Genetic Characteristics of Actinomycetes Isolates (GST, KP, KP11, Kp16, T24, T37 Code) by RFLP of NRPS Genes

Melati Aprilliana Ramadhani,Nanik Sulistyani

doi:10.12928/mf.v12i1.3020

Melati Aprilliana Ramadhani, Nanik Sulistyani

Open Access

https://doi.org/10.12928/mf.v12i1.3020

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Abstract

Actinomycetes is the one of antibiotic producing microorganisms. Actinomycetes isolates with GST, KP, KP11, KP16, T24, T37 have been isolated from soil. This study aims to determine the diversity of actinomycetes isolates by RFLP (Restriction Fragment Length Polymorphism) of NRPS (Non Ribosomal Peptide Synthetase) genes from GST, KP, KP11, KP16, T24, and T37 isolates. This study is divided into several steps of the DNA isolation from isolate GST, KP16, T24, T37, KP, KP11, PCR of the 16S rRNA and NRPS genes and cutting the NRPS genes fragmens with HaeIII enzyme to determine the diversity of isolates. Results of DNA isolate, PCR of 16S rRNA genes and NRPS genes, and RFLP of NRPS genes performed by agarose gel electrophoresis. The diversity of the isolates were analyzed by multivariate analysis UPGMA with MVSP 2.0 applications. The results showed the diversity of 6 actinomycetes isolates are T37 and GST have 64 % similarity, KP11 and KP16 have 96 % similarity, KP has 76 % similarity with KP11 and KP16, while T24 has 60 % similarity with KP and 12 % similarity with T37 and GST. Based on RFLP analysis of the NRPS genes, among of 6 isolates can divided into 4 groups are T37 and GST isolates as group 1, KP11 and KP16 isolates as group 2, KP isolate as group 3, and T24 isolate as group 4. Keyword : Actinomycetes, diversity, 16S rRNA genes, NRPS genes, RFLP

Highlights

NRPS (Nonribosomal peptide synthetase) is biosynthetic systems involved in the synthesis of a large number of important biologically active compounds produced by microorganisms, among others by actinomycetes (Sacido et al, 2005)
The objective of this research was to observe the diversity of 6 actinomycetes isolates with GST, KP16, T24, T37, KP, and KP11 code based on the RFLP of NRPS genes profile
NRPS genes was involved in production of secondary metabolites that are important in the pharmaceutical industry

Summary

Introduction

NRPS (Nonribosomal peptide synthetase) is biosynthetic systems involved in the synthesis of a large number of important biologically active compounds produced by microorganisms, among others by actinomycetes (Sacido et al, 2005). NRPS is multi-enzymatic, multidomain megasynthases involved in the biosynthesis of nonribosomal peptides. NRPS modules contain the activities corresponding to the condensation, adenylation, and thiolation steps involved in the recognition and condensation of the substrate. Additional domains (heterocyclase, N-methylase, epimerase, thioesterase, and reductase) are present depending on the requirements for the substrate activation, elongation, and modification (Sacido et al, 2005). Natural products screening programs have concentrated an intense effort in the discovery of biologically active metabolites produced by actinomycetes (Sacido et al, 2005)

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