Purification of nucleic acid
View Patent ↗The present invention relates to a simple and efficient method to isolate and purify nucleic acids, preferably genomic DNA, from complex samples compared with available methods, by using a ligand which relies on hydrogen bonding to purify the nucleic acids. Preferably the ligand is bound to magnetic beads/particles. More closely the method comprises adding a sample comprising nucleic acid to a polymer having neutral charge; reversibly binding said nucleic acid to said polymer by hydrogen bonding under pH conditions <5; washing said polymer; and eluting said nucleic acid from said polymer under conditions of pH >5. The method is very suitable for sample preparation of nucleic acids, for example for PCR applications.
1. A method for purification of nucleic acid comprising:
adding a sample comprising a nucleic acid to a polymer having neutral charge;
reversibly binding said nucleic acid to said polymer by hydrogen bonding under pH conditions <5 in the presence of a binding solution that does not contain polyoxyethylene based detergents;
washing said polymer with a washing solution; and
eluting said nucleic acid from said polymer under conditions of pH >5.
2. The method of claim 1 , wherein the polymer is polycarboxylated.
3. The method of claim 1 , wherein the polymer comprises 100-500 monomer units.
4. The method of claim 1 , wherein the polymer is polyacrylic acid, poly lactic acid or carboxymethyldextran.
5. The method of claim 1 , wherein polymers are used as ligands attached to a natural or synthetic solid phase or matrix, such as a bead, particle, membrane, filter, chip, sensor chip, for example a SPR chip, monolith, microfluidic device, pipette tip or any other surface.
6. The method of claim 5 , wherein the solid phase comprises magnetic beads.
7. The method of claim 5 , wherein the solid phase comprises a filter.
8. The method of claim 1 , wherein the binding of nucleic acid occurs at pH 1-5, preferably pH 2-4, and elution occurs at pH 5-14, preferably pH 7-10.
9. The method of claim 1 , wherein the nuclei acid is amplified, such as by PCR, directly after elution.
10. The method of claim 1 , wherein the sample comprises nucleic acids of different sizes and thus different hydrogen bonding strength which are separated from each other by adding a solution or flow of a medium affecting hydrogen bonding.
11. The method of claim 1 , wherein the eluted nucleic acids are larger than 200 bp.
12. The method of claim 1 , wherein the eluted nucleic acids are 10-40 kb and are used for PCR applications.
13. The method of claim 1 , wherein the eluted nucleic acids are directly sequenced after elution (without amplification).
14. The method of claim 1 , wherein the nucleic acid is used for vaccine applications.
15. The method of claim 1 , wherein the binding solution contains organic solvents at a concentration of less than 30%.
16. The method of claim 1 , wherein the washing solution contains organic solvents, or is a pure organic solvent.
17. The method of claim 15 , wherein the washing solution comprises an organic solvent at a concentration >30%.
18. The method of claim 17 , wherein said organic solvent is ethanol.