Salt Tolerance In Transgenic Plants Research Articles

Two rice cytosolic ascorbate peroxidases differentially improve salt tolerance in transgenic Arabidopsis

In order to determine the different roles of rice (Oryza sativa L.) cytosolic ascorbate peroxidases (OsAPXa and OsAPXb, GenBank accession nos. D45423 and AB053297, respectively) under salt stress, transgenic Arabidopsis plants over-expressing OsAPXa or OsAPXb were generated, and they all exhibited increased tolerance to salt stress compared to wild-type plants. Moreover, transgenic lines over-expressing OsAPXb showed higher salt tolerance than OsAPXa transgenic lines as indicated by root length and total chlorophyll content. In addition to ascorbate peroxidase (APX) activity, antioxidant enzyme activities of catalase (CAT), superoxide dismutase (SOD) and glutathione reductase (GR), which are also involved in the salt tolerance process, and the content of H2O2 were also assayed in both transgenic and wild-type plants. The results showed that the overproduction of OsAPXb enhanced and maintained APX activity to a much higher degree than OsAPXa in transgenic Arabidopsis during treatment with different concentrations of NaCl, enhanced the active oxygen scavenging system, and protected plants from salt stress by equilibrating H2O2 metabolism. Our findings suggest that the rice cytosolic OsAPXb gene has a more functional role than OsAPXa in the improvement of salt tolerance in transgenic plants.

Overexpression AtNHX1 confers salt-tolerance of transgenic tall fescue

Saline soil is a serious problem worldwide, and it is necessary to improve the salt tolerance of plants so as to avoid the progressive deterioration of saline soil. Here we report that over-expression of AtNHX1 improves salt tolerance in transgenic tall fescue. The AtNHX1 gene driven with CaMV35S promoter was constructed into the plant expression vector pGreen0229, and introduced into the embryonic calli of hypocotyls of tall fescue (Festuca arundinacea ) by particle bombardment. Regenerated plantlets were obtained by screening of herbicide (PPT, 2 mg/L), and the putative transformants were assayed by PCR and western blot analysis. 29 transgenic plants were obtained. The results indicated that the exogenous genes had been integrated into the genomes of transgenic plants, and AtNHX1 is expressed in the plants. There was remarkable salt tolerance in transgenic plants compared to control plants.

Expression of an Arabidopsis Vacuolar Sodium/Proton Antiporter Gene in Cotton Improves Photosynthetic Performance Under Salt Conditions and Increases Fiber Yield in the Field

Drought and salinity are two major limiting factors in crop productivity. One way to reduce crop loss caused by drought and salinity is to increase the solute concentration in the vacuoles of plant cells. The accumulation of sodium ions inside the vacuoles provides a 2-fold advantage: (i) reducing the toxic levels of sodium in cytosol; and (ii) increasing the vacuolar osmotic potential with the concomitant generation of a more negative water potential that favors water uptake by the cell and better tissue water retention under high soil salinity. The success of this approach was demonstrated in several plants, where the overexpression of the Arabidopsis gene AtNHX1 that encodes a vacuolar sodium/proton antiporter resulted in higher plant salt tolerance. Overexpression of AtNHX1 increases sodium uptake in vacuoles, which leads to increased vacuolar solute concentration and therefore higher salt tolerance in transgenic plants. In an effort to engineer cotton for higher drought and salt tolerance, we created transgenic cotton plants expressing AtNHX1. These AtNHX1-expressing cotton plants generated more biomass and produced more fibers when grown in the presence of 200 mM NaCl in greenhouse conditions. The increased fiber yield was probably due to better photosynthetic performance and higher nitrogen assimilation rates observed in the AtNHX1-expressing cotton plants as compared with wild-type cotton plants under saline conditions. Furthermore, the field-grown AtNHX1-expressing cotton plants produced more fibers with better quality, indicating that AtNHX1 can indeed be used for improving salt stress tolerance in cotton.

Overexpression of mtlD gene in transgenic Populus tomentosa improves salt tolerance through accumulation of mannitol

