Legume plants can establish symbiotic nitrogen fixation (SNF) with rhizobia mostly in root nodules, where rhizobia-infected cells are accompanied by uninfected cells in a mosaic pattern. Inside the mature nodules of the legume, carbon and nitrogen nutrients between host plant cells and their resident bacteria are actively exchanged. To elucidate the metabolite dynamics relevant for SNF in nodules, three tissues from a nodule of a model legume, Lotus japonicus, were isolated using laser microdissesction, and transcriptome analysis was done by an oligoarray of 60-mer length representing 21,495 genes. In our tissue-specific profiling, many genes were identified as being expressed in nodules in a spatial-specific manner. Among them, genes coding for metabolic enzymes were classified according to their function, and detailed data analysis showed that a secondary metabolic pathway was highly activated in the nodule cortex. In particular, a number of metabolic genes for a phenylpropanoid pathway were found as highly expressed genes accompanied by those encoding putative transporters of secondary metabolites. These data suggest the involvement of a novel physiological function of phenylpropanoids in SNF. Moreover, five representative genes were selected, and detailed tissue-specific expression was characterized by promoter-β-glucuronidase experiments. Our results provide a new data source for investigation of both nodule differentiation and tissue-specific physiological functions in nodules.