Construction and Genetic Analysis of Hybrid Strains between Apple and Tomato Pathotypes of Alternaria alternata by Protoplast Fusion
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The genetic controls of host-specific toxin (HST) biosynthesis and the pathogenicity of A. alternata pathogens have been limited by the asexual nature of the life cycle of these fungi. We used a protoplast fusion system for A. alternata to analyze the genetics of HST production and its relation to the specific pathogenicity of these pathogens. Drug-resistant transformants were isolated by genetic transformation, using vectors conferring resistance to hygromycin B and geneticin, for the A. alternata apple pathotype (AM-toxin producer) and A. alternata tomato pathotype (AAL-toxin producer), respectively. Protoplasts of the respective transformants were fused by electrofusion. The majority of resultant stable fusants produced both AM- and AAL-toxins and were pathogenic to susceptible cultivars of both apple and tomato. Pulsed-field gel electrophoresis analysis demonstrated that these fusants (or hybrids) carried small 1.7-and 1.1-Mb chromosomes, characteristic of the parental strains of the apple and tomato pathotypes, respectively. Detection of the AMT gene, involved in AM-toxin biosynthesis, by polymerase chain reaction revealed that all fusants pathogenic to apple maintained this gene. Microfluorimetry analysis using propidium iodide staining suggested that the fusants might be diploid.
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