Application of Phi29 DNA polymerase mediated whole genome amplification on single spores of arbuscular mycorrhizal (AM) fungi

Abstract

Genetic analysis of arbuscular mycorrhizal (AM) fungi relies on analysis of single spores. The low DNA content makes it difficult to perform large scale molecular analysis. We present the application of Phi29 DNA polymerase mediated strand displacement amplification (SDA) to genomic DNA extracted from single spores of Glomus and Gigaspora species to address this problem. The genome coverage of the SDA process was evaluated by PCR amplification of the beta-tubulin1 gene and part of the rDNA cluster present in AM fungi. The fidelity of SDA was evaluated further by sequencing the Glomus intraradices ITS1 variants to detect the four ITS1 variants previously identified for this fungus.