Method Of Concentrating And Purifying Nucleic Acid And Apparatus Therefor

Abstract

The conventional method of purifying and concentrating nucleic acids, because of dangerous chemicals, requires elaborate chemical equipment to result in restriction of environment available. Further, time-consuming operation is inevitable and high-speed centrifugation, etc. are needed to cause automation to be difficult and to cause high purification degree to be unattainable. Still further, in the purification method using a column/filter, application of dusty samples tends to invite clogging to lead to a drop of purification efficiency, and centrifugation or suction operation is needed to cause automation to be difficult. In this invention, surfactants (3,4) are adsorbed on impurity (2) contained in a sample, so that the impurity (2) conducts behavior different from that of nucleic acid (1) to thereby attain separation of the impurity (2) from the nucleic acid (1). Impurity (2) other than nucleic acid (1) is energized with cationic surfactant (4) and nonionic surfactant (3) and placed in an electric field to thereby effect separation and purification of the nucleic acid (1) for an analyte containing the impurity (2). Thus, the nucleic acid (1) is brought into the state of being concentrated or easily concentrated.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a structural diagram of concentration of nucleic acids by electrophoresis with existence of surfactants.

FIG. 2 is a structural diagram of an electrophoresis tank for first electrophoresis.

FIG. 3 is a structural diagram of an electrophoresis tank for second electrophoresis.

FIG. 4 is a structural diagram of a first electrophoresis tank.

FIG. 5 is a structural diagram of a second electrophoresis tank.

FIG. 6 is a perspective view of a sampling unit.

FIG. 7 is a plan view of the sampling unit.

FIG. 8 is a side view of the sampling unit.

FIG. 9 is a sectional side view of the sampling unit.

FIG. 10 is a perspective view of a connection part.

FIG. 11 is a sectional side view of the connection part.

FIG. 12 is a sectional side view of a filtration part.

FIG. 13 is a view of a separation unit while being assembled.

FIG. 14 is a graph of UV spectrum of collected liquid.

FIG. 15 is a structural diagram of an entire electrophoresis device.

FIG. 16 is a perspective view of an electrophoresis unit.

FIG. 17 is a sectional side view of the electrophoresis unit.

FIG. 18 illustrates diagrams of the electrophoresis unit at the former period of purification.

FIG. 19 illustrates diagrams of the electrophoresis unit at the latter period of purification.

FIG. 20 is a sectional side view of the electrophoresis unit while being assembled.

FIG. 21 is a perspective view of the electrophoresis unit while being assembled.

FIG. 22 illustrates a structure of a first block.

FIG. 23 illustrates a structure of a second block.

FIG. 24 is a front view of a gasket.

FIG. 25 is a graph showing a comparison result of purification of Neisseria gonorrhoeae genome.

FIG. 26 is a view of a gel containing samples showing a result of DNA concentration due to electrophoresis.

DESCRIPTION OF NOTATIONS

1 Nucleic Acid

2 Impurity

3 Nonionic Surfactant

4 Cationic Surfactant

5 Electrophoresis Tank

6 Sample Tank

7 Partition

BEST MODE FOR CARRYING OUT THE INVENTION

An object is to provide a nucleic acid concentration device having high purification rate without using dangerous chemical, and the object is achieved by using surfactant and electrophoresis.

EMBODIMENT 1

An embodiment of the present invention will be described.

First, configuration of an electrophoresis tank used for electrophoresis will be described.

FIG. 4 is a structural diagram of a first electrophoresis tank.

An electrophoresis tank 21 is divided into a cathode side tank 22 and an anode side tank 23 by partitions 24 and 25. Partitions 24 and 25 are disposed at a center portion of electrophoresis tank 21, and a sample unit 26 is fitted in partitions 24 and 25. Sample unit 26 projects at one end thereof into cathode side tank 22, and at the other end thereof into anode side tank 23. Sample unit 26 is blocked with gel on the cathode side thereof, and provided at a side surface thereof with an introduction hole for introducing a sample thereinto. The introduction hole is plugged during electrophoresis.

