發(fā)布時間：2018-04-09 10:44

  本文選題：幽門螺桿菌　切入點：外膜蛋白　出處：《浙江大學》2014年碩士論文

【摘要】：目的篩選并鑒定幽門螺桿菌外膜蛋白GroEL的優(yōu)勢T細胞和B細胞(T-B)聯(lián)合抗原表位。 方法采用PCR擴增幽門螺桿菌NCTC11637株groEL基因并構建其原核表達系統(tǒng),SDS-PAGE檢查目的重組蛋白rGroEL表達情況。Ni-NTA親和層析法提純rGroEL。采用激光共聚焦顯微鏡法檢測幽門螺桿菌NCTC11637和SS1株中GroEL分布。Triton X-114法提取上述菌株外膜蛋白樣本,采用Western Blot法檢測外膜蛋白樣本中的GroEL。采用專業(yè)生物信息學軟件預測GroEL的T-B聯(lián)合抗原表位并構建其噬菌體展示系統(tǒng)。采用Western Blot法和ELISA分別檢測含T-B聯(lián)合表位肽的重組噬菌體PⅢ蛋白和人工合成的T-B聯(lián)合表位肽的免疫反應性。 結果幽門螺桿菌NCTC11637和SSl株groEL基因核苷酸和氨基酸序列相似性高達97.68-99.63%。所構建原核表達系統(tǒng)能有效表達可溶性rGroEL。GroEL定位于幽門螺桿菌NCTC11637和SS1株外膜。GroEL168、GroEL258、GroEL288. GroEL365.GroEL396和GroEL438六個主要的預測T-B聯(lián)合抗原表位中,GroEL168顯示了很強的陽性雜交信號。ELISA結果顯示, GroEL168免疫反應性最強(P0.01),其次為GroEL258和GroEL288(P0.05)。 結論GroEL是幽門螺桿菌外膜蛋白。GroEL168是GroEL的優(yōu)勢T-B聯(lián)合抗原表位,可用于制備幽門螺桿菌多抗原肽疫苗。
[Abstract]:Objective to screen and identify the dominant T cells and B cell T-B associated epitopes of Helicobacter pylori outer membrane protein (GroEL).Methods the groEL gene of Helicobacter pylori NCTC11637 strain was amplified by PCR and its prokaryotic expression system was constructed to detect the rGroEL expression of the recombinant protein. Ni-NTA affinity chromatography was used to purify rGroelle.The distribution of GroEL in Helicobacter pylori NCTC11637 and SS1 strains was detected by laser confocal microscopy. Triton X-114 was used to extract the outer membrane protein samples from the above strains, and Western Blot method was used to detect the outer membrane protein samples.The T-B antigen epitopes of GroEL were predicted by bioinformatics software and its phage display system was constructed.The immunoreactivity of recombinant bacteriophage P 鈪，

本文編號：1726116