Journal of Applied Genetics

, Volume 50, Issue 1, pp 9–16 | Cite as

Expression pattern of the pre-prothaumatin II gene under the control of the CaMV 35S promoter in transgenic cucumber (Cucumis sativus L.) flower buds and fruits

  • M. Szwacka
  • E. Siedlecka
  • R. Zawirska-Wojtasiak
  • Ł. Wiśniewski
  • S. Malepszy
Original Article


Thaumatin II is an extremely sweet-tasting protein produced by fruits of the West African shrubThaumatococcus daniellii Benth, so it can be used in biotechnology to improve the tastes of various plant products. This study is concerned with the spatial and temporal aspects of expression of the 35S-pre-prothaumatin II chimeric gene in flower buds and fruits of transgenic cucumber (Cucumis sativus L.) line 225. The activity of the 35S promoter in organs of line 225 was compared with its activity in 2 other transgenic lines. The accumulation of recombinant thaumatin varied spatially in flower bud tissues of transgenic lines. We found that these differences in the spatial accumulation of transgenic protein concerned the ovary of female buds and the perianth of male buds. In contrast to flower parts, recombinant thaumatin was found in nearly all parts of the young fruit from the transgenic plants. The pre-prothaumatin II gene expression was detected at a very early developmental stage in male buds, and its pattern was rather conserved as the buds aged. The expression of the transgene was also detected in vascular tissues of examined organs but was undetectable in pollen grains, in agreement with the generally held view that the CaMV 35S promoter is virtually silent in pollen. Immunocytochemical analyses of sections of control organs revealed endogenous homolog(s) of thaumatin when using polyclonal antisera, but not when using monoclonal antibodies for recombinant thaumatin detection in transgenic cucumber.


