Double-sided chip for nucleic acid extraction for PCR test, cartridge comprising same, and nucleic acid extraction device using same

Abstract

The present invention relates to a double-sided chip for nucleic acid extraction for a PCR test, a cartridge comprising same, and a nucleic acid extraction device using same. The cartridge comprising the chip for nucleic acid extraction includes: a plate-shaped body unit comprising an absorption chamber for absorbing and removing, from a sample, inhibitors which inhibit PCR performance; a front cover coupled to one side of the body unit; and a rear cover coupled to the front cover with the body unit interposed therebetween, wherein the sample passes through the absorption chamber such that a nucleic acid is separated from the sample.

Description

Double-sided chip for nucleic acid extraction for PCR testing, cartridge containing same, and nucleic acid extraction device using same

[0001] The present invention relates to a double-sided chip for nucleic acid extraction for PCR testing, a cartridge containing the same, and a nucleic acid extraction device using the same. The present invention relates to a double-sided chip for nucleic acid extraction for PCR testing, a cartridge containing the same, and a nucleic acid extraction device using the same, which enables the rapid and easy extraction of nucleic acids by moving a biological sample within a single chip, effectively removing foreign substances from the biological sample, and easily separating nucleic acids and buffers.

[0002] Polymerase Chain Reaction (PCR) is a method for amplifying specific target genetic material. It is a technology that amplifies a small amount of genetic material with identical base sequences into a large quantity. PCR is used to diagnose various genetic diseases by amplifying human nucleic acids, and is also applied to bacterial, viral, and fungal nucleic acids for the diagnosis of infectious diseases.

[0003] In general, PCR is performed repeatedly through the following three steps. The three steps include: 1) a denaturing step in which a sample solution containing double-stranded DNA is heated to a specific temperature, for example, about 95°C, to separate the double-stranded DNA into single-stranded DNA; 2) an annealing step in which, after the denaturing step, an oligonucleotide primer having a sequence complementary to a specific base sequence to be amplified is provided to the sample solution, and the solution is cooled to a specific temperature, for example, 55°C, together with the separated single-stranded DNA, to bind the primer to the specific base sequence of the single-stranded DNA to form a partial DNA-primer complex; and 3) an extension (or amplification) step in which, after the annealing step, the sample solution is maintained at an active temperature of a DNA polymerase, for example, 72°C, to form double-stranded DNA based on the primer of the partial DNA-primer complex by a DNA polymerase. By repeating the above three steps several times, a target nucleic acid having a specific base sequence can be exponentially amplified. However, in the case of RNA, a step of reverse transcribing RNA into DNA may be performed prior to steps 1) to 3).

[0004] To perform this PCR, nucleic acids must be extracted from biological samples.

[0005] One example of a conventional nucleic acid extraction technique involves solubilizing a sample containing cells with SDS or proteinase K, then denaturing and removing the proteins with phenol to purify the nucleic acids. However, the phenol extraction method is not only time-consuming due to the numerous processing steps required, but also has the drawback of significantly low reliability, as the nucleic acid extraction efficiency is largely dependent on the researcher's experience and expertise. Recently, kits that utilize silica or glass fibers, which specifically bind to nucleic acids, have been used to address these issues. The silica and glass fibers have a low binding rate with proteins and cellular metabolites, allowing for the extraction of relatively high concentrations of nucleic acids. While this method offers the advantage of simplicity compared to the phenol method, it also requires the complete removal of chaotropic reagents and ethanol, which strongly inhibit enzymatic reactions such as polymerase chain reaction (PCR). This necessitates the complete removal of these substances, making the process very cumbersome and time-consuming.

[0006] In addition, in Korean Patent No. 10-0454869, DNA extraction using a cell lysis buffer was performed as follows. 1) After culturing animal cells, the turbidity containing the cells was centrifuged to recover the cells. 2) 300 μl of cell lysis buffer was added to the recovered cells. 3) After adding the cell lysis buffer, it was left at 70°C for 5 minutes. 4) In order to untangle denatured proteins and the like from the lysis step and allow them to pass through the filter, the lysed cells were pipetted 2-3 times. 5) After transferring the lysed cells to a filter made of silica membrane, centrifugation was performed at 13,000 rpm for 1 minute, the solution was discarded, and centrifugation was performed again. 6) 500 μl of washing buffer was added to the filter and centrifugation was performed. To increase efficiency, the solution was discarded and centrifuged once more to completely remove the washing buffer. 7) 200 ㎕ of elution buffer was added to the filter and centrifuged, and the eluted solution was centrifuged once more in the filter to obtain a larger amount of genomic DNA. 8) The extracted DNA was electrophoresed on a 12% agarose gel, stained with EtBr, and then observed.

