Abstract
We report here the simultaneous purification of chloroplast (cpDNA) and mitochondrial DNA (mtDNA) from green tissue and suspension cultures of carrot without organelle isolation. This method is based on isolating total nucleic acids from frozen tissue and separating the nuclear, chloroplast and mitochondrial fractions using sequential isopycnic sedimentation in two gradients of cesium chloride containing bisbenzamide. From 10 g of mature carrot leaves, 10 to 30μg of organelle DNA was consistently recovered from mature carrot leaves, while 30 to 50 μg was recovered from suspension cells. The method can be used to isolate chloroplast and mitochondrial DNAs from single plants without sacrificing the individual.
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Abbreviations
- cpDNA:
-
chloroplast DNA
- EtBr:
-
ethidium bromide
- mtDNA:
-
mitochondrial DNA
- PVPP:
-
polyvinylpolypyrrolidone
- DICA:
-
diethyldithiocarbamic acid
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Thorsteinson, K.E., Wolyn, D.J. Simultaneous purification of chloroplast and mitochondrial DNA without isolation of intact organelles from green carrot tissue and suspension cultures. Plant Mol Biol Rep 12, 26–36 (1994). https://doi.org/10.1007/BF02668661
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DOI: https://doi.org/10.1007/BF02668661