Studies On Overexpression Of BvNHX Gene In Sainfoin And Identification The Salt Tolerance Of The Transgenic Plants

Soil salinization is one of the main factors of the biological stress for the majority of higher plants,it causes serious influence on crops growth and development,especially.The results showed that the Na~+/H~+ antiporter of tonoplast membrane was induced by Na~+ in the cytoplasm by the action of H~+-ATP and H~+-PPase to reduce the toxicity of Na~+ to the cells.Under salt stress,maintain a stable pH and ion steady state play an important role.Previous studies have shown that sainfoin have limited salt tolerance,especially at the seedling stage,which is extremely sensitive to high salt.Therefore,it is very important to cultivate new varieties(lines)of sainfoin with strong salt tolerance,which is of great significance to the sustainable development of animal husbandry and the improvement and utilization of saline land in northwest China.In view of this,the study on the physiological response of salt stress was studied by using Agrobacterium tumefaciens-mediated transformation of Sugar beet BvNHX gene to sainfoin,and the salt tolerance was analyzed.The results obtained are as follows:1.Compared to the control(0 mmol/L),the plant biomass was significantly reduced in100 and 200 mmol/L.With the increase of salt concentration,the concentration of Na~+ in the shoot and the root of the sainfoin increased significantly.The concentration of Na~+ in the shoot under 50-200 mmol/L NaCl was 14-39 times higher than the control.Especially 200mmol/L NaCl,the root Na~+ net uptake rate increased by 35 times.Na~+ was significantly increased at 50 mmol/L NaCl,and the selectivity and transport capacity of K~+ and Na~+ were significantly decreased at the time of treatment.At the 9th day,compared with the control was decreased by 82% and 76%.The results showed that under the condition of 5-50 mmol/L NaCl,sainfoin was resistant to salt stress by maintaining its K~+ and Na~+ steady state balance.Under 100 and 200 mmol/L NaCl,a large amount of Na~+ was accumulated in shoot of sainfoin,regional and K~+ selective transport capacity is weak,leading to plant Na~+,K~+ steady state disorder,resulting in ion poisoning,thereby inhibiting its growth.2.The preliminary tissue culture system of sainfoin was established,the best explants was hypocotyls,the best callus medium was MS~+0.2 mg/L 6-BA~+0.5 mg/L NAA,and the somatic embryo differentiation medium was MS~+0.5 mg/L 6-BA ~+0.2 mg/L NAA.3.The BvNHX gene sainfoins were obtained by seedling infection method,it infected130 strains totally that survived 93 strains finally.By PCR and RT-PCR method that detected BvNHX gene transformed in sainfoin genome successfully and overexpressed,the transformation rate was about 10%,it obtained 9 strains of sainfoin which had overexpression of BvNHX gene.4.The fresh weight of transgenic(T0)plants was 27% and 44% higher than that of wild-type(WT)plants under 100 and 200 mmol/L NaCl,while there was no significant difference between the two groups under normal conditions(0 mmol/L).Under 200 mmol/L NaCl,the water content of T0 plants was 48% higher than that of WT plants.With the increase of NaCl concentration,Na~+ concentration in T0 and WT plants increased significantly.Under 200mmol/L NaCl,Na~+ in T0 plants was 24% lower than that of WT plants.The K~+ concentration in the shoots of T0 plants was 22% and 16% higher than that of WT plants at 100 and 200mmol/L NaCl,respectively.Under the 200 mmol/L NaCl,the ST value of T0 plants was no significant difference compared with 100 mmol/L Na Cl,and the WT plants decreased by 30%.It can be seen that overexpression of BvNHX gene in sainfoin increased the salt tolerance of plants.