[0007] Thus, conventional nucleic acid extraction methods have the advantage of being able to extract nucleic acids with high reproducibility in a short period of time and in a short period of time, consisting of the steps of 1) lysing cells by adding a cell lysis buffer to the cells; 2) transferring the lysed cells of step 1 to a filter to fix the nucleic acids; 3) washing the filter of step 2; and 4) recovering the nucleic acids from the filter. However, a centrifuge is used when transferring the lysed cells to the filter, washing the filter, or recovering the nucleic acids from the filter. Although the use of such a centrifuge is effective in shortening the time, it has the problem of low portability and mobility, and complicates the nucleic acid extraction process in the field.

[0008] Other methods include nucleic acid extraction using magnetic beads, syringes and filters, direct lysis buffer (DLB), and Trizol. However, magnetic bead extraction requires a magnet to fix the nucleic acid to the wall of the tube, and requires the use of a pump or valve (automated equipment) or multiple tips and pipettes (manual) to remove the solution. In addition, syringe and filter extraction methods have the problem that the filter can be damaged if a certain amount of force is applied during the syringe transfer process, making nucleic acid extraction difficult. Direct lysis buffer (DLB) extraction methods must be diluted by 1 / 10 because the DLB itself may contain PCR inhibitors, which significantly reduces sensitivity due to dilution. Furthermore, Trizol extraction methods have the problem of using harmful organic solvents such as phenol or chloroform.

[0009] Accordingly, there is a need for a device that can quickly and easily isolate nucleic acids from biological samples without using additional equipment.

[0010] The present invention aims to provide a double-sided chip for nucleic acid extraction for PCR testing, which can effectively remove foreign substances from a biological sample while moving the biological sample within a single chip without using additional equipment, easily separate nucleic acids and buffers, and quickly and easily extract nucleic acids, a cartridge including the same, and a nucleic acid extraction device using the same.

[0011] A cartridge including a nucleic acid extraction chip according to one aspect of the present invention comprises a plate-shaped body part including an absorption chamber for absorbing and removing an inhibitor that inhibits PCR performance from a sample; a front cover coupled to one side of the body part; and a rear cover coupled to the front cover with the body part interposed therebetween; characterized in that nucleic acids are separated from the sample as the sample passes through the absorption chamber.

[0012] In addition, the body part includes an injection chamber into which a biological sample to be examined is injected; a washing solution storage chamber which receives a washing solution for washing an absorbent filled in the absorption chamber; and a waste collection and nucleic acid separation chamber in which the washing solution that has washed the absorption chamber is collected, passes through the absorption chamber, and separates and discharges nucleic acid from which inhibitors have been removed, and it is preferable that the sample passes through the injection chamber, the washing solution storage chamber, the absorption chamber, and the waste collection and nucleic acid separation chamber in that order.

[0013] In addition, it is preferable that at least one of the washing liquid storage chamber, the absorption chamber, the waste collection and nucleic acid separation chamber is formed on one side of the body portion having a predetermined thickness, and the remaining parts of the washing liquid storage chamber, the absorption chamber, the waste collection and nucleic acid separation chamber that are not formed on the one side are formed on the other side opposite the one side.

[0014] In addition, it is preferable that the body part be provided with a nucleic acid chamber in which the nucleic acid separated through the waste collection and nucleic acid separation chamber is stored.

[0015] In addition, it is preferable that the body part includes an injection chamber having an injection port into which a biological sample to be tested is injected, the body part includes a cap that slides to open and close the injection port, and the cap includes a guide protrusion that is inserted into and guided by a sliding groove formed in the body part, and a sealing member that seals the edge of the injection port at a position where the injection port is closed.

