性蛋白,而在E.coli BL21/pET-28-CAV VP1和E.coli BL21/pET-28-CAV VP1Nd137的诱导表达产物中未见明显的表达蛋白出现。其中可溶性的CAV VP1Nd29蛋白将有可能用于研制CAV诊断抗原和亚单位疫苗研究。

According to the sequences of CAV VP1 published GenBank,a series of primers were designed. Four different segments of VP1 genes were amplified through PCR,and had been inserted into the multiple cloning site of expression plasmid vector pET28a.Four kinds of recombinant expression engineering bacterium(E.coli BL21/pET-28-CAVVP1,E.coli BL21/pET-28-CAVVPlNd29, E.coli BL21/pET-28-CAVVP1Nd56,E.coli BL21/pET-28-CAV VPlNd137) had been constructed.The expression of full VP1 and VP1 with a serial of N-terminus truncated of chicken infectious anemia virus(CAV) in recombinant E.coli were studied,the best way of expression were selected throgh SDS-PAGE electrophoresis.The results showed that E.coli BL21/ pET-28-CAV VPlNd29 can express a soluble protein of 44 ku,E.coli BL21/pET-28-CAV VPlNd56 can express an insoluble protein of 40 ku,but no visible expression protein can be seen in the product of induced recombinant E.coli BL21/pET-28-CAV VP1 and E.coli BL21/pET-28-CAV VP1Nd137.The soluble expression protein of CAV VP1Nd29 can be used in development of antigens, which will be used in the produce of diagnosis agent and subunit vaccine of CAV.