本文關(guān)鍵詞： 日本血吸蟲 IL-18 核酸疫苗 免疫　出處：《吉林農(nóng)業(yè)大學(xué)》2008年碩士論文　論文類型：學(xué)位論文

【摘要】： 日本血吸蟲病是一種危害嚴(yán)重的人畜共患寄生蟲病,目前仍以藥物(吡喹酮)治療為主。但是藥物治療不能阻斷血吸蟲病的傳播與流行。因此加強(qiáng)血吸蟲疫苗的研究是控制我國血吸蟲病的有效措施。 本研究應(yīng)用基因工程技術(shù)構(gòu)建了日本血吸蟲核酸疫苗pVAX-GST、pVAX-GST-FABP和pIRESneo-GST-FABP-Sj23,并且克隆了小鼠IL-18,構(gòu)建了真核表達(dá)載體pVAX-IL-18。以小鼠為動物模型評價了其免疫保護(hù)效果。 日本血吸蟲核酸疫苗的構(gòu)建運(yùn)用基因工程重組技術(shù)將日本血吸蟲保護(hù)性抗原基因GST、FABP、Sj23克隆入真核表達(dá)載體pVAX和pIRESneo中,通過酶切鑒定構(gòu)建成了日本血吸蟲真核表達(dá)載體pVAX-GST、pVAX-GST-FABP和pIRESneo-GST-FABP-Sj23。 小鼠IL-18真核表達(dá)載體的構(gòu)建應(yīng)用RT-PCR技術(shù)從小鼠脾淋巴細(xì)胞中擴(kuò)增了小鼠的白介素-18(IL-18),并將其克隆至pMD18-T載體,經(jīng)序列分析后將IL-18克隆至真核表達(dá)載體pVAX中,構(gòu)建了重組質(zhì)粒pVAX-IL-18。 動物免疫保護(hù)性試驗(yàn)將構(gòu)建的日本血吸蟲核酸疫苗及pVAX-IL-18以肌肉注射的方式免疫BALB/c小鼠,二次免疫后,用日本血吸蟲尾蚴分別攻擊感染小鼠,42天后剖殺沖蟲并計(jì)數(shù)。免疫試驗(yàn)結(jié)果表明,日本血吸蟲核酸疫苗均能誘導(dǎo)小鼠產(chǎn)生特異性的IgG抗體,pIRESneo-GST-FABP-Sj23+IL-18免疫組所誘導(dǎo)的IgG水平最高;且免疫組都產(chǎn)生了較高水平的IFN-γ,其中聯(lián)合IL-18組所誘導(dǎo)的IFN-γ明顯高于單獨(dú)免疫組(P0.01);在動物保護(hù)性實(shí)驗(yàn)中pIRESneo-GST-FABP-Sj23、pIRESneo-GST-FABP-Sj23+IL-18分別獲得了45.0%、69.8%的減卵率,40.0%、47.5%的減蟲率。上述結(jié)果表明日本血吸蟲多價核酸疫苗在動物免疫實(shí)驗(yàn)中獲得了較好的免疫保護(hù)效果,并且IL-18能增強(qiáng)日本血吸蟲核酸疫苗的免疫保護(hù)作用。
[Abstract]:Schistosomiasis japonicum is a serious zoonotic parasitic disease. At present, the drug (praziquantel) is still the main treatment, but the drug treatment can not stop the transmission and prevalence of schistosomiasis. Therefore, strengthening the research of schistosomiasis vaccine is an effective measure to control schistosomiasis in China. In this study, the nucleic acid vaccine pVAX-GST-FABP and pIRESneo-GST-FABP-Sj23 of Schistosoma japonicum were constructed by genetic engineering technique, and the mouse IL-18 was cloned, and the eukaryotic expression vector pVAX-IL-18 was constructed. Construction of Schistosoma japonicum Nucleic Acid Vaccine by genetic Engineering Recombinant technique, the protective antigen gene GST-FABPN-Sj23 of Schistosoma japonicum was cloned into eukaryotic expression vectors pVAX and pIRESneo. The eukaryotic expression vectors pVAX-GST-pVAX-GST-FABP and pIRESneo-GST-FABP-Sj23 of Schistosoma japonicum were constructed by restriction endonuclease digestion. Construction of murine IL-18 eukaryotic expression vector using RT-PCR technique, the mouse interleukin-18 IL-18G was amplified from mouse spleen lymphocytes and cloned into pMD18-T vector. After sequence analysis, IL-18 was cloned into eukaryotic expression vector pVAX, and a recombinant plasmid pVAX-IL-18 was constructed. The BALB/c mice were immunized with the constructed Schistosoma japonicum nucleic acid vaccine and pVAX-IL-18 by intramuscular injection. Schistosoma japonicum cercariae were used to kill and count Schistosoma japonicum in mice after 42 days of infection. The results of immunological test showed that the mice immunized with specific IgG antibody pIRESneo-GST-FABP-Sj23 IL-18 had the highest level of IgG. The IFN- 緯 in the combined IL-18 group was significantly higher than that in the single immunization group (P0.01), and in the animal protective experiment, pIRESneo-GST-FABP-Sj23 pIRESneo-GST-FABP-Sj23 IL-18 obtained 45.09.8% oocyte reduction rate and 47.5% worm reduction rate respectively. The polyvalent nucleic acid vaccine of Schistosoma japonicum has a good protective effect in animal immune experiment. And IL-18 can enhance the immune protection of Schistosoma japonicum nucleic acid vaccine.
【學(xué)位授予單位】：吉林農(nóng)業(yè)大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2008
【分類號】：R392

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