中国兽医科学2017，Vol.47Issue(6)：701-706,6.DOI:10.16656/j.issn.1673-4696.2017.06.005
鸡细胞周期蛋白依赖性激酶抑制蛋白p21单克隆抗体的制备
Generation of monoclonal antibody against chicken cyclin-dependent kinase inhibitor p21
摘要
Abstract
To prepare monoclonal antibody against chicken p21,the p21 gene was amplified by reverse transcription polymerase chain reaction(RT-PCR) and subcloned into the prokaryotic expression vector,pET-30a.Therecombinant plasmid,designated as pET-30a-p21,was transformed into E.coli BL21(DE3)competent cells.Separated by SDS-PAGE,the induced protein was shown to be expressed as inc]usionbodies.The positive bands were cut,ground and used as antigens to innunize BALB/c mice intraperitoneally for three times.The spleen was isolated and fused with SP2/0.Hybridoma clones that produced antibodies against p21 were screened by IFA,ELISA and limited dilution.The antibody levels were detected by indirect immunofluorescence assay (IFA),enzyme-linked immunosorbent assay (ELISA) and Western-blot.Twelve positive hybridoma clones were screened and identified as 1B12,1E12,1H7,2B5,2C5,C8,2D8,2F2,3C11,3D6,G7 and 4G6.The monoclonal antibody against p21 was prepared as a tool for the study of p21 functions.关键词
鸡/p21/单克隆抗体/原核表达Key words
chicken/p21/monoclonal antibody/prokaryotic expression分类
农业科技引用本文复制引用
李雪琪,连雪,鲍晨沂,孙海伟,陈鸿军,JUNG Yong-Sam,钱莺娟..鸡细胞周期蛋白依赖性激酶抑制蛋白p21单克隆抗体的制备[J].中国兽医科学,2017,47(6):701-706,6.基金项目
国家自然基金面上项目(31472218) （31472218）
江苏省自然科学青年基金项目(BK20140711) （BK20140711）
留学回国人员科研启动基金项目(教外司留[2014]1685号) （教外司留[2014]1685号）
中央高校基本业务费(Y0201300526,Y0201300527) （Y0201300526,Y0201300527）
江苏省优势学科专项经费 （）
