The use of diethyl pyrocarbonate and potassium permanganate as probes for strand separation and structural distortions in DNA.
Diethyl pyrocarbonate (DEPC) and potassium permanganate are useful reagents for detecting DNA distortions, especially melted regions. Unlike most other footprinting methods, these reagents can detect such distortions even within the regions of protein-DNA complexes normally protected in other footprinting techniques. Further, reactions are very robust, so that distorted regions can be detected even under conditions where efficiency of DNA-protein complex formation is not high. DEPC reacts with bases that are fully or partially unstacked in DNA, in the preferential order adenosine > guanine >> cytosine. Permanganate reacts strongly with thymine in unstacked regions of DNA, and exhibits only very weak reaction with guanine, cytosine, or adenine. The combination of both reagents gives excellent coverage of all sequence regions of DNA. Because reaction requires unstacking, the two reagents detect both melted regions and regions that are unstacked because of other distortions such as bending. Permanganate has the additional advantage that it can be utilized in living cells.
Department of Biochemistry and Molecular Biology, Colorado State University, 1870 Campus Delivery, Fort Collins, CO 80523, USA.
Brenda F
BF
Kahl
Marvin R
MR
Paule
D001483:Base Sequence; D004247:DNA / Q000737:chemistry* / Q000302:isolation & purification; D017931:DNA Primers / Q000737:chemistry; D015342:DNA Probes / Q000737:chemistry*; D004047:Diethyl Pyrocarbonate / Q000737:chemistry*; D008967:Molecular Biology / Q000379:methods*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011196:Potassium Permanganate / Q000737:chemistry*; D011401:Promoter Regions, Genetic / Q000235:genetics; D014158:Transcription, Genetic
D017931:DNA Primers; D015342:DNA Probes; D011196:Potassium Permanganate; D004247:DNA; D004047:Diethyl Pyrocarbonate