家蚕细小病毒样病毒（BmPLV-Z） VD1 ORF4的原核表达

对家蚕细小病毒样病毒（BmPLV-Z）VD1 ORF4理论上编码的氨基酸序列进行Blast搜索,结果表明其与DNA聚合酶B家族同源。通过PCR扩增其DNA聚合酶同源区（1 077 bp）,将扩增的目的DNA片段与原核表达载体pET30a进行连接,通过不同浓度的IPTG对捕获pET30a-1 077 bp重组质粒的大肠埃希菌进行诱导,对诱导产物进行SDS-PAGE电泳,结果表明其聚合酶同源区获得了表达;Western blot分析进一步确证诱导蛋白为带有6个组氨酸的融合蛋白。将割胶获得的目的蛋白免疫昆明小鼠制备其多抗,以纯化后的抗血清对VD1 ORF4全长序列和部分序列在原核中的漏扫描表达情况进行研究,SDS-PAGE和Western blot结果表明VD1 ORF4全长序列和部分序列的原核表达产物均一,都只有一条特异的目的蛋白带,说明了VD1 ORF4序列在原核表达系统中没有漏扫描表达的蛋白。

Prokaryotic Expression of Bombyx mori Parvo-Like Virus (BmPLV-Z) VD1ORF4 in E.coli

Theoretically coded amino acid sequence of BmPLV-Z VD1 ORF4 was carried out Blast searching,the results showed that it was homologous to DNA polymerase B family.The DNA polymerase fragment of the homologous domain(1 077 bp) in VD1 ORF4 was amplified by PCR.The amplified target DNA domain was linked to prokaryotic expression vector pET30a and induced E.coli that caught pET30a-1 077bp recombinant plasmids with different concentration of IPTG,and the induced products were carried out SDS-PAGE electrophoreses.T...