Molecular Cloning, Characterzation And Funcitonal Analysis Of SaVP1Genes In Spartina Anglica

Abiotic stresses, especially salinity, are the primary causes of crop loss. Recently, moreattention has been paid on the mechanism of ion homeostasis in plant cells. One of them isthe compartmentation of Na+into vacuoles, which can be achieved by the action of Na+/H~+antiporters on the vacuolar membranes. The electrochemical gradient of protons generatedby vacuolar H~+translocation enzymes H~+-PPase is the driving force for this process.Therefore, cloning and transforming the H~+-PPase will improve the plant salt tolerance.The main results were as follows:1. The full-length cDNA of halophyte Spartina anglica H~+-PPase gene was coined byRACE-PCR technique. The SaVP1cDNA sequence was2506bp, encoding667aminoacids.2. Homology analysis and molecular modeling revealed that the SaVP1stronglyresembled other H~+-PPase genes and the nucleic acid and amino acid levels. The cDNAsequence and amino acid sequence of the SaVP1gene were shared by94%and99%identity with those of the Leptochloa fusca respectively.3. The plant expressing vector pCAMBIA1301-SaVP1for transforming divot plantswas constructed. The recombinant vector subsequently introduced into Agrobacteriumtumefaciens (LBA4404) with freeze-thaw method. The leaf discs of tobacco plants wereinfected by Agrobacterium tumefaciens (LBA4404) and hygromycin-resistant plants wereregenerated on the selection medium containing hygromycin. PCR confirmed that obtainedthe positive transgenic tobacco plant.4. The salt tolerance test results showed that four transgenic tobacco lines could grownormally with300mM sodium chloride and they were dead with the400mM sodiumchloride, but the non-transgenic tobacco could not grow normally with200mM sodiumchloride.The results showed that SaVP1could improve the tobacco salt tolerance.