摘要
消化道途径转基因过程方便、快捷、易适应,可为基因治疗提供全新的模式。为了研究人 生长激素(bGH)基因的经消化道途径转基因过程,实验首先应用ECHO克隆系统。在供载体 pUni-hGH和宿主载体pcDNA4／TO-E的基础上,构建出hGH的哺乳动物表达载体pcDNA4-hGH;然 后结合酿酒酵母表达载体pESC-URA,构建出hGH的酵母-哺乳动物穿梭栽体pESC-CMV-hGH,测 序鉴定后转化酿酒酵母。用阳性重组酵母对小鼠进行灌胃免疫实验,间接ELISA方法在实验组 小鼠的血清中检测到抗hGH抗体的存在。结果证实hGH基因可通过消化道途径转进小鼠体细 胞并进行表达,初步证明了hGH的消化道基因治疗的可行性。
Digestive tract gene transfer bears the characteristics of readiness, swiftness and easily adaptive, therefore, may become a novel model for gene therapy. To investigate the digestive tract hGH gene transfer processes, ECHO cloning system has been employed. Donor plasmid vector（pUni-hGH） and acceptor vector （pcDNA4/TO-E） were used to construct the mammalian expression vector pcDNA4-hGH. Further recombinant process employs yeast expression vector pESC-URA to construct a shuttle vector of mammalian-yeast expression plasmid（pESC-CMV-hGH）. After verification of the reading frame by DNA sequencing, the confirmed constructs were used to transfect the yeast host cells. Positive yeast clones were identified and propagated. The yeasts hosting the hGH construct were used to feed the experimental mice. ELISA tests confirmed that there are anti-hGH antibody existed in the blood serum of the experimental animals. These results demonstrated that the foreign hGH genes could be transferred into the host digestive tract through the yeast carriers, which substantiates the applicability of the digestive tract gene therapy.
出处
《中国生物工程杂志》
CAS
CSCD
北大核心
2005年第B04期239-243,共5页
China Biotechnology
基金
国家自然科学基金资助项目:36890044
关键词
消化道途径
人生长激素基因
基因治疗
基因转移
Digestive tract Human growth hormone gene（hGH） Gene therapy Digestive tract gene transfer
