Isolation And Functional Gene Screening For Mangrove Actinomycetes

Mangroves which harbor abundant microorganisms are woody plant communities growing in intertidal zones of tropical and subtropical coast. The mangrove actinomycetes may form particular mechanisms of metabolism regulation and chemical defense for adapting such particular ecosystem. Their secondary metabolites will also become a new source of natural products. It's very important to investigate the diversity of mangrove actinomycetes, to screen functional genes and explore its potential resources.Samples were pretreated with two different methods: dry-heating and wet-heating. The pretreated samples were applied to there selective media adding with different antibiotics. 96 isolates were selected and primarily classified in to seven groups by their morphology. The most numbers and groups were obtained on the isolate GS1 medium, occupied 47.9%. The second was on the modified GS2 medium, occupied 33.3%. The least was on the HVA medium.The diversity of the mangrove actinomycetes was characterized by analyzing the 16S rDNA sequences. of the representative strains after RFLP. Analysis of the 16S rDNA sequences showed that these strains belong to 3 genera of Streptomyces, Saccharopolyspora and Nocardiopsis. Most stains distributed in 27 species of Streptomycetes sp. 3 isolates were suggested as novel species based on their 16S rDNA sequences.Sequence Guided Genetic screening was performed to evaluate the potential ability of synthesizing natural products. Three pairs of degenerate primers K1F/M6R,A3F/A7R,Halo-FW/Halo-RV were used to amplify the type I polyketide synthase (PKS-I) genes, non-ribosomal peptide synthetases (NRPS) genes and halogenase genes respectively. The results showed, there are 36 strains containing PKS-I gene, 40 strains containing NRPS gene and 11 strains containing halogenase gene. 6 PKS-I gene fragments exhibited less than 70% similarity with the existing sequence, and there are 6 strains have the three functional genes. Activity selection was carried out among 56 strains carrying PKS-1, NRPS and halogenase. Finally 21 active strains were selected, with the activity-ratio being 38.5%. Among these selected strains, IR of 12 strains was above 75%, IR of 5 strains was above 90%.The presented studies revealed that abundant marine actinomycetes resources exist in the sediment samples in Guangdong mangrove, and the functional gene screening can be powerful exploiting the potential microbial resources.