Genetic studies on the edible mushrooms Lentinula edodes and Pleurotus species
Classical and molecular genetic techniques were investigated as approaches to strain improvement in the edible Basidiomycete species Lentinula (syn. Lenitnus) edodes (Berk.) Pegler (the Shiitake mushroom) and Pleurotus spp. (oyster mushrooms). Central to the theme of breeding these species was the application of protoplasts. Attempts were made to develop a high efficiency isolation and regeneration system for both L. edodes and P. sajor-caju. Relatively low yields of protoplasts from L. edodes and their poor regenerative ability limited their use. Howewer, Pleurotus spp. were found to be quite amenable to protoplast isolation, yielding constantly high yields of viable protoplasts. Extensive mating-type analysis of monokaryotic isolates of L. edodus was carried out. Characterization of 17 commercial strains of L. edodes were found to have 9 different A factors and 10 different B factors, whilst four wild-type isolates from China were shown to have 8 different A factors and 8 different B factors. Analysis of several L. edodus strains revealed anomalous ratios of mating-type factors in basidiospore progeny which deviated from the expected 1: 1: 1: 1. Four strains having this imbalance had several A and B factors in common. Protoplasts were used to obtain neohaplonts from dikaryotic mycelia, however, the ratio of the two nuclear types often deviated from the 1:1 ratio of the dikayon. Transformation of P. sajor-caju was attempted by using several vectors containing bacterial dominant selectable genes under the control of fungal promotor and terminator sequences. Also, 'shot-gun' cloning of promotor sequences from P. sajor-caju was attempted by cloning random DNA fragments into a promotorless expression vector. No transformants were obtained by use of any of these vectors. Molecular karyotyping by clamped homogeneous electric field electrophoresis of several Pleurotus spp. revealed differences between species, and strains of P. sajor-caju in terms of chromosome size and number.