A cathode is inserted into cathode side tank 22, and an anode is inserted into anode side tank 23, so as to energize electrophoresis tank 21.

Another configuration of an electrophoresis tank will be described.

FIG. 5 is a structural diagram of a second electrophoresis tank.

For the second electrophoresis, an electrophoresis tank 21 is divided into a cathode side tank 22 and an anode side tank 23 by partitions 24 and 25. Partitions 24 and 25 are disposed at a center portion of electrophoresis tank 21, and a separation unit 32 is fitted in partitions 24 and 25. Separation unit 32 projects at one end thereof into cathode sided tank 22, and at the other end thereof into anode side tank 23. A cathode is inserted into cathode side tank 22, and an anode is inserted into anode side tank 23, so as to energize electrophoresis tank 21.

Separation unit 32 consists of mutually joined three parts, i.e., sampling unit 26, a connection part 33 and a filtration part 34. O-rings are interposed between sampling unit 26 and connection part 33, and between connection part 33 and filtration part 34, respectively, so as to ensure their joint, and prevent buffer liquid from leaking.

Sampling unit 26 is blocked at the cathode and anode sides thereof with gels, respectively. Filtration part 34 is provided with an ultrafiltration membrane.

Such electrophoresis tanks are used for concentrating nucleic acid.

First, for the first electrophoresis, the pretreated sample is poured into the introduction hole, and the introduction hole is plugged. Sample unit 26 is disposed in electrophoresis tank 21 so as to be slightly exposed at the upper surface thereof above liquid. Then, direct current is charged with voltage of 100 V so as to conduct electrophoresis for 20 minutes, thereby removing waste ion from the sample. The buffer liquid is prepared in pH 8.0 by 1×TAE solution, 40 mM Tris, 40 mM glacial acetic acid and 1 mM EDTA.

After the waste ion is removed from first electrophoresis tank 21, connection part 33 and filtration part 34 are connected to sample unit 26. O-rings are provided at respective joint portions so as to prevent leak of the liquid.

Connection part 33 is supplied therein with mixture liquid of 100% ethanol and 1×TAE, in which the ratio of 100% ethanol to 1×TAE is 6:4. Sampling unit 26 is supplied therein with TE-1 (10 mM Tris-HCl, 0.1 mM EDTA, pH 8.0). Filtration part 34 is supplied therein with TE-1.

Configurations of sampling unit 26, connection part 33 and filtration part 34 will be described.

First, configuration of sampling unit 26 will be described.

FIG. 6 is a perspective view of a sampling unit; FIG. 7 is a plan view of the sampling unit; FIG. 8 is a side view of the sampling unit; and FIG. 9 is a sectional side view of the sampling unit.

Sampling unit 26 is a container holding gel therein. A centrifugal filter made by Millipore, Microcon (trade mark) YM-3, is treated so that an existing inner ultrafiltration membrane is removed therefrom, and that a new hole having a diameter of 5 mm is bored therein, thereby serving as sampling unit 26. Any member can serve as sampling unit 26 if it has the same effect.

Sampling unit 26 includes a cylindrical body 41 and a base 43. Cylindrical body 41 is connected to base 43 and is bored with an introduction hole 42 for introducing a sample. Base 43 is a stepped circular column with a hole 44 opened at top and bottom surfaces thereof.

Gel 48, having a thickness of several millimeters, is disposed inside cylindrical body 41 so as to block the opening of cylindrical body 41. Therefore, a sample supplied from introduction hole 42 is supplied into the inside of gel 48.

Configuration of connection part 33 will be described.

FIG. 10 is a perspective view of the connection part; and FIG. 11 is a sectional side view of the connection part.

The centrifugal filter made by Millipore is also treated so that an existing inner ultrafiltration membrane is removed therefrom, and that a new hole having a diameter of 5 mm is bored therein, thereby serving as connection part 33.

