SITE-DIRECTED CHROMOSOMAL MARKING OF A FLUORESCENT PSEUDOMONAD ISOLATED FROM THE PHYTOSPHERE OF SUGAR-BEET - STABILITY AND POTENTIAL FOR MARKER GENE-TRANSFER

A plasmid-free, non-pathogenic, ribosomal RNA group 1 fluorescent pseu domonad, Pseudomonas fluorescens SBW25, was selected from the microflo ra of sugar beet (Beta vulgaris) and modified to contain constitutivel y expressed marker genes. By site directed homologous recombination a KX cassette [kanamycin resistance (kan(r)) and catechol 2,3 dioxygenas e (xylE)] and a ZY cassette [lactose utilization (lacZY, beta-galactos idase, lactose permease)] were introduced at least 1 Mbp apart on the 6.6 Mbp bacterial chromosome. Separate sites were selected to provide sensitive detection methods and allow assessments of marker gene stabi lity of the genetically modified micro-organism (GMM), SBW25EeZYGKX, w hen it colonized the leaves and roots of sugar beet plants following s eed inoculation.