A variety of genes involved in nodule development and function were induced during symbiotic root nodule development in alfalfa. We identified those genes by comparing the cDNA-amplified fragment length polymorphism (cDNA-AFLP) patterns of infected nodules and uninfected roots. Seventy-one transcript-derived fragments (TDFs) were isolated from about 3000 TDFs, which are enhanced or specifically expressed in nodule. The differential expression patterns of 61 genes were confirmed by reverse Northern hybridization or Northern hybridization analyses. Among these, 31 exhibited significant similarities to characterized database entries. These results suggest that the genes corresponding to the 31 TDFs may play important roles in signal transduction, gene expression and regulation, substance transportation or auto-regulation of nodulation during nodule development. The Northern hybridization analysis also indicates that the gene corresponding to TDF RX89 was not expressed in the nodule elicited by bacA mutant, presumably it is specially induced at the stage of nodule development when bacteroids begin to be differentiated.