CAJ | 학술논문

目的表达梅毒螺旋体(Treponema pal idum, Tp)重组蛋白Tp1038(rTp1038)，鉴定其抗原性，为进一步探讨其在梅毒血清学诊断中的价值奠定基础。方法PCR扩增Tp1038全长基因，构建含6个组氨酸(His)标签的原核表达重组体pET-28a(+)/Tp1038，转化宿主菌 E. coli诱导表达 rTp1038，以 Ni-NTA亲和层析柱纯化蛋白，SDS-PAGE分析蛋白表达形式与纯度，Western blot鉴定rTp1038的抗原性。结果成功构建了表达重组体pET-28a/Tp1038，经诱导宿主菌高效表达了一分子量大约为20 KDa的重组蛋白，以可溶性表达形式为主，纯化蛋白的纯度>98%；Western blot结果显示该纯化蛋白仅能被抗 His单抗和梅毒患者混合血清所特异性识别。结论高效表达了可溶性rTp1038，该重组蛋白有良好的抗原性和特异性。
목적표체매독라선체(Treponema pal idum, Tp)중조단백Tp1038(rTp1038)，감정기항원성，위진일보탐토기재매독혈청학진단중적개치전정기출。방법PCR확증Tp1038전장기인，구건함6개조안산(His)표첨적원핵표체중조체pET-28a(+)/Tp1038，전화숙주균 E. coli유도표체 rTp1038，이 Ni-NTA친화층석주순화단백，SDS-PAGE분석단백표체형식여순도，Western blot감정rTp1038적항원성。결과성공구건료표체중조체pET-28a/Tp1038，경유도숙주균고효표체료일분자량대약위20 KDa적중조단백，이가용성표체형식위주，순화단백적순도>98%；Western blot결과현시해순화단백부능피항 His단항화매독환자혼합혈청소특이성식별。결론고효표체료가용성rTp1038，해중조단백유량호적항원성화특이성。
Objective To express and identify recombinant protein Tp1038 (rTp1038) of Treponema pal idum (Tp), providing a basis for further investigation of its significance in syphilis serodiagnosis. Methods PCR was used to amplify ful length of Tp1038 gene. The prokaryotic expression recombinant pET-28a (+)/Tp1038 was constructed and transformed into E.coli BL21 for expressing rTp1038. rTp1038 were purified with Ni-NTA af inity chromatography. SDS-PAGE was used to analyse expression form and purity of rTp1038. Western blot was performed to identify antigenicity of rTp1038. Results The prokaryotic recombinant pET-28a (+)/Tp1038 was constructed successful y and an approximate 20 KDa recombinant protein was expressed ef iciently in E.coli with dominant soluble form and more than 98% purity. Western blot indicated that purified proteins were able to be recognized special y by ant-His monoclonal antibody and pooled sera from syphilis patients. Conclusions The soluble recombinant Tp1038 are expressed ef iciently in E.coli and have good antigenicity and specifity.