Abstract
Filamentous fungi naturally grow on solid surfaces, yet most genetic and biochemical analyses are still performed in liquid cultures. Here, we report a multiplexing platform using high-throughput photometric continuous reading that allows parallel quantification of hyphal growth and reporter gene expression directly on solid medium, thereby mimicking natural environmental conditions. Using this system, we have quantified fungal growth and expression of secondary metabolite GFP-based reporter genes in saprophytic Aspergillus and phytopathogenic Fusarium species in response to different nutrients, stress conditions and epigenetic modifiers. With this method, we provide not only novel insights into the characteristic of fungal growth but also into the metabolic and time-dependent regulation of secondary metabolite gene expression.
Highlights
Filamentous fungi naturally grow on solid surfaces, yet most genetic and biochemical analyses are still performed in liquid cultures
Phenotyping individual fungal clones for growth characteristics such as biomass accumulation is routinely done by the tedious procedure of harvesting mycelium accumulated in submerged cultures and quantification of the dry weight, or ‘quick and dirty’ by measuring the radial extension of colonies on plates containing solid medium
Both methods do not provide an accurate reflection of the fungal growth as liquid media are not the natural substrates, and radial hyphal extension does not necessarily represent biomass accumulation
Summary
Filamentous fungi naturally grow on solid surfaces, yet most genetic and biochemical analyses are still performed in liquid cultures. We report a multiplexing platform using high-throughput photometric continuous reading that allows parallel quantification of hyphal growth and reporter gene expression directly on solid medium, thereby mimicking natural environmental conditions. Using this system, we have quantified fungal growth and expression of secondary metabolite GFPbased reporter genes in saprophytic Aspergillus and phytopathogenic Fusarium species in response to different nutrients, stress conditions and epigenetic modifiers. We present for the first time a scalable platform for multipurpose, 96-well plate format phenotyping of fungi on solid substrates in response to a diverse set of genetic or environmental cues
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