Isolation And Characterization Of A Vacuolar Na～+/H～+ Antiporter Gene From Medicago Sativa

Salt stress in the soil has been existing for a long time, and the problem is more serious in recent years. High concentrations of salt can cause osmotic stress and ion toxicity, resulting in the damage for plant cells. Among the salt stress, Na~+ is the major cause of ion-specific damage in many plants. In order to tolerate high level of salt, plants compartment Na~+ into vacuoles or remove Na~+ out of cells from the cytoplasm through the operation of Na~+/H~+ antiporter to avoid the toxic effects of Na~+ in the cytosol and maintains osmotic balance. The compartmentation of Na~+ into vacuolar is achieved by the role of Na~+/H~+ antiporters on the vacuolar membranes. Vacuolar Na~+/H~+ antiporter is important in salt tolerance, and alfalfa is a kind of important leguminous plants. Therefore, an alfalfa (Medicago sativa cv: WL323) gene with high homology to Na~+/H~+ antiporter genes (named MsNHX1) was isolated by RT-PCR methods with homologous gene region and the Genbank accession number was AY513732. Sequence analysis showed the cDNA was 1623 bp in length and it encoded a protein of 541 amino acids. The deduced amino acid sequence showed high identities with other plant vacuolar type Na~+/H~+ antiporters, but had low identities with plasma membrane Na~+/H~+ antiporters. This indicated that the MsNHX1 encoded a putative vacuolar type Na~+/H~+ antiporter. Amino acid analysis with TMpred software indicated that the polypeptide chain contained 12 transmembrane motifs, which were related to the activity of the putative protein. Southern blot analysis revealed MsNHX1 exisited as a small gene family in the genome of tetraploid alfalfa. Transcripts of MsNHX1 were detected in alfalfa leaves and roots. In addition, the transcriptional level in root and leaf was upregulated when treated with 200...