Skip to main content
SpringerLink
Log in

Stable expression of Arabidopsis vacuolar Na+/H+ antiporter gene AtNHX1, and salt tolerance in transgenic soybean for over six generations

  • Articles
  • Gene Engineering
  • Published:
Chinese Science Bulletin

Abstract

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.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  1. Gonzalez N, Beemster G T S, Inzé D. David and Goliath: What can the tiny weed Arabidopsis teach us to improve biomass production in crops? Curr Opin Plant Biol, 2009, 12: 157–164

    Article  Google Scholar 

  2. Apse M P, Blumwald E. Na+ transport in plants. FEBS Lett, 2007, 581: 2247–2254

    Article  Google Scholar 

  3. Brett C L, Donowitz M, Rao R. Evolutionary origins of eukaryotic sodium/proton exchangers. Am J Physiol Cell Physiol, 2005, 288: C223–C239

    Article  Google Scholar 

  4. Zhao J, Cheng N H, Motes C M, et al. AtCHX13 is a plasma membrane K+ transporter. Plant Physiol, 2008, 148: 796–807

    Article  Google Scholar 

  5. Sze H, Padmanaban S, Cellier F, et al. Expression patterns of a novel AtCHX gene family highlight potential roles in osmotic adjustment and K+ homeostasis in pollen development. Plant Physiol, 2004, 136: 2532–2547

    Article  Google Scholar 

  6. Pardo J M, Cubero B, Leidi E O, et al. Alkali cation exchangers: Roles in cellular homeostasis and stress tolerance. J Biol Chem, 2006, 57: 1181–1199

    Google Scholar 

  7. Yokoi S, Quintero F J, Cubero B, et al. Differential expression and function of Arabidopsis thaliana NHX Na+/H+ antiporters in the salt stress response. Plant J, 2002, 30: 529–539

    Article  Google Scholar 

  8. Shi H Z, Wu S J, Zhu J K. Overexpression of a plasma membrane Na+/H+ antiporter improves salt tolerance in Arabidopsis. Nat Biotechnol, 2003, 21: 81–85

    Article  Google Scholar 

  9. An R, Chen Q J, Chai M F, et al. AtNHX8, a member of the monovalent cation: proton antiporter-1 family in Arabidopsis thaliana, encodes a putative Li+/H+ antiporter. Plant J, 2007, 49: 718–728

    Article  Google Scholar 

  10. Liu H, Wang Q Q, Yu M M, et al. Transgenic salt-tolerant sugar beet (Beta vulgaris L.) constitutively expressing an Arabidopsis thaliana vacuolar Na+/H+ antiporter gene, AtNHX3, accumulate more soluble sugar but less salt in storage roots. Plant Cell Environ, 2008, 31: 1325–1334

    Article  Google Scholar 

  11. Li H T, Liu H, Gao X S, et al. Knock-out of Arabidopsis AtNHX4 gene enhances tolerance to salt stress. Biochem Biophys Res Co, 2009, 382: 637–641

    Article  Google Scholar 

  12. He C, Yan J, Shen G, et al. Expression of an Arabidopsis vacuolar sodium/proton antiporter gene in cotton improves photosynthetic performance under salt conditions and increases fiber yield in the field. Plant Cell Physiol, 2005, 46: 1848–1854

    Article  Google Scholar 

  13. Wu Y Y, Chen Q J, Chen M, et al. Salt-tolerant transgenic perennial ryegrass (Lolium perenne L.) obtained by Agrobacterium tumefaciens-mediated transformation of the vacuolar Na+/H+ antiporter gene. Plant Sci, 2005, 169: 65–73

    Article  Google Scholar 

  14. Zhang H X, Blumwald E. Transgenic salt-tolerant tomato plants accumulate salt in foliage but not in fruit. Nat Biotechnol, 2001, 19: 765–768

    Article  Google Scholar 

  15. Zhang H X, Hodson J N, William J P, et al. Engineering salt tolerant Brassica plants: Characterization of yield and seed oil quality in transgenic plants with increased vacuolar sodium accumulation. Proc Natl Acad Sci USA, 2001, 98: 12832–12836

    Article  Google Scholar 

  16. Apse M P, Scottosanto J B, Blumwald E. Vacuolar cation/H+ exchange, ion homeostasis, and leaf development are altered in a T-DNA insertional mutant of AtNHX1, the Arabidopsis vacuolar Na+/H+ antiporter. Plant J, 2003, 36: 229–239

    Article  Google Scholar 

  17. Venema K, Belver A, Marin-Manzano M C, et al. A novel intracellular K+/H+ antiporter related to Na+/H+ antiporters is important for K+ ion homeostasis in plants. J Biol Chem, 2003, 278: 22453–22459

    Article  Google Scholar 

  18. Counillon L, Pouyssseur J. The expanding family of eukaryotic Na+/H+ exchangers. J Biol Chem, 2000, 275: 1–4

    Article  Google Scholar 

  19. Nass R, Cunningham K W, Rao R. Intracellular sequestration of sodium by a novel Na+/H+ exchanger in yeast is enhanced by mutations in the plasma membrane H+-ATPase. J Biol Chem, 1997, 272: 26145–26152

    Article  Google Scholar 

  20. Yamaguchi T, Apse M P, Shi H Z, et al. Topological analysis of a plant Na+/H+ antiporter reveals a luminal C terminus that regulates antiporter cation selectivity. Proc Natl Acad Sci USA, 2003, 100: 12510–12515

