Ishiura M, Uchida T, Okada Y
Department of Cell Biology, National Institute for Basic Biology, Aichi, Japan.
Cell Struct Funct. 1989 Aug;14(4):495-9. doi: 10.1247/csf.14.495.
Recombinant lambda phage DNA, encapsulated in phage particles and coprecipitated with calcium phosphate, efficiently transforms cultured mammalian cells without a requirement for carrier DNA. The present paper analyzes the stability of the transformants obtained by the phage transfer method. lambda phage particles containing recombinant DNA that includes the thymidine kinase (TK) gene of herpes simplex virus type 1 as a selective marker were introduced into Ltk- cells deficient in TK activity, and TK+ transformants were selected in HAT medium. To test the stability of the TK+ phenotype of the transformants, seven individual transformant clones were isolated, cultured in HAT selective medium and then in non-selective medium for various lengths of time. After such culture, transformants were allowed to develop colonies in both selective and non-selective medium. For all seven transformant clones, the numbers of colonies obtained in the two types of medium were almost identical, irrespective of whether or not each transformant clone had been previously cultured for 15 to 50 days in non-selective medium. This result suggests that most transformants obtained by the phage transfer method maintain the TK+ phenotype stably, for at least 50 days, when grown in non-selective medium.
包裹在噬菌体颗粒中并与磷酸钙共沉淀的重组λ噬菌体DNA，无需载体DNA就能有效地转化培养的哺乳动物细胞。本文分析了通过噬菌体转移法获得的转化体的稳定性。将含有重组DNA（其中包括1型单纯疱疹病毒的胸苷激酶（TK）基因作为选择标记）的λ噬菌体颗粒引入缺乏TK活性的Ltk-细胞中，并在HAT培养基中选择TK+转化体。为了测试转化体TK+表型的稳定性，分离出七个单独的转化体克隆，先在HAT选择培养基中培养，然后在非选择培养基中培养不同时长。经过这样的培养后，让转化体在选择培养基和非选择培养基中都形成菌落。对于所有七个转化体克隆，无论每个转化体克隆之前是否在非选择培养基中培养了15至50天，在两种类型培养基中获得的菌落数量几乎相同。这一结果表明，通过噬菌体转移法获得的大多数转化体在非选择培养基中生长时，至少50天能稳定维持TK+表型。
