CAJ | 학술논문

将DH5α/pGXmTSST基因工程菌用IPTG诱导表达,经超声波破碎、离心、收集上清再用谷胱甘肽琼脂糖凝胶柱（GS4B）亲和层析纯化金黄色葡萄球菌无毒诱变GST-mTSST-1融合蛋白.以SDS-PAGE分析该融合蛋白的表达量和纯度,经双向免疫琼脂扩散试验鉴定GST-mTSST-1融合蛋白的免疫活性.结果表明,GST-mTSST-1融合蛋白在重组菌中可溶性表达,通过GS4B胶亲和层析纯化,SDS-PAGE电泳分析得到纯度较高的目的蛋白,其相对分子质量约为47 000,与理论值一致.兔抗GST-mTSST-1抗血清可特异识别天然rTSST-1蛋白中与GST-mTSST-1融合蛋白相对应的抗原组份,证实GST-mTSST-1融合蛋白具有与野生型rTSST-1相同的表面抗原决定簇,具有较好的免疫原性.
장DH5α/pGXmTSST기인공정균용IPTG유도표체,경초성파파쇄、리심、수집상청재용곡광감태경지당응효주（GS4B）친화층석순화금황색포도구균무독유변GST-mTSST-1융합단백.이SDS-PAGE분석해융합단백적표체량화순도,경쌍향면역경지확산시험감정GST-mTSST-1융합단백적면역활성.결과표명,GST-mTSST-1융합단백재중조균중가용성표체,통과GS4B효친화층석순화,SDS-PAGE전영분석득도순도교고적목적단백,기상대분자질량약위47 000,여이론치일치.토항GST-mTSST-1항혈청가특이식별천연rTSST-1단백중여GST-mTSST-1융합단백상대응적항원조빈,증실GST-mTSST-1융합단백구유여야생형rTSST-1상동적표면항원결정족,구유교호적면역원성.
The E.coli DH5α cells harboring the pGXmTSST gene was induced by adding isopropyl-β-D-thiogalactoside（IPTG）,the bacteria were collected by centrifugation and were disrupted by sonication.The notoxic and mutant GST-mTSST-1 fusion protein of Staphylococcus aureus were isolated and purified by glutathione S-transferase agarose resin affinity chromatography column.The expression product and purity of GST-mTSST-1 fution protein were analyzed by sodium dodecylsulfate polyacrylanide gels（SDS-PAGE） electrophoresis,and the immune activity were tested by agar-gel double immunodiffussion essay.The results showed that the high purity and soluble GST-mTSST-1 fution protein were obtained by GS4B affinity chromatography and SDS-PAGE,its relative molecular mass was 47000.The polyclonal rabbit antiserum reacted readily with purified GST-mTSST-1 fution protein and rTSST-1,and the precipitation lines between GST-mTSST-1 and rTSST-1 were united each other.These results indicated that GST-mTSST-1 retained the same antibody-binding epitopes as wild-type rTSST-1and had well immunogenicity.