本文選題：人腸道病毒型 + 病毒樣顆粒��； 參考：《蠶業(yè)科學》2016年03期

【摘要】：人腸道病毒71型(human enterovirus 71,EV71)是近年來暴發(fā)危害較嚴重的手足口病的病原。利用家蠶桿狀病毒表達系統(tǒng)在家蠶卵巢培養(yǎng)細胞(Bm N)中共表達EV71病毒衣殼蛋白VP1與VP2,分析二者組裝成病毒樣顆粒(VLPs)的效率,比較用不同啟動子構建重組病毒的表達效果,并利用家蠶幼蟲進行2種蛋白質(zhì)的共表達及VLPs純化條件的研究。結果表明,重組病毒v Bm Bac-vp1-vp2與v Bm Bac-vp1-IRES-vp2均可介導EV71的VP1、VP2蛋白在Bm N細胞中表達,并能夠組裝成病毒樣顆粒,但采用Pph及Pp10啟動子的表達和組裝效率明顯高于Pph及IRES啟動子組合;將重組病毒v Bm Bac-vp1-vp2以1×10~4TCID_(50)/頭的劑量注射5齡起蠶,Western blotting檢測到VP1在幼蟲血淋巴中特異性表達,并且隨著感染時間的延長表達量增加;經(jīng)蔗糖梯度密度離心能夠有效地純化蠶體血淋巴中的病毒樣顆粒。研究結果為后續(xù)EV71疫苗研制奠定了一定的試驗基礎。
[Abstract]:Human enterovirus 71 (human enterovirus 71 / EV71) is the pathogen of hand, foot and mouth disease (HFMD) which is a serious outbreak in recent years. The EV71 capsid protein VP1 and VP2were co-expressed in silkworm ovary culture cells (BmNs) by using the baculovirus expression system of Bombyx mori. The efficiency of assembling them into virus-like granulocytes was analyzed, and the expression effects of recombinant viruses constructed by different promoters were compared. The co-expression of two proteins and the purification conditions of VLPs were studied by using silkworm larvae. The results showed that both vBm Bac-vp1-vp2 and vBm Bac-vp1-IRES-vp2 could mediate the expression of VP1pVP2 protein of EV71 in BMN cells, and could assemble virus-like particles. However, the expression and assembly efficiency of Pph and Pp10 promoters were significantly higher than those of Pph and IRES promoter combinations. The recombinant virus vBm Bac-vp1-vp2 was injected into the head of 5 instar silkworm at the dose of 1 脳 10~4TCID_(50)/. Western blotting was used to detect the specific expression of VP1 in haemolymph of larva, and the expression increased with the prolongation of infection time. The virus-like particles in hemolymph of silkworm body can be effectively purified by sucrose gradient density centrifugation. The results of the study laid a certain experimental foundation for the further development of EV71 vaccine.
【作者單位】： 江蘇科技大學;中國農(nóng)業(yè)科學院蠶業(yè)研究所;
【基金】：江蘇省自然科學基金項目(No.BK20151321) 江蘇省教育廳科技項目(No.11KJB180002)
【分類號】：R373.2;Q78

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