Hybridization of Candida albicans through fusion of protoplasts

Abstract

Protoplasts of complementing auxotrophs of Candida albicans can fuse in the presence of polyethylene glycol and generate prototrophic cells. The yields of prototrophs from fusion mixtures depend greatly on the particular combinations of auxotrophies involved but not on other features of the strain backgrounds of protoplasts. The initial cellular products of fusions isolated on selective media are heterokaryons which replicate slowly but also segregate single parental nuclei into blastospores in high frequency. Karyogamy within heterokaryons produces hybrid nuclei which, on segregation, give rise to rapidly growing, uninucleate substrains. Analyses of the substrains show that hybrid nuclei either stabilize as diploid or undergo random loss of chromosomes to stabilize at various levels of aneuploidy prior to segregation. Chromosome losses and radiation induced mitotic crossing-over can effect recombination for parental auxotrophic markers in hybrids; patterns of recombination for ader and arg markers provide the first documented example of chromosomal linkage in C. albicans. Thus, protoplast fusions offer opportunities otherwise unavailable for applying the incisive tools of genetic recombination to analysis of this important, asexual yeast.

Ploidy in the asporogenous yeast Candida maltosa, isolation of its auxotrophic mutants and their cell fusion.

“…The average intensity in (A) was calculated to be 19.8, 19.9 in (B) and 9.7 in (C). Cell fusion between auxotrophic strains As a few systems of protoplast fusion have succeeded in achieving genetic complementation for fungi imperfecti (13)(14)(15), we decided to use it in an attempt to determine whether the mutation has occurred in the same allele or not in Ade-and His mutants isolated as shown in Table 1. With the procedure described in MATERIALS AND METHODS, we obtained hybrids between an Ade-auxotroph and a His auxotroph of C. maltosa at a frequency of 5 x 10-3.…”

DNA content, kinetic complexity, and the ploidy question in Candida albicans.

“…Even the ploidy of C. albicans is in dispute (27,32,35,46). Several recent reports (27,28,45,46) conclude that at least some C. albicans strains are diploid, whereas results from other reports (32,35), are more simply interpreted in terms of haploidy.There have also been confficting reports of the DNA content of C. albicans strains; even values published during the last 2 years (35,39,46) disagree by nearly one order of magnitude. We present direct measurements of the total DNA content and sequence complexity of C. albicans.…”

“…In particular, little has been established concerning the nature and organization of Candida genomes, including those of the most extensively studied species, C. albicans. Even the ploidy of C. albicans is in dispute (27,32,35,46). Several recent reports (27,28,45,46) conclude that at least some C. albicans strains are diploid, whereas results from other reports (32,35), are more simply interpreted in terms of haploidy.…”

A Complementation Analysis by Parasexual Recombination of Candida albicans Morphological Mutants

“…Ten fusion products from each cross were transferred to fresh selective plates, and the plates incubated under the selective conditions required in each case. Under these conditions, colonies of more regular appearance and faster growth were recovered ; these were hybrids resulting from the combination of both parental genomes (Sarachek et al, 1981), that grew under the selective conditions as a result of complementation of the selective markers. On the other hand, the transfer of either fusion products or hybrids to complete (non-selective) medium led to the segregation of parental phenotypes.…”