As a chemist, I am working on hyaluronic acid (HA) hydrogels, which are DNA-modified. DNA is attached to HA via Thiol-Michael. However, due to the polyanionic character of HA, DNA forms aggregates during hydrogelation. I used 15 mM Phosphate Buffer and also tried 15 mM Phosphate Buffer 300 mM NaCl, but both didn't work.
Gernerally, what could I do to rule out this aggregation issues? Are there any additives that could be added?
Thanks for your help!
Hello Thomas, DNA aggregation refers to the attraction of multiple chains to one another to form a variety of condensed mesophases of complex structure. Multivalent counterions are crucially implicated because of their role in screening electrostatic repulsion between charged strands of DNA and mediating an effective attraction. As the multivalent ion concentration is raised above a certain threshold, DNA segments begin to aggregate, and precipitate from the solution. Above the aggregation threshold, the DNA concentration decreases gradually or abruptly, depending on various parameters such as the monovalent salt concentration and total DNA. So, the control lies mostly at manipulation of these two factors: DNA concentration and monovalent salt concentration. You should also watch out for the concentration of multivalent ions in your protocol.
Hi Thomas.
Maurice has a point. Have you looked at whether including EDTA helps?
Steingrimur Stefansson, might be a stupid question but why adding EDTA? If I got it correctly, EDTA coordinates to Mg2+ Ions to prevent DNA degradation by DNAses. I did not add any Mg to my solution and even if there is Mg present it should not affect DNA aggregation.
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