Efficient Extraction Method for High Quality Fungal RNA from Complex Lignocellulosic Substrates

Here we describe an efficient and reproducible method for the extraction of fungal RNA from complex lignocellulose containing materials. The fungal cells are snap-frozen and disrupted in chaotropic guanidinium thiocyanate buffer, after which the extracted RNA is isolated by using CsCl gradient ultracentrifugation. By lowering the pH of the extraction buffer, the procedure is also suitable for sample materials rich in humic acids. The method results in high quantity and quality RNA that is separated from endogenous contaminants (e.g., RNases) and substances derived from plant biomass (e.g., colored aromatic compounds). In addition, no further steps such as DNase treatment are needed. The extracted RNA is highly suitable for downstream gene expression analyses such as RNA sequencing.