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	鸡Rad51蛋白的体外表达与纯化及其抗血清的制备; Prokaryotic expression,purification of chicken Rad51 protein and generation of its antiserum
	刘宽灿 ; 王恒 ; 孟艳敏 ; 叶军 ; 袁立
	2011
关键词	鸡的Rad51 蛋白表达 抗体制备 Chicken Rad51 Protein expression Generation of antibody
英文摘要	目的:rAd51蛋白在双链断裂的dnA修复的同源重组中发挥重要作用。因而其过量表达在细胞内的基因操作中被用于在提高基因同源重组的效率。当前在鸡中还未有针对该蛋白的特异性抗体。为此我们对鸡的rAd51进行了体外表达与纯化,并制备了CrAd51抗体,为该蛋白质在鸡细胞中的功能研究及探讨其用于鸡细胞中提高同源重组的研究提供有效工具。方法:对鸡rAd51基因进行全长克隆并构建原核表达载体PET-28A(+),用IPTg诱导该蛋白在rOSSETA菌株中表达,纯化后注射bAlb/C小鼠获得抗体。结果:成功制备鸡rAd51特异多克隆抗体,ElISA显示该抗体血清效价达51 200,该抗体血清可以用于WESTErn等的蛋白检测技术。结论:针对鸡基因产物制备抗体的技术方法有效可行;首次成功获得鸡rAd51蛋白及多克隆抗体,为与该蛋白相关的研究提供了有效工具。; Objective:RAD51 protein plays a major role in homologous recombination of DNA double strand break repair.Overexpression of this protein has thus been used to enhance the homologous recombination efficiency in gene manipulations in cells.Currently,there is not yet an antibody available for this chicken protein.We therefore carried out in vitro expression,purification of the chicken Rad51,and tried to prepare the antibody.Methods:Full length of chicken Rad51 coding sequence was cloned and inserted into pET-28a(+) to generate a prokaryotic expression constructs;the expression was induced with IPTG in a Rosseta strain.After the purification,the gene product was procedurally injected into BALB/c mouse to generate antibody.Results:ELISA test showed that the titer of antiserum reached 51200;The antiserum had a good performance in Western blot for both sensitivity and specifity.Conclusion:The protocol adapted in this study works well in generating antiserum against chicken gene product;We have for the first time prepared an antiserum against chicken Rad51 providing a useful tool for studies on related to the chicken Rad51.; 科技部重大基础研究计划资助(2009CB941601)
语种	zh_CN
内容类型	期刊论文
源URL	[http://dspace.xmu.edu.cn/handle/2288/119076]
专题	生命科学－已发表论文
推荐引用方式 GB/T 7714	刘宽灿,王恒,孟艳敏,等. 鸡Rad51蛋白的体外表达与纯化及其抗血清的制备, Prokaryotic expression,purification of chicken Rad51 protein and generation of its antiserum[J],2011.
APA	刘宽灿,王恒,孟艳敏,叶军,&袁立.(2011).鸡Rad51蛋白的体外表达与纯化及其抗血清的制备..
MLA	刘宽灿,et al."鸡Rad51蛋白的体外表达与纯化及其抗血清的制备".(2011).

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