Applied Biochemistry and Biotechnology

, Volume 168, Issue 4, pp 761–769 | Cite as

Enhanced Green Fluorescent Protein Expression in Pleurotus ostreatus for In Vivo Analysis of Fungal Laccase Promoters

  • Antonella Amore
  • Yoichi Honda
  • Vincenza FaracoEmail author


The laccase family of Pleurotus ostreatus has been widely characterized, and studies of the genes coding for laccase isoenzymes in P. ostreatus have so far led to the identification of four different genes and the corresponding cDNAs, poxc, pox1, poxa1b and poxa3. Analyses of P. ostreatus laccase promoters poxc, pox1, poxa1b and poxa3 have allowed identification of several putative response elements, and sequences of metal-responsive elements involved in the formation of complexes with fungal proteins have been identified in poxc and poxa1b promoters. In this work, development of a system for in vivo analysis of P. ostreatus laccase promoter poxc by enhanced green fluorescent protein expression is performed, based on a poly ethylene glycol-mediated procedure for fungal transformation. A quantitative measurement of fluorescence expressed in P. ostreatus transformants is hereby reported for the first time for this fungus.


Fungi Laccases Transformation Promoters Fluorescence 



The authors are grateful to Prof. Giovanni Sannia Department of Chemical Sciences, University of Naples “Federico II”, Prof. Yitzhak Hadar and Mr. Tomer M. Salome of The Hebrew University of Jerusalem, Israel, and Prof. Ursula Kües and Dr. Martin Ruhl of Georg-August-University Göttingen, Germany, for their assistance and helpful discussions to make simple and efficient transformation protocol in P. ostreatus. The authors also thank Dr. Giuliana Napolitano of Department of Structural and Functional Biology, University of Naples “Federico II”for kindly making available the fluorescence microscope and her competences in exploiting it.


