Modern agriculture is dependent on the yearly application of large quantities of nitrogen fertilizer acquired via the chemical fixation of N 2 in the Haber–Bosch process. More sustainable agricultural systems take advantage of biological nitrogen fixation, particularly the symbiosis between nitrogen-fixing bacteria called rhizobia and leguminous plants. A long-term goal of symbiotic nitrogen-fixation (SNF) research is to optimize its use in agriculture by improving rhizobia or by engineering symbiotic relationships into nonlegumes. Using the model rhizobium
Sinorhizobium( Ensifer) meliloti, we establish that only 58 genes from the 1.35-Mb pSymA megaplasmid are required for effective SNF. This minimal SNF gene set, and genetic platform, have important implications for engineering approaches to optimize rhizobia inoculants and transfer symbiotic abilities to novel backgrounds.