TRAIL基因原核表达载体的构建、表达与抗体制备

Construction of Prokarytoic Expression Vector of TRAIL Gene and Preparation of Anti-TRAIL Antibody

目的:获取抗肿瘤坏死因子相关凋亡诱导配体(TRAIL)抗体。方法:利用PCR技术和基因重组技术将TRAIL基因构建于原核表达载体pPreTMH-Tb中,经酶切、测序鉴定证实构建完全正确后,通过IPTG诱导在大肠杆菌DH-5α中获得表达,并将TRAIL蛋白免疫家兔,通过Dot blot及Western blot检测抗TRAIL抗体的产生。结果:成功构建TRAIL基因原核表达载体,并在大肠杆菌DH-5α中获得表达,TRAIL蛋白表达量占菌体蛋白质总量的11.8％,Dot blot及Western blot检测证实获得了抗TRAIL抗体。结论:成功制备抗TRAIL抗体,这为TRAIL在真核细胞水平研究奠定了实验基础。

Objective: To obtain the anti-TRAIL antibody. Methods: A prokaryotic expression vector pPre-TMHTb was constructed by PCR and recombinant DNA techniques and was confirmed by enzymatic digestion and sequence analysis. TRAIL protein induced to express by IPTG in E. coli DH-5α. After TRAIL protein immunised rabbits, anti-TRAIL antibody was tested by Dot blot and Western blot. Results:A prokaryotic expression vector pPre-TMHTb was constructed and expressd in E. coli DH-5α. TRAIL protein was detected up to 11.8% of the total bacterial protein expressed and anti-TRAIL antibody was tested by Dot blot and Western blot. Conclusions: The anti-TRAIL antibody was obtained that provides an experimental basis for the studies of TRAIL in eukaryotic level.