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