本文選題：A組人輪狀病毒 + VP6��； 參考：《中國協(xié)和醫(yī)科大學(xué)》2008年碩士論文

【摘要】： 輪狀病毒(Rotavirus,RV)屬呼腸病毒科(Reoviridae)輪狀病毒屬,是引起人類、哺乳動物類和鳥類腹瀉的主要病原體。完整的RV病毒分為三層結(jié)構(gòu),VP6蛋白位于病毒的內(nèi)殼中間層,為第6基因片段編碼,由397個氨基酸組成,分子量約為44.8KDa。VP6蛋白是輪狀病毒中含量仍然最豐富的蛋白,約占病毒總蛋白量的51%,輪狀病毒內(nèi)殼由VP6蛋白構(gòu)成的260個致密的三聚體所形成。VP6在病毒結(jié)構(gòu)中具有物理受體的作用,在病毒基因轉(zhuǎn)錄中也具有重要作用。 本研究中的VP6基因克隆自臨床樣品中分離的A組輪狀病毒TB-Chen株。研究中,以VP6全基因?yàn)槟０?經(jīng)PCR擴(kuò)增,基因重組,將VP6蛋白編碼基因插入到質(zhì)粒pET中構(gòu)建了原核表達(dá)質(zhì)粒pET-VP6,并在大腸桿菌BL21(DE3)中高效表達(dá)重組VP6蛋白(rVP6)。rVP6占菌體總蛋白的27.4%。rVP6經(jīng)純化后免疫兔,獲得動物免疫血清抗體。Western blot檢測結(jié)果顯示,表達(dá)的rVP6能被抗-rVP6兔血清抗體特異性識別,能被抗-SA11和抗-Wa的兔免疫血清抗體特異性識別。免疫熒光結(jié)果顯示MA104細(xì)胞中的RVVP6抗原在RV感染3h后即可觀察到,在12h左右熒光強(qiáng)度達(dá)到最高。病毒的VP6蛋白大部份散布于細(xì)胞質(zhì)間,主要以顆粒狀的形式分布。中和實(shí)驗(yàn)結(jié)果顯示,抗-rVP6的兔血清抗體具有中和RV SA11和Wa株病毒的中和病毒感染活性。 這些結(jié)果為明確A組輪狀病毒VP6的生物學(xué)和免疫學(xué)特性及其在RV亞單位疫苗應(yīng)用開發(fā)中的價值提供了理論實(shí)驗(yàn)基礎(chǔ)。
[Abstract]:Rotavirus (Rotavirus RV) belongs to the family Reoviridae, which is the main pathogen causing diarrhea in humans, mammals and birds. The complete RV virus is divided into three layers, the VP6 protein is located in the middle layer of the virus's inner shell, which encodes the 6th gene fragment and consists of 397 amino acids. The molecular weight of the protein is about 44.8 KDa.VP6, which is still the most abundant protein in rotavirus. About 51% of the total protein of the virus, rotavirus inner shell is formed by 260 dense trimer composed of VP6 protein. VP6 has the function of physical receptor in the virus structure, and also plays an important role in viral gene transcription. The VP6 gene in this study was cloned from a group A rotavirus TB-Chen strain isolated from clinical samples. In this study, VP6 gene was amplified by PCR and the gene was recombined with VP6 as template. The prokaryotic expression plasmid pET-VP6 was constructed by inserting the VP6 protein coding gene into the plasmid pet. The recombinant VP6 protein (rVP6). RVP6 accounted for 27.4% of the total bacterial protein in E. coli BL21 (DE3). The results showed that the expressed rVP6 could be specifically recognized by anti-rVP6 rabbit serum antibody, and by anti-SA11 and anti-Wa rabbit immune serum antibody. The results of immunofluorescence showed that the RVVP6 antigen in MA104 cells could be observed 3 hours after RV infection, and the highest fluorescence intensity was observed at about 12 h. Most of the VP6 proteins were distributed in the cytoplasm and distributed in granular form. The neutralization results showed that the anti-rVP6 rabbit serum antibody had neutralizing virus infection activity of RV SA11 and WA strains. These results provide a theoretical and experimental basis for clarifying the biological and immunological characteristics of group A rotavirus VP6 and its value in the application and development of RV subunit vaccine.
【學(xué)位授予單位】：中國協(xié)和醫(yī)科大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2008
【分類號】：R392;R373

【參考文獻(xiàn)】

相關(guān)期刊論文 前6條

1 張春芳;賈立英;;亞洲地區(qū)輪狀病毒感染的流行病學(xué)概況[J];中國當(dāng)代兒科雜志;2006年01期

2 魏靜;李晉濤;支軼;唐艷;張小萍;王靖雪;吳玉章;;輪狀病毒結(jié)構(gòu)蛋白VP6 HLA-A*0201限制性CTL表位的初步鑒定[J];第三軍醫(yī)大學(xué)學(xué)報(bào);2006年06期

3 鞠月迎;;新型輪狀病毒疫苗研究進(jìn)展[J];世界臨床藥物;2006年06期

4 陳元鼎;劉曉;熊新宇;曹志亮;文喻玲;趙慶歡;余洋;尹興曉;李傳印;范耀春;;A組人輪狀病毒全基因組克隆和基因型分析[J];中國生物工程雜志;2008年02期

5 章青;王端可;葉新華;段招軍;杜曾慶;金玉;唐景裕;孫利偉;Duncan Steele;Roger Glass;方肇寅;;中國1998~2004年G_9型輪狀病毒分子流行病學(xué)研究[J];中國計(jì)劃免疫;2006年06期

6 何金生,王健偉,姜秀麗,王大燕,溫樂英,董京芳,屈建國,洪濤;輪狀病毒組特異性抗原VP6在重組腺病毒中的表達(dá)及其免疫學(xué)效果的研究[J];中華實(shí)驗(yàn)和臨床病毒學(xué)雜志;2002年02期

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