Connection part 33 includes a cylindrical body 41 and a base 43. Cylindrical body 41 is connected to base 43. Base 43 is a stepped circular column with a hole 44 opened at top and bottom surfaces thereof.

Gel 48, having a thickness of several millimeters, is disposed on a top surface of base 43 in cylindrical body 41 so as to prevent liquid from flowing between connection part 33 and sampling unit 26.

Configuration of filtration part 34 will be described.

FIG. 12 is a sectional side view of the filtration part.

The centrifugal filter made by Millipore is also treated so as to serve as filtration part 34.

Filtration part 33 includes a cylindrical body 41 and a base 43. Cylindrical body 41 is connected to a base 43. The length of cylindrical body 41 is as large as that of each of sampling unit 26 and connection part 33 minus about 5 mm. Base 43 is a stepped circular column with a hole 44 opened at top and bottom surfaces thereof.

An ultrafiltration membrane 49 is disposed on a top surface of base 43 in cylindrical body 41 so as to prevent leak of nucleic acid, thereby ensuring concentration of nucleic acid.

An example of operation for concentrating nucleic acids will be described.

Culture solution of Escherichia coli serves as a sample, which is subjected to the above-mentioned first and second electrophoreses so as to concentrate nucleic acids. The concentration of collected nucleic acids is measured by an absorbance measurement.

The sample is 100 μL culture solution of Escherichia coli DH5α.

100 μL solution of 1% Triton (trade mark) X-100 is added to the sample, and heated at a temperature of 96° C. for 10 minutes.

Afterward, 100 μl solution of 0.2% DPC is added so as to prepare a sample to be subjected to the electrophoreses.

0.5×TAE serves as buffer for the electrophoreses.

1×TAE is used for dissolving agarose.

The 1×TAE solution is prepared in pH 8.0 with 40 mM Tris, 40 mM glacial acetic acid.

Connection part 33 is left at rest so that the opening side of cylindrical body 41 faces upward. 1% agarose gel (SeaKem Gold agarose: TaKaRa) is poured into cylindrical body 41 from the opening side so as to accumulate to a thickness of 3-7 mm, and hardened.

Similar to connection part 33, sampling unit 26 is left at rest so that the opening side of cylindrical body 41 faces upward. 1% agarose gel (SeaKem Gold agarose: TaKaRa) is supplied into cylindrical body 41 from the open side so as to accumulate to a thickness of several millimeters. After the gel becomes hard, sampling unit 26 is vertically reversed so as to pour the gel hardened at the opening side into cylindrical body 41.

HU-6 (made by AR Brown) serves as the electrophoresis tank. MPSU-200 (made by AR Brown) serves as a power source.

The prepared sample to be subjected to electrophoreses is introduced into sampling unit 26 through introduction hole 42, and introduction hole 42 is plugged.

The electrophoresis tank is provided with putty so as to be divided into an anode side chamber and a cathode side chamber. 0.5×TAE is supplied into each of the anode and cathode side chambers.

Sampling unit 26 is fitted in the putty so as to be slightly exposed at an upper surface thereof above the buffer liquid.

Then, direct current is charged with voltage of 100V so as to conduct the first electrophoresis for 20 minutes.

Subsequently, connection part 33 and filtration part 34 are connected to sampling unit 26 after the first electrophoresis, so as to constitute the separation unit to be subjected to the second electrophoresis.

An example of operation for the second electrophoresis will be described.

Mixture solution of 100% ethanol and 1×TAE, in which the rate of 100% ethanol to 1×TAE is 6:4, is supplied in connection part 33.

TE-1 (10 mM Tris-HCL, 0.1 mM EDTA, pH 8.0) solution is supplied infiltration part 34.

Then, connection part 33 and filtration part 34 are connected to sampling unit 26, thereby assembling the separation unit.

FIG. 13 is a view of a separation unit while being assembled.