The mtlD gene encoding mannitol-1-phosphate dehydrogenase, which catalyzes the biosynthesis of mannitol from fructose, was cloned from Escherichia coli and transferred to poplar (Populus tomentosa Carr.) through Agrobacterium-mediated transformation. The transgenic plants were screened and selected on Murashige and Skoog (MS) medium containing 30-50 mg l(-1) kanamycin and verified by polymerase chain reaction (PCR) and Southern blotting. Expression of the gene led to synthesis and accumulation of mannitol in the transgenic plants. Gas chromatography and mass spectrometry (GC/MS) and capillary gas chromatography (GC) showed that transgenic plants accumulated much more mannitol in their tissues than the wild-type plants, whether cultured in vitro, or grown hydroponically or in the field. Increased salt tolerance of transgenic plants was observed both in vitro and in hydroponic culture. The transgenic buds rooted normally on MS medium containing 50 mM NaCl, whereas wild-type buds did not. In the 40-day hydroponic experiments, transgenic poplar plants survived in a 75-mM NaCl treatment, whereas the wild-type poplar plants tolerated only 25 mM NaCl. Under the same NaCl stress, stomatal conductance, transpiration rates and photosynthetic rates were all higher in transgenic plants than in wild-type plants, whereas cellular relative conductivity was lower. We demonstrated that the mtlD gene was expressed in transgenic poplar plants, resulting either directly or indirectly in mannitol accumulation and improved salt tolerance. The constant mannitol concentrations in transgenic plants during the NaCl treatments indicated that mannitol accumulation caused by the mtlD gene was not a consequence of NaCl stress. Height growth was reduced by about 50% in the transgenic plants compared with the wild-type plants in the absence of salt; however, relative growth rate was much less influenced by salt stress in transgenic plants than in wild-type plants. The stunted growth of the transgenic plants may in part explain their improved salt tolerance.

Salt-Regulated Mannitol Metabolism in Algae

Mannitol, one of the most widely occurring sugar alcohol compounds, is found in bacteria, fungi, algae, and plants. In these organisms the compound acts as a compatible solute and has multiple functions, including osmoregulation, storage, and regeneration of reducing power, and scavenging of active oxygen species. Because of the diverse functions of mannitol, introducing the ability to accumulate it has been a hallmark of attempts to generate highly salt-tolerant transgenic plants. However, transgenic plants have not yet improved significantly in their salt tolerance. Recently, we purified and characterized 2 enzymes that biosynthesize mannitol, mannitol-1-phosphate dehydrogenase (M1PDH) and mannitol-1-phosphate-specific phosphatase, from the marine red alga Caloglossa continua, which grows in estuarine areas where tide levels fluctuate frequently. The activation of Caloglossa M1PDH is unique in that it is regulated by salt concentration at enzyme level. In this review we focus on the metabolism of mannitol, mainly in marine photosynthetic organisms, and suggest how this might be applied to producing salt-tolerant transgenic plants.

Construction of plant expression vector and analysis of herbicide resistance and salt tolerance of transgenic tobacco

AbstractThe plant expression vector pCAMBIA1300-AtNHX1-als was constructed by inserting the herbicide resistance gene als of Arabidopsis thaliana into the plasmid pCAMBIA1300-AtNHX1, which contains the AtNHX1 gene encoding the Na+/H+ antiport from the vacuolar membrane of A. thaliana. Transgenic tobacco plants were obtained via Agrobacterium-mediated transformation. PCR and Southern blot assay indicated that genes als and AtNHX1 had been integrated into the genome of the transgenic plants. The herbicide resistance and salt tolerance of transgenic plants increased by about 1000-fold and by 1.5% NaCl concentration, respectively, compared with controls. Herbicide resistance of the T1 progeny was evaluated by spraying transgenic plants with different concentrations of Luhuanglong at the four-leaf stage. Controls gradually died under 100 mg/l Luhuanglong whereas 73% of the T1 plants still survived at 500 mg/l Luhuanglong. Thus the plant expression vector pCAMBIA1300-AtNHX1-als could be used to improve the herbicide resistance and salt tolerance of crops.

A heat treatment induced the expression of a Na +/H + antiport gene ( cNHX1) in citrus fruit

In previous studies, we showed that a short hot water treatment (at 62 °C for 20 s) increased chilling tolerance in grapefruit. In order to gain more insight into the molecular mechanisms involved in heat-induced chilling tolerance responses, we performed cDNA differential display analysis and isolated a 600-bp polymerase chain reaction (PCR) fragment whose expression was enhanced 24 h after the heat treatment, and that showed high sequence homology with the 5′ region of various NHX1 vacuolar Na +/H + antiporter genes. The full-length cDNA sequence of cNHX1 (for citrus NHX1) was further isolated by 3′-RACE; it had a total length of 2135 bp with an open reading frame of 1626 bp, and encoded a predicted polypetide of 542 amino acids with a molecular mass of 59.8 kDa. The cNHX1 protein shares 83, 78 and 56% identity with the other recently isolated Arabidopsis thaliana, Atriplex gmelini and Oryza sativa NHX1 proteins, respectively. It has been reported that the Arabidopsis AtNHX1 gene partially complemented a salt-sensitive yeast mutation and that overexpression of this gene significantly increased salt tolerance in transgenic plants. In the current study, we showed that the expression of the cNHX1 gene was temporarily induced by a heat treatment, and that its mRNA levels markedly increase during a 6-week cold storage period at 2 °C in heat-treated fruit but not in control untreated fruit, thus suggesting its possible involvement in heat-induced chilling tolerance responses. The expression of the cNHX1 gene was also markedly induced by salt stress, but not by other stresses, such as exposure to ethylene or UV irradiation.

