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Genetic transformation of Knufia petricola A95 - a model organism for biofilm-material interactions
(2014)
We established a protoplast-based system to transfer DNA to Knufia petricola strain A95, a melanised rock-inhabiting microcolonial fungus that is also a component of a model sub-aerial biofilm (SAB) system. To test whether the desiccation resistant, highly melanised cell walls would hinder protoplast formation, we treated a melanin-minus mutant of A95 as well as the type-strain with a variety of cell-degrading enzymes. Of the different enzymes tested, lysing enzymes from Trichoderma harzianum were most effective in producing protoplasts. This mixture was equally effective on the melanin-minus mutant and the type-strain. Protoplasts produced using lysing enzymes were mixed with polyethyleneglycol (PEG) and plasmid pCB1004 which contains the hygromycin B (HmB) phosphotransferase (hph) gene under the control of the Aspergillus nidulans trpC. Integration and expression of hph into the A95 genome conferred hygromycin resistance upon the transformants. Two weeks after plating out on selective agar containing HmB, the protoplasts developed cell-walls and formed colonies. Transformation frequencies were in the range 36 to 87 transformants per 10 µg of vector DNA and 106 protoplasts. Stability of transformation was confirmed by sub-culturing the putative transformants on selective agar containing HmB as well as by PCR-detection of the hph gene in the colonies. The hph gene was stably integrated as shown by five subsequent passages with and without selection pressure.
Life on the rocks: a simple genetically tractable model system to study fungus-material interactions
(2015)
Microcolonial fungi (MCF) are able to survive extreme environmental conditions. This study is focusing on responses to oxidative stress - using the rock fungus Knufia petricola strain A95. Comparative gene expression analyses of a wild type and a PKS mutant are used to identify genes which are especially regulated under oxidative stress conditions to help elucidate mechanisms of cell wall maturation and oxidative stress defence strategies.
Genetic manipulation of protective pigments in a rock-inhabiting model fungus Knufia petricola A95
(2016)
Black ascomycetous microcolonial fungi are persistent inhabitants of rock surfaces, but are especially conspicuous in hostile environments like cold and hot deserts. The unique robustness of MFCs is supported by protective pigments, like melanin and carotenoids. We study the role of these pigments in the stress resistance of the model rock fungus Knufia petricola (Chaetothyriales) strain A95.
Microcolonial fungi (MCF) are the most stress-resistant eukaryotes known to date. Black fungi are an interesting object to study mechanisms of stress resistance and to perform applied research to prevent material colonization and biodeterioration. Pigments, like melanin and carotenoids, have been proven to contribute to the unique robustness of MFCs. We study how these pigments ameliorate oxidative stress responses - one of the most significant environmental challenges encountered by MCF - using the model rock fungus Knufia petricola (Chaetothyriales) strain A95.