Competition for nodule occupancy on Phaseolus vulgaris by Rhizobium etli and Rhizobium tropici strains can be efficiently monitored in an ultisol during the early stages of growth using a constitutive GUS gene fusion

Abstract

The GUS gene fusion (pKW107) was transferred into 4 Rhizobium phaseoli strains (KIM5s, CIAT 895, CIAT 7202, CIAT 151) and 1 Rhizobium tropici strain (CIAT 899) in order to determine the nodulation competitiveness of these strains on common bean ( Phaseolus vulgaris) in soil. DNA was fingerprinted with the different mutants to verify correct integration of the glucuronidase gene in the strains' genomes. Co-inoculation trials in growth pouches proved that the GUS + mutant strains were unaltered in their nodulation capabilities. Plant studies in a non-sterile ultisol examined the value of GUS-marked Rhizobium strains for monitoring nodule occupancy under tropical conditions. At 14 and 21 days after planting (DAP), blue nodule color allowed consistently clear and reliable differentiation between nodules formed by GUS + rhizobia and indigenous soil rhizobia. At 30 DAP, however, expression of the GUS trait was inconsistent. Older nodules formed by the GUS + rhizobia were only a faint green after incubation in detection buffer, probably because of the restriction of gusA gene expression in the construct to free-living rhizobia. Nodules formed by GUS-harboring inocula strains were differentiated from ineffective green nodules formed by native rhizobia in 30 DAP nodules in microplates by crushing nodules before adding detection buffer. Data obtained by use of the GUS gene fusion were confirmed by data derived using the ELISA technique and antibiotic resistance markers. The GUS gene fusion was found to be a powerful tool to examine competitiveness of inocula strains in a tropical soil.