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Mitochondrial DNA from Euglena gracilis has been investigated in its chemical and physical properties. Its G + C content is equal to 25%; its buoyant density in a CsCl density gradient (1.690 g/cm3) is higher, by 5 mg/cm3, than expected for a bacterial DNA having the same base composition. The buoyant densities of denatured and renatured DNA are higher than that of native DNA by 10-12 mg/cm3 and 6 mg/cm3, respectively. The melting temperature, Tm, is 77 degrees C in standard saline citrate; the first derivative of the melting curve shows a striking multimodality. Degradation of the DNA by micrococcal nuclease indicates that about 40% of the DNA is formed by stretches lower than 10% in G + C. In all its properties the mitochondrial DNA from Euglena gracilis is strikingly similar to that of Saccharomyces cerevisiae.

Labelled chloroplast rRNAs from Spinacia oleracea were hybridized to restriction endonuclease digests of chloroplast DNA from Oenothera hookeri and Euglena gracilis, to mitochondrial DNA of Acanthamoeba castellanii, and to DNA of the E. coli rrn B operon in the transducing phage lambda rifd 18. The degree of homology is greatest for the 16S rRNA gene. Greater than 90% occurs between the two higher plant genes, 80% homology to the lower plant gene, 60%-70% homology to the bacterial gene, and 20% homology to the mitochondrial gene. The degree of hybridization varied considerably for the 23S and the 5S rRNA genes. Very high homology exists between the two higher plant genes, only about 50% homology for both the Euglena and bacterial genes, and no significant homology for the mitochondrial genes. These results show that any chloroplast (or E. coli) rRNA may be used as a probe to identify rRNA genes in other ctDNAs. Two RNA populations, each enriched for a different ctDNA-encoded mRNA, proved useful in the location of these genes on both higher plant ctDNAs. No significant hybridization was obtained using these probes to the Euglena ctDNA which seems to be too distantly related.

1. Chloroplast DNA was isolated from autotrophically and mixotrophically grown Euglena gracilis cells. 2. Aliquots of chloroplast DNA were mechanically degraded to an average molecular weight of 4-7 X 10(6) and G+C-rich DNA fragments (density 1.701 g/cm3) were separated from the bulk DNA (density 1.685 g/cm3) using preparative CsCl density gradients. 3. Total chloroplast DNA and its DNA subfractions, which first were characterized with respect to average G+C content and hybridization capacity for chloroplast rRNA, were hydrolysed with restriction endonucleases (endo R-EcoRI, end R-HindII, endoR-HindIII, endo R-HindII+III, endoR-Hpal, endo R-HpaII and endoR-HaeIII). The fragments were separated on gels under a variety of electrophoretic conditions. 4. With each enzyme tested, a rather large number of bands was obtained. In all cases, different banding patterns were obtained for total DNA, and the DNA subfractions. 5. Chloroplast DNA from autotrophically and mixotrophically grown cells gave identical banding patterns. 6. Digestion of total DNA with the endoR-HaeIII yielded 51-52 fragments separated in the gels in a total of 36 bands of which 11-12 bands were composed of 2-3 fragments as estimated by densitometry. The molecular weights of all fragments combined was 87 X 10(6) or 95% of the genome (92 X 10(6)). 7. Chloroplast RNA hybridized to 5.1% with total chloroplast DNA, equal to three RNA cistrons per genome (Mr92 X 10(6)). These cistrons are located on seven different types of endo R-HaeIII fragments. The hybridising fragments are preferentially found in the G+C-rich subfraction and in bands which are composed of 2-3 fragments.

Mitochondrial DNA from Ustilago cynodontis has been investigated in several of its properties. Its dG + dC content is equal to 33.5%; its buoyant density (1.698 g/cm3) is higher, by 5 mg/cm3, and its melting temperature (82.5 degrees C) is lower than expected for a bacterial DNA having the same base composition; the first derivative of its melting curve indicates a large compositional heterogeneity, its molarity of elution from hydroxyapatite is high, 0.28 M phosphate, and allows its partial separation from nuclear DNA. Degradation by micrococcal nuclease indicates that about 25% of the DNA is formed by stretches having no more than 15% dG + dC. Finally, the unit size of mitochondrial genome is about 50 X 10(6). In most of its properties, the mitochondrial genome of U. cynodontis presents strong analogies with that of Saccharomyces cerevisiae. A parallel investigation on mitochondrial DNA from Acanthamoeba castellanii which has as genome unit size of only 27 X 10(6), has shown that this shares with the former the dG + dC content (32.9%), the melting temperature (82.5 degrees C), a large compositional heterogeneity and a very similar pattern of micrococcal nuclease degradation; its buoyant density (1.692 g/cm3) and its molarity of elution from hydroxyapatite (0.25 M phosphate) are, however, normal, probably because of a different short-sequence pattern and the fact that its dA + dT-rich stretches are shorter, on the average.