  1. 1.
    Hublik, G., & Schinner, F. (2000). Enzyme and Microbial Technology, 27, 330–336.CrossRefGoogle Scholar
  2. 2.
    Moeder, M., Cajthaml, T., Koeller, G., Erbanová, P., & Sasek, V. (2005). Chemosphere, 61, 1370–1378.CrossRefGoogle Scholar
  3. 3.
    Jiang, G. X., Niu, J. F., Zhang, S. P., Zhang, Z. Y., & Xie, B. (2008). Bulletin of Environmental Contamination and Toxicology, 81, 1–6.CrossRefGoogle Scholar
  4. 4.
    Patel, H., Gupte, A., & Gupte, S. (2009). Applied Biochemistry and Biotechnology, 157, 367–376.CrossRefGoogle Scholar
  5. 5.
    Byss, M., Elhottová, D., Tříska, J., & Baldrian, P. (2008). Chemosphere, 73, 1518–1523.CrossRefGoogle Scholar
  6. 6.
    Giardina, P., Aurilia, V., Cannio, R., Marzullo, L., Amoresano, A., Siciliano, R., Pucci, P., & Sannia, G. (1996). European Journal of Biochemistry, 235, 508–515.CrossRefGoogle Scholar
  7. 7.
    Palmieri, G., Giardina, P., Bianco, C., Scaloni, A., Capasso, A., & Sannia, G. (1997). The Journal of Biological Chemistry, 272, 31301–31307.CrossRefGoogle Scholar
  8. 8.
    Palmieri, G., Cennamo, G., Faraco, V., Amoresano, A., Sannia, G., & Giardina, P. (2003). Enzyme and Microbial Technology, 33, 220–230.CrossRefGoogle Scholar
  9. 9.
    Giardina, P., Autore, F., Faraco, V., Festa, G., Palmieri, G., Piscitelli, A., & Sannia, G. (2007). Applied Microbiology and Biotechnology, 75, 1293–1300.CrossRefGoogle Scholar
  10. 10.
    Faraco, V., Ercole, C., Festa, G., Giardina, P., Piscitelli, A., & Sannia, G. (2008). Applied Microbiology and Biotechnology, 77, 1329–1335.CrossRefGoogle Scholar
  11. 11.
    Giardina, P., Palmieri, G., Scaloni, A., Fontanella, B., Faraco, V., Cennamo, G., & Sannia, G. (1999). Biochemical Journal, 341, 655–663.CrossRefGoogle Scholar
  12. 12.
    Giardina, P., Cannio, R., Martirani, L., Marzullo, L., Palmieri, G., & Sannia, G. (1995). Applied and Environmental Microbiology, 61, 2408–2413.Google Scholar
  13. 13.
    Pezzella, C., Autore, F., Giardina, P., Piscitelli, A., Sannia, G., & Faraco, V. (2009). Current Genetics, 55, 45–57.CrossRefGoogle Scholar
  14. 14.
    Palmieri, G., Giardina, P., Bianco, C., Fontanella, B., & Sannia, G. (2000). Applied and Environmental Microbiology, 66, 920–924.CrossRefGoogle Scholar
  15. 15.
    Faraco, V., Giardina, P., & Sannia, G. (2003). Microbiology, 149, 2155–2162.CrossRefGoogle Scholar
  16. 16.
    Thiele, D. J. (1992). Nucleic Acids Research, 20, 1183–1191.CrossRefGoogle Scholar
  17. 17.
    Mager, W. H., & De Kruij, V. A. J. (1995). Microbiology Reviews, 59, 506–531.Google Scholar
  18. 18.
    Rushmore, T. H., & Pickett, C. B. (1991). Methods in Enzymology, 206, 409–420.CrossRefGoogle Scholar
  19. 19.
    Li, G., Li, R., Liu, Q., Wang, Q., Chen, M., & Li, B. (2006). FEMS Microbiology Letters, 256, 203–208.CrossRefGoogle Scholar
  20. 20.
    Ding, Y., Liang, S., Lei, J., Chen, L., Kothe, E., & Ma, A. (2011). Microbiological Research, 166, 314–322.CrossRefGoogle Scholar
  21. 21.
    Raeder, U., & Broda, P. (1988). Methods in Enzymology, 161, 211–220.CrossRefGoogle Scholar
  22. 22.
    Chalfie, M., Tu, Y., Euskirchen, G., Ward, W. W., & Prasher, D. C. (1994). Science, 263, 802–805.CrossRefGoogle Scholar
  23. 23.
    Cubitta, A. B., Heima, R., Adamsa, S. R., Boydd, A. E., Grossa, L. A., & Tsien, R. Y. (1995). Trends in Biochemical Sciences, 20, 448–455.CrossRefGoogle Scholar
  24. 24.
    Lugones, L. G., Scholtmeijer, K., Klootwijk, R., & Wessels, J. G. H. (1999). Molecular Microbiology, 32, 681–689.CrossRefGoogle Scholar
  25. 25.
    Ma, B., Mayfield, M. B., & Gold, M. H. (2001). Applied and Environmental Microbiology, 67, 948–955.CrossRefGoogle Scholar
  26. 26.
    Burns, C., Gregory, K. E., Kirby, M., Cheung, M. K., Riquelme, M., Elliott, T. J., Challen, M. P., Bailey, A., & Foster, G. D. (2005). Fungal Genetics and Biology, 42, 191–199.CrossRefGoogle Scholar
  27. 27.
    Mooibroek, H., Kuipers, A. G. J., Sietsma, J. H., Punt, P. J., & Wessels, J. G. H. (1990). Molecular Genetics and Genomics, 222, 41–48.Google Scholar
  28. 28.
    Schuren, F. H. J., & Wessels, J. G. H. (1998). Current Genetics, 33, 151–156.CrossRefGoogle Scholar
  29. 29.
    Scholtmeijer, K., Wösten, H. A. B., Springer, J., & Wessels, J. G. H. (2001). Applied and Environmental Microbiology, 67, 481–483.CrossRefGoogle Scholar
  30. 30.
    Honda, Y., Matsuyama, T., Irie, T., Watanabe, T., & Kuwahara, M. (2000). Current Genetics, 37, 209–212.CrossRefGoogle Scholar
  31. 31.
    Salame, T. M., Yarden, O., & Hadar, Y. (2010). Microbial Biotechnology, 3, 93–106.CrossRefGoogle Scholar
  32. 32.
    Ma, B., Mayfield, M. B., Godfrey, B. J., & Gold, M. H. (2004). Eukaryotic Cell, 3, 579–588.CrossRefGoogle Scholar
  33. 33.
    Avis, T. J., Anguenot, R., Neveu, B., Bolduc, S., Zhao, Y., Cheng, Y., Labbe, C., Belzile, F., & Bélanger, R. R. (2008). Bioscience, Biotechnology, and Biochemistry, 72, 456–462.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  • Antonella Amore
    • 1
  • Yoichi Honda
    • 2
  • Vincenza Faraco
    • 1
    • 3
    Email author
  1. 1.Department of Chemical Sciences, University of Naples “Federico II”Complesso Universitario Monte S. AngeloNaplesItaly
  2. 2.Research Institute for Sustainable HumanosphereKyoto University GokashoUjiJapan
  3. 3.School of Biotechnological Sciences, University of Naples “Federico II”NaplesItaly

Personalised recommendations