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Efficient Transformation Procedure of a Newly Isolated Streptomyces sp. TN58 Strain Producing Antibacterial Activities

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Abstract

A new aerobic Gram-positive bacterium designated TN58 producing antibacterial activities against Gram-positive and Gram-negative bacteria was isolated from Tunisian soil. The nucleotide sequence of the 16S rRNA gene (1516 bp) of the TN58 strain showed high similarity (96–98%) to the Streptomyces 16S rRNA genes, especially with that of Streptomyces lavendulae which produces the anti-tumor compound mitomycin C, and the cyclic peptide antibiotic, complestatin. Cultural characteristic studies, alignment data of the 16S rRNA gene, and analysis of the nucleotide sequence of a 2.2 kb genomic DNA fragment from TN58 strongly suggested that this strain could be an actinomycete and most probably belongs to the genus Streptomyces. Study of the influence of different nutritional compounds on antibiotic production showed that the highest antibacterial activities were obtained when glycerol at 1% (w/v) was used as sole carbon source in the presence of potassium. In analytical conditions, the application to supernatant culture of the TN58 strain of various extraction and purification steps led to the isolation of two pure active molecules having a retention time of 38.6 and 50.2 min, respectively. TN58 strain was untransformable with the Streptomyces cloning vector pIJ702 via classical polyethylene glycol (PEG) protoplast transformation and previously described Streptomyces electroporation procedures. Transformation was rendered possible by the electroporation technique only after utilization of a preculture medium without sucrose and a regeneration plate containing a low sucrose concentration.

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Acknowledgments

This work was supported by the Tunisian government (Contract Programme CBS-LEMP). We are grateful to Dr. J.L. Pernodet (BGM Laboratory of the IGM Orsay, France) for providing DNA probes from S. ambofaciens and the M. luteus LB 14110 strain, and to Dr. A. Dhouib (CBS) for providing the four other indicator microorganisms. Thanks are also due to Mr. R. Rekik, Mr. R. Hmidi, Mr. E. Ben Messaoud, Dr. M. Ben Ali, and Dr. H. Chouayekh for their generous help.

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  1. Centre de Biotechnologie de Sfax, Laboratoire d’Enzymes et de Métabolites des Procaryotes, P.O. Box ‘‘K’’, 3038, Sfax, Tunisia

    Lotfi Mellouli, Ines Karray-Rebai, Samiha Sioud, Raoudha Ben Ameur-Mehdi, Belgacem Naili & Samir Bejar

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  1. Lotfi Mellouli
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  2. Ines Karray-Rebai
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  3. Samiha Sioud
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Correspondence to Lotfi Mellouli.

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Mellouli, L., Karray-Rebai, I., Sioud, S. et al. Efficient Transformation Procedure of a Newly Isolated Streptomyces sp. TN58 Strain Producing Antibacterial Activities. Curr Microbiol 49, 400–406 (2004). https://doi.org/10.1007/s00284-004-4292-9

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  • DOI: https://doi.org/10.1007/s00284-004-4292-9

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