DNA isolation and genome sequence of the 134-year-old holotype specimen of Boletus subvelutipes Peck.

Molecular characterization of type specimens is a powerful tool used in clarifying species identity/circumscription, as well as establishing the taxonomic and phylogenetic status of organisms in question. However, DNA sequencing of aged herbarium collections can be a challenge due to the quantity and quality of DNA still present in the specimens. Herein, we report a custom DNA isolation protocol suitable for processing minute quantities of old specimen tissue and its utilization via high-throughput sequencing technologies to obtain, for the first time, the genome assembly of the 134-year-old holotype of Boletus subvelutipes Peck, a North American fleshy pored mushroom of taxonomic and historical significance. A side-by-side evaluation of our DNA isolation method with that of a commercial "kit" by Qiagen is also presented. By relying on the type material, we have established the genetic identity of B. subvelutipes , as well as providing preliminary phylogenetic evidence for its generic affinities in Neoboletus within Boletaceae. The reference genome of the B. subvelutipes holotype provides a resource for future comparative genomic studies, taxonomic revisions in Boletaceae, and other evolutionary studies of fungi.