Improvement of cellulose degradation by cloning of endo-β-1, 3-1, 4 glucanase () gene from BTN7A strain.

The aim of this study is to construct a new recombinant strain able to degrade cellulose efficiently. The endo-β-1, 3-1, 4 glucanase () gene was cloned from BTN7A strain by using PCR technique. The specific primers of gene were deduced. Optimization of PCR mixture and program were identified. The nucleotide sequence of was placed in the public domain (GenBank accession number KM009051.1). The obtained DNA was cloned with pGEM®-T Easy Vector. The recombinant plasmid designated as Bgls-NRC-1 was transformed into DH5α. The successful cloning of the gene was tested either by PCR or by evaluating its expression in its new bacterial host. The gene was expressed efficiently in and the enzyme activity of the transformant was compared to the enzyme activity of the donor bacterial strain. The new constructs produce much higher enzyme yields than the donor bacterial strain, they produce about 29% and about 57% higher cellulase specific activity at 37 °C and 55 °C respectively. Optimization of cellulolytic activity of the new recombinant strain were described. The effect of minimal medium supplemented with CMC or cellulose, or complete medium (LB) on expression were tested, the order of cellulase activity production was CMC27.2 > cellulose 21.9 > LB 19.8 U/mg protein, respectively at 24 h. CMC was proved to be the best medium for cellulase production. Results also showed that double the initial inoculum resulted in more cellulase activities in all media.