I was wondering whether there are any chemical or enzymatic methods to remove a single nucleotide from the end of DNA. Analogous to Edman degradation but instead of a terminal amino acid it removes a single nucleotide.
It could be either 3 or 5 prime end
It is fine if the nucleotide gets chemically modified, the ejected nucleotide is not of any concern.
The use of exonuclease might remove more than one base which can be a problem.
There are chemical and enzymatic methods to remove a single nucleotide from DNA ends. The first one is Exonuclease III at 3’ end or nickase-based approaches (e.g., Nb.BsrDI nicking enzyme + exonuclease) can remove single 3’ nucleotides.
2nd, the T5 exonuclease (limited digestion) at 5’ end or phosphorothioate cleavage (chemical) can remove single 5’ nucleotides. While the chemical means is where the periodate oxidation (3’ end) or acid/base hydrolysis can degrade terminal bases selectively.
Indeed, there are a few approaches you can consider for this type of need:
If you're simply looking to remove a single nucleotide from either the 3' or 5' end—and don't care about preserving the released nucleotide—nucleases are a starting point. For example:
- Exonuclease I: Removes single-stranded DNA from the 3' end, though it may cut more than one base.
- T7 exonuclease or Exonuclease VII: Also active on double-stranded DNA but with less specificity.
To precisely stop at one nucleotide, you might consider combining several strategies:
Chemical approaches are also possible (though less common)—such as selective cleavage after base modification—but these are typically more complex. Hope this helps.
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