Construction of a Lotus japonicus late nodulin expressed sequence tag library and identification of novel nodule-specific genes.

摘要

A range of novel expressed sequence tags (ESTs) associated with late developmental events during nodule organogenesis in the legume Lotus japonicus were identified using mRNA differential display; 110 differentially displayed polymerase chain reaction products were cloned and analysed. Of 88 unique cDNAs obtained, 22 shared significant homology to DNA/protein sequences in the respective databases. This group comprises, among others, a nodule-specific homologue of protein phosphatase 2C, a peptide transporter protein, and a nodule-specific form of cytochrome P450 which were sequenced and full-copy or almost full-copy cDNA sequences have been submitted to GenBank under the accession numbers AF000402, AF000392 and AF000403, respectively. RNA gel-blotanalysis of 16 differentially displayed ESTs confirmed their nodule-specific expression pattern. The kinetics of mRNA accumulation of the majority of the ESTs analysed were found to resemble the expression pattern observed for the L. japonicus leghaemoglobin gene. These results indicate that the newly isolated molecular markers correspond to genes induced during late developmental stages of L. japonicus nodule organogenesis and provide important, novel tools for the study of nodulation.