WO2013180494A1

WO2013180494A1 - Multichannel device for distributing fluid, apparatus for extracting nucleic acid comprising same, and method for extracting nucleic acid by using same

Info

Publication number: WO2013180494A1
Application number: PCT/KR2013/004775
Authority: WO
Inventors: 김성우; 김덕중; 김선진; 이세현; 류호선
Original assignee: 나노바이오시스(주)
Priority date: 2012-05-30
Filing date: 2013-05-30
Publication date: 2013-12-05
Also published as: KR20130135111A

Abstract

One embodiment of the present invention relates to a multichannel device for distributing a fluid, an apparatus for extracting nucleic acid comprising same, and a method for extracting nucleic acid by using same. According to the present invention, a miniscule amount of the fluid can be rapidly distributed into equal amounts and injected into at least one very small inlet portion, the miniscule amount of the fluid can be accurately distributed into the at least one inlet portion by a single maneuver by a user, and nucleic acid extraction reaction time can be significantly reduced when carrying out a variety of biological reactions, which use a micromovement chip having the shape of a thin film, thereby rapidly progressing a series of various biological detection or analysis reactions.

Description

Multi-channel liquid dispensing apparatus, nucleic acid extracting apparatus comprising same, and nucleic acid extracting method using same

The present invention relates to a liquid dispensing device for dispensing a biological sample or a reagent and a liquid simultaneously and accurately to a thin microfluidic chip having a liquid inlet, a nucleic acid extracting device including the same, and a nucleic acid extracting method using the same.

Recently, a technique for extracting nucleic acids from biological samples such as cells, bacteria, or viruses to diagnose, treat, or prevent diseases at the genetic level has been widely used in connection with nucleic acid amplification reaction technology. In addition to the diagnosis, treatment, or prevention of diseases, there is a need for a technology for extracting nucleic acids from biological samples in various fields such as development of customized new drugs, forensic medicine, and detection of environmental hormones. As an example of the conventional nucleic acid extraction technology, there is a method of purifying nucleic acid by denatured protein with phenol after solubilizing a sample including cells by treatment with SDS or proteinase K. However, the phenol extraction method is not only time-consuming because many processing steps have to be performed, but also has a problem in that the nucleic acid extraction efficiency is highly dependent on the researcher's experience and experience, and thus the reliability is greatly reduced. Recently, in order to solve this problem, kits using silica or glass fibers that specifically bind to nucleic acids have been used. The silica or glass fiber has a low binding ratio with proteins and cellular metabolites, so that nucleic acids having a relatively high concentration can be obtained. This method has the advantage of being simpler than the phenol method. However, the use of chaotropic reagents or ethanol, which strongly inhibit enzymatic reactions such as polymerase chain reaction (PCR), requires the complete removal of these substances. This is very cumbersome and time consuming. Recently, a method of directly purifying nucleic acid using a filter has been disclosed in International Patent Publication No. 00/21973, which passes a sample through a filter to adsorb the cells to the filter, and then dissolves the cells adsorbed on the filter. After filtration, the nucleic acid adsorbed on the filter is washed and eluted. However, in order to elute the nucleic acid after adsorbing the cell to the filter, there is a problem in that the filter must be selected according to the type of the cell, and the devices used are large and complex, so that the researcher cannot easily use it.

In addition, a device for injecting a very small amount of liquid, such as a sample or a reagent, into the reaction vessel is essential when performing various biological reactions. Conventional reaction vessels are mostly tube-shaped, and moreover, multi-tubes and the like, which are equipped with a plurality of tubes having a very small volume, are also used. Thus, devices commonly used for injecting, mixing, or dispensing liquids, such as trace amounts of samples or reagents, into such reaction vessels are pipettes and tips. However, the pipette and tip are unsatisfactory in controlling the amount of liquid injected into the reaction vessel by the user's hand operation, and are particularly useful when injecting liquid into one or more inlets of a small size of a thin microfluidic chip. It is very cumbersome, requiring a tip with a fine outlet and a pipette designed accordingly. In addition, the pipette is very troublesome because it does not accurately and quickly distribute the same sample or reagent when the reaction is performed using a thin microfluidic chip having a plurality of reaction channels. Therefore, in a biological reaction using a thin-film microfluidic chip, it is possible to accurately and accurately distribute a very small amount of samples or reagents at the same time to one or more small inlets at the same time, and the user convenience is improved to perform the reaction quickly. There is a need for a liquid dispensing device.

One embodiment of the present invention can quickly and accurately distribute a small amount of liquid, such as a biological sample or reagent, to one or more inlets of a thin-film microfluidic chip having one or more reaction channels, and improves user convenience. To provide a multi-channel liquid distribution device, a nucleic acid extraction apparatus comprising the same, and a nucleic acid extraction method using the same.

A first embodiment of the present invention is a thin film substrate; A single liquid inlet disposed in one end region of the substrate; And at least one even liquid outlet disposed in the other distal region of the substrate and in fluid communication with the single liquid inlet, the channel having one end connected to the single liquid inlet side. The other end comprises at least one unit channel region having a channel pattern connected to the liquid outlet side, which is implemented to divide into two branches and distribute the flow rate in half. .

In the first embodiment of the present invention,

The liquid discharge port is implemented as 2 ^N , and the channel includes N unit channel regions formed from the single liquid inlet to the 2 ^N liquid outlet ports, and the N-th i-th unit channel region includes 2 ^i-1 openings. channel 3100 and the 2 ^i-1 from the start of the channel is divided into two branches each comprising a flow rate of 2 ⁱ of branch channel (3200) for dispensing a half, the start of the first unit channel domain channel side An end is connected to the single liquid inlet, and a branch channel end of the Nth unit channel region is connected to the 2 ^N liquid outlets, respectively, wherein N and i may be natural waters.

According to a second embodiment of the present invention, in a microfluidic chip 1 having a thin film shape having at least one even number of reaction channels having inlets and outlets at both ends, the at least one inlet may be provided. A multi-channel liquid dispensing device according to a first embodiment of the present invention for injecting liquid downward into one or more reaction channels, the liquid outlet having a liquid outlet corresponding to the number of one or more inlets; And fluid delivery means for fluidly connecting the at least one inlet of the microfluidic chip and the at least one liquid outlet of the multi-channel liquid dispensing device.

A third embodiment of the present invention is for extracting a nucleic acid from a biological sample, comprising an inlet, a channel region connected to the inlet, and an outlet connected to the channel region, wherein the channel region is introduced through the inlet. A microfluidic chip for extracting nucleic acids having a thin film shape having one or more reaction channels, including a heating unit configured to transfer heat obtained from the outside to the biological sample; A multi-channel liquid dispensing apparatus according to a first embodiment of the present invention, having a liquid outlet corresponding to the number of said at least one inlet; And fluid delivery means for fluidly connecting the one or more inlets of the microfluidic chip and the one or more liquid outlets of the multi-channel liquid dispensing device.

