The Arabidopsis vacuolar Na+/H+ antiporter gene,AtNHX1,was introduced into soybean by Agrobacterium-mediated transformation.Four independent kanamycin resistant lines were obtained.The result of PCR,Southern blotting ...The Arabidopsis vacuolar Na+/H+ antiporter gene,AtNHX1,was introduced into soybean by Agrobacterium-mediated transformation.Four independent kanamycin resistant lines were obtained.The result of PCR,Southern blotting and Northern blotting analyses demonstrated that the AtNHX1 gene was successfully inserted into the soybean genome and stably expressed in these kanamycin resistant lines.The stability of AtNHX1 expression and salt resistance were evaluated in the soybean transformants for over 6 generations.Two independently derived transgenic lines with high expression level of AtNHX1 were selected,and propagated to generation T5 in the absence of selection pressure.PCR and RT-PCR examinations revealed that AtNHX1 was highly expressed in all investigated transgenic T5 progenies.Furthermore,all transgenic T5 plants showed resistant to salt stress,same as those of homozygous T2 plants.Taken together,our results indicated that constitutive expression of AtNHX1 enhanced salt tolerance in soybean for over 6 generations,suggesting a great potential use of AtNHX1 for improving salt tolerance in plants by genetic engineering.展开更多
Soil salinity severely hampers agricultural productivity.Under salt stress,excess Na+accumulation causes cellular damage and plant growth retardation,and membrane Na+transporters play central roles in Na+uptake and ex...Soil salinity severely hampers agricultural productivity.Under salt stress,excess Na+accumulation causes cellular damage and plant growth retardation,and membrane Na+transporters play central roles in Na+uptake and exclusion to mitigate these adverse effects.In this study,we performed sos1 suppressor mutant(named sup)screening to uncover potential genetic interactors of SOS1 and additional salt tolerance mechanisms.Map-based cloning and sequencing identified a group of mutants harboring dominant gain-of-function mutations in the vacuolar Na+/H+antiporter gene AtNHX1.The gain-of-function variants of AtNHX1 showed enhanced transporter activities in yeast cells and increased salt tolerance in Arabidopsis wild type plants.Ion content measurements indicated that at the cellular level,these gain-of-function mutations resulted in increased cellular Na+accumulation likely due to enhanced vacuolar Na+sequestration.However,the gain-of-function suppressor mutants showed reduced shoot Na+but increased root Na+accumulation under salt stress,indicating a role of AtNHX1 in limiting Na+translocation from root to shoot.We also identified another group of sos1 suppressors with loss-of-function mutations in the Na+transporter gene AtHKT1.Loss-of-function mutations in AtHKT1 and gain-of-function mutations in AtNHX1 additively suppressed sos1 salt sensitivity,which indicates that the three transporters,SOS1,AtNHX1 and AtHKT1 function independently but coordinately in controlling Na+homeostasis and salt tolerance in Arabidopsis.Our findings provide valuable information about the target amino acids in NHX1 for gene editing to improve salt tolerance in crops.展开更多
基金supported by National Basic Research Program of China (Grant Nos. 2006CB100106 and 2010CB126600)National Natural Science Foundation of China (Grant Nos. 30571196, 0933ZF11C1 and 0933Z411C1)+1 种基金The Ministry of Science and Technology of China (Grant No. 2007AA10Z187)the Key Laboratory of Soybeam Biology in Chinese Education Ministry and Shanghai Key Laboratory of Bio-Energy Crops (Grant No. 08DZ2270800)
文摘The Arabidopsis vacuolar Na+/H+ antiporter gene,AtNHX1,was introduced into soybean by Agrobacterium-mediated transformation.Four independent kanamycin resistant lines were obtained.The result of PCR,Southern blotting and Northern blotting analyses demonstrated that the AtNHX1 gene was successfully inserted into the soybean genome and stably expressed in these kanamycin resistant lines.The stability of AtNHX1 expression and salt resistance were evaluated in the soybean transformants for over 6 generations.Two independently derived transgenic lines with high expression level of AtNHX1 were selected,and propagated to generation T5 in the absence of selection pressure.PCR and RT-PCR examinations revealed that AtNHX1 was highly expressed in all investigated transgenic T5 progenies.Furthermore,all transgenic T5 plants showed resistant to salt stress,same as those of homozygous T2 plants.Taken together,our results indicated that constitutive expression of AtNHX1 enhanced salt tolerance in soybean for over 6 generations,suggesting a great potential use of AtNHX1 for improving salt tolerance in plants by genetic engineering.
基金supported by the US Department of Agriculture[National Research Initiative project 2007–35100-18378 to H.S.].
文摘Soil salinity severely hampers agricultural productivity.Under salt stress,excess Na+accumulation causes cellular damage and plant growth retardation,and membrane Na+transporters play central roles in Na+uptake and exclusion to mitigate these adverse effects.In this study,we performed sos1 suppressor mutant(named sup)screening to uncover potential genetic interactors of SOS1 and additional salt tolerance mechanisms.Map-based cloning and sequencing identified a group of mutants harboring dominant gain-of-function mutations in the vacuolar Na+/H+antiporter gene AtNHX1.The gain-of-function variants of AtNHX1 showed enhanced transporter activities in yeast cells and increased salt tolerance in Arabidopsis wild type plants.Ion content measurements indicated that at the cellular level,these gain-of-function mutations resulted in increased cellular Na+accumulation likely due to enhanced vacuolar Na+sequestration.However,the gain-of-function suppressor mutants showed reduced shoot Na+but increased root Na+accumulation under salt stress,indicating a role of AtNHX1 in limiting Na+translocation from root to shoot.We also identified another group of sos1 suppressors with loss-of-function mutations in the Na+transporter gene AtHKT1.Loss-of-function mutations in AtHKT1 and gain-of-function mutations in AtNHX1 additively suppressed sos1 salt sensitivity,which indicates that the three transporters,SOS1,AtNHX1 and AtHKT1 function independently but coordinately in controlling Na+homeostasis and salt tolerance in Arabidopsis.Our findings provide valuable information about the target amino acids in NHX1 for gene editing to improve salt tolerance in crops.
