Ahmed Z U, Vining L C
J Bacteriol. 1983 Apr;154(1):239-44. doi: 10.1128/jb.154.1.239-244.1983.
Of seven chloramphenicol-producing actinomycetes examined, only Streptomyces venezuelae strain 13s contained extrachromosomal DNA detectable by agarose gel electrophoresis and cesium chloride-ethidium bromide density gradient centrifugation. The single 17-megadalton plasmid present in this strain was indistinguishable from plasmid pUC3 previously isolated from mutagenized cultures. Strains selected for their inability to produce chloramphenicol after treatment with acriflavine or ethidium bromide still contained a plasmid that had the same electrophoretic mobility as plasmid pUC3 and yielded similar fragments when digested with restriction endonucleases. By regenerating protoplasts of strain 13s and screening for isolates lacking extrachromosomal DNA, strain PC51-5 was obtained. The absence of plasmid pUC3 sequences in this strain was confirmed by Southern hybridization using 32P-labeled plasmid as a probe. Since the plasmidless strain produced as much chloramphenicol as did the parent strain, pUC3 contains neither structural nor regulatory genes for antibiotic production. Evidence from electrophoretic analysis of BamHI digests of total cellular DNA from wild-type and dye-treated nonproducing progeny indicated that acriflavine caused structural changes in the chromosome.
在检测的7株产氯霉素放线菌中，只有委内瑞拉链霉菌13s菌株含有可通过琼脂糖凝胶电泳和氯化铯-溴化乙锭密度梯度离心检测到的染色体外DNA。该菌株中存在的单一17兆道尔顿质粒与先前从诱变培养物中分离出的质粒pUC3无法区分。在用吖啶黄素或溴化乙锭处理后因无法产生氯霉素而被挑选出的菌株仍含有一种质粒，其电泳迁移率与质粒pUC3相同，用限制性内切酶消化时产生相似的片段。通过再生13s菌株的原生质体并筛选缺乏染色体外DNA的分离株，获得了PC51-5菌株。使用32P标记的质粒作为探针进行Southern杂交，证实了该菌株中不存在质粒pUC3序列。由于无质粒菌株产生的氯霉素与亲本菌株一样多，因此pUC3既不包含抗生素生产的结构基因也不包含调控基因。对野生型和经染料处理的不产生产物的后代的总细胞DNA的BamHI消化产物进行电泳分析的证据表明，吖啶黄素导致了染色体的结构变化。
