Genomic DNA extraction from minimal amount of dried mushroom samples

Abstract

Aims: To develop a simple and rapid genomic DNA extraction technique for dried (≈ 1 year old) mushroom fruiting bodies that yields high-quality DNA, suitable for use by post-graduate institutions. Method: Small amounts (0.04 g) of pulverized dried mushroom sample were incubated in a Tris /EDTA/SDS lysis buffer (100mM:10mM: 2%) at 65°C to lyse the chitinous fungal cell walls. Genomic DNA purification was performed using chloroform isoamyl alcohol (24:1), and DNA was precipitated using 100% ethanol. Results: Genomic DNA was successfully extracted under 70 minutes from 16 samples morphologically identified as Panaeolus, Copelandia, Gymnopilus, Pluteus and Favolus species. DNA concentrations were on average of 696.9ng/µL. PCR successfully amplified the ITS-5.8S region. The protocol has been successfully used by numerous post-graduate students in our research programme. Conclusion: The rapid and easy protocol produced high-quality genomic DNA void of any inhibitors that is suitable for downstream molecular implications across multiple mushroom genera. Noticeably, this method requires only minute quantities (0.04g) of starting material and is ideal for student training in higher academic institutions.