30
Jan
Gene Silencing Technologies in the Basidiomycetes Coprinus cinereus and Agaricus bisporus
Post Transcriptional Gene Silencing (PTGS) methods such as RNAi have emerged as exciting new technologies in the repression of gene expression. In this report, we describe the development of gene silencing technologies within Agaricus bisporus and Coprinus cinereus.
Using C. cinereus we describe the utilization of green fluorescent protein (GFP) in the evaluation of three different silencing cassettes, both in terms of ease of construction and in silencing capabilities. All three constructs, untranslatable sense orientated, antisense and hairpin, were transformed into a GFP expressing C. cinereus strain. Lines exhibiting complete silencing, and strains exhibiting partial repression of GFP were recovered from transformations with all three cassettes.
In A. bisporus we chose the endogenous serine protease gene as a target for silencing. Serine protease has been implicated in post-harvest and age-related senescence of sporophores. On harvesting, mushrooms degenerate rapidly to give browned caps and loss of texture in the fruit body, and such problems can dramatically reduce sale ability of the mushrooms. Silenced lines have been generated and these show a range of biological effects depending on the degree of silencing. Suppression of genes involved in these pathways could increase mushroom shelf-life and profitability for mushroom growers, or help to further elucidate the complex biochemical pathways involved in post-harvest degradation.
Gene silencing would appear to be an effective tool for the study of gene function in these fungi, and is of particular use where the dikaryotic or diploid nature of species precludes effective gene disruption.
