本文選題：遲緩愛(ài)德華氏菌 + 保護(hù)性抗原��； 參考：《中國(guó)科學(xué)院研究生院（海洋研究所）》2010年博士論文

【摘要】： 遲緩愛(ài)德華氏菌(Edwardsiella tarda)是一種魚(yú)類(lèi)病原菌,其感染魚(yú)類(lèi)后引起遲緩愛(ài)德華菌病(edwardsiellosis),給養(yǎng)殖業(yè)造成嚴(yán)重的經(jīng)濟(jì)損失。 本研究中,應(yīng)用IVIAT技術(shù),我們從病魚(yú)體內(nèi)分離的遲緩愛(ài)德華氏菌菌株TX01中篩選得到Eta21、Eta6和FliC三個(gè)抗原蛋白。其中Eta21與其他菌株中一個(gè)潛在的多肽酶相似性較高;Eta6與大腸桿菌中的Ecotin前體蛋白同源;FliC是一種鞭毛蛋白。以牙鲆(Japanese flounder)作為模式動(dòng)物的研究中,發(fā)現(xiàn)重組Eta21蛋白能夠有效的保護(hù)牙鲆抵抗遲緩愛(ài)德華氏菌的侵染;重組Eta6蛋白具有一定的保護(hù)效應(yīng);重組FliC蛋白沒(méi)有明顯的保護(hù)效應(yīng)。 以eta6和fliC基因構(gòu)建的DNA疫苗pETA6和pFliC的相對(duì)保護(hù)效率分別為50%和33%。與pETA6的保護(hù)效應(yīng)相比,將eta6基因和fliC基因連在一起構(gòu)建的融合DNA疫苗pCE6的保護(hù)效應(yīng)顯著提高。同時(shí),將前期鑒定出的遲緩愛(ài)德華氏菌TX01的抗原基因et18連接到fliC基因的后面,構(gòu)建了融合DNA疫苗pCE18。與能單純表達(dá)Et18的DNA疫苗pEt18相比,pCE18免疫牙鲆后可以產(chǎn)生更好的保護(hù)效應(yīng)。pEta6和pCE6免疫后,牙鲆體內(nèi)產(chǎn)生了特異性的抗體,而且其體內(nèi)一些與固有免疫和獲得性免疫相關(guān)基因的表達(dá)水平也獲得了提高。相對(duì)于pEta6,pCE6誘導(dǎo)的上述基因表達(dá)水平提高的幅度更為明顯。 為提高Eta21的保護(hù)效應(yīng),我們將eta21基因與胞外瓊膠酶agaV的分泌片段基因融合,構(gòu)建了重組質(zhì)粒pTAET21。以含有此重組質(zhì)粒的大腸桿菌DH5α作為活菌疫苗,其保護(hù)效應(yīng)較之重組蛋白Eta21的保護(hù)能力有明顯的提高。
[Abstract]:Edwardsiella tardaa is a kind of fish pathogen, which causes Edwardsiella tarda disease after it is infected with fish, and causes serious economic loss to the aquaculture industry. In this study, three antigenic proteins, Eta21, Eta6 and FliC, were isolated from the Eta21 Eta6 and FliC strains of Edwardian tardy strain TX01 by using IVIAT technique. There is a high similarity between Eta21 and a potential polypeptidase in other strains. Eta6 is a flagellin with Ecotin precursor protein homologous in Escherichia coli. Using Japanese flounder as a model animal, it was found that the recombinant Eta21 protein could effectively protect Paralichthys olivaceus from infection by Edwarder, the recombinant Eta6 protein had some protective effect, and the recombinant FliC protein had no obvious protective effect. The relative protective efficiency of DNA vaccine pETA6 and pFliC constructed by eta6 and fliC gene were 50% and 33% respectively. Compared with the protective effect of pETA6, the protective effect of the fusion DNA vaccine pCE6 constructed by linking eta6 gene and fliC gene was significantly improved. At the same time, the previously identified TX01 antigen gene et18 was linked to the fliC gene, and the fusion DNA vaccine pCE18 was constructed. Compared with the DNA vaccine pEt18, which can express Et18 alone, pCE18 can produce better protective effect after immunization. PEta6 and pCE6 can produce specific antibody in flounder. Moreover, the expression level of some genes related to innate immunity and acquired immunity were also improved. Compared with pEta6, pCE6 induced gene expression increased more significantly. In order to improve the protective effect of Eta21, we fused the eta21 gene with the secreting fragment of extracellular agarose enzyme agaV and constructed the recombinant plasmid pTAET21. The protective effect of Escherichia coli DH5 偽 containing the recombinant plasmid as a live vaccine was significantly higher than that of the recombinant protein Eta21.
【學(xué)位授予單位】：中國(guó)科學(xué)院研究生院（海洋研究所）
【學(xué)位級(jí)別】：博士
【學(xué)位授予年份】：2010
【分類(lèi)號(hào)】：R392.1

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