点突变ras癌基因全cDNA真核表达重组体的构建及诱导NIH3T3细胞转化的研究

将活化癌基因Ｔ２４－ｒａｓ的全ｃＤＮＡ序列正向插入真核载体ｐＭＡＭｎｅｏ，构建成重组质粒ｐＭＡＭｎｅｏ－Ｔ２４－ｒａｓ．将该质粒转染ＮＩＨ３Ｔ３细胞，通过药物筛选，建成细胞系３Ｔ３（Ｔ２４）．Ｓｏｕｔｈｅｒｎ杂交证明外源Ｔ２４－ｒａｓ基因已整合于受体细胞染色体中．３Ｔ３（Ｔ２４）细胞表现出形态学方面的明显变化：具失去接触抑制能力，且在裸鼠体内致瘤等恶性行为．本文构建的Ｔ２４－ｒａｓ基因真核表达重组体和建立的转化细胞系可用于肿瘤的诊断、预防、治疗及抗肿瘤药物的筛选、评估等研究．

STUDIES ON THE MALIGNANT TRANSFORMATION OF NIH3T3 CELLS AFTER INTRODUCING RECOMBINANT PLASMID OF FULLLENGTH ACTIVATED RAS ONCOGENE

The recombinant plasmid pMAMneo-T24-ras was constructed by inserting the fulllength cDNA of mutant T24-ras into mammalian expressive vector pMAMneo.The transformed cell line,3T3(T24)was obtained by introducing pMAMneo-T24-ras plasmid into NIH3T3 cells and selecting with G418. Southern blot analysis showed that exogenous T24-ras oncogene was integrated into cell genome.In addition,the transformed cell line not only showed the morphological changes but also possessed the malignant behavior of loss of contact inhibition and tumorigenesis in nude mouse. Eukaryotic recombinant containing activated ras oncogene and corresponding transformed cell line which were constructed and established respectively can be used for study on blocking oncogene expression by antisense RNA,inhibiting malignant changes of cells and selecting antitumor drugs and so on.