To assemble the separation unit, sampling unit 26, connection part 33 and filtration part 34 are aligned in the same direction, and O-rings 51 are interposed between connection part 33 and filtration part 34, and between connection part 33 and filtration part 34, respectively, so as to prevent leak of liquid from the separation unit.

The electrophoresis tank is provided with putty so as to be divided into an anode chamber side and a cathode side chamber, and 0.5×TAE is supplied into each of the anode and cathode side chambers.

The assembled separation unit is disposed in the putty so as to place the end of sampling unit 26 side on the cathode side, and place filtration part 34 on the anode side.

Then, direct current is charged with voltage of 200 V so as to conduct the second electrophoresis for 240 minutes.

Subsequently, nucleic acid solution is collected from filtration part 34 and the absorbance thereof is measured, so that the concentration of the collected nucleic acids is calculated based on UV spectrum.

FIG. 14 is a graph of UV spectrum of collected liquid.

The calculated concentration of nucleic acid is 32.3 ng (6.7*106 copy/μL).

The concentration of nucleic acids is calculated so that the absorbance in 260 nm (A260) is multiplied by proper coefficient of the nucleic acid property, and by an optical length of the cell (mm), and divided by 10.

Also, the purity of the collected nucleic acid is calculated based on the UV spectrum.

The calculated purity of nucleic acid is 1.91.

The purity is calculated so that the absorbance in 260 nm (A260) is divided by the absorbance in 280 nm (A280). If the sample is 100% DNA, the calculated value becomes about 1.8. If the sample is 100% RNA, the calculated value becomes 2.0. The value of A280 is a reflex of the amount of protein and phenol contained in the measurement target. If the absorbance ratio is greatly less than 1.5, it should be considered that the sample is contaminated with monomeric substance such as protein.

EMBODIMENT 2

An electrophoresis device according to a second embodiment will now be described.

FIG. 15 is a structural diagram of an entire electrophoresis device.

An electrophoresis device 50 comprises an electrophoresis tank 50b in which an electrophoresis unit 51 is disposed so as to concentrate nucleic acids contained in a sample. Electrophoresis tank 50b includes buffer tanks 55 and 58 and a cooling water tank 57. Buffer tanks 55 and 56 are filled therein with buffer liquid for electrophoresis. Cooling water tank 57 is filled therein with cooling water, such as iced water, for cooling electrophoresis unit 51. An anode 53 is disposed in buffer tank 55, and a cathode 54 is disposed in buffer tank 56. Electrophoresis unit 51 projects at opposite ends thereof into respective buffer tanks 55 and 56. A sample is introduced into electrophoresis unit 51, and anode 53 and cathode 54 are energized therebetween so as to concentrate nucleic acids in the sample.

Partitions 52 separate cooling water tank 57 from buffer tanks 55 and 56. Partitions 52 and electrophoresis tank 50b are made of isolative material. Partitions 52 may be made of putty such as silicone bulking agent or epoxy resin. Electrophoresis unit 51 is cooled at both sides thereof by the cooling water through partitions 52 so as to cancel heat generation during the electrophoresis, thereby ensure purification of a sample at a stable temperature environment.

Electrophoresis unit 51 will now be described.

FIG. 16 is a perspective view of an electrophoresis unit; FIG. 17 is a sectional side view of the electrophoresis unit; FIG. 18 illustrates diagrams of the electrophoresis unit at the former period of purification; and FIG. 19 illustrates diagrams of the electrophoresis unit at the latter period of purification.

Electrophoresis unit 51 is mainly made of isolative material such as acrylic resin. Plural members are fastened together with bolts so as to constitute a single body of electrophoresis unit 51. Electrophoresis unit 51 is provided therein with a columnar space 51b along the longitudinal direction thereof, and provided with two holes disposed perpendicular to the longitudinal direction of space 51b. One hole is an introduction hole 67 for introducing a sample into space 51b, and the other is an extraction hole 66 through which the concentrated sample is collected from space 51b. Introduction hole 67 and extraction hole 66 are opened to space 51b.

