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Development of transformation system for Trichophyton rubrum by electroporation of germinated conidia
Anita Dobrowolska1, Pawel Staczek
1Department of Genetics of Microorganisms, University of Lodz, Banacha 12/16, 90-237, Lodz, Poland. anidobro@biol.uni.lodz.pl
Abstract:
Dermatophytes are the fungi that can cause infections of skin, hair, and nails due to their ability to utilize keratin. The genetic transformation systems of dermatophytes were successfully applied to Trichophyton mentagrophytes and Microsporum canis. Here we describe the procedure for genetic transformation of Trichophyton rubrum by electroporation of their germinated conidia. A linearized transformation vector (pCHSH75-Pch/GFP/TtrpC) containing bacterial hygromycin B phosphotransferase gene (hph) and green fluorescent protein gene (egfp) was introduced into the germinated conidia of T. rubrum by electroporation. PCR reaction analysis showed that egfp gene was integrated randomly and Southern blotting analysis demonstrated a single integration of hph gene into the chromosomal DNA of randomly selected transformant. In this work we report the efficient transformation and selection of the stable T. rubrum transformants.
Insights
Researchers developed an efficient method to genetically transform Trichophyton rubrum, a common cause of fungal infections. This breakthrough enables further study of dermatophyte fungi and the development of new antifungal treatments.
Area of Science:
- Medical Mycology
- Molecular Biology
- Fungal Genetics
Background:
- Dermatophytes are fungi that cause skin, hair, and nail infections by utilizing keratin.
- Existing genetic transformation systems are established for Trichophyton mentagrophytes and Microsporum canis.
- Trichophyton rubrum transformation is crucial for understanding dermatophyte pathogenesis.
Purpose of the Study:
- To establish an efficient genetic transformation protocol for Trichophyton rubrum.
- To introduce specific genes (hygromycin B phosphotransferase and green fluorescent protein) into T. rubrum.
- To analyze the integration and stability of introduced genes in T. rubrum transformants.
Main Methods:
- Germinated conidia of T. rubrum were subjected to electroporation.
- A linearized transformation vector (pCHSH75-Pch/GFP/TtrpC) carrying hph and egfp genes was used.
- Polymerase Chain Reaction (PCR) and Southern blotting were employed for analysis.
Main Results:
- Successful introduction of the egfp gene into T. rubrum via electroporation.
- Random integration of the egfp gene into the T. rubrum genome was confirmed by PCR.
- Single integration of the hph gene into the chromosomal DNA was demonstrated by Southern blotting.
Conclusions:
- An efficient and stable genetic transformation system for Trichophyton rubrum has been established.
- This method allows for the selection of stable T. rubrum transformants.
- The developed protocol facilitates further research into dermatophyte genetics and antifungal strategies.
