BACILLUS THURINGIENSIS TRANSFORMATION

摘要

The present invention describes a process that for the first time renders possible a direct and targeted genetic manipulation of Bacillus thuringiensis and the closely related B. cereus using recombinant DNA technology. The invention comprises a process for inserting and cloning DNA sequences in gram positive bacteria selected from the group consisting of Bacillus thuringiensis and Bacillus cereus, comprising: (a) isolating the DNA to be introduced; (b) cloning the thus isolated DNA in a cloning vector that is capable of replicating in a bacterial host cell selected from the group consisting of Bacillus thuringiensis and Bacillus cereus cells in a heterologous cloning system; (c) directly introducing the thus cloned vector DNA into intact cells of said bacterial host cell via electroporation at a transformation rate which overcomes the restriction barrier present in the said bacterial cells; and (d) cultivating the thus transformed bacterial cells and isolating the thus cloned vector DNA. The present invention also relates to the construction of plasmids and "shuttle" vectors and to the B. thuringiensis strains that have been transformed therewith. Also described is a process for the direct cloning, expression and identification of genes in B. thuringiensis and in the closely related B. cereus.