多发性骨髓瘤恶性相关基因快速表达克隆法的建立y.PDF

7 1 生命科学研究 Vol.7 No. 1 2003 3 Life Science Research Mar. 2003 田菁燕, 胡维新, 钟 慧 ( , 410078) : 为从多发性骨髓瘤细胞中克隆与鉴定恶性 关基因并试图阐明其发病分子机制, 建立了一种改进的 快速表达克隆法, 简单快捷直接从多发性骨髓瘤细胞株ARH 77 cDNA 文库中克隆恶性 关基因. 其特点是 常用的表达克隆法与经典的DNA 介导的NIHP3T3 转染实验 结合, 直接从cDNA 表达文库中克隆恶性 关基 因. 首先建立高质量的cDNA 表达文库, 将 cDNA 表达文库分成几 个基因池, 转染NIHP3T3 细胞; 将转化活性最 强的基因池再分成几 个亚基因池, 转染NIHP3T3 细胞; 将转化活性最强的亚基因池再分成次级亚基因池, 直到 获得有明显转化活性的单 个cDNA 克隆. 该技术亦可用于从其他肿瘤细胞中克隆恶性 关基因. : 基因克隆; 表达克隆法; 恶性 关基因; 多发性骨髓瘤 : Q786 : A : 1007 7847(2003) 01 0042 06 An Improved Expression Technique for Quick Cloning of Tumor Associated Gene from Human Multiple Myeloma TIAN Jing yan,HU Wei xin, ZHONG Hui (Molecular B iology R esearch Center ,X iangy a Medical College , Central South Univ er sity , Changsha 410078,H unan , China) A stract: The cloning and identification of a tumor associated gene from human multiple myeloma ( MM) is one of the most important issues to elucidate itsmolecular mechanisms and to establish a novel strategy against it. SinceMM develops as a sporadic disease, linkage analysis is not available for tumor associated gene cloning.We report an ex pression screening technique to clone the tumor associated genes from human MM and this method can be used for the cloning of tumor associated gene from other malignant cells. mRNA was isolated from humanMM cell line ARH 77 and then cDNA was synthesized,which was linked to a kind of special adaptors and amplified by polymerase chain reaction (PCR) .The library was constructed by ligating cDNA with eukaryote expression vector pcDNA3. 1 ( + ) through E