Molecular cloning of a mouse DNA repair gene that complements the defect of group-A xeroderma pigmentosum.
Molecular cloning of a mouse DNA repair gene that complements the defect of group-A xeroderma pigmentosum.
复制标题
小鼠 DNA 修复基因的分子克隆，可补充 A 组色素性干皮病的缺陷。
DOI：
--
复制
发表时间：
1989
影响因子：
11.1
通讯作者：
Y. Okada
中科院分区：
文献类型：
--
作者：
K. Tanaka;I. Satokata;Z. Ogita;T. Uchida;Y. Okada
为了分离负责着色性干皮病 (XP) 互补 A 组的基因，将质粒 pSV2gpt 和来自小鼠胚胎的基因组 DNA 共转染到 XP2OSSV 细胞（A 组 XP 细胞系）中。从约1.6 X 10(5) pSV2gpt 转化的XP 菌落中分离出两个原代抗UV XP 转染子。将pSV2gpt和来自初级转染子的基因组DNA再次共转染到XP2OSSV细胞中，并通过筛选约4.8×10(5)个pSV2gpt转化的XP集落获得次级抗UV XP转染子。二次转染子保留了较少的小鼠重复序列。将补充 XP2OSSV 细胞缺陷的小鼠基因从二次转染子的基因组中克隆到 EMBL3 载体中。克隆的 DNA 的转染还赋予另一组 A XP 细胞系以 UV 抗性，但不赋予 C、D、F 或 G 组的 XP 细胞系。以克隆的小鼠 DNA 修复基因的亚片段作为探针对 Poly(A)+ RNA 进行 Northern 印迹分析，结果显示在供体小鼠胚胎和二次转染子中转录了大约 1.0 千碱基的 mRNA，并且大约 1.0 和大约 1.3 千碱基的 mRNA 被转录。在正常人类细胞中转录，但在 A 组 XP 细胞的三种菌株中均未检测到这些 mRNA。这些结果表明，克隆的DNA修复基因对A组XP具有特异性，并且可能是A组XP人类基因的小鼠同源物。
For isolation of the gene responsible for xeroderma pigmentosum (XP) complementation group A, plasmid pSV2gpt and genomic DNA from a mouse embryo were cotransfected into XP2OSSV cells, a group-A XP cell line. Two primary UV-resistant XP transfectants were isolated from about 1.6 X 10(5) pSV2gpt-transformed XP colonies. pSV2gpt and genomic DNA from the primary transfectants were again cotransfected into XP2OSSV cells and a secondary UV-resistant XP transfectant was obtained by screening about 4.8 X 10(5) pSV2gpt-transformed XP colonies. The secondary transfectant retained fewer mouse repetitive sequences. A mouse gene that complements the defect of XP2OSSV cells was cloned into an EMBL3 vector from the genome of a secondary transfectant. Transfections of the cloned DNA also conferred UV resistance on another group-A XP cell line but not on XP cell lines of group C, D, F, or G. Northern blot analysis of poly(A)+ RNA with a subfragment of cloned mouse DNA repair gene as the probe revealed that an approximately 1.0 kilobase mRNA was transcribed in the donor mouse embryo and secondary transfectant, and approximately 1.0- and approximately 1.3-kilobase mRNAs were transcribed in normal human cells, but none of these mRNAs was detected in three strains of group-A XP cells. These results suggest that the cloned DNA repair gene is specific for group-A XP and may be the mouse homologue of the group-A XP human gene.