[0016] In addition, the absorption chamber includes a first absorption chamber, a second absorption chamber, and a third absorption chamber that are connected in the vertical direction, and the first, second, and third absorption chambers include a resin to which the inhibitor is adsorbed, and a mesh-structured barrier is provided between the first absorption chamber and the second absorption chamber, and between the second absorption chamber and the third absorption chamber, so that the sample sequentially passes through the first, second, and third absorption chambers, and the resin is preferably prevented from moving by the barrier.

[0017] In addition, the body part is provided with a first tube passage connecting the injection chamber and the washing liquid storage chamber; a second tube passage connecting the washing liquid storage chamber and the absorption chamber; and a third tube passage connecting the absorption chamber and the sink waste collection and nucleic acid separation chamber; and includes a valve that moves up and down with respect to the body part to simultaneously open or close the first tube passage, the second tube passage, and the third tube passage, and it is preferable that the rear cover includes a valve exposure hole exposing the valve so as to transmit a pressure force that moves the valve upward.

[0018] In addition, it is preferable that an air injection hole is formed in the injection chamber through which air is injected by an air pump, the sample is pushed and moved toward the absorption chamber by the air pressure provided through the air injection hole, and a docking part connected to the air pump is provided in the body part.

[0019]

[0020] Meanwhile, a nucleic acid extraction device using a cartridge according to another aspect of the present invention is characterized by including a plate-shaped body part including an absorption chamber for absorbing and removing an inhibitor that inhibits PCR performance from a sample; a front cover coupled to one side of the body part; and a rear cover coupled to the front cover with the body part therebetween; a cartridge for separating nucleic acid from a sample as the sample passes through the absorption chamber; and a mounting part including an insertion part into which the cartridge is inserted and mounted, and a door provided in the insertion part for opening and closing the insertion part.

[0021] Here, the body part includes an injection chamber into which a biological sample to be examined is injected; a washing solution storage chamber which receives a washing solution for washing an absorbent filled in the absorption chamber; and a waste collection and nucleic acid separation chamber in which the washing solution that has washed the absorption chamber is collected, passes through the absorption chamber, and separates and discharges nucleic acid from which inhibitors have been removed, and it is preferable that the sample passes through the injection chamber, the washing solution storage chamber, the absorption chamber, and the waste collection and nucleic acid separation chamber in that order.

[0022] Here, at least one of the washing liquid storage chamber, the absorption chamber, the waste collection and nucleic acid separation chamber is preferably formed on one side of the body portion having a predetermined thickness, and the remaining parts of the washing liquid storage chamber, the absorption chamber, the waste collection and nucleic acid separation chamber that are not formed on the one side are preferably formed on the other side opposite the one side.

[0023] Here, it is preferable that the body part is provided with a nucleic acid chamber in which the nucleic acid separated through the waste collection and nucleic acid separation chamber is stored.

[0024] Here, it is preferable that the body part includes an injection chamber having an injection port into which a biological sample to be tested is injected, the body part includes a gap that opens and closes the injection port by sliding, and the cap includes a guide protrusion that is inserted into a sliding groove formed in the body part and is guided, and a sealing member that seals the edge of the injection port at a position where the injection port is closed.

[0025] Here, the body part is provided with a first tube passage connecting the injection chamber and the washing liquid storage chamber; a second tube passage connecting the washing liquid storage chamber and the absorption chamber; and a third tube passage connecting the absorption chamber and the sink waste collection and nucleic acid separation chamber; and includes a valve that moves up and down with respect to the body part to simultaneously open or close the first tube passage, the second tube passage, and the third tube passage, and the mounting part is provided with a pressure transmission means for transmitting a pressure to move the valve upward, and the rear cover preferably includes a valve exposure hole exposing the valve so that the pressure transmission means pressurizes the valve.

[0026] Here, an air injection hole is formed in the injection chamber through which air is injected by an air pump, and the sample is pushed and moved toward the absorption chamber by the air pressure provided through the air injection hole. It is preferable that a docking part connected to the air pump is provided in the body part, and a pump connection part inserted into the docking part is provided to which the air pump is connected to the mounting part.

[0027]

[0028] Meanwhile, a double-sided chip for nucleic acid extraction for PCR testing according to another aspect of the present invention is characterized by including: an input chamber into which a biological sample to be tested is input; an absorption chamber containing an absorbent that absorbs and removes an inhibitor that inhibits PCR performance from the sample; a nucleic acid chamber containing nucleic acids from which inhibitors have been removed by passing through the absorption chamber; and a body part including the input chamber, the absorption chamber, and the nucleic acid chamber so that the sample sequentially passes through the input chamber, the absorption chamber, and the nucleic acid chamber.

