Santerre R F, Allen N E, Hobbs J N, Rao R N, Schmidt R J
Gene. 1984 Oct;30(1-3):147-56. doi: 10.1016/0378-1119(84)90115-x.
Novel bacterial resistance genes were cloned and expressed as dominant selection markers in mammalian cells. Escherichia coli genes coding for resistance to the aminocyclitol antibiotics hygromycin B (Hm) and G418 were cloned into the eukaryotic expression plasmid pSV5GPT [Mulligan and Berg, Proc. Natl. Acad. Sci. USA 78 (1981) 2072-2076]. Mouse cells normally sensitive to 100 micrograms/ml Hm were transformed with these plasmids and selected in 200 micrograms/ml Hm. Transformants resistant to as much as 1 mg/ml Hm and 500 micrograms/ml G418 were isolated. Cell extracts contained an acetyltransferase activity capable of acetylating G418 and an Hm aminocyclitol phosphotransferase activity. Plasmid DNA sequences were identified by Southern blot analysis of high Mr DNA isolated from transformed cells.
新型细菌耐药基因被克隆，并作为哺乳动物细胞中的显性选择标记进行表达。编码对氨基环醇类抗生素潮霉素B（Hm）和G418耐药的大肠杆菌基因被克隆到真核表达质粒pSV5GPT中[Mulligan和Berg，《美国国家科学院院刊》78（（1981年）2072 - 2076]。用这些质粒转化对100微克/毫升Hm正常敏感的小鼠细胞，并在200微克/毫升Hm中进行选择。分离出对高达1毫克/毫升Hm和500微克/毫升G418耐药的转化体。细胞提取物含有能够乙酰化G418的乙酰转移酶活性和Hm氨基环醇磷酸转移酶活性。通过对从转化细胞中分离的高分子量DNA进行Southern印迹分析来鉴定质粒DNA序列。