In a third embodiment of the present invention,

A chip outlet region end mounting portion configured to be fixedly mounted to at least one outlet region end of the microfluidic chip, at least one upward liquid inlet corresponding to an upper end of at least one outlet portion of the microfluidic chip, and at least one It may further comprise a liquid storage container having at least one liquid storage chamber in fluid communication with the upward liquid inlet.

The microfluidic chip may include a first filter disposed in a first channel region connected to the inlet, and disposed in a second channel region connected to the heating unit and capable of passing a material having a size corresponding to a nucleic acid. Can be.

The microfluidic chip has a heating unit disposed in a first channel region connected to the inlet, and disposed in a second channel region connected to the heating unit, and having a first filter through which a material having a size corresponding to a nucleic acid can pass therethrough. And a nucleic acid separation unit disposed in a third channel region connected to the first filter and having a nucleic acid binding material capable of specifically binding to the nucleic acid.

The microfluidic chip has a heating unit disposed in a first channel region connected to the inlet, and disposed in a second channel region connected to the heating unit, and having a first filter through which a material having a size corresponding to a nucleic acid can pass therethrough. And a nucleic acid separation unit disposed in a third channel region connected to the first filter and having a nucleic acid binding material capable of specifically binding to the nucleic acid, and disposed in a fourth channel region connected to the nucleic acid separation unit. It may be provided with a second filter capable of passing a substance of a size corresponding to the nucleic acid.

The microfluidic chip may include a nucleic acid separation unit including a heating unit disposed in a channel region connected to the inlet unit and disposed in a channel region connected to the heating unit and provided with a nucleic acid binding material capable of specifically binding to the nucleic acid. Can be.

The microfluidic chip is provided with a nucleic acid separation unit which is disposed in the channel region connected to the inlet, the nucleic acid binding material is disposed in the channel region connected to the heating portion and is provided with a nucleic acid binding material that can specifically bind to the nucleic acid. And a second filter disposed in a channel region connected to the nucleic acid separation unit and capable of passing a material having a size corresponding to the nucleic acid.

A fourth embodiment of the present invention includes the steps of providing a nucleic acid extracting apparatus according to a third embodiment of the present invention; Injecting a biological sample or reagent into said nucleic acid extraction microfluidic chip through said multi-channel liquid dispensing device and fluid delivery means; And extracting nucleic acids from the biological sample by driving the nucleic acid extracting microfluidic chip.

A fifth embodiment of the present invention includes the steps of providing a nucleic acid extracting apparatus according to a third embodiment of the present invention; Injecting a biological sample or reagent into said nucleic acid extraction microfluidic chip through said multi-channel liquid dispensing device and fluid delivery means; Extracting nucleic acids from the biological sample by driving the nucleic acid extracting microfluidic chip; And storing the nucleic acid extraction product in a liquid storage chamber of the liquid storage container.

One embodiment of the present invention relates to a multi-channel liquid dispensing apparatus, a nucleic acid extracting apparatus comprising the same, and a nucleic acid extracting method using the same, and accordingly, in performing various biological reactions using a thin-film microfluidic chip, The same amount of liquid can be rapidly dispensed and injected into one or more small inlets, and only one user can accurately distribute the amount of liquid to the one or more inlets, furthermore nucleic acid extraction reaction time It is expected to significantly shorten the speed of the subsequent various biological detection or analytical reactions.

1 illustrates a multi-channel liquid dispensing apparatus according to one embodiment of the present invention.

2 to 3 show unit channel regions of the channels of the multi-channel liquid dispensing apparatus according to FIG. 1.

4 to 5 schematically show a microfluidic chip according to an embodiment of the present invention.

6 illustrates a multi-channel liquid dispensing injection device according to one embodiment of the present invention.

7 to 10 illustrate in detail the microfluidic chip according to an embodiment of the present invention, and shows a nucleic acid extraction method using the same.

11-13 illustrate a liquid storage container according to one embodiment of the present invention.

FIG. 14 illustrates a flow path of a liquid such as a biological sample or a reagent in a state in which a microfluidic chip and a liquid storage container are combined according to an embodiment of the present invention.

FIG. 15 illustrates a flow path of a liquid, such as a biological sample or reagent, in conjunction with a multi-channel liquid dispensing device, microfluidic chip, and liquid storage container in accordance with one embodiment of the present invention.

16 to 17 show the results of nucleic acid extraction experiments using the nucleic acid extracting apparatus and the nucleic acid extracting apparatus according to an embodiment of the present invention, respectively.

Hereinafter, with reference to the accompanying drawings will be described in detail an embodiment of the present invention. The description set forth below is only for easily understanding the embodiments of the present invention, and is not intended to limit the protection scope of the present invention from such description.

1 shows a multi-channel liquid dispensing apparatus 2 according to one embodiment of the invention.

According to FIG. 1, the multi-channel liquid dispensing apparatus 2 includes a substrate 1000 having a thin film shape; A single liquid inlet 2000 disposed in one end region of the substrate; And at least one even liquid outlet 4000 disposed in the other distal region of the substrate and in fluid communication with the single liquid inlet via channel 3000. At least one unit channel region having one end connected to the single liquid inlet side and the other end divided into two branches and configured to distribute the flow rate at half, having a channel pattern connected to the liquid outlet side.

The substrate 1000 has a microcavity, that is, a channel, configured to receive a very small amount of liquid and to allow the contained liquid to move. The substrate 1000 may be embodied in a thin film shape such as a thin plate to save an amount of a sample or a reagent in a biological or biochemical reaction requiring a very small amount of liquid, and to adjust a minute amount. Since the substrate 1000 serves as a base of the single liquid inlet 2000, the channel 3000, and the plurality of liquid outlets 4000 to be described later, the substrates 1000 support these

modules

2000, 3000, and 4000. The materials may be easily manufactured, for example, plastics, silicon, metals, ceramics, and the like, but are not limited thereto.

