Genetic Transformation Analysis Pipeline

Putative transformants were analysed as follows: Δpks4.2 transformants were sub-cultivated three times on MM agar medium and the spore colour formation compared to the wild-type strain. In case of questionable phenotypes, strains were further subjected to diagnostic PCR. Integration of Psnc1::gfp::snc1 was analysed using fluorescence microscopy. In brief, colonies cultivated on selective agar MM medium containing 15 mM acetamide for 24 h at 37 °C and fluorescence images were taken using an inverted TCS SP8 (Leica, Germany) as described earlier [30 (link)]. Most colonies with GFP-secretory vesicle signals had correct integration of the donor DNA at the snc1 locus as checked by diagnostic PCR (~ 99%). Consequently, transformants with GFP- secretory vesicle signals, were considered snc1 targeted. For Δpks4.1, Δalp1 and Δptf1 diagnostic PCR was done on the corresponding locus. For primer sequence information, see Additional file 5.