Skip to main content
Log in

Enhanced ascorbic acid accumulation in transgenic potato confers tolerance to various abiotic stresses

  • Original Research Paper
  • Published:
Biotechnology Letters Aims and scope Submit manuscript

Abstract

l-Ascorbic acid (Vitamin C, AsA) is an important component of human nutrition. Plants and several animals can synthesize their own ascorbic acid, whereas humans lack the gene essential for ascorbic acid biosynthesis and must acquire from their diet. In the present study, we developed transgenic potato (Solanum tuberosum L. cv. Taedong Valley) over-expressing l-gulono-γ-lactone oxidase (GLOase gene; NCBI Acc. No. NM022220), isolated from rat cells driven by CaMV35S constitutive promoter that showed enhanced AsA accumulation. Molecular analyses of four independent transgenic lines performed by PCR, Southern and RT-PCR revealed the stable integration of the transgene in the progeny. The transformation frequency was ca. 7.5% and the time required for the generation of transgenic plants was 6–7 weeks. Transgenic tubers showed significantly enhanced AsA content (141%) and GLOase activity as compared to untransformed tubers. These transgenics were also found to withstand various abiotic stresses caused by Methyl Viologen (MV), NaCl or mannitol, respectively. The T1 transgenic plants exposed to salt stress (100 mM NaCl) survived better with increased shoot and root length when compared to untransformed plants. The elevated level of AsA accumulation in transgenics was directly correlated with their ability to withstand abiotic stresses. These results further demonstrated that the overexpression of GLOase gene enhanced basal levels of AsA in potato tubers and also the transgenics showed better survival under various abiotic stresses.

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

Access this article

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

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  • Agius F, Lamothe RG, Caballero JL, Blanco JM, Botella MA, Valpuesta V (2003) Engineering increased vitamin C levels in plants by over-expression of a d-galacturonic acid reductase. Nat Biotechnol 21:177–181

    Article  CAS  PubMed  Google Scholar 

  • Arnon DI (1949) Copper enzyme in isolated chloroplasts: polyphenol oxidase in Beta vulgaris. Plant Physiol 24:1–15

    Article  CAS  PubMed  Google Scholar 

  • Chen Z, Todd E, Ling YJ, Chang SC, Gallie DR (2003) Increasing vitamin C content of plants through enhanced ascorbate recycling. Proc Natl Acad Sci USA 100:3525–3530

    Article  CAS  PubMed  Google Scholar 

  • Conklin PL, Barth C (2004) Ascorbic acid, a familiar small molecule intertwined in the response of plants to ozone, pathogens, and the onset of senescence. Plant Cell Environ 27:959–970

    Article  CAS  Google Scholar 

  • Crowell EF, Mitchell JM, David SD (2008) Accumulation of vitamin E in potato (Solanum tuberosum) tubers. Transgenic Res 17:205–217

    Article  CAS  PubMed  Google Scholar 

  • Dabrowski K (1990) Gulonolactone oxidase is missing in teleost fish-the direct spectrophotometric assay. Biol Chem Hoppe-Seyler 37:207–214

    Google Scholar 

  • Davey MW, Gilot C, Persiau G, Østergaard J, Han Y, Bauw GC, Van Montagu MC (1999) Ascorbate biosynthesis in Arabidopsis cell suspension culture. Plant Physiol 121:535–543

    Article  CAS  PubMed  Google Scholar 

  • Doyle JJ, Doyle JL (1987) A rapid DNA isolation procedure for small quantities of fresh tissue. Phytochem Bull 19:11–15

    Google Scholar 

  • Fan L, Xheng S, Xuemin W (1997) Antisense suppression of phospholipase d-γ retards abscisic acid- and ethylene-promoted senescence of postharvest Arabidopsis leaves. Plant Cell 9:2183–2196

    Article  CAS  PubMed  Google Scholar 

  • Han JS, Kozukue N, Young KS, Lee KR, Friedman M (2004) Distribution of ascorbic acid in potato tubers and in home-processed and commercial foods. J Agric Food Chem 52:6516–6521

    Article  CAS  PubMed  Google Scholar 

  • Hemavathi, Upadhyay CP, Ko EY, Nookaraju A, Kim HS, Heung JJ, Oh MO, Reddy AC, Chun SC, Kim DH, Park SW (2009) Over-expression of strawberry d-galacturonic acid reductase in potato leads to accumulation of vitamin C with enhanced abiotic stress tolerance. Plant Sci 177(5) (in press)

  • Jain AK, Nessler CL (2000) Metabolic engineering of an alternative pathway for ascorbic acid biosynthesis in plants. Mol Breed 6:73–78