A collection tank 71 and a sample tank 72 are disposed inside electrophoresis unit 51, so that introduction hole 67 is connected to collection tank 71, and sampling hole 66 is connected to sample tank 72. A gel wall 64 is disposed between sample tank 72 and collection tank 71. Another gel wall 64 is also disposed on the cathode side of sample tank 72. An ultrafiltration membrane 65 is disposed on the anode side of collection tank 71. Namely, sample tank 72 is formed between two gel walls 64, and collection tank 71 is formed between gel wall 64 and ultrafiltration membrane 65.

A purification process by electrophoresis unit 51 will be described.

To purify a sample, space 51b in electrophoresis unit 51 is full of buffer for electrophoresis. Sample tank 72 and collection tank 71 are also full of buffer. As shown in FIG. 18(a), after the sample is introduced into sample tank 72, electrophoresis unit 51 is energized between opposite ends thereof. Here, the sample contains target nucleic acids, impurity, and nucleic acids which are smaller than the target nucleic acids.

When electrophoresis unit 51 is energized, as shown in FIG. 18(b), nucleic acids 1 and waste electrolyte 2b move toward the anode, and impurity 2 moves toward the cathode. By the energization for a certain period, as shown in FIG. 19(c), nucleic acids 1 and waste electrolytes 2b reach collection tank 71 through gel wall 64. By further energization, as shown in FIG. 19(d), waste electrolytes 2b having small molecular weight pass ultrafiltration membrane 65 so as to be exhausted from collection tank 71. Target nucleic acids 1 remain in collection tank 71.

In this way, in electrophoresis unit 51, target nucleic acids 1 can be easily separated from the sample by use of the electrophoresis and the ultrafiltration membrane.

Individual parts in electrophoresis units 51 will be described.

FIG. 20 is a sectional side view of the electrophoresis unit while being assembled; FIG. 21 is a perspective view of the electrophoresis unit while being assembled; FIG. 22 illustrates a structure of a first block; FIG. 23 illustrates a structure of a second block; and FIG. 24 is a front view of a gasket.

Electrophoresis unit 51 is an assemble comprising a first block 61, second blocks 62, third blocks 63, gaskets 73 and an ultrafiltration membrane 65. Each of first block 61, second blocks 62 and third blocks 63 are bored at the axial center portion thereof with a hole opened at front and rear surfaces thereof so as to form a space 51b of electrophoresis unit 51, and each of blocks 61, 62 and 63 is bored with at four corners with respective holes opened at front and rear surfaces thereof so as to be fastened to another block by bolts.

First block 61 serves as an end portion of electrophoresis unit 51. Second blocks 62 serve as sample unit 72 and collection unit 71. Each of third blocks 63 holds gel wall 64. Each of gaskets 73 is disposed between each neighboring pair of the blocks so as to prevent cooling water from entering the inside of electrophoresis unit 51. Ultrafiltration membrane 65 is sandwiched between gaskets 73 and 73 at one end portion of electrophoresis unit 51.

As shown in FIG. 22, first block 61 is bored at the axial center portion thereof with a hole 61b serving as a part of space 51b of electrophoresis unit 51. Further, first block 61 is bored at four comers thereof with holes 61c into which respective bolts are inserted.

As shown in FIG. 23, second block 62 is bored at the axial center portion thereof with a hole 62b serving as a part of space 51b of electrophoresis unit 51. Second block 62 is also bored at four comers thereof with holes 62c into which respective bolts are inserted. Further, second block 62 is bored with a vertical hole 62d opened to hole 62b. Hole 62d serves as either introduction hole 67 or extraction hole 66 of electrophoresis unit 51.

Third block 63 is equivalent to first block 61 thinned in the fore-and-aft direction. Third block 63 is provided at the axial center portion thereof with a hole for holding gel wall 64 therein.

