Purification of Fungal High Molecular Weight Genomic DNA from Environmental Samples
Sequencing of a high number of fungal genomes has become possible due to the development of next generation sequencing techniques (NGS). The most recent developments aim to sequence single-molecule long-reads in order to improve genome assemblies, but consequently needs higher quality (minimum >20 kbp) DNA as starting material. However, environmental-derived samples from soil, wood, or litter often contain phenolic compounds, pigments, and other molecules that can be inhibitors for reactions during sequencing library construction. In this chapter, we propose an optimized protocol allowing the preparation of high quality and long fragment DNA from different samples (mycelium, fruiting body, soil) compatible with the current sequencing requirements.
KeywordsCTAB-based DNA extraction Sucrose density gradient ultracentrifugation Fruiting body
Our research was financed by the “Institut National de la Recherche Agronomique” (INRA), the Région Grand Est and the lab of excellence ARBRE (ANR-11-LABX-0002-01). The protocols were developed at the INRA Grand Est-Nancy Ecogenomics facilities. Part of our research was sponsored by the Genomic Science Program of the US Department of Energy, Office of Science, Biological and Environmental Research (under contract DE-AC05-00OR22725) and the US Department of Energy (DOE) Joint Genome Institute (JGI; Office of Science of the US Department of Energy). We would like to thank Francis Martin for helpful discussions and input on this chapter.
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