Zdzienicka M Z, Verhaegh G W, Jongmans W, Morolli B, Jaspers N G, Oshimura M
MGC-Department of Radiation Genetics and Chemical Mutagenesis, University of Leiden, The Netherlands.
Int J Radiat Biol. 1994 Dec;66(6 Suppl):S189-95.
In order to isolate a human gene complementing the defect in A-T-like hamster cell mutants, the mutants were used as recipients for genomic DNA transfection, using either HeLa chromosomal DNA or DNA from a human cosmid library. Three primary transformants with an intermediate X-ray sensitivity and almost normal sensitivity to MMS, but retaining radioresistant DNA synthesis (RDS), were obtained. To identify the human chromosome that complements the defect in the A-T-like mutants, and to assess the degree of complementation for survival and RDS, microcell-mediated chromosome transfer was used. At least 20 independent hybrid clones between the mutant and each one of the human chromosomes 1, 2, 4, 5, 15, 17 or 18 were isolated. All hybrid clones remained X-ray sensitive, except one with chromosome 4, and another with chromosome 15, both showing an intermediate X-ray sensitivity. By using in situ hybridization we found that this partial correction was due to the presence of a mouse chromosome. In these two hybrids containing the mouse chromosome together with human chromosome 4 or 15, RDS was fully complemented only in the hybrid with chromosome 4 but not in the one containing chromosome 15, suggesting that RDS and X-ray sensitivity may be complemented independently.
为了分离出可弥补类共济失调毛细血管扩张症（A-T）仓鼠细胞突变体缺陷的人类基因，将这些突变体用作基因组DNA转染的受体，转染所用的DNA来源为HeLa染色体DNA或人类黏粒文库的DNA。获得了三个具有中等X射线敏感性且对甲基磺酸甲酯（MMS）几乎具有正常敏感性，但保留了抗辐射DNA合成（RDS）能力的初级转化体。为了鉴定可弥补类A-T突变体缺陷的人类染色体，并评估其对存活和RDS的互补程度，采用了微细胞介导的染色体转移技术。在突变体与人类1号、2号、4号、5号、15号、17号或18号染色体中的每一条之间，至少分离出了20个独立的杂交克隆。除了一个含有4号染色体的克隆和另一个含有15号染色体的克隆表现出中等X射线敏感性外，所有杂交克隆对X射线仍然敏感。通过原位杂交，我们发现这种部分校正归因于一条小鼠染色体的存在。在这两个同时含有小鼠染色体以及人类4号或15号染色体的杂交克隆中，只有含有4号染色体的杂交克隆中的RDS得到了完全互补，而含有15号染色体的杂交克隆中的RDS未得到完全互补，这表明RDS和X射线敏感性可能是独立互补的。
