MUTATION, SELECTION, AND PROTOPLAST FUSION OF THE ERGOT FUNGUS
Abstract
Strain improvement of industrially important fungi such as the ergot fungus which have no sexual reproduction in culture is possible through mutation and selection as well as somatic recombination via the parasexual cycle. Apparently the limiting factor in the parasexual cycle is the formation of a heterokaryon through hyphal anastomosis; however, this can be manipulated by forming heterokaryons through protoplast fusion. Lytic enzymes from the snail, Helix pomatia were used for the formation of protoplasts of strains of the ergot fungus carrying auxotrophic deficiencies to serve as markers for the isolation of fusion hybrids. Several attempts to improve the alkaloid production of the ergot fungus, strain SD 58 based on mutation and selection techniques were made. N-methyl-N'-nitro-N-nitrosoguanidine for mutation and a color reaction with Salkowski's reagent for selection were used. A twofold increase of alkaloid production and an improved vegetative development of the strains were obtained from already high producing strains. A 60 percent increase of alkaloid production was also obtained from some of the auxotrophic revertant strains. In the case of protoplast fusion an intraspecific fusion of strain SD 58 resulted in a more than fourfold increase of alkaloid in one hybrid. Hybrids had multinucleated cells which was not the normal state for strain SD 58. Sclerotialike cell structures which were found only in high producing hybrids and/or the heterokaryotic state might be responsible for the high alkaloid productivity. Some hybrids lacked the alkaloid induction effect by tryptophan which is a pronounced regulatory mechanism well documented in strain SD 58. Hybrids of an ergotamine producing strain of Claviceps purpurea (ATCC 20102) and strain SD 58, showed an ability to produce both clavine and peptide alkaloids but in lesser amount compared with the parent strains. Alkaloid induction by tryptophan in the hybrids showed a slight effect indicating that the regulatory mechanisms of alkaloid production in the hybrids were influenced by both parents, since tryptophan inhibits alkaloid production in the ergotamine parent strain.
Degree
Ph.D.
Subject Area
Microbiology
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