Thacker J, Stretch A, Stephens M A
Mutat Res. 1977 Feb;42(2):313-26. doi: 10.1016/s0027-5107(77)80033-x.
The induction of mutation to purine analogue resistance was assessed in Chinese hamster V79-4 cells exposed to gamma-radiation. After irradiation, the cells were grown in non-selective medium for different time intervals before respreading into medium containing 0.5-0.7 mug/ml thioguanine. In some experiments colonies arising in thioguanine-medium were counter-selected in medium containing the glutamine analogue azaserine, which distinguishes mutants with very little activity of the enzyme hypoxanthine-guanine phosphoribosyl transferase. Only these mutants were increased in frequency by radiation, the maximum measured frequencies occuring in cells respread after 2 days growth in non-selective medium. With longer intervals of post-irradiation growth in non-selective medium a fraction of the induced mutants was lost, and after large doses of radiation it is doubtful if the maximum frequency observed after 2 days post-irradiation growth represents the true induced frequency. The detection of freshly-induced mutants seemed to depend upon the dilution and decay of products formed from the genes prior to their mutation by radiation, since (a) selection of mutants in a higher concentration of thioguanine (2 mug/ml) increased the post-irradiation growth interval required to detect the maximum frequency of induced mutants, and (b) with increasing duration of post-irradiation growth in the non-selective medium, induced mutant cells were able progressively to overcome the growth-limiting effects of the analogue, to give large colonies when respread in thioguanine-medium. The radiosensitivities of 7 isolated mutant lines were indistinguishable from that of wild type cells, but the mutants were at a slight disadvantage when grown in competition with wild type cells. This disadvantage was consistent with the expected fitness of mutants relative to wild type cells calculated from estimates of the spontaneous mutation rate and the mean spontaneous mutation frequency. The induction of mutation to thioguanine resistance was non-linear with dose, yielding induced frequencies per rad of 5-10(-8) to 3-10(-7), but a plot of induced mutation frequency against log surviving fraction gave an approximately linear relationship. The same linear relationship holds for recently-published data on human and mouse cell cultures, so that all three mammalian cell types exhibit the same fixed probability of mutation induction relative to the extent of inactivation caused by ionising radiation.
在中国仓鼠V79 - 4细胞中，评估了暴露于γ射线后对嘌呤类似物抗性的诱变情况。辐照后，细胞在非选择性培养基中培养不同时间间隔，然后再接种到含有0.5 - 0.7微克/毫升硫鸟嘌呤的培养基中。在一些实验中，在硫鸟嘌呤培养基中形成的菌落，在含有谷氨酰胺类似物重氮丝氨酸的培养基中进行反向选择，重氮丝氨酸可区分次黄嘌呤 - 鸟嘌呤磷酸核糖转移酶活性极低的突变体。只有这些突变体的频率因辐射而增加，在非选择性培养基中生长2天后再接种的细胞中测得的最大频率出现。在非选择性培养基中辐照后生长间隔更长时，一部分诱导突变体会丢失，大剂量辐射后，辐照后生长2天观察到的最大频率是否代表真正的最大频率值得怀疑。新诱导突变体的检测似乎取决于辐射诱变前基因形成的产物的稀释和衰减，因为：(a) 在较高浓度的硫鸟嘌呤（2微克/毫升）中选择突变体，会增加检测到诱导突变体最大频率所需的辐照后生长间隔；(b) 随着在非选择性培养基中辐照后生长时间的增加，诱导突变体细胞能够逐渐克服类似物的生长限制作用，当再接种到硫鸟嘌呤培养基中时形成大菌落。7个分离的突变株系的放射敏感性与野生型细胞无法区分，但与野生型细胞竞争生长时，突变体略有劣势。这种劣势与根据自发突变率估计值和平均自发突变频率计算出的突变体相对于野生型细胞的预期适应性一致。对硫鸟嘌呤抗性的诱变与剂量呈非线性关系，每拉德的诱导频率为5×10⁻⁸至3×10⁻⁷，但诱导突变频率与对数存活分数的关系图呈现出近似线性关系。最近发表的关于人类和小鼠细胞培养的数据也呈现出相同的线性关系，因此，相对于电离辐射引起的失活程度，所有三种哺乳动物细胞类型都表现出相同的固定诱变概率。
