A nolC-lacZ gene fusion in Rhizobium fredii facilitates direct assessment of competition for nodulation of soybean

Abstract

A constitutively expressed chromosomal gene, nolC, regulates the ability of Rhizobium fredii strain USDA257 to nodulate certain agronomically improved soybean cultivars. Other advanced cultivars, as well as unimproved varieties such as 'Peking', form nitrogen-fixing nodules with parental strain USDA257 and with nolC mutants. We have used 257DH512, a derivative of USDA257 that contains a nolC-lacZ fusion, to directly measure strain competition for nodule occupancy. Seedlings of the soybean cultivar 'Peking' were inoculated with mixtures containing various ratios of 257DH512 and R. fredii USDA208; nodules were harvested 14 days later and stained histochemically with X-gal, a chromogenic substrate for the β-galactosidase product of lacZ. We were able to stain and evaluate batches of nearly 500 nodules in just 4 h, without the use of expensive equipment. Under the conditions of these experiments, USDA208 was considerably more competitive than 257DH512, even when it had been present in the inoculum at a numerical disadvantage. We suggest that gene fusions can be used to greatly simplify the assessment of nodulation competitiveness in rhizobia. Key words: competition, nolC-lacZ fusion, soybean – Rhizobium fredii, nodule occupancy.