Isolation and Haploidization of Heterozygous Diploid Strains in a Species of Humicola

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Abstract
SUMMARY Stable auxotrophic mutants were induced in Humicola sp. strain 20-31 by irradiation with far ultraviolet light. Mixed inoculation of different auxotrophs on to minimal medium agar produced prototrophic mycelia in all seven combina- tions tested with a frequency of 0.5 to 2.5 %. These prototrophic mycelia were stable when subcultured by macerated hyphae, single hyphal tips, single aleurio- spores or single phialospores, and contained nuclei of approximately twice the volume of those in strain 20-3 I. They were more sensitive to growth inhibition by p-fluorophenylalanine (FPA) than 20-3 I and its auxotrophic derivatives and, at concentrations of 140 and 160 pg FPA/ml, gave rise to more vigorously growing sectors. The mycelia from these sectors segregated for morphological and auxo- trophic markers differentiating the parent auxotrophs and contained nuclei similar in size to strain 20-31. When reinoculated on to FPA media, these segregants showed the higher tolerance characteristic of strain 20-31 and failed to segregate further. It is concluded that in mixed culture, auxotrophic mutant derivatives of Humicola sp. strain 20-31 readily form somatic heterozygous diploids and that these are induced to haploidize by treatment with FPA. A difference in pigmenta- tion which segregates spontaneously in strain 20-31 is attributed to a single chro- mosomal gene and the six markers studied are assigned to at least four linkage groups. Parasexual genetic analysis may be of value in taxonomic studies of imperfect fungi.

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