Characterization of DNA from the Basidiomycete Schizophyllum commune

• Published in: Experimental mycology Vol. 4; no. 2; pp. 123 - 134
• Main Authors: Ullrich, Robert C., Droms, Kurt A., Doyon, Joel D., Specht, Charles A.
• Format: Journal Article
• Language: English
• Published: Elsevier Inc 1980
• Subjects: characterization of genome; Basidiomycete; Schizophyllum commune; DNA reassociation; homokaryon; dikaryon
• ISSN: 0147-5975; 1878-4399
• DOI: 10.1016/0147-5975(80)90015-8

We have examined the whole-cell DNA of homokaryons and dikaryons of Schizophyllum commune by physical methods and reassociation kinetics. DNA with a single-strand length of 2800 bases was routinely isolated with urea—phosphate buffers and hydroxyapatite. Melting profiles indicate the bulk of the DNA to consist of a 57 ± 1.22% guanine plus cytosine content (GC) with a small amount of DNA melting at temperatures indicative of 22.1% GC. Analytical ultracentrifugations in CsCl gradients revealed a single band with buoyant density 1.718 g/ml equating to 59% GC. On the other hand, cesium chloride isopycnic centrifugation in the preparative ultracentrifuge disclosed a light satellite (ϱ = 1.687 g/ml, 27% GC) constituting about 2% of the whole-cell DNA. Analyses of the kinetics of reassociation of the whole-cell DNA by three different methods estimate the DNA of an S. commune homokaryotic cell to be 8.6 times that of an Escherichia coli cell (mean, 3.6 × 10 7 nucleotide pairs, NTP; range of experimental values 3.1 − 4.1 × 10 7 NTP). The dikaryotic cell would contain approximately twice this amount of DNA. We detect no other differences between the DNA of homokaryotic and dikaryotic cells as described within the limits of these methods. The reassociation data are best described with a single unique component accounting for 80–90% of the DNA and with a single repetitive fraction of 10 ± 2% of whole-cell DNA. Some ambiguity surrounds the repetition frequency and sequence complexity of the repetitive fraction because it does not reassociate well as a separate fraction (i.e., it does not reassociate with the anticipated rate constant, nor does it approach completion of the reaction) after being isolated by denaturation, reassociation, and isolation on hydroxyapatite.