The multi-channel liquid dispensing device 2 is for simultaneously distributing the same amount of sample or reagent to a reaction vessel having one or more inlets in only one operation. Thus, the multi-channel liquid dispensing device 2 may comprise one, ie a single liquid inlet 2000 and a plurality of, for example, one or more even liquid outlets 4000 in fluid communication therewith. . More specifically, the single liquid inlet 2000 is disposed in one end region of the substrate 1000, and the one or more even liquid outlets 4000 are disposed in the other end region of the substrate, which are channels The fluid is connected in fluid communication via a 3000 to implement a structure in which liquid can move from the single liquid inlet 2000 to the one or more even liquid outlets 4000. Meanwhile, the single liquid inlet 2000 and one or more even number of liquid outlets 4000 may be disposed upward from one surface of the thin film substrate 1000, or may be disposed on both surfaces of the thin film substrate 1000. It may be arranged upward and downward, respectively. For reference, the bottom figure of FIG. 1 shows that a single liquid inlet 2000 and one or more even liquid outlets 4000 of the multi-channel liquid dispensing device 2 are both disposed on one surface of the thin film-shaped substrate 1000. It illustrates an upward arrangement.

2 to 3 show the unit channel region 3500 of the channel 3000 of the multi-channel liquid dispensing apparatus 2 according to FIG. 1.

According to FIG. 2, the channel 3000 of the multi-channel liquid dispensing device 2 has one end connected to the single liquid inlet 2000 side, and the other end is divided into two branches so that the flow rate is 1/2. One or more unit channel regions 3500, which are implemented to dispense and have a channel pattern connected to the

liquid outlets

4000a and 4000b, respectively. The unit channel region 3500 may be an arbitrarily divided region of the channels 3000 that are continuously connected in fluid communication from the single liquid inlet 2000 to the one or more even liquid outlets 4000. Thus, the unit channel region 3500 may be implemented in one or more in the multi-channel liquid distribution device 2 to form a continuous flow path between the single liquid inlet 2000 and the liquid outlet 4000. . 2 is assumed to be one unit channel region 3500. One end of the one unit channel region 3500 is directly connected to the single liquid inlet 2000, and the other end is directly connected to the one or more even numbers, that is, two

liquid outlets

4000a and 4000b. In this case, when a user injects a predetermined amount F of liquid into the single liquid inlet 2000, the injected liquid moves along a channel connected to one end of the unit channel region 3500, and the other end of the liquid is injected (F). The flow rate is divided by 1/2 along the bifurcated channel at 1 / 2F and moves to the two

liquid outlets

4000a and 4000b. In this case, the general electrical circuit formula can be used to dispense exactly 1/2 of the flow rate dispensed. That is, since the liquid introduced into the single liquid inlet 2000 has the same physical properties, the cross-sectional area of the two divided channels is compared with the cross-sectional area of one channel before the split in consideration of the electric circuit formula and the resistance thereof. This can be achieved through

Furthermore, the multi-channel liquid dispensing apparatus 2 can easily determine the number of liquid dispensing if the number of the one or more even liquid outlets 4000 is determined in advance. Specifically, the liquid discharge port is embodied in 2 ^N , the channel comprises N unit channel region formed from the single liquid inlet to the 2 ^N liquid outlet, wherein the i-th unit channel region below N is 2 ^i- is divided into two streams respectively from ^one start channel and the 2 ^i-1 of the start channel, including, but 2 ⁱ of branch channel for distributing a flow rate of 1/2, the first start of the second unit of the channel region is the channel-side ends It is connected to a single liquid inlet, the branch channel end of the N-th unit channel region is each connected to the 2 ^N liquid outlet, wherein N and i can implement a multi-channel liquid distribution device is natural water. For example, according to FIG. 3, if four liquid outlets 4000 are determined (4000a, 4000b, 4000c, 4000d), the channel 3000 is divided into four liquid outlets 4000a, from the single liquid inlet 2000. And two unit channel regions 3500 and 3500 'up to 4000b, 4000c, and 4000d, and the first unit channel region 3500 of the unit channel region includes one start channel 3100 and the one start channel ( 2

branch channels

3200 and 1 / 2F, each divided into two branches from 3100, for dividing the flow rate in half, wherein the second unit channel region 3500 'of the unit channels includes two starting channels. 4 branch channels 3200 'and 1 / 4F, each divided into two branches from 3100' and the two starting channels 3100 ', distributing the flow rate in half, in this case the first An end of the start channel 3100 side of the unit channel region 3500 is connected to the single liquid inlet 2000 and the second unit channel region 35 00 ') end of branch channel 3200' is connected to the four

liquid outlets

4000a, 4000b, 4000c, 4000d.

4 to 5 schematically show a microfluidic chip 1 according to an embodiment of the invention.

The microfluidic chip 1 has one or more reaction channels 70 which are used for various reactions, for example biological or biochemical reactions, in which such reactions occur. 4 to 6, the microfluidic chip 1 is provided with eight reaction channels 70, but is not limited thereto. The reaction channel 70 has an inlet 10 and an outlet 60 at both ends, and a liquid such as a biological sample or a reagent is introduced into the reaction channel 70 through the inlet 10. And liquid, such as the biological reaction product or waste, is discharged through the outlet 60. The microfluidic chip 1 may be implemented in a thin film shape, such as a thin plate, and may include a space for accommodating a small amount of liquid. The microfluidic chip 1 can be usefully used for biological reactions using very small amounts of liquids, for example biological samples, and reagents for extracting nucleic acids therefrom. The detailed structure and use of the microfluidic chip 1 will be described later.

According to FIG. 6, a multi-channel liquid dispensing injection device according to an embodiment of the present invention has a thin-film microfluidic chip having one or more even reaction channels having inlets and outlets at both ends thereof. The liquid outlet port (4000) according to (1), for injecting liquid downwardly through the one or more inlets 10 into the one or more reaction channels, which matches the number of one or more inlets (10, 8). Said multi-channel liquid dispensing device (2); And fluid delivery means 4 for fluidly connecting the one or more inlets 10 of the microfluidic chip 1 and the one or more liquid outlets 4000 of the multi-channel liquid dispensing device 2. do. In this case, the microfluidic chip 1 may be used for nucleic acid extraction reaction, polymerase chain reaction (PCR), etc., and according to an embodiment of the present invention, the multi-channel liquid dispensing device 2 and the fluid delivery means 4 The liquid delivered to the microfluidic chip 1 through) may be a sample or a reagent necessary for each reaction. The fluid delivery means 4 is related to the arrangement direction of at least one even number of liquid outlets 4000 of the multi-channel liquid dispensing device 2 according to an embodiment of the invention. According to FIG. 6, since the liquid discharge direction of the liquid discharge port 4000 is upward, and the inlet part 10 of the thin-film microfluidic chip 1 is also upward, the multi-channel In order to transfer the liquid from the liquid dispensing device 2 to the microfluidic chip 1, there is a case where a separate fluid delivery means 4 is required for fluidly connecting them. However, although not shown, if the liquid outlet 4000 of the multi-channel liquid dispensing device 2 according to an embodiment of the present invention is implemented downward, the bottom of the one or more even number of liquid outlet 4000 If there is a region in which the inlet region of the microfluidic chip 1 is mounted so that the downward liquid outlet 4000 and the inlet of the microfluidic chip 1 can be closely connected, the separate fluid transfer means as described above ( 4) Of course it may not be required.

