Integrative vector for stable transformation and expression of a β-1,3-glucanase gene in Clavibacter xyli subsp. cynodontis

Clavibacter xyli subsp. cynodontis is an endophytic Gram-positive bacterium that is able to colonize the xylem tissue of various plants. We constructed an integrative vector that can carry foreign genes into a repetitive chromosomal element of C. xyli subsp. cynodontis. Using this vector, we transformed C. xyli subsp. cynodontis with a gene coding for an endoglucanase enzyme under the control of a strong promoter that was previously isolated from the genome of C. xyli subsp. cynodontis. The transformed bacteria efficiently expressed active glucanase and secreted it into the culture medium. The vector has the advantage of being stably maintained in the chromosome; the transformations were maintained without antibiotic selection both in in vitro culture and in bacteria growing in maize xylem.