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Fungal Genet Biol. 2005 Nov;42(11):904-13.

High frequency transformation of Cryptococcus neoformans and Cryptococcus gattii by Agrobacterium tumefaciens.

Author information

1
Laboratory of Clinical Infectious Diseases, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD 20892, USA.

Abstract

Cryptococcus neoformans and Cryptococcus gattii are the caus-ative agents of cryptococcal meningoencephalitis and are amenable to genetic manipulations, making them important models of pathogenic fungi. To improve the efficiency of Agrobacterium tumefaciens mediated transformation (ATMT) in C. neoformans, we optimized various co-cultivation conditions including incubation time and temperature, and bacteria to yeast ratio. ATMT was also applied to both serotypes (B and C) of C. gattii. Transformation efficiency by ATMT in C. neoformans was comparable to either electroporation or biolistic transformation and gave superior efficiencies in serotypes B and C, but unlike Saccharomyces cerevisiae, adenine auxotrophy did not increase ATMT efficiency in C. neoformans or C. gattii. All transformants tested were stable, with a majority containing only a single T-DNA insertion; however, homologous recombination was not observed. Additionally, we isolated adenine auxotrophs containing a single T-DNA insertion in the ADE2 gene for representative serotype B and C strains.

PMID:
16260158
DOI:
10.1016/j.fgb.2005.07.003
[Indexed for MEDLINE]

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