Rapid step-gradient purification of mitochondrial DNA |
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Authors: | Cornelius Welter Eckart Meese Nikolaus Blin |
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Institution: | (1) Institute of Human Genetics, University of the Saar, D-6650 Homburg/Saar, FRG |
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Abstract: | A convenient modification of the step gradient (CsCl/ethidium bomide) procedure is described. This rapid method allows isolation of covalently closed circular DNA separated from contaminating proteins, RNA and chromosomal DNA in ca. 5 h. Large scale preparations can be performed for circular DNA from eukaryotic organelles (mitochondria). The protocol uses organelle pelleting/NaCl-sarcosyl incubation steps for mitochondria followed by a CsCl step gradient and exhibits yields equal to the conventional procedures. It results in DNA sufficiently pure to be used for restriction endonuclease analysis, subcloning, 5 -end labeling, gel retention assays, and various types of hybridization. |
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Keywords: | Mitochondria extrachromosomal DNA CsCl gradient DNA label restriction |
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