Molecular characterization of thermophilic Streptomyces using amplified 16s ribosomal DNA restriction analysis (ARDRA)

Abstract

The present study aims to characterize the 19 thermophilic Streptomyces isolates on the basis of their restriction profile obtained by Amplified Ribosomal DNA Restriction Analysis (ARDRA) method. Nineteen thermophilic Streptomyces isolates, characterized and identified on the basis of cultural, morphological, physiological and biochemical characteristics using PIB Win (Probabilistic identification of Bacteria) software, have been further characterized by Amplified Ribosomal DNA Restriction Analysis method. DNA of all the 19 selected isolates and two reference Streptomyces cultures of MTCC ( S. phaeochromogenes (MTCC 2614) and S. thermovulgaris (MTCC 1822) were isolated and the 16S rRNA gene was amplified using primer F1 (5'-AGAGTTTGATCITGGCTCAG-3’; I= inosine) and primer R5 (5'-ACGGITACCTTGTTACGACTT-3'). Two restriction endonuclease enzymes i.e. Eco R I and Hae III were used for restriction digestion. The 16S rDNA were amplified by PCR using specific primers and the amplified product were cleaved separately by both endonucleases, yielding 4 to 5 bands by Hae III and 2 to 3 bands by Eco R I. Digestion with two endonucleases indicate that Hae III enzyme is better than Eco R I for differentiating Streptomyces isolates.