Molecular Cloning,Expression And Salt Tolerance Analysis Of SeHSP90Gene From Halophyte Salicornia Europaea

Salt stress is one of the major abiotic stress in the nature, and many higher plants are serious damaged by the high salinity in soil which impaired the growth and development of the plants. Therefore, it has important practical significance for salt-tolerant crop cultivation, analyse the physiological and molecular mechanisms of the response to salt stress. Researches have shown that heat shock protein (HSP) plays an important role in the tolerance response to salinity stress. HSP90is an important member of HSP family and Saliconia europaea is one of the most famous salt-tolerant plants. This research paid attention to the HSP90gene of Saliconia europaea, the corresponding protein and the role of the protein played in the salt stress tolerance.Gene segment of1142bp in the middle of SeHSP90was amplified from Saliconia europaea by Degenerate PCR. cDNA sequence of SeHSP90(GenBank:KC181862) about2461bp was isolated using RACE technology. The open reading frame was2091bp and it encoded a697-amino-acid protein with a molecular weight of79kDa.Bioinformatics analysis showed that SeHSP90was out of the control of protein transport system after synthesis and it was anchored in the cytoplasm finally. It has feature sequence of protein family; physical and chemical properties analysis show that the hydrophilic SeHSP90for protein isoelectric point is5.01, there are three kinds of phosphorylated amino acid and it does not exist transmembrane structure and signal peptide sequence; Homology analysis indicated that SeHSP90share highly conserved amino acid sequence with different species of HSP90and it has the closest relationship with Capsicum chinense.The prokaryotic expression vector pET-SeHSP90was constructed by double digestion technique and transformed into BL21(DE3). Protein electrophoresis assay showed expression protein stripe size was in line with expectations and it suggested SeHSP90could be expressed in BL21(DE3). The IPTG concentration optimization experiments showed that the optimum IPTG concentration was0.025mmol/L for BL21(DE3) with SeHSP90.Salt resistance analysis inferred that the growth of recombinant strain (pET-SeHSP90) was significantly better than that of control strain. And growth advantage is more significant with salinity increase. When salt concentration were5%,6%,6.5%,7%, the growth of recombinant strain were about1.5,1.7,2.5and3times against the control strain, respectively. These have important implications for further studies on the molecular mechanisms of resistance to adversity in Saliconia europaea.