Method of separation using aromatic thioether ligands
View Patent ↗The present invention is a method of separating nucleic acid molecules from contaminants, such as proteins, in a solution and isolating one or more desired nucleic acid molecules, which method comprises the following steps (a) providing an aqueous adsorption solution, which includes nucleic acids and a salt that forms lyotropic ions when dissolved; (b) passing said solution over a matrix to adsorb the nucleic acids onto the matrix, said matrix including an aromatic ring moiety and at least one thioether moiety; (c) passing an aqueous eluent over said matrix to desorb the nucleic acid molecules therefrom, which eluent includes a salt that forms lyotropic ions and a gradient of increasing ionic strength originating from an increasing concentration of a salt that forms less lyotropic ions when dissolved than the ones present in said aqueous adsorption solution; and (d) isolating a fraction comprising the desired nucleic acid molecules.
1. A method of separating nucleic acid molecules from contaminants in a solution and isolating one or more desired nucleic acid molecules, which method comprises the following steps
(a) providing an aqueous adsorption solution, which includes nucleic acids and a salt that forms lyotropic ions when dissolved;
(b) passing said solution over a matrix to adsorb the nucleic acids onto the matrix, said matrix including an aromatic ring moiety and at least one thioether moiety;
(c) passing an aqueous eluent over said matrix to desorb the nucleic acid molecules therefrom, which eluent includes a salt that forms lyotropic ions and a gradient of increasing ionic strength originating from an increasing concentration of a salt that forms less lyotropic ions when dissolved than the ones present in said aqueous adsorption solution; and
(d) isolating a fraction comprising the desired nucleic acid molecules.
2. The method of claim 1 , wherein the fraction isolated is plasmid DNA, which is essentially free from proteineous components and/or RNA and/or endotoxins.
3. The method of claim 1 , wherein the lyotropic and less lyotropic ions of steps (a) and (c) are anions formed of the respective salts.
4. The method of claim 1 , wherein the salt in step (a) is ammonium sulphate or sodium sulphate.
5. The method of claim 4 , wherein the salt in the adsorption solution is at a concentration below about 3 M.
6. The method of claim 1 , wherein the salt dissolved in the eluent is an alkali salt, such as sodium chloride or potassium chloride.
7. The method of claim 6 , wherein the salt in the eluent is at a concentration no greater than about 3 M.
8. The method of claim 1 , wherein the matrix is comprised of a carrier to which an aromatic thioether ligand has been coupled, wherein in said ligand the aryl group is selected from the group consisting of pyridyl, phenyl, benzyl, toluyl, phenethyl, naphtyl, imidazolyl, pyrazolyl, pyrazinyl, pyrimidinyl, pyridazinyl, piperidinyl, morpholinyl, piperazinyl, indolyl, quinolinyl, and purinyl groups.
9. The method of claim 8 , wherein the ligand has been coupled to the carrier via the thioether moiety.
10. The method of claim 1 , further comprising the step of washing said matrix after step (b).