    Article  CAS  Google Scholar 

  • Kwon SY, Jeong YJ, Lee HS, Kim JS, Cho KY, Allen RD (2002) Enhanced tolerances of transgenic tobacco plants expressing both superoxide dismutase and ascorbate peroxidase in chloroplasts against methyl viologen mediated oxidative stress. Plant Cell Environ 25:873–882

    Article  Google Scholar 

  • Kwon SY, Choi SM, Ahn YO, Lee HS, Lee HB, Park YM, Kwak SS (2003) Enhanced stress-tolerance of transgenic plants expressing a human dehydroascorbate reductase gene. J Plant Physiol 160:347–353

    Article  CAS  PubMed  Google Scholar 

  • Lechno S, Zamski E, Tel-Or E (1997) Salt stress-induced responses in cucumber plants. J Plant Physiol 150:206–211

    CAS  Google Scholar 

  • Loewus FA (1963) Tracer studies on ascorbic acid formation in plants. Phytochemistry 2:109–128

    Article  CAS  Google Scholar 

  • Lorence A, Chevone BI, Mendes P, Nessler CL (2004) Myo-inositol oxygenase offers a possible entry point into plant ascorbate biosynthesis. Plant Physiol 134:1200–1205

    Article  CAS  PubMed  Google Scholar 

  • Mano J, Ohno C, Domae Y, Asada K (2001) Chloroplastic ascorbate peroxidase is the primary target of methylviologen-induced photooxidative stress in spinach leaves: its relevance to monodehydroascorbate radical detected with in vivo ESR. Biochem Biophys Acta 1504:275–287

    Article  CAS  PubMed  Google Scholar 

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

    Article  CAS  Google Scholar 

  • Oba K, Fukui M, Imai Y, Iriyama S, Nogami K (1994) l-Galactono-γ-lactone dehydrogenase: partial characterization, induction of activity and role in the synthesis of ascorbic acid in wounded white potato tuber tissue. Plant Cell Physiol 35:473–478

    CAS  Google Scholar 

  • Pallanca JE, Smirnoff N (1999) Ascorbic acid metabolism in pea seedlings. A comparison of d-glucose, l-sorbosone, and l-galactono-1,4-lactone as ascorbate precursors. Plant Physiol 120:453–461

    Article  CAS  PubMed  Google Scholar 

  • Sambrook J, Fritsch EF, Maniatis T (1989) Molecular cloning: a laboratory manual, 2nd edn. Cold Spring Harbour Laboratory Press, Cold Spring Harbor, pp 9.31–9.32

    Google Scholar 

  • Shalata A, Tal M (1998) The effect of salt stress on lipid peroxidation and antioxidants in the leaf of the cultivated tomato and its wild salt-tolerant relative Lycoperscion pennellii. Physiol Plant 104:169–174

    Article  CAS  Google Scholar 

  • Smirnoff N (1996) The function and metabolism of ascorbic acid in plants. Ann Bot 78:661–669

    Article  CAS  Google Scholar 

  • Smirnoff N (2001) l-Ascorbic acid biosynthesis. Vitam Horm 61:241–266

    Article  CAS  PubMed  Google Scholar 

  • Tokunaga T, Miyahara K, Tabata K, Esaka M (2005) Generation and properties of ascorbic acid-overproducing transgenic tobacco cells expressing sense RNA for l-galactono-1,4-lactone dehydrogenase. Planta 220:854–863

    Article  CAS  PubMed  Google Scholar 

  • Ulloa RM, Mac-Intosh GC, Melchiorre M, Mentaberry AN, Dallari P, Moriconi DN, Tellez-Inon MT (1997) Protein kinase activity in different stages of potato (Solanum tuberosum L.) microtuberization. Plant Cell Rep 16:426–429

    CAS  Google Scholar 

  • Wheeler GL, Jones MA, Smirnoff N (1998) The biosynthesis pathway of vitamin C in higher plants. Nature 393:365–369

    Article  CAS  PubMed  Google Scholar 

  • Wolucka BA, Van Montagu M (2003) GDP-mannose 3′,5′-epimerase forms GDP-l-gulose, a putative intermediate for the de novo biosynthesis of vitamin C in plants. J Biol Chem 278:47483–47490

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This research was supported by the Konkuk University research fund. The research fellowship to Hemavathi as “Research Fellow” from Konkuk University is gratefully acknowledged.

Author information

Authors and Affiliations

Authors

Corresponding authors

Correspondence to Chandrama Prakash Upadhyaya, Doo Hwan Kim or Se Won Park.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Hemavathi, Upadhyaya, C.P., Akula, N. et al. Enhanced ascorbic acid accumulation in transgenic potato confers tolerance to various abiotic stresses. Biotechnol Lett 32, 321–330 (2010). https://doi.org/10.1007/s10529-009-0140-0

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10529-009-0140-0

Keywords

Navigation