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Nucleic Acids Research, 1986, Vol. 14, No. 11 4401-4420
© 1986

Articles

Separation of chromosomal DNA molecules from C.albicans by pulsed field gel electropboresis

R. G. Snell and R. J. Willrins*,*

Department of Physics PO Box 56, Dunedin, New Zealand *Department of Molecular Carcinogenesis Laboratory of the Department of Biochemistry, University of Otago PO Box 56, Dunedin, New Zealand

*To whom correspondence should be addressed

Received April 16, 1986. Accepted May 20, 1986.

Modifications have been made to standard pulse field gel eledrophoresis (PFGE) systems to enable very large DNA molecules to be resolved. The single most important modification was to elevate the temperature of ekctrophoresls to 35°C. This enabled the largest SaccharomTces cerevlslat chromosome to be reprodbcibly resolved. More impressively, it enabled the DNA of Candida albkans to be clearly resolved Into six bands, a feat which was very difficult at lower temperatures. Even so, optimal resolution could only be obtained by carefully adjusting field voltages and switching times. The DNA from the two largest C. albkans chromosomes, which was estimated to be at least 5–10Mbp in size, ran somewhat anomalously, giving fuzzy bands which did not migrate in the direction of the average electric field. That the highestmolecnlar weight band was a distinct chromosome was demonstrated by specific hybridisation to the C. albkans ADE2 gene probe. With further fine tuning, the PFGE system described here should be capable of resolving DNA from the smallest human chromosomes.


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