7 to 10 illustrate in detail the microfluidic chip according to an embodiment of the present invention, and shows a nucleic acid extraction method using the same. 7 to 10, the microfluidic chip according to an embodiment of the present invention can be used for nucleic acid extraction. 7 to 10, the microfluidic chip is referred to as "microfluidic chip for nucleic acid extraction".

The microfluidic chip for nucleic acid extraction is a structure for extracting nucleic acid, that is, an inlet, an outlet, a channel connecting the inlet and the outlet, a first filter, and a first filter. 2 refers to a microchip that has a standard such as a filter in millimeters or micrometers.

According to FIG. 7A, the nucleic acid extracting microfluidic chip according to an embodiment of the present invention has an inlet 10, a channel region 70 connected to the inlet 10, and an outflow connected to the channel region 70. Including a portion 60, the channel region 70 includes a heating portion 20 implemented to transfer heat obtained from the outside to the biological sample introduced through the inlet portion 10, 7b to Various modules for efficiently extracting nucleic acids from biological samples, such as 7g, may be provided.

In the microfluidic chip for nucleic acid extraction according to an embodiment of the present invention shown in FIG. 7B, a heating unit 20 is disposed in a first channel region connected to the inlet unit 10, and connected to the heating unit 20. The microfluidic chip for nucleic acid extraction according to an embodiment of the present invention shown in FIG. 7C includes a first filter 30 disposed in a second channel region and capable of passing a material having a size corresponding to nucleic acid. The heating unit 20 is disposed in the first channel region connected to the inlet unit 10, and the first channel region is disposed in the second channel region connected to the heating unit 20 to pass a material having a size corresponding to the nucleic acid. A nucleic acid separation unit 40 having a filter 30 and disposed in a third channel region connected to the first filter 30 and having a nucleic acid binding material 45 capable of specifically binding to the nucleic acid. Can be provided, for nucleic acid extraction according to an embodiment of the present invention shown in Figure 7d In the microfluidic chip, a heating unit 20 is disposed in a first channel region connected to the inlet 10, and is disposed in a second channel region connected to the heating unit 20, and passes through a material having a size corresponding to a nucleic acid. And a nucleic acid binding material 45 (bead) disposed in a third channel region connected to the first filter 10 and capable of specifically binding to the nucleic acid. And a second filter 50 disposed in a fourth channel region connected to the nucleic acid separator 40 to pass a material having a size corresponding to the nucleic acid. In the microfluidic chip for nucleic acid extraction according to an embodiment of the present invention illustrated in FIG. 7E, the heating unit 20 is disposed in the first channel region connected to the inlet unit 10, the heating unit 20. Disposed in a second channel region associated with the nucleic acid and capable of passing a substance of a size corresponding to the nucleic acid. A nucleic acid separation having a first filter 30 and a nucleic acid binding material 45 (membrane) disposed in a third channel region connected to the first filter 10 and capable of specifically binding to the nucleic acid. And a second filter 50 disposed in a fourth channel region connected to the nucleic acid separation unit 40 and capable of passing a material having a size corresponding to the nucleic acid. In the microfluidic chip for nucleic acid extraction according to an embodiment of the present invention illustrated in FIG. 7f, a heating unit 20 is disposed in a channel region connected to the inlet unit 10, and a channel region connected to the heating unit 20 is provided. Nucleic acid isolating portion 40 is disposed and is provided with a nucleic acid binding material 45, which can specifically bind to the nucleic acid (40, membrane), a nucleic acid according to an embodiment of the present invention shown in Figure 7g Extraction microfluidic chip has a channel region connected to the inlet 10 The nucleic acid separation unit 40 is disposed in the heating unit 20, the nucleic acid separation unit 40 is disposed in the channel region connected to the heating unit 20, and provided with a nucleic acid binding material 45 that can specifically bind to the nucleic acid. And a second filter 50 disposed in a channel region connected to the nucleic acid separator 40 and capable of passing a material having a size corresponding to the nucleic acid.

The biological sample is a biological material including a nucleic acid such as DNA or RNA, and may be, for example, a liquid sample including animal cells, plant cells, pathogens, fungi, bacteria, viruses, and the like, but is not limited thereto.

The inlet 10 is a portion into which the biological sample or the solution for nucleic acid extraction is introduced into the microfluidic chip, and the outlet 60 is a nucleic acid obtained from the biological sample, a solution for nucleic acid extraction, Other waste (waste) and the like is discharged to the outside the microfluidic chip. In this case, if necessary, the inlet 10 and the outlet 60 may serve as outlets and inlets, respectively. The solution for nucleic acid extraction includes all the solutions required for nucleic acid extraction, and may be, for example, distilled water, a nucleic acid binding buffer, an elution buffer, or the like. On the other hand, the inlet 10 and the outlet 60 is connected in fluid communication via the channel 70, the heating unit 20, the first filter 30, the nucleic acid separation unit to be described in detail below Components 40, the second filter 50, and the like may be connected to the channel 70 to perform each function. The channel 70 may be implemented in various standards, but the width and depth of the channel are preferably implemented in the range of 0.001 to 10 millimeters (mm), but are not limited thereto. Meanwhile, the first, second, third, and fourth channel regions, which will be described below, refer to a sequential arrangement from the inlet portion 10 to the outlet portion 60, and to a specific position in the channel 70. It is not limited.

The heating part 20 is a part in which heat is applied from the outside to a solution (including a biological sample) introduced through the inlet part 10 and is disposed in a first channel region connected to the inlet part 10. For example, when a sample containing cells, bacteria, or viruses is introduced through the inlet 10, when the cells, bacteria, or viruses reach the heating unit 20, about 80 to 100 degrees are instantaneously. Heated to (° C.) to destroy the outer wall of the cell, bacteria or virus and release the cell material to the outside (cell lysis). The heating unit 20 may be supplied with heat in a contact or non-contact manner from the heating module 600 of the nucleic acid extraction apparatus to be described below.

