本文關(guān)鍵詞：多價(jià)血吸蟲(chóng)蛋白疫苗的優(yōu)選與原核表達(dá)研究　出處：《華中科技大學(xué)》2012年碩士論文　論文類(lèi)型：學(xué)位論文

  更多相關(guān)文章： 日本血吸蟲(chóng) 原核表達(dá) 蛋白疫苗 生物信息學(xué)分析

【摘要】：血吸蟲(chóng)病是一種世界性的傳染性疾病，，它嚴(yán)重危害著人類(lèi)的生命健康，阻礙社會(huì)發(fā)展，常規(guī)的預(yù)防和治療手段難以遏制形勢(shì)日益嚴(yán)峻的血吸蟲(chóng)疫情，研制血吸蟲(chóng)疫苗是預(yù)防血吸蟲(chóng)病最有效的方法之一。目前血吸蟲(chóng)蛋白疫苗和血吸蟲(chóng)核酸疫苗是主要的兩種疫苗形式。血吸蟲(chóng)核酸疫苗能夠?yàn)閷?shí)驗(yàn)動(dòng)物提供一定的免疫保護(hù)力，然而其安全性在短時(shí)間內(nèi)還無(wú)法得到確認(rèn)。與核酸疫苗相比，蛋白疫苗的技術(shù)較為成熟，且其安全性得到驗(yàn)證，因此血吸蟲(chóng)蛋白疫苗有可能首先實(shí)行產(chǎn)業(yè)化。血吸蟲(chóng)蛋白疫苗有單價(jià)蛋白疫苗和多價(jià)蛋白疫苗形式，生產(chǎn)多價(jià)蛋白疫苗可以減少生產(chǎn)成本和簡(jiǎn)化生產(chǎn)工藝。然而，利用基因工程技術(shù)生產(chǎn)多價(jià)蛋白疫苗時(shí)，是否因?yàn)榈鞍兹诤现笃芜^(guò)大而影響其合成是一個(gè)需要研究的問(wèn)題。為此，本研究建立了血吸蟲(chóng)蛋白疫苗原核表達(dá)系統(tǒng)，對(duì)單價(jià)蛋白疫苗和多價(jià)蛋白疫苗的原核表達(dá)進(jìn)行了深入的研究，取得了以下實(shí)驗(yàn)結(jié)果： (1)根據(jù)WHO提供的血吸蟲(chóng)疫苗候選抗原以及利用生物信息學(xué)方法分析抗原的疏水性和高級(jí)結(jié)構(gòu)，篩選出合適的血吸蟲(chóng)抗原基因（Sj26, SjGAPDH,Sj26.GAPDH）來(lái)進(jìn)行研究。 (2)克隆到了日本血吸蟲(chóng)單價(jià)抗原基因Sj26和SjGAPDH以及融合抗原基因Sj26.GAPDH，利用分子生物學(xué)方法將抗原基因連接到原核表達(dá)載體上，構(gòu)建成原核表達(dá)系統(tǒng)，經(jīng)酶切驗(yàn)證和測(cè)序驗(yàn)證，表明構(gòu)建成功。通過(guò)鎳離子層析柱的親和層析來(lái)對(duì)目的蛋白進(jìn)行分離純化，并且摸索出鎳離子層析柱的最佳洗脫條件。 (3)利用酶聯(lián)免疫吸附測(cè)定(ELISA)對(duì)分離得到的單價(jià)日本血吸蟲(chóng)抗原蛋白和融合的抗原蛋白進(jìn)行定量分析。統(tǒng)計(jì)學(xué)分析表明，融合抗原蛋白的原核表達(dá)量與單價(jià)抗原蛋白的原核表達(dá)量沒(méi)有明顯差異，從而對(duì)血吸蟲(chóng)蛋白疫苗的優(yōu)選作出了初步判斷。
[Abstract]:Schistosomiasis is a worldwide infectious disease, which seriously endangers human life and health, hinders the development of society, and it is difficult to contain the increasingly severe situation of schistosomiasis by conventional prevention and treatment. Development of schistosomiasis vaccine is one of the most effective methods to prevent schistosomiasis. At present, Schistosoma protein vaccine and schistosomiasis nucleic acid vaccine are two main forms of vaccine. Schistosoma nucleic acid vaccine can provide certain amount of vaccine for laboratory animals. Immune protection. However, its safety can not be confirmed in a short time. Compared with nucleic acid vaccine, protein vaccine technology is more mature, and its safety has been verified. Therefore, it is possible that the protein vaccine of Schistosoma japonicum can be industrialized first. The protein vaccine of Schistosoma japonicum can be produced in the form of monovalent protein vaccine and polyvalent protein vaccine. The production of polyvalent protein vaccines can reduce production costs and simplify production processes. However, when using genetic engineering techniques to produce multivalent protein vaccines. It is a problem that whether the protein fusion fragment is too large to affect its synthesis. Therefore, the prokaryotic expression system of Schistosoma japonicum protein vaccine has been established in this study. The prokaryotic expression of monovalent protein vaccine and polyvalent protein vaccine was studied. The following results were obtained: 1) according to the candidate antigen of Schistosoma japonicum vaccine provided by WHO and the analysis of hydrophobicity and advanced structure of antigen by bioinformatics, the suitable gene of Schistosoma japonicum antigen (Sj26) was selected. SjGAPDH Sj26. GAPDH). The monovalent antigen genes Sj26 and SjGAPDH of Schistosoma japonicum and the fusion antigen gene Sj26.GAPDH were cloned. The prokaryotic expression system was constructed by ligating the antigen gene to the prokaryotic expression vector by molecular biological method. The expression system was confirmed by restriction endonuclease digestion and sequencing. The results showed that the target protein was separated and purified by affinity chromatography of nickel ion chromatography column, and the optimum elution conditions were found out. (3) the monovalent Schistosoma japonicum antigen protein and fusion antigen protein were quantitatively analyzed by enzyme linked immunosorbent assay (Elisa). There was no significant difference between the prokaryotic expression of fusion antigen protein and that of monovalent antigen protein, so the preliminary judgement was made on the optimal selection of Schistosoma japonicum protein vaccine.
【學(xué)位授予單位】：華中科技大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2012
【分類(lèi)號(hào)】：R532.21;R392.11