[0029]

[0030] Meanwhile, a double-sided chip for nucleic acid extraction for PCR testing according to another aspect of the present invention comprises: a plate-shaped body having a predetermined thickness; an absorption chamber formed on one surface of the body or the other surface opposite the one surface, and containing an absorbent that absorbs and removes an inhibitor that inhibits PCR performance from the sample; a waste collection and nucleic acid separation chamber formed on one surface of the body or the other surface opposite the one surface, and collecting a washing liquid that washes the absorption chamber, and passing through the absorption chamber to separate and discharge nucleic acids from which inhibitors have been removed.

[0031] A double-sided chip for nucleic acid extraction for PCR testing according to an embodiment of the present invention, a cartridge including the same, and a nucleic acid extraction device using the same provide the effect of effectively removing foreign substances from a biological sample, easily separating nucleic acids and buffers, and quickly and easily extracting nucleic acids while moving the biological sample within a single chip without using additional equipment.

[0032] In addition, the present invention simplifies the nucleic acid separation procedure by eliminating the use of additional equipment such as a centrifuge for nucleic acid separation and easily separating nucleic acids from a single chip, and improves user convenience by making it easy to carry and move to the field.

[0033] Figure 1 is a perspective view of a double-sided chip for nucleic acid extraction according to one embodiment of the present invention.

[0034] Figure 2 is a cross-sectional view of the main part of Figure 1.

[0035] Figure 3 is a perspective view of Figure 1 from a different angle.

[0036] Figure 4 is a perspective view of the opposite side of Figure 1.

[0037] Figure 5 is a drawing showing the path along which the sample flows sequentially in the double-sided chip, indicated by numbers.

[0038] Figure 6 is a drawing showing the chambers through which the sample passes in order.

[0039] Figure 7 is a plan view of Figure 4.

[0040] Figure 8 is a plan view of Figure 3.

[0041] Figure 9 is a perspective view of Figure 1 from a different angle.

[0042] Figure 10 is a drawing showing the nucleic acid extraction from the nucleic acid chamber.

[0043] Figure 11 is a drawing showing a cartridge including a double-sided chip;

[0044] Figure 12 is an exploded perspective view of Figure 11.

[0045] Figure 13 is a drawing showing a state in which a double-sided chip is attached to a rear cover.

[0046] Figure 14 is a drawing showing a state in which the valve closes the tubular path.

[0047] Figure 15 is a drawing showing a state in which the valve opens the tubular path.

[0048] Figure 16 is a drawing showing the valve rising by a pressure transmission means.

[0049] Figure 17 is a drawing showing the cap sliding to open and close the injection chamber and the bottom of the cap.

[0050] Fig. 18 is a perspective view of the cap.

[0051] Fig. 19 is a front view of Fig. 19,

[0052] Figure 20 is a drawing showing a cartridge mounted on a nucleic acid extraction device.

[0053] Figure 21 is a drawing showing how a pump connection part is connected to a cartridge.

[0054] Hereinafter, various embodiments of the present invention will be described in connection with the accompanying drawings. Various embodiments of the present invention may have various modifications and various embodiments, and thus specific embodiments are illustrated in the drawings and related detailed descriptions are provided. However, this is not intended to limit the various embodiments of the present invention to specific embodiments, but should be understood to include all modifications and / or equivalents or substitutes included in the spirit and technical scope of the various embodiments of the present invention. In connection with the description of the drawings, similar reference numerals have been used for similar components.

[0055] Expressions such as “includes” or “may include” that may be used in various embodiments of the present invention indicate the existence of the disclosed function, operation, or component, etc., and do not limit one or more additional functions, operations, or components, etc. In addition, in various embodiments of the present invention, it should be understood that terms such as “includes” or “has” are intended to specify the existence of a feature, number, step, operation, component, part, or combination thereof described in the specification, and do not exclude in advance the possibility of the existence or addition of one or more other features, numbers, steps, operations, components, parts, or combinations thereof.