The first filter 30 serves to distinguish between passing materials and non-passing materials by sizes in the fluid flow direction, and may be, for example, a structure having pores of a predetermined size. In one embodiment of the present invention, the first filter 30 is disposed in the second channel region connected to the heating unit 20, it is implemented to pass a material of a size corresponding to the nucleic acid. The first filter 30 collects a material having a size larger than that of the nucleic acid in the dissolution product generated by the heating in the heating unit 20 in the heating unit 20, but the nucleic acid and the material having a corresponding size passes through It is moved to the nucleic acid separation unit 40 to be described below. The first filter 30 may be implemented in various standards, but may include pores having a diameter in the range of 0.1 to 0.4 micrometers (μm), and have a thickness in the range of 0.01 to 10 millimeters (mm). It is desirable to have. More preferably, the first filter 30 has a pore having a diameter of 0.2 micrometer (μm), but preferably has a thickness of 0.01 to 0.5 millimeters (mm).

The nucleic acid separation unit 40 is for selectively separating the nucleic acid from a nucleic acid or a substance having a size corresponding thereto. According to FIG. 7, the nucleic acid separation unit 40 is a space between the first filter 30 and the second filter 50 to be described below, and the nucleic acid binding material 45 capable of specifically binding to the nucleic acid. ) Is provided. The nucleic acid binding material 45 may be any material that can specifically bind to the nucleic acid. The nucleic acid binding material 45 has a nucleic acid binding functional group attached thereto, and may be, for example, silica (SiO 2) beads, biotin, strptavidin attachment beads, or a membrane. The bead or membrane to which the nucleic acid binding functional group is attached may be implemented in various standards, but preferably has a diameter within the range of 0.001 to 20 millimeters (mm). In addition, the nucleic acid separation unit 40 may include a bead or membrane to which the nucleic acid binding functional group is attached in various contents and specifications, but preferably within a range of 1 microgram (μg) to 200 mg (mg). After the nucleic acid is specifically bound to the nucleic acid binding material 45, the foreign matter is removed by washing the inside of the nucleic acid separation part 40, and the nucleic acid binding part 40 of the target nucleic acid-nucleic acid binding material 45 is removed. Only the complex remains. Thereafter, when an elution buffer is provided to the nucleic acid separation unit 40, the target nucleic acid may be separated from the complex.

The second filter 50 serves to distinguish the passing material and the non-passing material by size through the pore in the fluid flow direction. For example, the second filter 50 may be a structure having a pore having a predetermined size. have. In one embodiment of the present invention, the second filter 50 is disposed in the fourth channel region connected to the nucleic acid separation unit 40, it is implemented to pass a material of a size corresponding to the nucleic acid. The second filter 50 collects the nucleic acid binding material 45 in the nucleic acid separation unit 40, but passes the nucleic acid separated from the nucleic acid binding material 45 to the outlet 60. Let's do it. The second filter 50 may be implemented in various standards, but having a pore having a diameter in the range of 0.1 to 100 micrometers (μm), but having a thickness in the range of 0.01 to 0.5 millimeters (mm). desirable. More preferably, the second filter 50 has a pore having a diameter of 0.2 micrometer (μm), but preferably has a thickness of 0.3 millimeter (mm).

8 is a cross-sectional view of the microfluidic chip for nucleic acid extraction according to an embodiment of the present invention.

According to Figure 8, it can be seen a cross-sectional view of the microfluidic chip for nucleic acid extraction according to an embodiment of the present invention. Microfluidic chip for nucleic acid extraction according to an embodiment of the present invention is a silver first plate (100); A second plate (200) disposed on the first plate and having a channel (70) including the first to fourth channel regions; And a third plate 300 disposed on the second plate 200 and having the inlet 10 and the outlet 60 disposed thereon. Nucleic acid extraction microfluidic chip according to an embodiment of the present invention may be implemented in a variety of materials, preferably may be implemented in a plastic material. As such, when the plastic material is used, the heat transfer efficiency may be increased only by adjusting the thickness of the plastic, and the manufacturing process may be simplified, thereby greatly reducing the manufacturing cost. Meanwhile, the first plate 100 and the third plate 300 may include polydimethylsiloxane (PDMS), cyclo olefin copolymer (COC), polymethyl methacrylate (PMMA), Material selected from the group consisting of polycarbonate (PC), polypropylene carbonate (PPC), polyether sulfone (PES), and polyethylene terephthalate (PET), and combinations thereof The second plate 200 includes polymethylmethacrylate (PMMA), polycarbonate (PC), cycloolefin copolymer (COC), polyamide (PA), Polyethylene (PE), polypropylene (PP), polyphenylene ether (PPE), polystyrene (PS), polyoxymethylene (POM), polyether Polyetheretherketone (PEEK), polytetrafluoroethylene (PTFE), polyvinylchloride (PVC), polyvinylidene fluoride (PVDF), polybutylene terephthalate (PBT) It may include a thermoplastic resin or a thermosetting resin material selected from the group consisting of, fluorinated ethylene propylene (FEP), perfluoroalkoxyalkane (PFA), and combinations thereof. In addition, the inlet portion of the third plate is implemented in the range of 0.1 to 5.0 millimeters (mm) in diameter, the outlet portion is implemented in the range of 0.1 to 5.0 millimeters (mm) in diameter, the thickness of the first plate and the third plate Is implemented within the range of 0.01 to 20 millimeters (mm), the thickness of the second plate may be implemented within the range of 30 micrometers (μm) to 10 millimeters (mm). In addition, the microfluidic chip for nucleic acid extraction according to an embodiment of the present invention may be implemented as one or more inlets, outlets, and channels connecting them, if necessary, in this case from one or more biological samples on one chip Nucleic acid can be extracted, and nucleic acid can be extracted quickly and efficiently.

9 is a schematic diagram of a nucleic acid extraction apparatus equipped with a microfluidic chip for nucleic acid extraction according to an embodiment of the present invention.

According to Figure 9, the nucleic acid extracting apparatus according to an embodiment of the present invention is a nucleic acid extracting microfluidic chip (1) already described; A chip mounting module 500 implemented to mount the microfluidic chip 1; A heating module 600 implemented to apply heat to the heating unit 20 of the microfluidic chip 1 mounted on the chip mounting module 500; And a solution for extracting nucleic acids into the microfluidic chip 1 by being connected to the inlet 10 and / or the outlet 60 of the microfluidic chip 1 mounted on the chip mounting module 500. It may include a fluid control module 700 implemented to introduce and / or to discharge the solution present in the microfluidic chip (1) to the outside.

The nucleic acid extraction apparatus is a device implemented to perform all the steps for nucleic acid extraction in the state in which the microfluidic chip 1 according to an embodiment of the present invention, the chip mounting module 500, In addition to the heating module 600 and the fluid control module 700, it may further include various modules required for extracting other nucleic acids. In addition, the nucleic acid extracting apparatus according to an embodiment of the present invention can be implemented so that all steps can be implemented in an automated manner, the nucleic acid amplification reaction can proceed immediately after nucleic acid extraction in conjunction with the polymerase chain reaction (PCR) apparatus have.