Engineering salt tolerance in crop plants

Environmental stress caused by salinity is a serious factor limiting the productivity of agricultural crops, which are predominantly sensitive to the presence of high concentrations of salts in the soil. Although drainage and the supply of high quality water can solve the problem, these measures are extremely costly and not applicable in extensive agriculture. Salt has a detrimental effect on plants because it causes a water deficit, which results in osmotic stress, and because of the effects of excess sodium ions on key biochemical processes. To tolerate high levels of salts, plants should be able to use ions for osmotic adjustment and to accumulate these ions in the vacuole to keep sodium away from the cytosol. It has been shown previously that overexpression of AtNHX1 (a vacuolar Na+–H+ antiport) in Arabidopsis plants, allowed the transgenic plants to grow in 200 mm NaCl, suggesting that crop plants with improved salt tolerance could be engineered.Hong-Xia Zhang and Eduardo Blumwald 1xTransgenic salt tolerant tomato plants accumulate salt in the foliage but not in the fruits. Zhang, H-X. and Blumwald, E. Nat. Biotechnol. 2001; 19: 765–768Crossref | PubMed | Scopus (627)See all References1 now report the generation of salt-tolerant transgenic tomato plants. Whereas wild-type plant growth was severely inhibited by salt, the transgenic plants overexpressing an Arabidopsis vacuolar Na+–H+ antiport were able to grow and produce fruit in the presence of 200 mm NaCl (Fig. 1Fig. 1). The sodium and chloride content of the leaves of the transgenic plants was 20-fold higher that in untransformed plants. This increase in sodium content in the transgenic plants correlated with an increase in the amount of antiport protein and an increase in Na+–H+ antiport activity in the tonoplast vesicles of transgenic plants compared with the untransformed plants. More significantly, whereas the leaves of the transgenic plants accumulated high sodium and chloride concentrations, the tomato fruits displayed low sodium and chloride content.Fig. 1(a) Control and (b) transgenic tomato plants growing in the presence of 200 mm NaCl. (Photographs courtesy of Eduardo Blumwald.)View Large Image | Download PowerPoint SlideIn summary, these results show that the accumulation of sodium into the vacuoles, mediated by the vacuolar Na+–H+ antiport, allows crop plants to minimize the toxic effects of salinity. In spite of many years of conventional breeding for salt tolerance, only a few varieties, with only limited salt tolerance, have been released. Salt tolerance is a complex trait, and a long list of salt-responsive genes has been reported. One school of thought has concluded that salt tolerance will be achieved only after pyramiding several characteristics into a single genotype, where each one alone could not confer a significant increase in salt tolerance. However, Zhang and Blumwald's work shows that it is possible to generate crop plants that can tolerate high salinity levels with the modification of a single gene. This result suggests that with the incorporation of transgenic modification, breeding of salt-tolerant crops will be possible with far fewer target traits than has been anticipated. A substantial area of irrigated land has been damaged by the accumulation of salt, and the process of soil salinization is a permanent threat. The generation of salt-tolerant transgenic plants is an essential step toward solving the problem of feeding the world's rapidly growing population.

Salt tolerance of transgenic rice (Oryza sativa L.) withmtlD gene andgutD gene

Southern blot analysis indicated thatmtlD gene (encoding mannitol-1-phosphate dehydrogenase) andgutD gene (encoding glucitol-6-phosphate dehydrogenase) had been integrated into the rice genome mediated byAgrobacterium tumefaciens LBA4404(pBIGM). The expression of the above two genes in transgenic rice plants was demonstrated by Northern blot analysis and enzymatic activity assay. Analysis of sugar alcohol showed that transgenic rice plants could produce and accumulate mannitol and sorbitol. The salt tolerance of transgenic plants was much higher than that of their controls.

Transfer ofE. coli gutD gene into maize and regeneration of salt-tolerant transgenic plants

GutD gene, encoding a key enzyme (glucitol-6-phosphate dehydrogenase) of sugar alcohol metabolic pathway inE. coli, was transferred into maize. Results of Southern and Western blotting analysis certified that this gene had integrated and been expressed in transgenic maize plants and their progeny. The synthesis and accumulation of sorbitol were detected in transgenic maize plants and a preliminary nutrient solution culture experiment showed that gutD transgenic maize plants had an increased tolerance to salt stress compared with nontransgenic ones.