3 562-565 基础研究 4 肝脏特异性基因疫苗研究.pdfVIP

现代食品科技 Modern Food Science and Technology 2010, Vol.26, No.6 肝脏特异性基因疫苗研究 张利涛，刘慧琳，曹以诚 （华南理工大学生物科学与工程学院，广东广州510006 ） 摘要：本研究构建了肝脏特异性表达的载体，为进一步构建肝脏特异性的siRNA载体做前期准备。通过PCR方法扩增得到 人Alpha-1抗胰蛋白酶（human-α1-anti- Trypsin ）启动子片段（以下简称hAAT启动子）。回收纯化后克隆到pCDNA6载体中，同时以EGFP作为标记蛋白。将构建好的 载体分别转染到肝癌细胞HePG2 、乳腺癌细胞MDA-MB- 231和肺癌细胞A549 中，验证其在肝脏中特异性表达的能力。然后在荧光显微镜下观察。结果表明：载体在肝癌细胞中能很好 的表达EGFP ，在肺癌细胞A549 、乳腺癌细胞MDA-MB-231 中不表达EGFP 。说明采用human-α1-anti- Trypsin启动子可以实现目的基因在肝癌细胞中特异性表达，这项研究应用于抗肝癌、肝炎的siRNA基因治疗，将有利于显著提 高治疗效果，减少对身体的副作用。 关键词：特异性表达；HepG2；hAAT启动子；EGFP 文章篇号：1673-9078(2010)6-562-565 Studies on Liver-specific Gene Vaccine ZHANG Li-tao, LIU Hui-lin, CAO Yi-cheng (School of Bioscience Bioengineering, South China University of Technology, Guangzhou 510006, China) Abstract: Liver-special expression vector was constructed for the further preparation of the liver-special expression siRNA. The hAAT promoter was amplified by PCR, which was purified and cloned to pCDNA6 vector. EGFP was the marker protein. pCDNA6- hAAT-EGFP and pCDNA6-CMV-EGFP were transfect to HepG2, MDA-MB-231, A549 cell lines, to validate the specificity of the vector. Then, these cell lines were observed by the fluorescence microscope. Result showed that, pCDNA6-hAAT-EGFP can express EGFP in the HepG2 cell line. However, no EGFP was expressed in MDA-MB-231 and A549 cell line. The vector with hAAT promoter can achieve specific expression in HepG2 cell line. It will dramatically improved the therapeutic effect,and reduced adverse effect in siRNA treatent. Key words: specific expression; HepG2; hAAT promoter; EGFP 562 肝脏的疾病，如肝癌、肝炎是严重影响人 公司，DM