Nucleic acid fractionation by counter-migration capillary electrophoresis
First Claim
1. A method of fractionating a mixture of different molecular weight nucleic acid fragments, comprisingloading a liquid sample containing the nucleic acid fragments into one end of a microcapillary tube filled with a fluid electrolyte solution, where the inner wall surface of the tube contains negatively charged groups, one end of the tube is placed in fluid communication with an anodic reservoir containing a polymer solution of an uncharged polymer having a molecular weight of at least about 50,000 daltons, and the other tube end is placed in communication with an cathodic reservoir, andapplying a voltage between the anodic and cathodic reservoirs which is effective to draw said polymer solution into and through the tube by electroosmotic flow, at a fluid flow rate in the tube which is greater, in the direction of the cathodic reservoir, than the molecular-weight dependent rates of migration of such nucleic acid fragments, in the direction of the anodic reservoir, such larger molecular weight nucleic acid fragments move more rapidly toward the cathodic reservoir.
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Abstract
A method of fractionating nucleic acid fragments is disclosed. The method is based on counter migration of the fragments in an upstream direction through a polymer solution which is moving by electroosmotic flow in a downstream direction. Fractionation of selected-size nucleic acid fragments can be enhanced by reducing the difference between the electroosmotic flow rate and the migration rates of the selected-size fragments.
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Citations
19 Claims
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1. A method of fractionating a mixture of different molecular weight nucleic acid fragments, comprising
loading a liquid sample containing the nucleic acid fragments into one end of a microcapillary tube filled with a fluid electrolyte solution, where the inner wall surface of the tube contains negatively charged groups, one end of the tube is placed in fluid communication with an anodic reservoir containing a polymer solution of an uncharged polymer having a molecular weight of at least about 50,000 daltons, and the other tube end is placed in communication with an cathodic reservoir, and applying a voltage between the anodic and cathodic reservoirs which is effective to draw said polymer solution into and through the tube by electroosmotic flow, at a fluid flow rate in the tube which is greater, in the direction of the cathodic reservoir, than the molecular-weight dependent rates of migration of such nucleic acid fragments, in the direction of the anodic reservoir, such larger molecular weight nucleic acid fragments move more rapidly toward the cathodic reservoir.
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16. A method of fractionating a mixture of different molecular weight nucleic acid fragments, comprising
loading a liquid sample containing the nucleic acid fragments into one end of a microcapillary tube filled with a fluid electrolyte solution, where the inner wall surface of the tube contains negatively charged groups, one end of the tube is placed in fluid communication with an anodic reservoir containing a polymer solution of an uncharged polymer having a molecular weight of at least about 50,000 daltons, and the other tube end is placed in communication with an cathodic reservoir, and applying a pulsed voltage between the anodic and cathodic reservoirs which is effective to draw said polymer solution into and through the tube by electroosmotic flow, at a fluid flow rate in the tube which is greater, in the direction of the cathodic reservoir, than the molecular-weight dependent rates of migration of such nucleic acid fragments, in the direction of the anodic reservoir, such larger molecular weight nucleic acid fragments move more rapidly toward the cathodic reservoir, and adjusting the frequency of the pulsed voltage to increase the migration rates of the selected molecular weight fragments with respect to the rates in a constant electrical field of the same field strength.
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18. A method of fractionating a mixture of different molecular weight nucleic acid fragments, comprising
loading a liquid sample containing the nucleic acid fragments into one end of a microcapillary tube filled with a fluid electrolyte solution, where the inner wall surface of the tube contains negatively charged groups, one end of the tube is placed in fluid communication with an anodic reservoir containing a polymer solution containing an intercalating agent and an uncharged polymer having a molecular weight of at least about 50,000 daltons, and the other tube end is placed in communication with an cathodic reservoir, and applying a voltage between the anodic and cathodic reservoirs which is effective to draw said polymer solution into and through the tube by electroosmotic flow, at a fluid flow rate in the tube which is greater, in the direction of the cathodic reservoir, than the molecular-weight dependent rates of migration of such nucleic acid fragments, in the direction of the anodic reservoir, such larger molecular weight nucleic acid fragments move more rapidly toward the cathodic reservoir.
Specification