Genetic manipulations of Glomerella graminicola (anamorph Colletotrichum graminicola)
Abstract
Glomerella graminicola is a plant-pathogenic fungus which in its anamorphic stage (Colletotrichum graminicola) causes anthracnose disease on maize and numerous other grasses. The objective of my thesis research was to characterize various aspects of sexual reproduction in G. graminicola, with the intention of developing practical methods for conducting a classical genetic analysis of the fungus. The results of my study included the development of a technique for conducting and analyzing crosses among isolates of Glomerella from maize. A Glomerella stage produced by sorghum isolates of the fungus was discovered, and methods for conducting crosses with it were also developed. For the maize isolates, methods were developed for tetrad analysis and for study of the inheritance of cytoplasmic factors. Maternal inheritance of mitochondria was demonstrated by studying segregation of mitochondrial restriction fragment length polymorphisms. Techniques were developed for genetic linkage mapping of G. graminicola with molecular markers, specifically random amplified polymorphic DNA (RAPDs). Preliminary maps were generated, and it was tentatively estimated that G. graminicola has nine linkage groups. Maize and sorghum isolates of Glomerella, although generally fertile among themselves, were not interfertile. Further comparisons of the morphologies of their anamorphic and teleomorphic stages, mitochondrial DNA restriction fragment length polymorphisms, and RAPD fingerprints led me to conclude that the two groups are sufficiently different to deserve separate species designation. I have proposed the name Glomerella sorghi for the sorghum species. I also established a method for cotransformation and characterized its performance, and I used the technique to generate a pyrimidine auxotrophic mutant by producing a targeted gene disruption at the PYR1 locus by homologous integration of a copy of the PYR1 gene which lacked the 3$\sp\prime$ and 5$\sp\prime$ ends. The ability to perform a sexual genetic analysis, as well as to use cotransformation to make targeted gene disruptions, should expedite future studies of molecular determinants of pathogenicity in this fungus.
Degree
Ph.D.
Advisors
Hanau, Purdue University.
Subject Area
Plant pathology
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