Lai L W, Hart I M, Patterson D
Eleanor Roosevelt Institute for Cancer Research, Denver, Colorado 80206.
Genomics. 1991 Feb;9(2):322-8. doi: 10.1016/0888-7543(91)90260-l.
Somatic hybrids between human cells and the Chinese hamster ovary (CHO) K1 mutant, Ade -H cells, were selected for purine prototrophy by growth in adenine-free medium. The Ade -H mutant is defective in the enzyme adenylosuccinate (AMPS) synthetase (ADSS; EC 6.3.4.4), which carries out the first of a two-step sequence in the biosynthesis of AMP from IMP, and therefore requires exogenous adenine for growth. The presence of the long arm of human chromosome 1 in the hybrids is 100% concordant for the ability to grow in adenine-free medium and restoration of the enzyme activity. Hybrid segregants that lose the ability to grow in adenine-free medium lose all or a portion of chromosome 1 and enzyme activity. Southern blot hybridization with a chromosome 1-specific probe, BCMI, confirms the existence of human chromosome 1 in these hybrids. Analysis of a human/CHO translocation chromosome that arose in one of the hybrids suggests that the gene correcting the defect lies in the region 1 cen-1q12. In summary, we have shown by cytogenetics, segregant analysis, biochemical assay, and Southern blot analysis that human chromosome 1, most likely in the region 1cen-1q12, corrects the defect in ADSS-deficient mutant Ade-H cells.
通过在无腺嘌呤培养基中培养，筛选出了人细胞与中国仓鼠卵巢（CHO）K1突变体Ade -H细胞之间的体细胞杂种，以获得嘌呤原养型。Ade -H突变体在腺苷琥珀酸（AMPS）合成酶（ADSS；EC 6.3.4.4）方面存在缺陷，该酶在从肌苷酸（IMP）生物合成AMP的两步序列中执行第一步，因此需要外源腺嘌呤才能生长。杂种中人类1号染色体长臂的存在与在无腺嘌呤培养基中生长的能力以及酶活性的恢复100%一致。失去在无腺嘌呤培养基中生长能力的杂种分离株会丢失全部或部分1号染色体以及酶活性。用1号染色体特异性探针BCMI进行的Southern印迹杂交证实了这些杂种中存在人类1号染色体。对其中一个杂种中出现的一条人/CHO易位染色体的分析表明，纠正缺陷的基因位于1cen-1q12区域。总之，我们通过细胞遗传学分析、分离株分析、生化测定和Southern印迹分析表明，人类1号染色体，很可能在1cen-1q12区域，纠正了ADSS缺陷型突变体Ade -H细胞中的缺陷。