anti-thaumatin antibody CaMV 35S promoter Cucumis sativus L. floral organs immunocytochemistry pre-prothaumatin II transgenic cucumber lines transgene expression 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Bartoszewski G, Niedziela A, Szwacka M, Niemirowicz-Szczytt K, 2003. Modification of tomato taste in transgenic plants carrying a thaumatin gene from Thaumatococcus daniellii Benth. Plant Breed 122: 347–351.CrossRefGoogle Scholar
  2. Baulcombe DC, Saunders GR, Bevan MW, Mayo MA, Harrison BD, 1986. Expression of biologically active viral satellite RNA from the nuclear genome of transformed plants. Nature 321: 446–449.CrossRefGoogle Scholar
  3. Benfey PN, Ren L, Chua N-H, 1990. Tissue-specific expression from CaMV 35S enhancer subdomains in early stages of plant development. The EMBO J 9: 1677–1684.Google Scholar
  4. Beynon RJ, Dowswell P, Stephen AG, Tomkinson NP, 1994. Structural studies of thaumatins. In: Witty M, Higginbotham JD, eds. Thaumatin. CRC Press, Boca Raton-Ann Arbor-London-Tokyo: 123–133.Google Scholar
  5. Bhullar S, Datta S, Advani S, Chakravarthy S, Gautam T, Pental D, Burma PK, 2007. Functional analysis of cauliflower mosaic virus 35S promoter: re-evaluation of the role of subdomains B5, B4 and B2 in promoter activity. Plant Biotechnol J 5: 696–708.CrossRefPubMedGoogle Scholar
  6. Cordero de Mesa M, Santiago-Doménech N, Pliego-Alfaro F, Quesada MA, Mercado JA, 2004. The CaMV 35S promoter is highly active on floral organs and pollen of transgenic strawberry plants. Plant Cell Rep 23: 32–38.Google Scholar
  7. Dixon DC, Klessig DF, 1995. Immunolocalization of proteins in fixed and embedded plant tissues. In: Maliga P, Klessig DF, Cashmore AR, Gruissen W, Varner JE eds. Methods in Plant Molecular Biology: a Laboratory Manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor: 95–100.Google Scholar
  8. Hough CAM, Edwardson JA, 1978. Antibodies to thaumatin as a model of the sweet taste receptor. Nature 271: 381–383.CrossRefPubMedGoogle Scholar
  9. Iglesias VA, Moscone EA, Papp I, Neuhuber F, Michalowski S, Phelan T, et al. 1997. Molecular and cytogenetic analyses of stably and unstably expressed transgene loci in tobacco. The Plant Cell 9: 1251–1264.CrossRefPubMedGoogle Scholar
  10. Jayasankar S, Li ZJ, Gray DJ, 2003. Constitutive expression of Vitis vinifera thaumatin-like protein after in vitro selection and its role in anthracnose resistance. Functional Plant Biol 30: 1105–1115.CrossRefGoogle Scholar
  11. Liljeroth E, Marttila S, von Bothmer R, 2005. Immunolocalization of defence-related proteins in the floral organs of barley (Hordeum vulgare L.). J Phytopathol 153: 702–709.CrossRefGoogle Scholar
  12. Mascarenhas JP, Hamilton DA, 1992. Artifacts in the localization of GUS activity in anthers of petunia transformed with a CaMV35S-GUS construct. The Plant J 2: 405–408.CrossRefGoogle Scholar
  13. Matzke AJ, Matzke MA, 1998. Position effects and epigenetic silencing of plant transgenes. Curr Opin Plant Biol 1: 142–148.CrossRefPubMedGoogle Scholar
  14. Niemirowicz-Szczytt K, 1993. Hodowla roślin dyniowatych. In: Wydawnictwo SGGW, Hodowla Roślin Warzywnych: 185–236.Google Scholar
  15. Pret’ová A, Obert B, Wetzstein HY, 2001. Leaf developmental stage and tissue location affect the detection of â-glucuronidase in transgenic tobacco plants. Biotechnol Lett 23: 555–558.CrossRefGoogle Scholar
  16. Pressey R, 1997. Two isoforms of NP24: A thaumatin-like protein in tomato fruit. Phytochemistry 44: 1241–1245.CrossRefPubMedGoogle Scholar
  17. Saito S, Fujii N, Miyazawa Y, Yamasaki S, Matsuura S, Mizusawa H, Fujita Y, Takahashi H, 2007. Correlation between development of female flower buds and expression of the CS-ACS2 gene in cucumber plants. J Exp Bot 58: 2897–2907.CrossRefPubMedGoogle Scholar
  18. Slootstra JW, De Geus P, Haas H, Verrips CT, Meloen RH, 1995. Possible active site of the sweet-tasting protein thaumatin. Chem Senses 20: 535–543.CrossRefPubMedGoogle Scholar
  19. Szwacka M, Morawski M, Burza W, 1996. Agrobacterium tumefaciens-mediated cucumber transformation with thaumatin II cDNA. J Appl Genet 37A: 126–129.Google Scholar
  20. Szwacka M, Krzymowska M, Osuch A, Kowalczyk ME, Malepszy S, 2002a. Variable properties of transgenic cucumber plants containing the thaumatin II gene from Thaumatococcus daniellii. Acta Physiol Plantarum 24: 173–185.CrossRefGoogle Scholar
  21. Szwacka M, Piworun E, Ozdoba A, Szeląg T, Urbańczyk-Wochniak E, 2002b. Spatial and temporal expression of the sweet protein (thaumatin II) gene in transgenic cucumber plants. 10th IAPTC&B Congress on Plant Biotechnology 2002 and Beyond, Orlando Florida, June 23–28: 42-A.Google Scholar
  22. Tagashira N, Plader W, Filipecki M, Yin Z, Wiśniewska A, Gaj P, et al. 2005. The metabolic profiles of transgenic cucumber lines vary with different chromosomal locations of the transgene. Cell Mol Biol Lett 10: 697–710.PubMedGoogle Scholar
  23. Terada R, Shimamoto K, 1990. Expression of CaMV35S-GUS gene in transgenic rice plants. Mol Gen Genet 220: 389–392.CrossRefGoogle Scholar
  24. Thomas CE, Inaba T, Cohen Y, 1987. Physiological specialization in Pseudoperonospora cubensis. Phytopathol 77: 1621–1624.CrossRefGoogle Scholar
  25. Urbanczyk-Wochniak E, Filipecki M, Przybecki Z, 2002. A useful protocol for in situ RT-PCR on plant tissues. Cell Mol Biol Lett 7: 7–18.PubMedGoogle Scholar
  26. van der Leede-Plegt LM, van de Ven BCE, Bino RJ, van der Salm TPM, van Tunen A.J, 1992. Introduction and differential use of various promoters in pollen grains of Nicotiana glutinosa and Lilium longiflorum. Plant Cell Rep 11: 20–24.CrossRefGoogle Scholar
  27. van der Wel H, Bel WJ, 1980. Enzymatic properties of the sweet-tasting proteins thaumatin and monellin after partial reduction. Eur J Biochem 104: 413–418.CrossRefGoogle Scholar
  28. van Loon LC, Rep M, Pieterse CMJ, 2006. Significance of inducible defense-related proteins in infected plants. Annu Rev Phytopathol 44: 135–162.CrossRefGoogle Scholar
  29. Wilkinson JE, Twell D, Lindsey K, 1997. Activities of CaMV 35S and nos promoters in pollen: implication for field release of transgenic plants. J Exp Bot 48: 265–275.CrossRefGoogle Scholar
  30. Witty M, Harvey WJ, 1990. Sensory evaluation of transgenic Solanum tuberosum producing r-thaumatin II. N Zealand J Crop Hort Sci 18: 77–80.Google Scholar

Copyright information

© Institute of Plant Genetics, Polish Academy of Sciences, Poznan 2009

Authors and Affiliations

  • M. Szwacka
    • 1
  • E. Siedlecka
    • 1
  • R. Zawirska-Wojtasiak
    • 2
  • Ł. Wiśniewski
    • 1
  • S. Malepszy
    • 1
  1. 1.Department of Plant Genetics, Breeding and BiotechnologyWarsaw University of Life SciencesWarszawaPoland
  2. 2.Department of Food Science and NutritionPoznan University of Life SciencesPoznanPoland

Personalised recommendations