Whole Genome Amplification as an Innovative Approach for Genomic Studies in Lichenized Fungi

Abstract Molecular sequence data has transformed the field of lichen taxonomy and provided crucial insight into evolutionary relationships. However, DNA obtained from standard extraction methodologies may not yield sufficient high-quality DNA to successfully perform high-throughput sequencing. Furthermore, standard DNA extraction protocols often, require the destructive use of large portions of lichen thalli and apothecia. For crustose lichens, in particular, obtaining enough high-quality DNA for subsequent genomic sequencing becomes challenging considering the small amount of source material. A further problem arises by the presence of non-target microorganisms, often co-symbionts within the lichen thalli, which can decrease the coverage of sequence reads from the mycobiont and photobiont of interest. In this paper, we present Whole Genome Amplification (WGA) as a suitable technique to recover large quantities of high-quality DNA from limited amounts of lichen thallus material or fractions of a single apothecium and minimize the quantity of non-target reads. Two WGA kits were tested: Qiagen Repli-G Single-Cell Kit and Jena Bioscience Direct WGA kit, and results were compared to CTAB-extracted DNA. DNA obtained from the three methods, was used to assemble and annotate a total of 18 mitochondrial genomes from the order Lecanorales. Of these, ten mitogenomes were fully and eight mitogenomes were partially assembled. The resulting mitogenomes showed consistent gene position patterns and phylogenetic placement was largely concordant with data previously published. We demonstrate that this alternative method can be reliably used for obtaining enough high-quality DNA material for genomic research on lichens.