[0056] When a component is referred to as being "connected" to another component, it should be understood that the component may be directly connected to the other component, but that there may also be a new component between the component and the other component. Conversely, when a component is referred to as being "directly connected" or "directly connected" to another component, it should be understood that no new component exists between the component and the other component.

[0057] The terms used in the various embodiments of the present invention are used solely to describe specific embodiments and are not intended to limit the various embodiments of the present invention. Singular expressions include plural expressions unless the context clearly indicates otherwise.

[0058] Unless otherwise defined, all terms used herein, including technical or scientific terms, have the same meaning as commonly understood by one of ordinary skill in the art to which the various embodiments of the present invention belong.

[0059] Terms defined in commonly used dictionaries should be interpreted as having a meaning consistent with their meaning in the context of the relevant technology, and will not be interpreted in an idealized or overly formal sense unless explicitly defined in various embodiments of the present invention.

[0060]

[0061] The present invention relates to a double-sided chip for nucleic acid extraction for PCR testing, and relates to a chip and device for removing inhibitors that interfere with PCR from a biological sample and isolating and obtaining nucleic acids for performing PCR on the sample. Of course, the double-sided chip for nucleic acid extraction according to the present invention can be utilized not only in cases where nucleic acid extraction is required for the diagnosis, treatment, or prevention of disease, but also in various fields such as new drug development and detection of environmental hormones, where nucleic acid extraction from a sample is required.

[0062] First, the term "PCR (polymerase chain reaction)" used herein refers to a reaction that amplifies a specific target nucleic acid molecule using a thermostable DNA polymerase. In addition to the DNA polymerase, PCR may also use a reaction mixture containing primers (forward primer, reverse primer), which are oligonucleotides capable of specifically hybridizing to the target nucleic acid, a deoxynucleotide triphosphate mixture (dNTP mixture), and divalent ions such as Mg2+.

[0063] A 'primer' is used to initiate a PCR reaction, and refers to an oligonucleotide or polynucleotide that hybridizes complementarily to template DNA. Primers for a PCR reaction may be a pair of forward primers (or sense primers) selected from the sense strand of the nucleic acid molecule being amplified in the same direction as the genetic code progression, and reverse primers (or antisense primers) selected from the antisense strand complementary to the sense strand.

[0064] "Sample" refers to genetic material, such as nucleic acid, that is the target for amplification, or a biological solution containing such genetic material. "Reaction reagent" is intended to detect the target genetic material and may include fluorescent dyes, primers, and the like. Primers may be a pair of primers, each 15-30 bp long, capable of binding to both ends of a specific region of the target gene. Furthermore, DNA polymerase uses an enzyme that retains its activity even at high temperatures, such as 90°C or higher.

[0065]

[0066] Hereinafter, a preferred embodiment according to the present invention will be described in detail with reference to the attached drawings.

[0067] According to one embodiment of the present invention, a double-sided chip (1000) for nucleic acid extraction is provided, which forms a channel and a chamber on both sides of a body part (10) through which a biological sample flows, so that an inhibitory substance (hereinafter referred to as an inhibitor) that inhibits PCR is adsorbed and removed while passing through the channel and chamber, and only nucleic acids are collected and separated in a nucleic acid chamber (300). According to this embodiment, the body part (10) is formed in a plate shape having a predetermined thickness, and an injection chamber (100), an absorption chamber (200), a nucleic acid chamber (300), a washing solution storage chamber (400), and a weight collection and nucleic acid separation chamber (500) to be described later are provided in the body part (10).

[0068]

[0069] 1. Injection chamber (100)

[0070]

[0071] A biological sample to be tested is introduced into the injection chamber (100). Before dispensing the sample into the injection chamber, the sample is pretreated to allow nucleic acid separation. The pretreatment involves injecting a predetermined lysis buffer, such as a lysis buffer, into the biological sample to destroy the cell wall and release the nucleic acid. The lysis buffer is a lysis buffer used to destroy the cell wall. The sample with the nucleic acid exposed is introduced into the injection chamber (100).

[0072] As illustrated in FIGS. 1 and 2, the injection chamber (100) is formed on one side of the body portion (10) and forms a space in the vertical direction, with the width becoming narrower as it goes downward. An injection port (101) into which the sample is injected is provided on the upper side of the injection chamber (100), and an outlet port (102) through which the sample is discharged is provided on the lower side. According to the present embodiment, an inclined guide portion (103) is formed to guide the sample to gather toward the outlet port (102).

