The Response Of Creeping Bentgrass(Agrostis Stolonifera L.)under Salt Stress And Functional Study Of AsNHX2 Gene

Creeping bentgrass(Agrostis stolonifera L.),a cool season turfgrass,belongs to Poaceae Agrostisand has choiceness leaf texture,characteristic of low pruning and frost resistance.Creeping bentgrass is the most widely used cool-season grass in urban green land,golf,bowling and other high quality lawns.However,high water consumption and poor salinity tolerance limits the wider application of creeping bentgrass.Currently,it is little known about the physiological and molecular responses of creeping bentgrass under salt stress,cloning and functional analysis of salt tolerance related genes,and the reflectance.In this study,the photosynthesis,antioxidant metabolism,antioxidant enzymes gene expression and leaf hyperspectral characteristics of penncross and seaside ? under salt stress wereinvestigated.NHX2 gene was cloned from penncross and its function was further explored.The results of the study are as below:Under salt stress,turfgrass quality,chlorophyll content,net photosynthetic rate,and enzyme activity of CAT,APX,POD were higher in seaside ? than penncross.Interestingly,the SOD activity was lower in the tolerant cultivar at the initial stage but increased to a higher level than penncross at the later stage.qRT-PCR was employed to determine the five ROS enzyme genes' expression characters.The results showed that the expression level of CytCu/ZnSOD and POD were lower at 3 h but higher at 12 h in seaside ? than penncross.APX expressed more abundantly from the 3rd hour after treatment in seaside ? than penncross one.FeSOD and CAT expressed more abundantly in seaside ? among all the sampling times.The inversion model was established by regression analysis between the electrolyte leakage and characteristic parameters of hyperspectral reflectance of the leaves about penncross and seaside ?.The electrolyte leakage with the 541nm,617nm and 651nm band reflectance values,541nm,617nm band and 651 nm,718nm band of NDVI,667nm,668nm band of NDVI,green edge amplitude and green edge area had high correlation.Using these parameters as the independent variables,the inversion model constructed by partial least squares regression method can be used to inverse the electrolyte leakage.AsNHX2 gene was cloned from penncross by RACE methods.The ORF of AsNHX2 is 1617bp and belongs to the NHX class ? family.qRT-PCR analysis showed that the expression of AsNHX2 gene was the highest in the new leaves.It could be up-regulated by NaCl,low temperature and SA.35S::AsNHX2:YFP plasmid was constructed and transformed to the Nicotiana benthamiana L.leaf by Agrobacterium mediated transformation.The subcellular localization results showed that the AsNHX2 gene was localized in the vacuole membrane.AsNHX2-transgenic tobacco plants were generated by Agrobacterium mediated transformation.Compared with the control,the AsNHX2-overexpressing tobacco plants grew better under salt treatment.It could be closely related to the better absorbing ability of K+,higher photosynthetic efficiency,better degradation of reactive oxygen species and up-regulation of the expression of NtSOD,NtCAT,NtDREB4 and NtP5CS.In summary,this study proved that there are differences between the seaside ? and penncross in both molecular and physiological mechanisms.The indicators measured in this study were lower in the initial stage but increased to higher level in seaside ? than penncross under salt stress.The model constructed by partial least squares regression method could be used to inverse the electrolyte leakage validly.The AsNHX2 was localized in the vacuole membrane,and could play positive roles in the salt resistance progress in plants.