Isolation and physicochemical characterization of mitochondrial DNA from cultured cells ofPetunia hybrida.

Mitochondrial DNA ofPetunia hybrida was purified from cell suspension cultures. Up to 50% of the DNA could be isolated as supercoiled DNA molecules by CsCl-ethidium bromide density gradient centrifugation. The DNA purified from DNase-treated mitochondria bands at a single buoyant density of 1.760 gcm(-3) in neutral density gradients and runs on agarose gels as a single band with an apparent molecular weight exceeding 30 megadaltons (Md). Summing of the restriction endonuclease fragment lengths indicates a mitochondrial genome size of at least 190 Md. Electron microscopic analysis reveals the presence of a heterogeneous population of circular DNA molecules, up to 60 Md in size. Small circular DNA molecules, ranging in size from 2-30 Md are present, but unlike in cultured cells of other plant species they do not form discrete size classes and furthermore, they constitute less than 5% of the total DNA content of the mitochondria. The restriction endonuclease patterns of mitochondrial DNA do not qualitatively alter upon prolonged culture periods (up to at least two years).