Abstract
Genome-wide mutagenesis is a powerful method for identifying new genes that contribute to a phenotype of interest. For many fungal pathogens of plants and animals, Agrobacterium tumefaciens-mediated transformation (ATMT) serves as an efficient insertional mutagen. In Histoplasma capsulatum, the T-DNA element transferred by Agrobacterium stably integrates into the genome, and the majority of mutants contain single copies of the inserted sequence. The T-DNA sequence facilitates the determination of the genomic sequence flanking the insertion through hemi-specific PCR techniques, plasmid rescue, or inverse PCR. We present optimized procedures for generating insertional mutants in H. capsulatum using Agrobacterium-mediated transformation and using this for forward and reverse genetic approaches.
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Zemska, O., Rappleye, C.A. (2012). Agrobacterium-Mediated Insertional Mutagenesis in Histoplasma capsulatum . In: Brand, A., MacCallum, D. (eds) Host-Fungus Interactions. Methods in Molecular Biology, vol 845. Humana, Totowa, NJ. https://doi.org/10.1007/978-1-61779-539-8_4
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DOI: https://doi.org/10.1007/978-1-61779-539-8_4
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Publisher Name: Humana, Totowa, NJ
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