Genomics and metagenomics technologies to recover ribosomal DNA and single-copy genes from old fruitbody and ectomycorrhiza specimens

Abstract

High-throughput sequencing (HTS) has become a standard technique for genomics, metagenomics and taxonomy, but these analyses typically require large amounts of high-quality DNA that is difficult to obtain from uncultivable organisms including fungi with no living culture or fruit-body representatives. By using 1 ng DNA and low coverage Illumina HiSeq HTS, we evaluated the usefulness of genomics and metagenomics tools to recover fungal barcoding genes from old and problematic specimens of fruit-bodies and ectomycorrhizal (EcM) root tips. Ribosomal DNA and single-copy genes were successfully recovered from both fruit-body and EcM specimens typically <10 years old (maximum, 17 years). Samples with maximum obtained DNA concentration <0.2 ng μl-1 were sequenced poorly. Fungal rDNA molecules assembled from complex mock community and soil revealed a large proportion of chimeras and artefactual consensus sequences of closely related taxa. Genomics and metagenomics tools enable recovery of fungal genomes from very low initial amounts of DNA from fruit-bodies and ectomycorrhizas, but these genomes include a large proportion of prokaryote and other eukaryote DNA. Nonetheless, the recovered scaffolds provide an important source for phylogenetic and phylogenomic analyses and mining of functional genes. Copyright Leho Tedersoo et al. This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.