The microfluidic chip 1 for nucleic acid extraction is as described above.

The chip mounting module 500 is a portion on which the microfluidic chip 1 is mounted. The chip mounting module 500 may be implemented in various ways corresponding to the shape of the contact surface of the microfluidic chip 1.

The heating module 600 is a module for supplying heat to the heating unit 20 of the microfluidic chip 1 when the microfluidic chip 1 is mounted on the chip mounting module 500. The heating module 600 may be implemented in various ways, but a contact heating block is preferable.

The fluid control module 700 is connected to the inlet part 10 and / or the outlet part 60 of the microfluidic chip 1 mounted on the chip mounting module 500 to be inside the microfluidic chip 1. It is a module implemented to introduce a solution for nucleic acid extraction and / or to discharge the solution existing in the microfluidic chip (1) to the outside. The fluid control module 700 may include various components, for example, a microchannel that is a fluid movement passage, a pneumatic pump providing a driving force for fluid movement, a valve for controlling opening and closing of fluid movement, and a nucleic acid. It may further include a storage chamber containing a variety of solutions required for nucleic acid extraction, such as binding buffer, elution buffer, silica gel (silica gel), distilled water (DW).

Meanwhile, the nucleic acid extracting apparatus according to an embodiment of the present invention is an electronic control module (not shown) for automatically controlling the microfluidic chip 1, the heating module 600, and the fluid control module 700. ) May be further included. The electronic control module can precisely control the respective modules so that the quantitative nucleic acid can be extracted from the microfluidic chip 1 according to a pre-stored program. The prestored program includes, for example, a program relating to a series of steps relating to a nucleic acid extraction method which will be described in detail below.

10 is a flowchart of a nucleic acid extraction method according to an embodiment of the present invention. Specifically, FIGS. 10A to 10D illustrate various nucleic acid extraction methods based on the microfluidic chip 1 for nucleic acid extraction according to an embodiment of the present invention.

According to Figure 10a, a method for extracting a nucleic acid from a biological sample according to an embodiment of the present invention comprises the steps of providing a microfluidic chip for nucleic acid extraction according to Figure 7f (microfluidic chip providing step); Introducing a biological sample selected from the group consisting of cells, bacteria, and viruses through an inlet of the microfluidic chip (biological sample introduction step); Moving the introduced biological sample to a heating part of the microfluidic chip and then heating the heating part of the microfluidic chip to dissolve the biological sample (biological sample dissolution step); Separating the nucleic acid from the soluble material through a nucleic acid binding material (membrane) (nucleic acid separation step); As an optional step, the step of removing foreign matters generated in the nucleic acid separation process (foreign matter removing step); And extracting the nucleic acid through the outlet after moving the nucleic acid to the outlet (nucleic acid extraction step).

According to Figure 10b, a method for extracting a nucleic acid from a biological sample according to an embodiment of the present invention comprises the steps of providing a microfluidic chip for nucleic acid extraction according to Figure 7b or 7c (microfluidic chip providing step); Introducing a biological sample selected from the group consisting of cells, bacteria, and viruses through an inlet of the microfluidic chip (biological sample introduction step); Moving the introduced biological sample to a heating part of the microfluidic chip and then heating the heating part of the microfluidic chip to dissolve the biological sample (biological sample dissolution step); The material obtained from the dissolution step is transferred to the first filter of the microfluidic chip and then passed through the first filter, and removing the material not passed through the first filter (filtration step through the first filter) ); Separating the nucleic acid from the material passing through the first filter (nucleic acid separation step); As an optional step, the step of removing foreign matters generated in the nucleic acid separation process (foreign matter removing step); And extracting the nucleic acid through the outlet after moving the nucleic acid to the outlet (nucleic acid extraction step).

According to Figure 10c, a method for extracting a nucleic acid from a biological sample according to an embodiment of the present invention comprises the steps of providing a microfluidic chip for nucleic acid extraction according to Figure 7g (microfluidic chip providing step); Introducing a biological sample selected from the group consisting of cells, bacteria, and viruses through an inlet of the microfluidic chip (biological sample introduction step); Moving the introduced biological sample to a heating part of the microfluidic chip and then heating the heating part of the microfluidic chip to dissolve the biological sample (biological sample dissolution step); Separating the nucleic acid from the soluble material through a nucleic acid binding material (bead) (nucleic acid separation step); As an optional step, the step of removing foreign matters generated in the nucleic acid separation process (foreign matter removing step); Separating the nucleic acid from the nucleic acid binding material, passing the separated nucleic acid to the second filter and passing it through a second filter (filtration through a second filter); And extracting the nucleic acid through the outlet after moving the nucleic acid to the outlet (nucleic acid extraction step).

According to Figure 10d, a method for extracting a nucleic acid from a biological sample according to an embodiment of the present invention comprises the steps of providing a microfluidic chip for nucleic acid extraction according to Figure 7d or 7e (microfluidic chip providing step); Introducing a biological sample selected from the group consisting of cells, bacteria, and viruses through an inlet of the microfluidic chip (biological sample introduction step); Moving the introduced biological sample to a heating part of the microfluidic chip and then heating the heating part of the microfluidic chip to dissolve the biological sample (biological sample dissolution step); The material obtained from the dissolution step is transferred to the first filter of the microfluidic chip and then passed through the first filter, and removing the material not passed through the first filter (filtration step through the first filter) ); Separating the nucleic acid from the soluble material through a nucleic acid binding material (bead or membrane) (nucleic acid separation step); As an optional step, the step of removing foreign matters generated in the nucleic acid separation process (foreign matter removing step); Separating the nucleic acid from the nucleic acid binding material, passing the separated nucleic acid to the second filter and passing it through a second filter (filtration through a second filter); And extracting the nucleic acid through the outlet after moving the nucleic acid to the outlet (nucleic acid extraction step).

