Preliminary Analysis Of RtV-ATPase B Subunit Gene And Rtnhx Gene Expression From Reaumuria Trigyna

Reaumuria trigyna is peculiar to China,which has strong resistance to salt and drought stress.Reaumuria trigyna can discharge of salt ions through salt gland. In the transcriptome sequencing database, a large number of genes associated with ion transport expression increased under salt stress.Such as HKT、SOS、NHX、KUP、 AKT and HAK etc.Na+/H+reverse transporters are located in the vacuole membrane.the Na+can be dividing into vacuole to Prevent the cytoplasm from high Na+poison, balance the ion and penetration balance to improve the salt tolerance of plants through this protine.ATPase provide most energy of ion transport and other physiological activities.So the V-H+-ATPase is also very important to plants salt tolerance.The B subunit plays an important role in this enzyme because it not only contains the regulatory site,but also involved in the formation of the catalytic sites.Cloning NHX and V-H+-ATPase B subunit from Reaumuria trigyna,and verify its function under aviotic stress.which will lay the foundation of reveal the felationship of ion transport and its resistance for Reaumuria trigyna.The main results are as follow:1.Sequence analysised the full length of RtNHX and RtV-ATPase. RtNHX contains an open reading frame (ORF) with the length of1662bp and encodes a protein with553amino acids. The ORF length of RtV-ATPase is996bp, which encodes a protein with331amino acids.2.The result of RT-PCR showed that under different concentrations of NaCl stress,RtV-ATPase gene expression level gradually increased from100mM to300mM,and expression level highest in300mM.RtNHX gene expression level highest in200mM,and then gradually reduced.These two genes expression levels was lowest in400mM.3.The prokaryotic expression vectors pET-32a-RtNHX and pET-32a-V-ATPase were constructed and transformed into E. coli. The fusion proteins in recombinant E. coli cells were expressed after IPTG inducing by SDS-PAGE electrophoresis analysis.4.The recombinant strain’s tolerance of NaCl significantly higher than the empty vector cell.But these two bacterial strain tolerance of Sorbitol was not obvious.5.The eukaryotic expression vector pPZP221-RtV-ATPase was constructed and transformed into Arabidopsis.Laying the foundation for functional identification.