engleski

Transposon Tn5424 induces amplifications in Streptomyces rimosus

Tn5424 is a transposon carrying the thiostrepton-resistance gene (tsr) based on the insertion sequence IS466 from Streptomyces coelicolor A3(2). It can be introduced into Streptomyces strains using derivatives of the temperature-sensitive replication plasmid pSG5. The original plasmid pUKG403 that transfers transposon Tn5424 was not convenient for use in Streptomyces rimosus because the plasmid-borne kanamycin resistance can not be selected in this species. In order to solve this problem an erythromycin resistance gene (mls) was introduced into the plasmid to give pPZG200. Selection at high temperature lead to loss of erythromycin resistance, but a certain proportion of the population retained the transposon thiostrepton-resistance marker indicating that transposition into the chromosome had occurred. When total DNA from these strains was examined by restriction analysis all turned out to carry high level DNA amplification of 1.5 kb and 9.0 kb in length. The amplified DNA hybridised to a tsr probe. The total DNA from all transposants hybridised to a cosmid clone pPZG25 (carrying the oxytetracycline cluster) showing the same pattern as in parental strain.

Streptomyces; transposon; DNA amplification

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