The Cloning Of A Vacuolar Na~+/H~+ Antiporter CarNHX1 Gene From Chickpea And The Study Of Enhancing The Salt Tolerance Of Tobacco

Chickpea (Cicer arietinum Linn.) is a Leguminous herb, originated in Western Asia and the Near East, is the world's cultivated area of the larger beans and riched in nutritional value. It is of great practical significance to the northwest, southwest, and other vast arid and damp-arid areas of China with dinstinctive characteristic of tolerance to drought and salinity. Mainly distributed in the plant vacuole membrane, the plant Na+/H+ antiporter is responsible for the Na+ transmembrane transport and functions as one of the main mechanisms for plants to fight against salt stress. This study offers references to researching work of the molecular mechanism of salt-tolerance and molecular breeding in the future through analysis of gene functions as well as gene regulation and modulation of Na+/H+ antiporter gene.1 CarNHX1 of Chickpea were cloned with RT-PCR and homology cloning. The RT-PCR and a Real-time PCR assay showed that the level of CarNHX1 transcriptions was up-regulated obviously in the seedlings which indicated that CarNHX1 are related to salt tolerance of the plant.2. For further study of CarNHX1 salt-tolerance mechanism, the promoter of CarNHX1 was cloned with Genome walking technology and 5'upstream 1655 bp fragment was obtained. By means of bioinformatics, the structure and function of CarNHX1 were estimated thoroughly. Gene sequence analysis showed that CarNHX1 belongs to Na+/H+ antiportor family. Further analysis showed that the CarNHX1 was a transmembrane protein in plasmalemma with signal peptide. Analysis showed that promoter sequence contains high levels of transcriptional response elements, common starting devices and upstream regulatory elements.3. The recombinant vector pCAMBIA1301-CarNHX1 was transfored into the wild tobacco, thanks to the Agrobacterium-mediated leaf disc transformation method. Then, screen and identify transgenic positive tobacco through the Kan resistance and PCR. Salt resistance of transgenic tobacco and related physiological and biochemical indexes were assayed. The results indicate that the transgenic tobacco is, me extent, resist to some of high salt abiotic stress.