人tumstatin基因的克隆表达及其鉴定.pdfVIP

8·36 · 肿瘤防治研究 2007 年第 34 卷第 11 期 人 t u m st atin 基因的克隆表达及其鉴定 颜廷东 , 宋娜玲 , 赵启仁 , 刘家慧 , 贺 　欣 , 褚丽萍 , 张春明 Cloning of Human tumstatin Gene and Expression of Its Recombinant Protein YAN Tingdong , SON G Naling , ZHAO Qiren , L IU J iahui , H E Xin , C HU Liping , ZHAN G Chun ming Key L aboratory of M olecul a r N uclea r M e d ici ne , I ns ti t ute of R a d i ation M e d ici ne , Chi nese A ca demy of M e d ical S ciences an d Pek i ng Union M e d ical Col leg e , Ti anj i n 300 192 , Chi na Corresp on d i ng A ut hor : Z HA O Qiren , Em ai l :z haoqi ren98 @y ahoo. com. cn Abstract :Objective 　 To clone human t umst atin gene , exp ress it s recombinant p rot ein for furt her re search . Methods 　Tot al RN A was ext ract ed from H E K293 cell . The t um st atin gene wa s amp lified by R TPCR , t hen cloned into p MD 19T and sequenced . Sub sequently , t he t um st atin gene wa s cloned into t he p E T2 8a and t ran sformed into E. col i BL 2 1 (D E3) where it wa s induced to exp ress by IP T G. The p roduct s were identified by SD SPA GE and West ern blot . Results 　R TPCR p roduct wa s about 750bp , it s sequence wa s t he same a s t hat of t um st atin report ed . The exp ression vector p E T2 8at um st atin was con st ruct ed successfully , and t here wa s a new p rot ein band about Mr 29 KD on SD SPA GE. The ratio of t he exp ressed p roduct to tot al bact erial p rot ein s was 30 %. The result of West ern blot demon st rat ed t hat t he exp ressed p roduct wa s t he t um st atin p rot ein . Conclusion 　Human t um st atin gene was cloned and it s recombinant p rot ein s were exp ressed successfully in t hi s st udy . Key words :t um st atin ; Tumor angio genesi s inhibitor ; Gene cloning and exp ressing 摘 　要 : 目的　重组表达人 t um st atin ,为人 t umst atin 的肿瘤受体显像奠定基础 。方法 　提取 H E K293 细胞总 RN A ,通过 R TPCR