[0073] According to the present embodiment, the sample introduced into the injection chamber (100) moves to the washing liquid storage chamber (400) through the discharge port (102) formed at the bottom of the injection chamber (100) by air pressure injected from the top. The air is provided from a pump connected to the lower side of the body part (10), and the air provided by the pump is supplied to the upper side wall of the injection chamber (100). An air injection hole (104) into which air is injected by the pump is formed in the injection chamber (100). According to the present embodiment, the sample can be pushed and moved toward the absorption chamber (200) by the air pressure provided to the air injection hole (104).

[0074]

[0075] 2. Absorption chamber (200)

[0076]

[0077] The absorption chamber (200) is provided to absorb and remove inhibitors that inhibit PCR from the sample. The inhibitors include salts, proteins, and other intracellular chemicals contained in the sample, and act as factors that interfere with the amplification of nucleic acids by the substances during PCR, and are removed by the absorbent contained in the absorption chamber (200). Since the inhibitors contained in the sample are adsorbed and removed by the absorbent, and only the nucleic acids can be discharged and collected downward, the nucleic acids can be separated simply and quickly. The present invention provides a new concept separation device that separates nucleic acids without the process of fixing nucleic acids using a predetermined adsorption filter or the like to separate nucleic acids from a conventional sample and re-separating the nucleic acids from the adsorption filter.

[0078] According to the present embodiment, the absorption chamber (200) includes first, second, and third absorption chambers (210, 220, and 230). The first, second, and third absorption chambers (210, 220, and 230) are filled with spherical resin particles for absorbing an inhibitor. The spherical resin particles absorb the inhibitor. According to the present embodiment, the first, second, and third absorption chambers (210, 220, and 230) are arranged in a vertical direction, and the sample passes through the first, second, and third absorption chambers (210, 220, and 230) sequentially.

[0079] According to the present embodiment, a mesh-structured barrier is formed at each boundary of the first, second, and third absorption chambers (210, 220, 230) to prevent the resin contained in one chamber from moving to another chamber. According to the present embodiment, a fastening groove (203) into which the barrier is inserted is formed between the first absorption chamber (210) and the second absorption chamber (220) and between the second absorption chamber (220) and the third absorption chamber (230). In addition, according to the present embodiment, a fastening groove (203) is also formed between the absorption chamber inlet (201) and the first absorption chamber (210) and between the third absorption chamber (230) and the absorption chamber outlet (202), so that a barrier is installed to prevent the resin from leaking.

[0080] The above first, second, and third absorption chambers (210, 220, and 230) may be selected to include any one of a cathodic anion exchange resin, a positive cation exchange resin, and a chelating resin. The chelating resin has a weak anode and has a property of adsorbing (capturing) metal divalent ions. In addition, the anion exchange resin, chelating resin, and cation exchange resin are formed by including a spherical resin, and the inhibitor is adsorbed to the spherical resin. The diameter of the spherical resin may be formed to be approximately 0.18 mm to 0.3 mm.

[0081] The above absorption chamber (200) has three chambers, each having a polarity, so that an inhibitor having a polarity in the biological sample can be absorbed and removed in the first, second, and third absorption chambers (210, 220, and 230). According to the present embodiment, the first absorption chamber (210) includes an anion exchange resin, the second absorption chamber (220) includes a chelate resin, and the third absorption chamber (230) includes a cation exchange resin. As described above, the first, second, and third absorption chambers (210, 220, and 230) are arranged sequentially from top to bottom, so that the sample passes through the anion exchange resin, the chelate resin, and the cation exchange resin sequentially.

[0082] According to the present embodiment, the volumes of the anion exchange resin, the chelate resin, and the cation exchange resin can be prepared to be substantially 1:1:1. Of course, the volume ratio of the anion exchange resin, the chelate resin, and the cation exchange resin is not limited to 1:1:1. Since the substances included in the biological sample are polar, they can be absorbed in each layer while passing through the first, second, and third absorption chambers (210, 220, and 230), and only the nucleic acid can flow downward. The anion exchange resin can include at least one selected from TMA (Trimethylamine), DMEA (Dimethylethanolamine), and Tertiary Amine, and the chelate resin can include at least one selected from carboxyl groups (-COOH), iminodiacetate acid chelates, and the like. In addition, the cation exchange resin can include at least one selected from sulfate groups (-SO3H), carboxyl groups (-COOH), and sulfonic acid groups (-SO3H).

