Nalbantoglu J, Goncalves O, Meuth M
J Mol Biol. 1983 Jul 5;167(3):575-94. doi: 10.1016/s0022-2836(83)80099-0.
To determine the types of gene structural alterations causing deficiency of adenine phosphoribosyl transferase (aprt) activity in spontaneous and chemically induced mutations of cultured somatic cells, we analyzed the restriction enzyme cleavage patterns of aprt gene sequences in mutant strains selected from Chinese hamster ovary cells. Patterns of aprt-containing fragments in Southern blots were mostly unchanged in our collection of 280 ethyl methane sulfonate-induced and spontaneous aprt- mutants, suggesting that base-pair changes or other alterations below our limit of resolution on agarose gels (approximately 50 base-pairs) are responsible for the great majority of mutations at the aprt locus. Occasionally, these mutations could be localized when they resulted in the loss or gain of a restriction enzyme site and the generation of new fragments of predictable size. Deletions of aprt-containing sequences were detected in only eight of 119 spontaneous mutants and in only one ethyl methane sulfonate-induced mutant. An insertion of 300 base-pairs near the 5' end of the aprt structural gene was found in one spontaneous aprt- strain. This insertion mutant was stable with a reversion frequency of less than 2 X 10(-7). Several unstable aprt- mutants were detected in our collection, but these had no observable alterations of aprt coding or flanking sequences.
为了确定在培养的体细胞自发突变和化学诱导突变中导致腺嘌呤磷酸核糖转移酶（aprt）活性缺乏的基因结构改变类型，我们分析了从中国仓鼠卵巢细胞中筛选出的突变株中aprt基因序列的限制性内切酶切割模式。在我们收集的280个甲磺酸乙酯诱导的和自发的aprt突变体中，Southern印迹中含aprt片段的模式大多未发生变化，这表明碱基对变化或其他低于我们在琼脂糖凝胶上分辨率极限（约50个碱基对）的改变是aprt基因座上绝大多数突变的原因。偶尔，当这些突变导致限制性酶切位点的丢失或获得以及产生可预测大小的新片段时，这些突变可以定位。在119个自发突变体中仅检测到8个含有aprt序列的缺失，在甲磺酸乙酯诱导的突变体中仅检测到1个。在一个自发的aprt突变株中发现了在aprt结构基因5'端附近插入300个碱基对。这个插入突变体是稳定的，回复频率小于2×10^(-7)。在我们的收集中检测到几个不稳定的aprt突变体，但这些突变体的aprt编码或侧翼序列没有可观察到的改变。
