Cloning and Characterization of a DExD/H box RNA Helicase Gene from Medicago sativa L.

Based on an EST in the SSH library of Medicago sativa L.,a full length of 1678 bp cDNA was isolated by RACE.The cDNA sequence was predicted to contain a 1281 bp ORF and code a protein of 426 amino acids,which is homology to Arabidopsis thaliana DEAD-box ATP-dependent RNA helicase 56(RH56) and RH15.The gene was predicted to be a DExD/H box RNA helicase gene and named as MsRH(GenBank accession No.JX508648).Transient expression of MsRH-GFP fusion in onion epidermis cell indicated that the MsRH localized in nucleus.To investigate the function of MsRH,the plant over-expression vector pBI-MsRH was constructed and transferred into tobacco by Agrobacterium LBA4404.After PCR and RT-PCR analysis of five kanamycin resistant plants,35S:MsRH was confirmed to insert into tobacco genome and transcribe mRNA successfully.After treated with 250 mM NaCl for seven days,the proline contents of transgenic plants were lower than those of control plants(WT),and the malondialdehyde contents and relative electric conductivity were higher than those of control tobacco(WT).Taken together,these results demonstrated that MsRH increased the salt sensitivity of tobacco plants under salt stress.