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Being that it is relatively simple to extract DNA from a variety of sources, what further technique can be used to separate the two complementary strands in solution?

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closed as off-topic by canadianer, David, theforestecologist, Bryan Krause, kmm May 31 '17 at 0:23

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  • $\begingroup$ Heating the solution? I doubt if you know about PCR... $\endgroup$ – another 'Homo sapien' May 29 '17 at 7:36
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There is usually a denaturation step in a typical PCR protocol, see e.g. here, which serves exactly this purpose: the strands are thermally divided (typically at 95 °C for a few seconds). As you can see, this is a very common process, but the strands only remain denatured if kept at high temperatures.

Room-temperature strand separation in regular buffer solutions will be very hard, because annealing (the opposite process, where two complementary strands pair up) is thermodynamically favourable. More permanent separation might be obtained by dissolving DNA in particular solutions, as described here, where DMSO seems particularly effective. This results in separated strands at room temperature, but both strands will clearly still be present. One might think about separating the two with various methods (e.g. HPLC), but I doubt these solutions are compatible with regular HPLC columns.

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  • $\begingroup$ Are there nucleases that could recognize a sequence on the unwanted strand and cut it? That turn the long unwanted strand into 2 shorter strands. You could then separate them from the long desired strand by electrophoresis. $\endgroup$ – user137 May 28 '17 at 10:33
  • $\begingroup$ definitely yes! you can check here for a list of nucleases that cleave single stranded substrates. I think the problem would then be that even the shorter fragments (at least one if not both) would re-anneal... $\endgroup$ – WaTTacK May 28 '17 at 20:35
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I've added this answer because the comment on the accepted answer suggests that they are interested in isolating separated strands of a dsDNA.

Single stranded DNAs can be separated by polyacrylamide gel electrophoresis of denatured dsDNA.

End-labelling of dsDNA followed by strand separation was a key technique when sequencing DNA by the Maxam-Gilbert method (chemical sequencing). The figure below is taken from this paper.

strand separation

For this experiment dsDNA fragments were end-labelled (radioactive label) then the strands were separated by heating to 65 °C in 10M urea. The sample was cooled rapidly and then loaded on to a polyacrylamide gel. (Because the two strands of dsDNA will usually have different base compositions they will be resolved during electrophoresis.) The DNA bands were then visualised by autoradiography. As explained in the paper the absence of a second band for the 165 bp fragment is an artefact of the labelling method (i.e. one strand was unlabelled and the visible band was ssDNA.). As you can see, except for the smallest fragment, individual strands were resolved by this method. Once separated these ssDNAs can be isolated from the gel slices.

However, I don't think this method would be useful for large DNAs because the gel wouldn't resolve them well enough.

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