ISOLATION OF CHINESE-HAMSTER CELL MUTANTS DEFICIENT IN DIHYDROFOLATE-REDUCTASE ACTIVITY
ISOLATION OF CHINESE-HAMSTER CELL MUTANTS DEFICIENT IN DIHYDROFOLATE-REDUCTASE ACTIVITY
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DOI：
10.1073/pnas.77.7.4216
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发表时间：
1980-01-01
期刊：
影响因子：
--
通讯作者：
CHASIN, LA
中科院分区：
文献类型：
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作者：
URLAUB, G;CHASIN, LA
在诱变和暴露于高比活性[3 H]脱氧尿苷作为选择剂后，分离缺乏二氢叶酸还原酶（四氢叶酸脱氢酶，7，8-二氢叶酸：NADP+氧化还原酶; EC 1.5.1.3）活性的中国仓鼠卵巢细胞突变体。完全缺陷的突变体不能从野生型细胞开始分离，但可以容易地从含有野生型水平的二氢叶酸还原酶活性的一半的推定杂合子中选择。使用[3 H]脱氧尿苷和甲氨蝶呤降低细胞内二氢叶酸还原酶活性，从野生型细胞中选择杂合子。完全缺陷突变体需要甘氨酸、嘌呤和胸苷生长;这种表型在细胞杂交中对野生型是隐性的。分离出回复突变体，其中一种产生热不稳定的二氢叶酸还原酶活性。这些突变体可能是有用的代谢研究有关癌症化疗和精细结构的基因突变作图，通过使用可用的分子探针，该基因。
Mutants of Chinese hamster ovary cells lacking dihydrofolate reductase (tetrahydrofolate dehydrogenase, 7,8-dihydrofolate:NADP+ oxidoreductase; EC 1.5.1.3) activity were isolated after mutagenesis and exposure to high-specific-activity [3H]deoxyuridine as a selective agent. Fully deficient mutants could not be isolated starting with wild-type cells, but could readily be selected from a putative heterozygote that contains half of the wild-type level of dihydrofolate reductase activity. The heterozygote was selected from wild-type cells using [3H]deoxyuridine together with methotrexate to reduce intracellular dihydrofolate reductase activity. Fully deficient mutants require glycine, a purine and thymidine for growth; this phenotype is recessive to wild type in cell hybrids. Revertants were isolated, one of which produces a heat-labile dihydrofolate reductase activity. These mutants may be useful for metabolic studies relating to cancer chemotherapy and for fine-structure genetic mapping of mutations by using available molecular probes for this gene.