【摘要】：采用酵母雙雜交系統(tǒng)篩選與禽流感病毒AM2相互作用的宿主蛋白,可為研究AM2蛋白的生物學(xué)新功能提供依據(jù)。將檢測(cè)無(wú)毒性與自激活作用的誘餌質(zhì)粒pGBKT7-AM2與淋巴文庫(kù)質(zhì)粒進(jìn)行融合雜交,然后對(duì)經(jīng)選擇性培養(yǎng)基篩選到的陽(yáng)性克隆進(jìn)行酶切鑒定和測(cè)序分析,陽(yáng)性克隆進(jìn)行酵母回轉(zhuǎn)試驗(yàn)驗(yàn)證。經(jīng)酵母雙雜交篩選與驗(yàn)證最終獲得2個(gè)與AM2相互作用的蛋白。選擇線粒體細(xì)胞色素C氧化酶亞單位Ⅱ(COXⅡ)蛋白進(jìn)行進(jìn)一步研究,構(gòu)建真核表達(dá)載體pcDNA3.0-HA-COXⅡ,并進(jìn)行GST-pull down和免疫熒光試驗(yàn)驗(yàn)證。結(jié)果表明,COXⅡ與AM2存在相互作用,且在細(xì)胞質(zhì)中存在共定位現(xiàn)象,為進(jìn)一步研究禽流感病毒的致病機(jī)理和病毒-宿主相互作用機(jī)制提供了理論基礎(chǔ)。
[Abstract]:Yeast two-hybrid system was used to screen host proteins interacting with AM2 of avian influenza virus, which could provide basis for the study of new biological functions of AM2 protein. The bait plasmid pGBKT7-AM2, which is nontoxic and self-activated, was fused with the lymphoid library plasmid, and the positive clones screened by selective culture medium were digested and sequenced. The positive clones were tested by yeast rotation test. Two proteins interacting with AM2 were obtained by yeast two-hybrid screening and verification. The mitochondrial cytochrome C oxidase subunit 鈪，

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