本文選題：細(xì)菌噬菌體 + 免疫球蛋白G��； 參考：《安徽醫(yī)科大學(xué)學(xué)報(bào)》2013年08期

【摘要】：目的使用人和羊不同物種IgG來誘導(dǎo)噬菌體展示免疫球蛋白結(jié)合分子單結(jié)構(gòu)域隨機(jī)組合文庫進(jìn)行體外分子進(jìn)化篩選,判斷不同的Fc段構(gòu)像是否具有特異的結(jié)合優(yōu)勢,同時(shí)探索其優(yōu)勢結(jié)合序列的組合特點(diǎn)。方法通過PCR獲得金黃色葡萄球菌蛋白A(SpA)的A、B、D、E和鏈球菌(C和G群)蛋白G(SpG)的B2、B3各單結(jié)構(gòu)域片段,構(gòu)建由該6個(gè)片段隨機(jī)組合的噬菌體展示文庫。再分別使用人和羊IgG對文庫進(jìn)行體外親和篩選,以期能夠獲得具有特異優(yōu)勢結(jié)合能力的組合分子,并在Phage ELISA的水平上進(jìn)一步驗(yàn)證篩選結(jié)果。結(jié)果成功構(gòu)建了符合進(jìn)行體外進(jìn)化篩選要求的噬菌體展示單結(jié)合結(jié)構(gòu)域隨機(jī)組合文庫后,經(jīng)過6輪人IgG誘導(dǎo)篩選,文庫的展示片段大小統(tǒng)一為3個(gè)結(jié)構(gòu)域組合分子,E-B-B3;經(jīng)過6輪羊IgG誘導(dǎo)篩選,文庫的展示片段大小也統(tǒng)一為4個(gè)結(jié)構(gòu)域組合分子,D-A-B3-B3。Phage ELISA進(jìn)一步驗(yàn)證最終組合序列的對人和羊IgG的結(jié)合能力。結(jié)論首次得到兩種天然SpA、SpG分子中不存在的,且分別特異地與人和羊IgG具有強(qiáng)結(jié)合作用的新型組合分子E-B-B3、D-A-B3-B3,在人和羊IgG的純化和檢測具有很大的應(yīng)用前景。
[Abstract]:Objective to use IgG from different species of human and sheep to induce phage display immunoglobulin binding molecular single domain random combination library for in vitro molecular evolution screening to determine whether different FC conformation has specific binding advantage. At the same time, it explores the combination characteristics of its advantage combination sequence. Methods the single domain fragments of staphylococcus aureus protein Agna spa E and streptococcal G group C and G group were obtained by PCR, and the phage display library was constructed by random combination of the six fragments. Then human and sheep IgG were used to screen the library in vitro in order to obtain the combination molecules with specific dominant binding ability and to further verify the screening results at the level of Phage Elisa. Results the phage display mono-binding domain random combination library was successfully constructed after six rounds of human IgG induction screening. The displayed fragment size of the library was unified into three domain assemblage molecules, E-B-B3, which were screened by six rounds of sheep IgG induction. The displayed fragment size of the library was also unified into four domain combinators, D-A-B3-B3.Phage Elisa, to further verify the binding ability of the final combination sequence to human and sheep IgG. Conclusion for the first time, two kinds of natural SpAG-free molecules, E-B-B3D-A-B3-B3, which are non-existent and strongly bound to human and sheep IgG, are obtained for the first time, which have great application prospect in the purification and detection of human and sheep IgG.
【作者單位】： 安徽醫(yī)科大學(xué)病理生理學(xué)教研室;第二軍醫(yī)大學(xué)微生物學(xué)教研室 上海市醫(yī)學(xué)生物防護(hù)重點(diǎn)實(shí)驗(yàn)室;安慶醫(yī)藥高等專科學(xué)校;
【基金】：安徽省教育廳自然基金(編號:KJ2010A117) 國家自然科學(xué)基金項(xiàng)目(編號:30872405、30872246、30972632、30972799) 上海市基礎(chǔ)研究重點(diǎn)項(xiàng)目(編號:08JC1405200)
【分類號】：R392.7

【參考文獻(xiàn)】

相關(guān)期刊論文 前5條

1 劉文俊;趙祖國;呂世靜;晏雙雙;;噬菌體展示技術(shù)在抗體親和力成熟中的應(yīng)用進(jìn)展[J];免疫學(xué)雜志;2012年07期

2 徐容,沈毅佒,鄧松華,蔡春曉,陳秋莉,賈建安,王錦紅,潘欣,潘衛(wèi);噬菌體展示protein A及protein L Ig結(jié)合單結(jié)構(gòu)域隨機(jī)組合文庫及Ig親和篩選[J];生物化學(xué)與生物物理進(jìn)展;2005年06期

3 葛宜兵;楊旭芳;杜哲明;龐強(qiáng);曹潔;陳秋莉;王錦紅;張華群;廖文婷;祁培培;劉超;章萍萍;鄧松華;潘衛(wèi);;噬菌體展示HIV-1 Tat38-61堿性區(qū)51和55位隨機(jī)突變體文庫的構(gòu)建[J];生物工程學(xué)報(bào);2011年05期

4 祁培培;丁瑩瑩;吳莉莉;陳秋莉;王錦紅;劉超;廖文婷;張婧;曹潔;潘衛(wèi);;人IgG四亞類對噬菌體展示Ig結(jié)合蛋白單結(jié)構(gòu)域隨機(jī)組合文庫的體外進(jìn)化篩選[J];生物工程學(xué)報(bào);2012年09期

5 徐容,潘衛(wèi),沈毅,陳秋莉,潘欣,馮春芝,戚中田,劉彥君,鄧松華;新型噬菌體展示載體pCANTAB5S的構(gòu)建[J];安徽醫(yī)科大學(xué)學(xué)報(bào);2004年02期

【共引文獻(xiàn)】

相關(guān)期刊論文 前10條

1 楊華,李連青,王蓓霞,徐容,沈毅s，

本文編號：2016895