Nucleic acid analyzing method
To provide a method for analyzing a nucleic acid according to high throughput microcapillary electrophoresis during microchip electrophoresis under non-steady electric field, the method being capable of detecting polymorphism of a large number of genes at a high speed. The present invention is useful for the diagnosis and treatment of diseases such as detection of gene diseases and application to Taylor-made therapy.
1. A method for analyzing a nucleic acid in a microcapillary electrophoresis comprising the steps of:
(a) carrying out electric field inversion during electrophoresis on a microchip, thereby separating each of the nucleic acids having different physicochemical properties, and
(b) detecting the nucleic acid separated by the above step (a), wherein an effective length in electrophoresis is 0.5 to 70 mm.
2. The method for analyzing a nucleic acid according to claim 1 , wherein a forward/backward time weight in the electric field inversion is 1/1 to 10/1.
3. The method for analyzing a nucleic acid according to claim 1 , wherein the electric field inversion is carried out by applying an electric field at a frequency of at least 10 Hz.
4. The method for analyzing a nucleic acid according to claim 1 , wherein the electric field has the strength of |10| to |100000| (absolute value) V/cm.
5. The method for analyzing a nucleic acid according to claim 1 , wherein the microchip is a microchip comprising a sample-injection member, a channel for sample analysis and a reservoir for an electrode.
6. The method for analyzing a nucleic acid according to claim 5 , wherein the microchip is a chip comprising an upper plate and a lower plate, wherein:
(A) the lower plate has thereon two orthogonal channels of 1 to 200 μm in width and 0.5 to 100 μn in depth,
(B) the upper plate has four reservoirs of 0.5 to 4 mm in both diameter and depth, and
(C) the reservoirs as defined in (B) are arranged at positions corresponding to each end of the channels as defined in (A), and wherein an electric field can be applied to the reservoir.
7. The method for analyzing a nucleic acid according to claim 6 , wherein the channel holds a separation medium containing at least one member selected from the group consisting of methyl cellulose, hydroxypropyl methyl cellulose (HPMC), hydroxyethyl cellulose (HEC), hydroxypropyl cellulose (HPC), polyethylene glycol (PEG), polyethylene oxide (PEO), polyacrylamide (PAA), polyvinyl pyrrolidone (PVP), dextran and agarose.
8. The method for analyzing a nucleic acid according to claim 7 , wherein pH of the separation medium is 1 to 12.
9. The method for analyzing a nucleic acid according to claim 7 or 8 , wherein the separation medium contains 1% by weight of methyl cellulose, of which buffer is at least one member selected from the group consisting of Tris-borate buffer, Tris-acetate buffer, TAE (Tris-acetate, EDTA) buffer, TBE (Tris-borate, EDTA) buffer, Tris-hydrochloric acid buffer and phosphate buffer.
10. The method for analyzing a nucleic acid according to claim 1 , wherein a physicochemical property is at least one member selected from the group consisting of nucleic acid conformation polymorphism, molecular weight and higher-order structure.
11. The method for analyzing a nucleic acid according to claim 1 , wherein the nucleic acid in the step (b) is detected by at least one member selected from the group consisting of ultraviolet/visible light absorption detection, fluorescence detection, differential refractive index detection, thermo-optical detection, circular dichroism detection, electrochemical detection and electroconductivity detection.
12. The method for analyzing a nucleic acid according to claim 1 , wherein the effective length in electrophoresis is 0.5 to 35mm.
13. The method for analyzing a nucleic acid according to claim 1 , wherein the nucleic acids having different physicochemical properties are from 20 nucleotides to about 100 nucleotides in length.