    Article  Google Scholar 

  21. Yamaguchi T, Aharon G S, Sottosanto J B, et al. Vacuolar Na+/H+ antiporter cation selectivity is regulated by calmodulin from within the vacuole in a Ca2+- and pH-dependent manner. Proc Natl Acad Sci USA, 2005, 102: 16107–16112

    Article  Google Scholar 

  22. Apse M P, Aharon G S, Snedden W S, et al. Salt tolerance conferred by overexpression of a vacuolar Na+/H+ antiport in Arabidopsis. Science, 1999, 285: 1256–1258

    Article  Google Scholar 

  23. Chen L H, Zhang B, Xu Z Q. Salt tolerance conferred by overexpression of Arabidopsis vacuolar Na+/H+ antiporter gene AtNHX1 in common buckwheat (Fagopyrum esculentum). Transgenic Res, 2008, 17: 121–132

    Article  Google Scholar 

  24. Xue Z Y, Zhi D Y, Xue G P, et al. Enhanced salt tolerance of transgenic wheat (Tritivum aestivum L.) expressing a vacuolar Na+/H+ antiporter gene with improved grain yields in saline soils in the field and a reduced level of leaf Na+. Plant Sci, 2004, 167: 849–859

    Article  Google Scholar 

  25. Murashige T, Skoog F. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol Plant, 1962, 15: 473–497

    Article  Google Scholar 

  26. Liu H K, Wei Z M. A method for sterilizing mature seeds of soybean. Plant Physiol Commun, 2002, 38: 261–262

    Google Scholar 

  27. Liu H K, Yang C, Wei Z M. Efficient Agrobacterium tumefaciens-mediated transformation of soybeans using an embryonic tip regeneration system. Planta, 2004, 219: 1042–1049

    Article  Google Scholar 

  28. Sambrook J, Russell D W. Molecular Cloning: A Laboratory Manual. 3rd ed (Vols. 1–3). Cold Spring Harbor: Cold Spring Harbor Laboratory Press, 2001

    Google Scholar 

  29. Fiehn O, Kopka J, Trethewey R N, et al. Identification of uncommon plant metabolites based on calculation of elemental compositions using gas chromatography and quadrupole mass spectrometry. Anal Chem, 2000, 72: 3573–3580

    Article  Google Scholar 

  30. Hoagland D R, Arnon D I. The water culture method for growing plants without soil. California Agricultural Experiment Station Circular 371. The college of Agriculture University of California. Berkerley, CA, USA, 1950

    Google Scholar 

  31. Wang B S, Zhao K F. Comparison of extractive methods of Na+, K+ in wheat leaves. Plant Physiol Commun, 1995, 31: 50–52

    Google Scholar 

  32. Fukuda A, Nakamura A, Tagiri A, et al. Function, intracellular localization and the importance in salt tolerance of a vacuolar Na+/H+ antiporter from rice. Plant Cell Physiol, 2004, 45: 146–159

    Article  Google Scholar 

  33. Ohta M, Hayashi Y, Nakashima A, et al. Introduction of a Na+/H+ antiporter gene from Atriplex gmelini confers salt tolerance to rice. FEBS Lett, 2002, 532: 279–282

    Article  Google Scholar 

  34. Fukada-Tanaka S, Inagaki Y, Yamaguchi T, et al. Color-enhancing protein in blue petals: Spectacular morning glory blooms rely on a behind-scenes proton exchanger. Nature, 2000, 407: 581

    Article  Google Scholar 

  35. Xia T, Apse M P, Aharon G S, et al. Identification and characterization of a NaCl-inducible vacuolar Na+/H+ antiporter in Beta vulgaris. Plant Physiol, 2002, 116: 206–212

    Article  Google Scholar 

  36. Wu C A, Yang G D, Meng Q W, et al. The cotton GhNHX1 gene encoding a novel putative tonoplast Na+/H+ antiporter plays an important role in salt stress. Plant Cell Physiol, 2004, 45: 600–607

    Article  Google Scholar 

  37. Zorb C, Noll A, Karl S, et al. Molecular characterization of Na+/H+ antiporters (ZmNHX) of maize (Zea mays L.) and their expression under salt stress. J Plant Physiol, 2005, 162: 55–66

    Article  Google Scholar 

Download references

Author information

Author notes

  1. TianXing (YiZhou) Li

    Present address: Department of Chemistry and Life Sciences, Chuxiong Normal University, Chuxiong, 675000, China

Authors and Affiliations

  1. College of Life Sciences, Yunnan University, Kunming, 650091, China

    TianXing (YiZhou) Li

  2. National Key Laboratory of Plant Molecular Genetics, Shanghai Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, 200032, China

    TianXing (YiZhou) Li, Yue Zhang, Hua Liu, YuTing Wu & HongXia Zhang

  3. Key Laboratory of Soybean Biology in Chinese Education Ministry, Northeast Agricultural University, Harbin, 150030, China

    WenBin Li

Authors
  1. TianXing (YiZhou) Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yue Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Hua Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. YuTing Wu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. WenBin Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. HongXia Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to HongXia Zhang.

Additional information

Contributed equally to this work

About this article

Cite this article

Li, T.(., Zhang, Y., Liu, H. et al. Stable expression of Arabidopsis vacuolar Na+/H+ antiporter gene AtNHX1, and salt tolerance in transgenic soybean for over six generations. Chin. Sci. Bull. 55, 1127–1134 (2010). https://doi.org/10.1007/s11434-010-0092-8

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11434-010-0092-8

Keywords

Search

Navigation