[0083] An absorption chamber outlet (202) is formed at the lower side of the third absorption chamber, and an inhibitor is removed from the sample through the absorption chamber outlet (202) and the remaining nucleic acid is discharged downward. According to the present embodiment, the sample moves inside the chip at a speed of 15 μl / sec or less, and as a result, the flow rate of the nucleic acid discharged through the absorption chamber outlet (202) becomes 15 μl per second or less. The flow rate per unit time of the nucleic acid discharged through the absorption chamber outlet (202) can be controlled by appropriately forming the cross-sectional area of ​​the absorption chamber outlet (202) through which the nucleic acid passes. As the cross-sectional area of ​​the above absorption chamber outlet (202) becomes relatively larger, the discharge amount of the nucleic acid may increase. However, since the speed at which the sample passes through the absorption chamber (200) relatively increases, it may not be possible to secure enough time for the inhibitor to be sufficiently adsorbed in the first, second, and third absorption chambers (210, 220, 230). Therefore, it was confirmed that the inhibitor could be sufficiently removed when the discharge flow rate of the nucleic acid was adjusted to 15 μL or less per second.

[0084]

[0085] 3. Nucleic acid chamber (300)

[0086]

[0087] The nucleic acid chamber (300) receives nucleic acids from which inhibitors have been removed by passing through the absorption chamber (200). As illustrated in FIGS. 1 and 2, according to the present embodiment, the nucleic acid chamber (300) has a space whose width narrows from the top to the bottom. As illustrated in FIG. 10, a nucleic acid chamber inlet (310) through which nucleic acids are introduced is formed in the nucleic acid chamber (300). The nucleic acid collected in the nucleic acid chamber (300) can be extracted using a device such as a pipette (330) through an extraction port (320) formed opposite the nucleic acid chamber inlet (310).

[0088] According to the present embodiment, the sample passes through the injection chamber (100), the absorption chamber (200), and the nucleic acid chamber (300) sequentially, and the body part (10) is provided with the injection chamber (100), the absorption chamber (200), and the nucleic acid chamber (300), so that the process of separating and storing nucleic acid from the sample is possible in one body part (10).

[0089]

[0090] 4. Washing fluid storage chamber (400)

[0091]

[0092] The washing liquid storage chamber (400) is a chamber provided between the injection chamber (100) and the absorption chamber (200), and accommodates a washing liquid for washing the absorbent filled in the absorption chamber (200). The absorption chamber (200) is filled with a polar resin, and each chamber is filled with a preservation buffer to maintain the polarity of the first, second, and third absorption chambers (210, 220, 230) forming the absorption chamber (200). The washing liquid is provided to wash the preservation buffer.

[0093] According to the present embodiment, the washing liquid storage chamber (400) is formed on one side of the body portion (10), and is formed on the same side as the side on which the absorption chamber (200) is formed. When the sample introduced into the injection chamber (100) is pushed out by air pressure, the washing liquid stored in the washing liquid storage chamber (400) moves toward the absorption chamber (200) by the movement pressure of the sample and air, and the absorbent of the absorption chamber (200) is washed first.

[0094] As illustrated in FIG. 4, the washing liquid storage chamber (400) includes a plurality of washing liquid flow paths (440) arranged in parallel in one direction so that the washing liquid can flow. As illustrated in FIG. 7, the washing liquid flows into the upper washing liquid storage chamber inlet (450), passes through the washing liquid flow path (440), and then moves to the absorption chamber (200) through the washing liquid storage chamber outlet (460). The washing liquid flows in a zigzag pattern from upper to lower along the washing liquid flow path (440). According to the present embodiment, the washing liquid flow path (440) includes a first path (401), a second path (402), and a third path (403) having different lengths.

