[Cloning and expression of Clostridium thermocellum F7 cellulase genes in Escherichia coli and Bacillus subtilis cells]

Abstract

The Clostridium thermocellum total DNA Sau 3A fragments' library was constructed on the basis of shuttle vector pMK4 for the Escherichia coli - Bacillus subtilis. 14 clones with endoglucanase activity and one with beta-glucosidase activity were selected in E. coli cells. Recombinant plasmids pCE were characterized by structural instability of various degree in B. subtilis cells. The results of the physical mapping, analysis of gene products in E. coli mini-cells as well as the DNA-DNA blot hybridization have led to conclusion on cloning of 7 individual genes for endoglucanases. Up to 3 polypeptides of various molecular weight corresponding to the products of cel gene were revealed in E. coli mini-cells containing the recombinant plasmids. The hybridization analysis demonstrated considerable homology of the majority of cel genes.