According to FIG. 11, the liquid storage container 5000 is for storing and storing a reaction product after the reaction by the microfluidic chip 1 is completed, and at least one outlet portion of the microfluidic chip 1 ( 60) a chip outlet region end mounting portion 5100 implemented such that the region end is fixedly mounted, and one or more upward liquid suction ports 5200 respectively corresponding to upper ends of the one or more outlet portions 60 of the microfluidic chip 1. And one or more liquid storage chambers 5300 in fluid communication with the one or more upward liquid inlets 5200. 12 to 13 illustrate a process in which the liquid discharged through one or more outlets 60 of the microfluidic chip 1 moves in the liquid storage container 5000. For example, after the nucleic acid extraction reaction in the microfluidic chip 1 is completed, the liquid storage container 5000 is fixedly mounted to the chip outlet region end mounting portion 5100, and contains a solution containing a desired nucleic acid ( When E1, E2 is discharged through the one or more outlets 60, the nucleic acid-containing solution E1, E2 is introduced through one or more upward liquid inlet 5200 of the liquid storage container 5000, The liquid storage container 5000 moves through the channel (F1, F2) to reach the one or more liquid storage chambers 5300 (S1, S2). Thereafter, the liquid storage container 5000 may be separated from the microfluidic chip 1 to separately store and store the nucleic acid-containing solution, and may further utilize the nucleic acid-containing solution in subsequent nucleic acid detection and analysis. Will be. FIG. 14 illustrates a flow path of a liquid such as a biological sample or a reagent in a state in which the microfluidic chip 1 and the liquid storage container 5000 are combined according to an embodiment of the present invention. In this case, the driving force for the continuous movement of the liquid in the microfluidic chip 1 and the liquid storage container 5000 is connected to one or more inlets 10 of the microfluidic chip 1. Multi-channel liquid dispensing device 2 according to one embodiment, or any pump or syringe or the like. 15 is a flow path of a liquid such as a biological sample or a reagent in a state in which the multi-channel liquid dispensing apparatus 2, the microfluidic chip 1, and the liquid storage container 5000 are combined according to an embodiment of the present invention. Shows.

Under the premise of the above-described nucleic acid extracting apparatus, an embodiment of the present invention can provide a convenient, fast and efficient ultra-fast nucleic acid extracting method. For example, the first nucleic acid extracting method may include providing the nucleic acid extracting apparatus described above; Injecting a biological sample or reagent into said nucleic acid extraction microfluidic chip through said multi-channel liquid dispensing device and fluid delivery means; And extracting nucleic acids from the biological sample by driving the nucleic acid extracting microfluidic chip, wherein the second nucleic acid extracting method comprises the steps of providing the nucleic acid extracting apparatus described above; Injecting a biological sample or reagent into said nucleic acid extraction microfluidic chip through said multi-channel liquid dispensing device and fluid delivery means; Extracting nucleic acids from the biological sample by driving the nucleic acid extracting microfluidic chip; And storing the nucleic acid extraction product in a liquid storage chamber of the liquid storage container.

Hereinafter, in Examples 1 to 2, compared to other nucleic acid extracting apparatuses (Qiagen), while extracting the nucleic acid from the biological sample to determine the yield and the running time of the nucleic acid extract, and further through the polymerase chain reaction (PCR) The result reliability of the extract was again confirmed.

실시예 1. 핵산 추출의 산출량 및 진행시간 확인Example 1. Confirmation of yield and run time of nucleic acid extraction

First, DNA is extracted using a general tube included in a third-party product and a nucleic acid extracting microfluidic chip 1 according to an embodiment of the present invention, and then the yield and duration of the tuberculosis strain cells. Confirmed.

Nucleic acid extraction step using a third-party nucleic acid separation device is as follows. Tuberculosis strain cells were prepared, and the tuberculosis strain cells were mixed with 6% NaOH and 4% NaLC in a 1: 1: 1 ratio to prepare a sample solution. The sample solution was then centrifuged to remove supernatant (10 min, 7500 rpm, 4 ° C.). Thereafter, 20 μl Proteinase K was added to the sample solution, and the sample solution was left at 56 ° C. until it became clear. Then, 200 μl AL buffer was added to the sample solution, mixed for 15 seconds, and left at 56 ° C. for 10 minutes. The sample solution was then transferred to a column and centrifuged for 1 minute (8000 rpm). Then, 500 μl AW 1 buffer was added and centrifuged for 1 minute (8000 rpm). Thereafter, 500 µl AW 2 buffer was added and centrifuged for 1 minute (14,000 rpm). Then centrifuged again for 1 minute (14,000 rpm). Thereafter, the column was placed in a new tube, and 200 µl of AE buffer was added, followed by standing for 3 minutes. Thereafter, DNA was eluted after centrifugation for 1 minute. As a result, about 100 μl of the final DNA product was obtained and it took about 30 minutes to obtain the final DNA product.

Subsequently, nucleic acid was extracted from the same tuberculosis strain cells using a multi-channel liquid dispensing apparatus 2, a fluid delivery means 4, and a nucleic acid extracting microfluidic chip 1 according to an embodiment of the present invention. The detailed process is as follows.

Tuberculosis strain cells were prepared, and the tuberculosis strain cells were mixed with 6% NaOH and 4% NaLC in a 1: 1: 1 ratio to prepare a sample solution. Thereafter, the microfluidic chip for nucleic acid extraction (25 x 72 x 2 mm, silica beads (OPS Diagnostics, LLC), filter (Whatman)) according to Figure 7 was introduced into the inlet. After the introduction of 300 μl of silica gel and 1X DNA binding buffer at the inlet of the microfluidic chip according to the embodiment of the present invention, the heating portion of the microfluidic chip according to the embodiment of the present invention is 95 ° C. Heated rapidly. Thereafter, waste in the sample solution was removed through the inlet of the microfluidic chip according to the embodiment of the present invention, and 100 µl of an elution buffer was introduced. In this case, a reagent was introduced into the nucleic acid extraction microfluidic chip using the multi-channel liquid dispensing apparatus 2 and the fluid delivery means 4 according to one embodiment of the present invention. Then, the final product was obtained through the outlet of the microfluidic chip according to one embodiment of the present invention (using a liquid storage container according to one embodiment of the present invention), and as a result, about 100 μl of the final DNA product was obtained. It took about 5 minutes to get the final DNA product.

As a result of the experiment, when the multi-channel liquid distribution device 2, the fluid delivery means 4, and the nucleic acid extraction microfluidic chip 1 according to an embodiment of the present invention, the amount of the nucleic acid extraction product is maintained as it is. On the other hand, unlike conventional nucleic acid extraction methods, the total time required can be significantly reduced.

실시예 2. 타사 제품 및 본 발명의 일 실시예에 따른 핵산 추출 방법에 의해 획득된 각각의 DNA 산물의 중합효소 연쇄반응(PCR) 결과Example 2 Polymerase Chain Reaction (PCR) Results of Each DNA Product Obtained by a Third Party Product and a Nucleic Acid Extraction Method According to an Example of the Present Invention

In order to secure the reliability of the DNA product obtained in Example 1, a polymerase chain reaction (PCR) was performed based on the DNA product.

