New source of cellulase production using a metagenomic technique

Safaa M Ali ,Nadia A Soliman

doi:10.21608/nrmj.2019.66751

Abstract

The cellulase enzymes with high effectiveness under conditions agreeable to the industrial processes necessities are one of the keys for the successful development of chemical and drug synthesis. The soil metagenome is an affluent source for the discovery of new natural products. The objective of the current study was to identify the isolated functional gene(s) of the cellulase enzyme by using metagenomics. The plan of work composed of collection of different soil samples, isolation of total DNA, fragmentation, cloning, and expression of the isolated gene(s) in the suitable host microorganism. The total genomic DNA was extracted using a kit (QIAGEN), and then digested by different restriction enzymes BamHI. The digested fragments ranging from ~300-5000 bp were ligated, cloned into pUC19 vector, and then transformed into Escherichia coli DH5α. The resulting clones were screened as cellulase producers using a qualitative method. The positive clones which showed hydrolysis on the plate were screened once more in Luria-Bertani (LB) medium. The plasmids were isolated and then tested using universal primer (M13), to detect the fragment size and sequence for the Polymerase Chain Reaction (PCR) products. This study establishes an effortless and professional method for cloning of recent cellulase genes through ecological metagenomes. In the outlook, the metagenomic guide approachs may be functional to the elevated selection of novel cellulase from the environment.

Highlights

Functional genomics is being used to search for specific activities of an organism without the need for culturing this organism that harbor the pathways involved (Gillespie et al, 2002)
The incidence of the positive cellulase clones obtained from the collected soil samples using the metagenomic technique, was evaluated through the isolation of DNA
Results after ligation reveled that about 96 clones are recovered from the digested DNA using the Bam HI, 84 clones are selected from the DNA digested with HindIII, 110 clones are selected from the DNA digested with EcoRI, and about 49 clones are selected from DNA digested with SalI

Summary

Introduction

Functional genomics is being used to search for specific activities of an organism without the need for culturing this organism that harbor the pathways involved (Gillespie et al, 2002). Most of the microorganisms responsible for nutrients cycling in the environment have to be cultivated, and this could include those species responsible for the degradation of cellulose It has recently been reported by Fernández-Arrojo et al, (2010); Ferrer et al, (2016) that the discovery of new enzymes through metagenomics has enormous potential to obtain a wide range of useful biocatalysts. It is possible to identify novel cellulase genes free of DNA amplification, by following a direct metagenome sequencing approach to provide genes that can be cloned, expressed and classified prior to future use All these are possible without the need for any data on the organisms from which they originated (David et al, 2012). The aim of this study was to isolate functional gene(s) of the cellulase enzyme by using metagenomics technique

Objectives

Methods

Results

Conclusion