Culture techniques are vital in both traditional and modern fungal taxonomy. Establishing sexual-asexual links and synanamorphs, extracting DNA and secondary metabolites are mainly based on cultures. However, it is widely accepted that a large number of species are not sporulating in nature while others cannot be cultured. Recent ecological studies based on culture-independent methods revealed these unculturable taxa, i.e., dark taxa. Recent fungal diversity estimation studies suggested that environmental sequencing plays a vital role in discovering missing species. However, Sanger sequencing is still the main approach in determining DNA sequences in culturable species. In this paper, we summarize culture-based and culture-independent methods in the study of ascomycetous taxa. High-throughput sequencing of leaf endophytes, leaf litter fungi and fungi in aquatic environments is important to determine dark taxa. Nevertheless, currently, naming dark taxa is not recognized by the ICN, thus provisional naming of them is essential as suggested by several studies.
Keywords: consolidated species concept; fungal diversity; molecular taxonomy; morphological characters; polyphasic approach; traditional taxonomy.