猪带绦虫TSO45W-4B基因在长双歧杆菌中的表达

Expression of Taenia solium TSO45W-4B gene in bifidobacterium longum

摘要:

目的：在成功构建猪带绦虫大肠杆菌-双歧杆菌穿梭表达质粒pGEX-TSO45W-4B的基础上，研究猪带绦虫TSO45W-4B基因在长双歧杆菌中的表达情况。方法：将猪带绦虫大肠杆菌-双歧杆菌穿梭表达质粒pGEX-TSO45W-4B电转化入长双歧杆菌，异丙基-β-D-硫代半乳糖苷（isopropyl-β-d-thiogalactoside，IPTG）诱导表达，十二烷基硫酸钠聚丙烯酰胺凝胶电泳（sodium dodecyl sulfatepolyacrylamide gel electrophoresis，SDS-PAGE）和Western blot分析表达情况。结果：酶切、聚合酶链反应（poly－merase chain reaction，PCR）和测序证实，重组质粒pGEX-TSO45W-4B成功转入长双歧杆菌。SDS-PAGE显示，重组蛋白相对分子质量（Mr）约为40 kD，经谷胱甘肽转移酶（glutathione S-transferase，GST）亲和层析后可获得纯度为85%以上的重组蛋白。Western blot显示，重组蛋白能被兔抗TSO45W-4B血清、囊虫病猪血清和囊虫病患者血清所识别。结论：猪带绦虫TSO45W-4B基因能够在长双歧杆菌中获得表达，表达的重组蛋白具有特异的抗原性。

Abstract:

Objective：To observe the expression of the gene of TSO45W-4B of Taenia solium in bifidobacterium longum（B.longum） on the basis of successful construction of the escherichia coli（E.coil）-bifidobacterium shuttle plasmid pGEX-TSO45W-4B of Taenia solium. Methods：The shuttle expression plasmid pGEX-TSO45W-4B of Taenia solium was electroporated into B.longum. After induc－tion with isopropyl-β-d-thiogalactoside，the expression of the recombinant protein was identified by sodium dodecyl sulfatepolyacry－lamide gel electrophoresis（SDS-PAGE） and Western blot. Results：Restriction analysis，polymerase chain reaction and sequencing showed that the recombinant plasmid pGEX-TSO45W-4B was successfully transformed into B.longum. As demonstrated by SDS-PAGE，the relative molecular mass（Mr） of the expressed recombinant protein was approximately 40 kD，and the purity of the recom－binant protein was 85% after purification withglutathione S-transferase affinity chromatography. The rabbit antiserum of TSO45W-4B，cysticercosis swine serum and cysticercosis patients serum could bind to the recombinant protein in Western blot assay. Conclusion：The gene of TSO45W-4B of Taenia solium could be expressed in B.longum and the expressed recombinant protein shows specific antigenicity.