The polymerase chain reaction (PCR) was used as a third-party product (Bio-Rad: CFX connect equipment). PCR samples and reagents for carrying out the polymerase chain reaction (PCR) include 10 microliters (μl) of real-time PCR mixed solution (TOYOBO SYBR qPCR mix), 2 microliters (μl) forward primer, and reverse primer (Reverse). A total of 20 microliters (μl) were prepared, including 2 microliters (μl) of Primer, 10 μM, 1 microliter (μl) of template DNA (1 ng), 5 microliters (μW) of distilled water (DW), and the like. . Then, the pre-denaturation step was performed at 95 ° C. and 30 sec (1 cycle), followed by the denaturation step at 95 ° C. and 5 sec, and the anealing & extension step at 72-65 ° C. and 30 sec (40). cycle).

Table 1 shows real-time PCR results (Ct values) for nucleic acid extraction products, and FIG. 16 is a graph measuring real-time PCR results for nucleic acid extraction products obtained using a nucleic acid extraction method by fluorescence for each PCR cycle. 17 is a photograph of gel electrophoresis of the final PCR product. The graph curve of FIG. 16 is the PCR result curve (X-axis: period, Y-axis: fluorescence) of the DNA product by each nucleic acid extraction method.

Table 1

Classification (gDNA copies / rxn)	Ct value
Non template (1)	-
Plasmid DNA (1 × 10 ⁵ copies) (2)	18.32
1 × 10 ⁶ (3)	16.77
1 × 10 ³ (4)	26.00

(1) is a negative control and (2) is a positive control.

Through the PCR results, the nucleic acid extraction method using the nucleic acid extraction apparatus according to an embodiment of the present invention significantly reduces the time required for the reaction by significantly reducing the nucleic acid extraction step while maintaining or improving the result reliability of the nucleic acid extraction product. I could confirm that I could.

Claims

A thin film substrate;

A single liquid inlet disposed in one end region of the substrate; And

At least one even liquid outlet disposed in the other distal region of the substrate and in fluid communication with the single liquid inlet;

As containing,

The channel comprises one or more unit channel regions, one end of which is connected to the single liquid inlet side and the other end of which is divided into two and has a channel pattern connected to the liquid outlet side to implement a half flow rate. That is,

Multi-channel liquid dispensing device.
The method of claim 1,

The liquid discharge port is implemented as 2 N , and the channel includes N unit channel regions formed from the single liquid inlet to the 2 N liquid outlet ports, and the N-th i-th unit channel region includes 2 i-1 openings. channel and the 2 i-1 from the start of the channel is divided into two branches each comprising a 2 i of branch channels for distributing the flow rate of 1/2, the first unit of the start of the channel section a channel-side terminal is the single liquid inlet And the branch channel ends of the Nth unit channel region are respectively connected to the 2 N liquid outlets, wherein N and i are natural water.
In the thin-film microfluidic chip having one or more even reaction channels having inlets and outlets at both ends, the liquid is injected downward into the one or more reaction channels through the one or more inlets. As,

A multi-channel liquid dispensing device according to claim 1 or 2, having a liquid outlet corresponding to the at least one inlet number; And

Fluid delivery means for fluidly connecting at least one inlet of the microfluidic chip and at least one liquid outlet of the multi-channel liquid dispensing device;

A multi-channel liquid dispensing injection device comprising a.
A nucleic acid for extracting a nucleic acid from a biological sample, comprising an inlet, a channel region connected to the inlet, and an outlet connected to the channel region, wherein the channel region is heat obtained from the outside of the biological sample introduced through the inlet. A microfluidic chip for extracting nucleic acids having a thin film-like shape having one or more reaction channels, including a heating unit implemented to deliver the;

A multi-channel liquid dispensing device according to claim 1 or 2, having a liquid outlet corresponding to said at least one inlet number; And

Fluid delivery means for fluidly connecting at least one inlet of the microfluidic chip and at least one liquid outlet of the multi-channel liquid dispensing device;

A nucleic acid extraction apparatus comprising a.
The method of claim 4, wherein

A chip outlet region end mounting portion configured to be fixedly mounted to at least one outlet region end of the microfluidic chip, at least one upward liquid inlet corresponding to an upper end of at least one outlet portion of the microfluidic chip, and at least one And a liquid storage container having at least one liquid storage chamber in fluid communication with an upward liquid inlet.
The method of claim 4, wherein

The microfluidic chip may include a heating unit disposed in a first channel region connected to the inlet, and disposed in a second channel region connected to the heating unit, and including a first filter configured to pass a material having a size corresponding to a nucleic acid. Characterized in that, the nucleic acid extracting device.
The method of claim 4, wherein

The microfluidic chip has a heating unit disposed in a first channel region connected to the inlet, and disposed in a second channel region connected to the heating unit, and having a first filter through which a material having a size corresponding to a nucleic acid can pass therethrough. And a nucleic acid separation unit disposed in a third channel region connected to the first filter and having a nucleic acid binding material capable of specifically binding to the nucleic acid.
The method of claim 4, wherein

The microfluidic chip has a heating unit disposed in a first channel region connected to the inlet, and disposed in a second channel region connected to the heating unit, and having a first filter through which a material having a size corresponding to a nucleic acid can pass therethrough. And a nucleic acid separation unit disposed in a third channel region connected to the first filter and having a nucleic acid binding material capable of specifically binding to the nucleic acid, and disposed in a fourth channel region connected to the nucleic acid separation unit. And a second filter capable of passing a substance of a size corresponding to the nucleic acid.
The method of claim 4, wherein

The microfluidic chip has a heating unit disposed in a channel region connected to the inlet unit, and a nucleic acid separation unit including a nucleic acid binding material disposed in the channel region connected to the heating unit and specifically capable of specifically binding to the nucleic acid. Characterized in that, the nucleic acid extracting device.
The method of claim 4, wherein

The microfluidic chip is provided with a nucleic acid separation unit which is disposed in the channel region connected to the inlet, the nucleic acid binding material is disposed in the channel region connected to the heating unit and is provided with a nucleic acid binding material capable of specifically binding to the nucleic acid. And a second filter disposed in a channel region connected to the nucleic acid separation unit and capable of passing a material having a size corresponding to the nucleic acid.
Providing a nucleic acid extracting apparatus according to claim 4;

Injecting a biological sample or reagent into said nucleic acid extraction microfluidic chip through said multi-channel liquid dispensing device and fluid delivery means; And

Extracting nucleic acids from the biological sample by driving the nucleic acid extracting microfluidic chip;

Including, nucleic acid extraction method.
Providing a nucleic acid extracting apparatus according to claim 5;

Injecting a biological sample or reagent into said nucleic acid extraction microfluidic chip through said multi-channel liquid dispensing device and fluid delivery means;

Extracting nucleic acids from the biological sample by driving the nucleic acid extracting microfluidic chip; And

Storing the nucleic acid extract product in a liquid storage chamber of the liquid storage container;

Including, nucleic acid extraction method.