Cloning and Functional Characterization of Three Superoxide Dismutases Genes from Halophyte Salicornia europaea and Thellungiella halophila
In order to study the halophyte salt-tolerance mechanism and compare the salt tolerance of superoxide dismutase (SOD) from different halophytes,we cloned the full-length cDNAs of manganese (Mn) and Cu/Zn SOD from Salicornia europaea by RACE method.Sequence analysis indicated that the S.europaea MnSOD gene (SeMSD) (GenBank accession number:JQ061158) comprised an open reading frame of ′699 bp,encoding a polypeptide of 233 amino acids with a predicted molecular mass of 25.7 kD.Correspondingly,the Cu/ZnSOD gene (SeCSD,GenBank accession number:JQ061160) consisted of an open reading frame of 684 bp which encoded a protein of 228 amino acids with a predicted molecular mass of 23.3 kD.According to these two obtained cDNA sequences and Thellungiella halophila MnSOD gene (ThMSD)published on GenBank (EF140719),the prokaryotic expression vectors pET30a-SeMSD,pET30a-SeCSD and pET30a-ThMSD were constructed,and the target proteins were expressed successfully in BL21 Escherichia coli.Through optimizating the concentration of Isopropyl β-D-Thiogalactopyranoside (IPTG),the salt tolerance of these bacteria with the three superoxide dismutases were tested under 6.5 ％ and 7.5 ％NaCl.The results demonstrated that the recombinants BL21 (pET30a-SeMSD) and BL21 (pET30a-ThMSD) strains showed better tolerance to salinity stress in comparison with the control stains BL21 (pET30a),but the recombinant BL21 (pET30a-SeCSD) strains displayed no salt tolerance increase.
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