Gene disruption by biolistic transformation in serotype D strains of Cryptococcus neoformans.

Published

Journal Article

Gene disruption by biolistic transformation in serotype D strains of Cryptococcus neoformans. Fungal Genetics and Biology 29, 38-48. Cryptococcus neoformans is an opportunistic fungal pathogen with a defined sexual cycle and well-developed genetic and molecular approaches. Two different transformation systems have been developed, and a number of genes have been disrupted by homologous recombination. However, the frequency of homologous recombination achieved by these approaches has differed dramatically between strains of the A and D serotypes. Transformation by electroporation in serotype D strains results in homologous recombination at frequencies of 1/1000 to 1/100,000, whereas transformation by the biolistic method has resulted in gene disruption at frequencies between 2 and 50% in serotype A strains. We find that gene disruption by homologous recombination can be achieved in the congenic serotype D strain series by biolistic transformation with frequencies of approximately 1 to 4%. By this approach, we have readily disrupted the genes encoding a MAPK homolog (CPK1), the calcineurin A catalytic subunit (CNA1), and a G protein alpha subunit (GPA1). By physical and genetic methods, we show that these mutations result from targeted recombination events without ectopic integrations. Because genetic approaches can be applied in the congenic serotype D strains, our observations represent a significant advance in molecular approaches to understand the physiology and virulence of this important human pathogen.

Full Text

Duke Authors

Cited Authors

  • Davidson, RC; Cruz, MC; Sia, RA; Allen, B; Alspaugh, JA; Heitman, J

Published Date

  • February 2000

Published In

Volume / Issue

  • 29 / 1

Start / End Page

  • 38 - 48

PubMed ID

  • 10779398

Pubmed Central ID

  • 10779398

International Standard Serial Number (ISSN)

  • 1087-1845

Digital Object Identifier (DOI)

  • 10.1006/fgbi.1999.1180

Language

  • eng

Conference Location

  • United States