In the Southern blot method, for example, a solution of NaOH is used to denature the DNA in the sample. I find this counterintuitive since I expected that $\text{Na}^+$ cations would neutralize the negative charges of phosphate groups, hence stabilizing the double helix.

  • $\begingroup$ Both high and low pH can affect H-bonding especially in complexes formed by bases such as pyridine, quinoline, imidazole (ref). The bond is most stable when pH = pKa. I am not really sure if this is the case here. Perhaps alkaline denaturation is preferred because it won't cause hydrolysis. This question can be migrated to chemistry SE if no suitable answer is posted. $\endgroup$
    Commented Feb 25, 2015 at 5:31

1 Answer 1


The pKas of (neutral) guanine and thymine are 9-10 (ref). At high pH (>~10), those bases will be deprotonated and exist as negatively charged conjugate bases. As the deprotonated species, part of the G/C and A/T hydrogen bonding networks are eliminated. In the figure below, green dotted lines represent the hydrogen bonds that explain the observed base pairing. Red dotted lines represent disrupted interactions.

enter image description here

  • 1
    $\begingroup$ I can add figures if that would help. $\endgroup$
    – jerepierre
    Commented Feb 25, 2015 at 17:21
  • $\begingroup$ That would be great - it illustrates the answer better. $\endgroup$
    – Chris
    Commented Feb 25, 2015 at 18:29
  • $\begingroup$ @jerepierre the figures you added were great. Thank you very much $\endgroup$
    – El Cid
    Commented Feb 26, 2015 at 1:10

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