Gasket 73 is a cruciform sheet when viewed in front, made of silicone coat having a thickness of 0.5 mm in this embodiment, and provided at the center portion thereof with a hole 73b. As shown in FIG. 24, gasket 73 is cut off at four comers thereof so as to allow passing of respective bolts for fastening the blocks. Hole 73b is diametrically as large as each of hole 61b of first block 61 and holes 62b of second blocks 62.

Ultrafiltration membrane 65 is larger than hole 73b, so as to be sandwiched between gaskets 73 and 73.

A test of concentrating, purifying and separating nucleic acids by means of the electrophoresis device according to the invention will be described.

Comparison Test 1

A comparison test 1 is comparison between purification of Neisseria gonorrhoeae genome from cultured urine contaminated with Neisseria gonorrhoeae by using the present electrophoresis device and purification of the same by the method using magnetic silica beads.

The operation using the electrophoresis device of the present embodiment will be described.

Neisseria gonorrhoeae is cultured in chocolate culture medium EX (made by Nissui Pharmaceutical Co., Ltd.) at a temperature of 37° C. for 2 days.

The obtained colony is suspended in physiological saline so as to have an absorbance of OD530=0.18, and then, diluted by physiological saline so as to be a 1/100 solution. In this way, a diluted germ solution is obtained.

The 1.2 μL diluted germ solution is added to 60 μL mixed urine of a healthy person. Further, 60 μL liquefied buffer A, having composition as indicated in Table 1, is added to the urine, so as to provide a mixture solution.

The mixture solution is heated at a temperature of 96° C. for 10 minutes. A 100 μL part of the mixture solution serves as a sample, which is introduced into the present electrophoresis device and subjected to electrophoresis for 60 minutes by a voltage of 150 V.

A 5 μL part of the collected 100 μL sample is subjected to polymerase chain reaction (PCR), and its fluorescence intensity is measured.

The prescription of the PCR treatment is indicated in Table 2.

Here, the size of first block 61 is determined to have a width of 20 mm, a height of 20 mm and a thickness of 5 mm, with hole 61b having a diameter of 5 mm. Second block 62 is determined in size similar to first block 61, and further, hole 62d has a diameter of 2 mm. Third block 63 has the same width and height as those of first block 61, and has a thickness of 2.5 mm. Agarose gels serve as the gel walls. SeaKem Gold agarose (made by TaKaRa) is used as the agarose. 10 mM Tris-HCl (pH 8.0) serves as the solution liquid. YM-100 (made by Millipore) serves as the ultrafiltration membrane. Gasket 73 has a thickness of 0.5 mm.

The operation using magnetic silica beads will be described.

MagExtractor (made by Toyobo) serves as the magnetic silica beads.

The same operation as the above-mentioned operation using the electrophoresis is performed till the mixture liquid is heated at a temperature of 96° C. for 10 minutes. A 100 μL part of the mixture liquid is extracted as a sample, and treated according to the protocol of MagExtractor. A 5 μL part of the collected 100 μL sample is subjected to PCR treatment, and its fluorescence intensity is measured.

With respect to the heating cycles of the PCR treatment, as indicated in Table 3, retention at a temperature of 50° C. for 2 minutes is performed, and then retention at a temperature of 95° C. for 2 minutes is performed, and subsequently, fifty cycles, each of which is combination of retention at a temperature of 95° C. for 10 seconds and retention at a temperature of 56° C. for 60 seconds, are performed.

TABLE 1
Composition of Liquefied Buffer A
2% (weight)  TritonX-100
0.1%         Cetyl Tri-Methyl Ammonium
             Chloride (CTAC)
10 mM        Tris-HCl (pH 8.0)
10 mM        EDTA (pH 8.0)
300 mM       NaCl
16.6 copy/μL Human Genome

TABLE 2
Prescription of PCR
Prescription for One Reaction
	H20	15.15	μL
	10X Gene Taq Buffer	2.5	μL
	10 mM AUGC	0.5	μL
	100 mM MgCl2	0.375	μL
	5 μM F-D-NG-R1-32	1	μL
	100 μM NG-F3405-20	0.125	μL
	100 μM NG 3526-20R	0.125	μL
	2 U/μL UNG	0.1	μL
	5 U/μL Gene Taq NT	0.125	μL
		20	μL + 5 μL Recovered Sample

