Heefner D L, Squires C H, Evans R J, Kopp B J, Yarus M J
J Bacteriol. 1984 Aug;159(2):460-4. doi: 10.1128/jb.159.2.460-464.1984.
Clostridium perfringens 11268 CDR (Rifr Tcs), the strain transformed in our experiments, was generated by curing a spontaneous, rifampicin-resistant mutant of C. perfringens 11268 (Rifr Tcr). High-temperature growth yielded tetracycline-sensitive, rifampicin-resistant cells which no longer contained pCW3, a 42.8-kilobase plasmid. The tetracycline-sensitive, rod-shaped cell was then converted to an L-phase variant by growth in the presence of penicillin G (10 micrograms/ml) and 0.4 M sucrose. After several passages, the antibiotic was removed from the medium, and cells continued to grow as L-phase variants. Another large plasmid, pJU124 (38.8 kilobases), which confers tetracycline resistance, was used for transformation. Transformation of L-phase variants of C. perfringens 11268 CDR (Rifr Tcs) was mediated by polyethylene glycol. Transformation frequency is a nonlinear function of DNA concentration. Restriction analysis showed that the plasmid isolated from the transformants was identical to that supplied. Stable L-phase variants do not revert to rod-shaped cells, but autoplasts can be both transformed and reverted.
产气荚膜梭菌11268 CDR（Rifr Tcs），即我们实验中转化的菌株，是通过去除产气荚膜梭菌11268（Rifr Tcr）的一个自发利福平抗性突变体中的质粒而产生的。高温培养产生了对四环素敏感、对利福平抗性的细胞，这些细胞不再含有pCW3，一种42.8千碱基的质粒。然后通过在含有青霉素G（10微克/毫升）和0.4 M蔗糖的条件下培养，将对四环素敏感的杆状细胞转化为L相变体。经过几次传代后，从培养基中去除抗生素，细胞继续作为L相变体生长。另一个赋予四环素抗性的大质粒pJU124（38.8千碱基）用于转化。产气荚膜梭菌11268 CDR（Rifr Tcs）的L相变体的转化由聚乙二醇介导。转化频率是DNA浓度的非线性函数。限制性分析表明，从转化子中分离出的质粒与提供的质粒相同。稳定的L相变体不会回复为杆状细胞，但原生质体既能被转化也能回复。
