人癌胚抗原转基因小鼠的建立

发布时间：2018-06-02 01:54

  本文选题：显微注射 + CEA　； 参考：《郑州大学》2006年硕士论文

【摘要】：转基因动物技术的出现是20世纪生物技术领域中的重大事件，而生物技术将是21世纪的支柱产业之一，是经济和科技发展的关键技术。转基因技术使人们能从整体、器官、细胞、分子多层次，从发育时间和组织生理空间多角度，立体多维地研究基因，并可按人类的意愿改造和修饰物种。 癌胚抗原(carcinoembryonic antigen，CEA)是胚胎发展中产生的抗原之一，是一种最常见的肿瘤相关抗原，是目前国际上公认的肿瘤标记物。其基因定位于人类19号染色体长臂(19q13.1～13.2)，分子量约为180kd，为免疫球蛋白超家族的成员。CEA在正常消化道黏膜仅少量分泌，而90％的消化道恶性肿瘤、胰腺癌，，70％非小细胞肺癌和50％乳腺癌有CEA的高表达，故CEA可作为这些肿瘤检测和治疗的靶分子。 本实验以小鼠为研究对象，用显微注射法将包含小鼠乳清酸蛋白基因启动子(约2.4kb)、人癌胚抗原的编码序列(约2.1kb)及polyA信号的转基因构件转入原核期小鼠胚胎，制备了CEA转基因小鼠，旨在研究CEA与乳腺癌等肿瘤发生、发展的关系，为以后制作小鼠乳腺生物反应器提供经验和参考。主要进行了以下工作： 提纯pWA-CEA质粒，分别进行HindⅢ单酶切、HindⅢ和Xho Ⅰ双酶切、SacⅡ和KpnⅠ双酶切鉴定。对构建成功的pWA-CEA质粒用SacⅡ和KpnⅠ双酶切得到目的基因(约4.9kb)，胶回收并纯化目的片断，溶于TE(10mmol／L Tris-HCl，pH7.4；0.2mmol／L EDTA)中，注射用DNA浓度为3μg／ml，储存-20℃备显微注射用。将雌鼠做超排卵处理，与正常雄鼠交配，取其受精卵，培养至单细胞双原
[Abstract]:The emergence of transgenic animal technology is an important event in the field of biotechnology in the 20th century, and biotechnology will be one of the pillar industries in the 21st century and the key technology for the development of economy and science and technology. Transgenic technology enables people to study genes in a three-dimensional and multidimensional manner from the perspectives of whole, organ, cell and molecular levels, development time and tissue physiological space, and can modify and modify species according to human wishes. Carcinoembryonic antigen (CEA) is one of the antigens produced in embryonic development. It is one of the most common tumor-associated antigens and is an internationally recognized tumor marker. Its molecular weight is about 180 kd.The gene is located in human chromosome 19, 19q13.1 and 13.2kd. it is a member of the immunoglobulin superfamily, which secretes only a small amount of CEA in the normal gastrointestinal mucosa, while 90% of the malignant tumors of the digestive tract. 70% of non-small cell lung cancer and 50% of breast cancer have high expression of CEA, so CEA can be used as a target molecule for the detection and treatment of these tumors. In this study, mice were studied. Transgenic components containing mouse whey acid protein gene promoter (about 2.4 kb), human carcinoembryonic antigen coding sequence (about 2.1 kb) and polyA signal were transferred into prokaryotic mouse embryos by microinjection. CEA transgenic mice were prepared to study the relationship between CEA and carcinogenesis and development of breast cancer, and to provide experience and reference for the production of mouse mammary gland bioreactor in the future. The main tasks were as follows: PWA-CEA plasmids were purified and identified by single enzyme digestion of Hind 鈪

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