Quantification of the arbuscular mycorrhizal fungus Glomus intraradices in host tissue using real-time polymerase chain reaction

A rapid method to quantify the colonization of the arbuscular mycorrhizal fungus (AMF) Glomus intraradices in planta using quantitative real-time polymerase chain reaction (qRT-PCR) technique. Specific PCR primers for the fungus (28S rDNA sequence) and host root tissue (chitinase and chalcone synthase gene) were developed and their respective specificity determined. The plant specific primers for Lycopersicon esculentum, Medicago truncatula amplified linearly over a concentration range of: 6.4 pg to 20 ng. The G. intraradices-specific primer amplified as low as 1 pg of its target DNA, which allowed us to detect a single spore of the fungus. High degrees of correlation were obtained when threshold cycle (Ct) was plotted against vesicular, hyphal and total colonization using microscopically quantified host roots. This is the first report of the application of the qRT-PCR technique for quantification of AMF colonization in planta. The success of its application should open up the possibility of its wider application in AM research.