TABLE 3
Heating Cycles
50° C. 2 min
95° C. 2 min
95° C. 10 Sec
       {close oversize brace} 50 Cycles
56° C. 60 Sec

FIG. 25 is a graph showing a comparison result of purification of Neisseria gonorrhoeae genome. In FIG. 25, the wide line represents the purification result of the present method, and the narrow line represents the purification result of the magnetic silica beads method. The dotted line represents the result of negative control.

Consequently, detection of nucleic acid purified by the present method is earlier than that by the magnet silica beads method. This result means that the efficiency of purification and separation of nucleic acid by the present method is higher than that by the magnetic silica beads method.

Concentration Test

An operation for concentrating DNA by use of the present electrophoresis device will now be described.

In the electrophoresis device of the present embodiment, the capacity of the sample tank is 200 μL, and the capacity of the collection tank is 50 μL.

100 μL λ/HindIII DNA (2.7 ng/μL) is mixed with 100 μL liquefied buffer A so as to prepare a sample. The sample is introduced into the sample tank, and subjected to electrophoresis with a voltage of 100 V for 30 minutes so as to be concentrated.

Subsequently, 2 μL, 4 μL, 6 μL and 8 μL samples are extracted from either of the sample before concentration and the sample after concentration, and subjected to gel electrophoresis. Further, a 10 μL sample is extracted from the collected sample after concentration so as to be subjected to the electrophoresis.

FIG. 26 is a view of a gel containing samples showing a result of DNA concentration due to electrophoresis.

In FIG. 26, the samples before concentration belong to a group A, and the samples after concentration belong to a group B. As shown in FIG. 26, in comparison between the samples having the same amount, it is noticed that a band of the sample after concentration is clearer than a band of the sample before concentration.

In FIG. 26, the band state of the 2 μL collected sample matches the band state of the 6 ,L sample before concentration.

In this way, the electrophoresis device of the present embodiment effects concentration of DNA.

INDUSTRIAL APPLICABILITY

The present invention, whose operability or device structure is simple, is suitable to various applications such as an inspection device for automatically concentrating and inspecting nucleic acids.

Claims

1. A method for concentrating and purifying nucleic acid by use of electrophoresis, characterized in that electric charge on impurity contained together with nucleic acid in a sample is adjusted before the sample is placed into an electric field so as to concentrate and purify the nucleic acid.

2. A method for concentrating and purifying nucleic acid by use of electrophoresis, characterized in that cationic surfactant is added into a sample containing nucleic acid so as to adjust electric charge on impurity contained in the sample, and then the sample is placed in an electric field and subjected to electrophoresis so as to concentrate and purify nucleic acid.

3. A method for concentrating and purifying nucleic acid by use of electrophoresis, characterized in that cationic surfactant and nonionic surfactant are added into a sample containing nucleic acid so as to adjust electric charge on impurity contained in the sample, and then the sample is placed in an electric field and subjected to electrophoresis so as to concentrate and purify nucleic acid.

4. The method for concentrating and purifying nucleic acid according to claim 3, wherein the cationic surfactant is adsorbed on the matter other than nucleic acid so as to adjust electric charge on the matter, and the adsorption degree of the cationic surfactant is adjusted by the amount of the added nonionic surfactant.

5. A device for concentrating and purifying nucleic acid, characterized in that nonionic surfactant and cationic surfactant are added to a sample, and the sample is subjected to electrophoresis so as to concentrate and purify nucleic acid on an anode side.

6. A device for concentrating and purifying nucleic acid by use of electrophoresis, characterized by a container having side surfaces made of isolative material is partitioned into a sample introduction chamber and a nucleic acid collection chamber by an electro-conductive separation member for prevention of expansion, and the container is connected at an end thereof to an electrode through a buffer tank.