CHARACTERIZATION AND SUBCLONING OF A NEW HUMAN ONCOGENE

新人类癌基因的表征和亚克隆

• 批准号: 4692483
• 负责人: G VECCHIO
• 金额: --
• 依托单位: DIVISION OF CANCER EPIDEMIOLOGY AND GENETICS
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/4692483
• 关键词: B lymphocyte; clone cells; gene expression; genetic library; genetic mapping; human tissue; lymphoma; lymphosarcoma; messenger RNA; molecular oncology; nucleic acid sequence; oncogenes

A new human oncogene derived from the DNA of the spleen of a patient with a poorly differentiated B lymphocytic lymphoma has recently been cloned in cosmids in our laboratory. We have begun characterization of two cosmid clones by restriction endonuclease mapping of the oncogene. The first clone, designated C-8-X-1-1, has been shown to contain both human and mouse (NIH/3T3)-derived sequences. The human sequences of the C-8-X-1-1 (which had previously been shown to be devoid of biological activity, i.e., transformation of NIH/3T3 cells), have been shown to overlap partially with the human sequences contained in the cosmid clone C-14-1-2, which was shown to possess transforming activity. This last clone has been partially characterized as far as the enzyme restriction map is concerned and several fragments of it have been purified and subcloned either in pBR322 or in pAT-153. Some of the fragments, which are almost completely free of repetitive human sequences (Alu-negative), have been used to screen a normal human genomic library in order to detect sequences homologous to the new oncogene in normal human DNA. Three phages containing sequences homologous to some of the isolated fragments have been isolated so far and their further characterization is currently being performed. The Alu-negative fragments isolated from the cloned oncogene, as well as some Alu-containing sequences, also isolated by us, are being used to detect the mRNA for the gene in transfected NIH/3T3 cells and will be used to screen fresh normal and tumor tissues for expression of the newly isolated oncogene.

一个新的人类癌基因来源于一个患有癌症的病人的脾脏DNA， 低分化B淋巴细胞淋巴瘤最近被克隆在 在我们的实验室里。 我们已经开始对两种粘粒进行表征 通过癌基因的限制性内切酶图谱的克隆。 第一 一个命名为C-8-X-1-1的克隆已被证明含有人和小鼠两种基因 (NIH/3 T3）-衍生序列。 C-8-X-1-1的人类序列（ 先前已显示缺乏生物活性，即， NIH/3 T3细胞的转化），已显示与 粘粒克隆C-14-1-2中所含的人类序列， 具有转化活性。 最后一个克隆体已经部分地 就酶限制性内切酶图谱而言， 其片段已被纯化并亚克隆到pBR 322或 PAT-153 有些碎片几乎完全没有 重复的人类序列（阴性），已被用于筛选 正常人基因组文库，以检测与正常人基因组文库同源的序列。 正常人类DNA中的新癌基因。 三个含环序列 与一些分离的片段同源的基因已经分离出来， 目前正在对其进行进一步的鉴定。 的 从克隆的癌基因中分离出的阴性片段，以及一些 我们也分离出了含有DNA的序列， 在转染的NIH/3 T3细胞中表达该基因的mRNA，并将用于筛选 新鲜的正常和肿瘤组织，用于表达新分离的 癌基因