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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 <jats:italic>Trichoderma harzianum<\/jats:italic> 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 (<jats:italic>hph<\/jats:italic>) gene under the control of the <jats:italic>Aspergillus nidulans trp<\/jats:italic> C. Integration and expression of <jats:italic>hph<\/jats:italic> 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 \u03bcg of vector DNA and 10<jats:sup>6<\/jats:sup> 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 <jats:italic>hph<\/jats:italic> gene in the colonies. The <jats:italic>hph<\/jats:italic> gene was stably integrated as shown by five subsequent passages with and without selection pressure.<\/jats:p>","DOI":"10.1186\/s13568-014-0080-5","type":"journal-article","created":{"date-parts":[[2014,11,3]],"date-time":"2014-11-03T13:43:38Z","timestamp":1415022218000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":31,"title":["Genetic transformation of Knufia petricola A95 - a model organism for biofilm-material interactions"],"prefix":"10.1186","volume":"4","author":[{"given":"Steffi","family":"Noack-Sch\u00f6nmann","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tanja","family":"Bus","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ronald","family":"Banasiak","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nicole","family":"Knabe","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"William J","family":"Broughton","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"H","family":"Den Dulk-Ras","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Paul JJ","family":"Hooykaas","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Anna A","family":"Gorbushina","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2014,11,4]]},"reference":[{"key":"80_CR1","first-page":"156","volume":"50","author":"ZA Avakian","year":"1981","unstructured":"Avakian ZA, Karavaiko GI, Mel\u2019nikova EO, Krutsko VS, Ostroushko II: Role of microscopic fungi in the process of weathering of pegmatite deposit rocks and minerals. 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