Isolation, characterization, and expression patterns of a DMC1 homolog from the basidiomycete Pleurotus ostreatus.

Here we describe the isolation of a Pleurotus ostreatus gene PoDMC1. The predicted amino acid sequence of the oyster mushroom gene is 62% identical to the yeast DMC1 and 60% identical to human DMC1. The highest degree of amino acid identity (88%), however, was shown with Coprinus CoLIM15, a DMC1 homolog recently found in Coprinus cinereus. The exact matching of sizes and positions of most introns in both basidiomycete genes underlines the close relationship between these DMC1 orthologs. The RecA homolog DMC1 from yeast and its orthologs from other species have been reported to be meiosis specific and essential for sporulation. Here we show that PoDMC1 is exclusively expressed in the lamellae/basidiospore fraction of fruit bodies and not in somatic cells of fruiting bodies or in vegetative mycelium. Furthermore, the gene is not expressed in the lamellae/basidiospore fraction of a nonsporulating mutant of P. ostreatus. Since one of the major problems in cultivating the oyster mushroom is the abundant sporulation that causes allergic reactions in man, PoDMC1 could be an important target gene in constructing sporeless Pleurotus strains.

Transformation of the cultivated mushroom Agaricus bisporus (Lange) using T-DNA from Agrobacterium tumefaciens.

Agrobacterium tumefaciens is known to transfer parts of its tumor-inducing plasmid, the T-DNA, to plants, yeasts and filamentous fungi. We have used this system to transform germinating basidiospores and vegetative mycelium of a commercial strain of the cultivated basidiomycete Agaricus bisporus. Analysis of transformants shows that the T-DNA integrates at random sites into the host genome and that the selection marker is stable during mitosis and meiosis. The Agrobacterium system allows the transformation of both homokaryons and heterokaryons of A. bisporus. Also, both karyotypes of an heterokaryon can be transformed simultaneously. Furthermore, this is the first report on the transformation of vegetative mycelium of a commercial strain of A. bisporus.

Abr1, a transposon-like element in the genome of the cultivated mushroom Agaricus bisporus (Lange) Imbach.

A 300-bp repetitive element was found in the genome of the white button mushroom, Agaricus bisporus, and designated Abr1. It is present in approximately 15 copies per haploid genome in the commercial strain Horst U1. Analysis of seven copies showed 89 to 97% sequence identity. The repeat has features typical of class II transposons (i.e., terminal inverted repeats, subterminal repeats, and a target site duplication of 7 bp). The latter shows a consensus sequence. When used as probe on Southern blots, Abr1 identifies relatively little variation within traditional and present-day commercial strains, indicating that most strains are identical or have a common origin. In contrast to these cultivars, high variation is found among field-collected strains. Furthermore, a remarkable difference in copy numbers of Abr1 was found between A. bisporus isolates with a secondarily homothallic life cycle and those with a heterothallic life cycle. Abr1 is a type II transposon not previously reported in basidiomycetes and appears to be useful for the identification of strains within the species A. bisporus.

Molecular karyotype of the white rot fungus Pleurotus ostreatus.

The white rot fungus Pleurotus ostreatus is an edible basidiomycete with increasing agricultural and biotechnological importance. Genetic manipulation and breeding of this organism are restricted because of the lack of knowledge about its genomic structure. In this study, we analyzed the genomic constitution of P. ostreatus by using pulsed-field gel electrophoresis optimized for the separation of its chromosomes. We have determined that it contains 11 pairs of chromosomes with sizes ranging from 1.4 to 4.7 Mbp. In addition to chromosome separation, the use of single-copy DNA probes allowed us to resolve the ambiguities caused by chromosome comigration. When the two nuclei present in the dikaryon were separated by protoplasting, analysis of their karyotypes revealed length polymorphisms affecting various chromosomes. This is, to our knowledge, the clearest chromosome separation available for this species.