【參考文獻(xiàn)】

相關(guān)期刊論文 前10條

1 萬(wàn)中原,李雍龍,韓家俊,石佑恩;日本血吸蟲(chóng)多價(jià)分子疫苗的保護(hù)性免疫力研究[J];解放軍廣州醫(yī)高專(zhuān)學(xué)報(bào);1998年01期

2 鄭江;我國(guó)血吸蟲(chóng)病防治應(yīng)堅(jiān)持以社會(huì)措施為主導(dǎo)的策略[J];國(guó)外醫(yī)學(xué).流行病學(xué)傳染病學(xué)分冊(cè);2005年01期

3 付譯節(jié);陳建平;楊莉;;血吸蟲(chóng)疫苗研究新進(jìn)展[J];國(guó)際生物制品學(xué)雜志;2007年06期

4 馮思玲;;生物信息學(xué)技術(shù)研究[J];信息技術(shù);2009年05期

5 林金祥,李莉莎;血吸蟲(chóng)病防治的回顧與現(xiàn)狀[J];海峽預(yù)防醫(yī)學(xué)雜志;2005年01期

6 劉彥,肖建華;血吸蟲(chóng)核酸疫苗研究進(jìn)展[J];寄生蟲(chóng)與醫(yī)學(xué)昆蟲(chóng)學(xué)報(bào);2003年02期

7 王學(xué)敏;李碧俠;任守文;;豬CAPN2基因的生物信息學(xué)分析[J];江蘇農(nóng)業(yè)科學(xué);2008年01期

8 王轉(zhuǎn)梅;陳崇順;王軼;曹偉杰;鄒愛(ài)蘭;馬文曄;張娟;;煙草品種Xanthi NN堿性β-1,3-葡聚糖酶基因的克隆與生物信息學(xué)分析[J];江蘇農(nóng)業(yè)科學(xué);2009年01期

9 欒德琴;白云峰;包文斌;陳國(guó)宏;;基因本體論研究進(jìn)展及在生物信息數(shù)據(jù)庫(kù)中的應(yīng)用[J];江蘇農(nóng)業(yè)科學(xué);2009年04期

10 趙群;;基于生物信息學(xué)對(duì)主要蜥蜴科動(dòng)物系統(tǒng)發(fā)生關(guān)系的研究[J];江蘇農(nóng)業(yè)科學(xué);2009年04期

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