Transformation Of The Na~+/H~+ Antiporter PeNHX3 From Populus Euphratica Into Arabidopisis

Populus euphratica is a tree halophyte that exists in saline desert areas in northwest China. P. euphraticais highly tolerant to salt and drought stress. Thus, exploring how P. euphratica deals with salt will be helpful for the development of salt tolerant crops.Studies have shown that Na+, K+/H+ antiporters play a significant role in Na/K+ transport and salt resistance in plants. We found that the Na+/H+ antiporter PeNHX3 from P. euphratica mediated the transport of Na+, K+ and Li+ in a yeast growth assay.Using domain-switch analysis, we found that the TM11 of PeNHX3 wascrucial to Li+ transport.In this study, we analyzed the ion transport activity, subcellular localization and salt tolerance of PeNHX3 using gene cloning, genetic transformation and domain-switch analysis. Our goal is to explore the role of PeNHX3 in ion transport and slat tolerance.The major results are listed below:1. We transformed the PeNHX3 gene into Col-0, Atnhxl and Atsosl seedlings and obtained the transgenic seedlings using the methods of gene cloning, vector construction and Agrobacterial transformation.2. The subcellular localization of PeNHX3 was observed using the tobacco transient expression system. The result showed that PeNHX3 was mainly localized in tonoplasts.3. Plant growthanalysis showed that PeNHX3 enhanced salt resistance of Col-0 and Atnhxl’mutant seedlings but not Atsosl, indicating that PeNHX3’s role is transferring Na+ into the vacuole to reduce Na+levels in cytosol. Osmotic stress analysis showed that PeNHX3 improved tolerance ofAtnhx to mannitol stress. In addition, we found that PeNHX3 increasedtolerance of Col-0 and Atnhx1 to Li+ stress, indicating that PeNHX3 mediates Li+ transport in Arabidopsis.4. In order to verify the in plantafunction of PeNHX3 TM11 domain in regulating Li+ transport, we transformed the two domain-switched genes, AtNHX1-PeNHX3-C399 and AtNHX1-PeNHX3-C537, into Col-0 and AtnhxI to analyze their ion transport activities and salt tolerance. Presently, T2 transgenic seedlings are growing for screening.