[0095] According to the present embodiment, the washing liquid storage chamber (440) includes a first storage unit (410) configured with a plurality of first channels (401) having a first length to store the washing liquid, a second storage unit (420) configured with a plurality of second channels (402) shorter than the first length to store the washing liquid and provided on the lower side of the first storage unit (410), and a third storage unit (430) configured with a plurality of second channels (403) shorter than the second length to store the washing liquid and provided on the lower side of the second storage unit (430). According to the present embodiment, the first, second, and third storage units (410, 420, 430) are sequentially provided from the upper side to the lower side. The first, second, and third storage units (410, 420, and 430) are each formed with a plurality of horizontal flow paths (401, 402, and 403) in a zigzag manner so that sufficient washing liquid can be stored.

[0096]

[0097] 5. Waste collection and nucleic acid separation chamber (500)

[0098]

[0099] The waste collection and nucleic acid separation chamber (500) is provided to collect the washing liquid discharged from the washing liquid storage chamber (400) at the rear end of the absorption chamber (200) and to separate and discharge the nucleic acid. The nucleic acid separated by the waste collection and nucleic acid separation chamber (500) is collected in the nucleic acid chamber (300). The waste collection and nucleic acid separation chamber (500) may be formed on one or the other surface of the body portion (10).

[0100] Therefore, according to the present invention, at least one of the absorption chamber (200), the washing liquid storage chamber (400), and the waste collection and nucleic acid separation chamber (500) may be formed on at least one of one surface of the body part (10) having a predetermined thickness and the other surface opposite thereto. That is, at least one of the absorption chamber (200), the washing liquid storage chamber (400), and the waste collection and nucleic acid separation chamber (500) may be formed on one surface of the body part (10) having a predetermined thickness, and the remainder of the absorption chamber (200), the washing liquid storage chamber (400), and the waste collection and nucleic acid separation chamber (500) that are not formed on the one surface may be formed on the other surface opposite thereto. According to the present embodiment, the washing liquid storage chamber (400) and the absorption chamber (200) are formed on one surface of the body part (10), and the waste collection and nucleic acid separation chamber (500) is formed on the other surface of the body part (10).

[0101] However, the absorption chamber (200) may be formed on one side or the other side of the body part (10), and the waste collection and nucleic acid separation chamber (500) may also be formed on one side or the other side of the body part (10). That is, in one embodiment, the absorption chamber (200) and the waste collection and nucleic acid separation chamber (500) may be formed on the same side of the body part (10).

[0102]

[0103] Referring to FIGS. 8 and 9, the waste collection and nucleic acid separation chamber (500) is provided to collect waste and separate nucleic acids by having the washing liquid passing through the absorption chamber (200) fill the waste chamber (510) and subsequently moving the nucleic acids flowing through the inflow path (570) through the nucleic acid separation path (530) to separate them.

[0104] The above waste collection and nucleic acid separation chamber (500) includes an inlet passage (570), a washing liquid collection passage (520), a nucleic acid separation passage (530), and a waste chamber (510).

[0105] The above inflow path (570) extends from top to bottom and is a path through which washing liquid and nucleic acid are introduced. According to the present embodiment, the washing liquid is first introduced into the inflow path (570), and the nucleic acid then flows in following the washing liquid. The inflow path (570) includes a curved path (560) having a predetermined curvature at its end.

[0106] The washing liquid collection channel (520) extends downward from the end of the inflow channel (570) and is connected so that the washing liquid flows. According to the present embodiment, the washing liquid collection channel (520) is connected to a curved channel (560) provided at the end of the inflow channel (570) and extends downward.

[0107] The nucleic acid separation channel (530) extends from the end of the inflow channel (570) in a direction different from that of the washing solution collection channel (520), thereby forming a channel through which the nucleic acid flows. According to the present embodiment, the nucleic acid separation channel (530) is connected to the curved channel (560) and extends upward in the opposite direction of the washing solution collection channel (520). The nucleic acid separation channel (530) is provided with a resistance protrusion (590) protruding from the inner surface. The resistance protrusion (590) functions to induce the washing solution to move toward the washing solution collection channel (520) downward when the washing solution is initially introduced.

[0108] According to the present embodiment, a resistance portion (580) that generates resistance when the washing liquid flows is formed on the rounded inner surface where the inflow channel (570) and the nucleic acid separation channel (530) are connected to each other, that is, on the inner surface of the curved channel (560). According to the present embodiment, the resistance portion (580) may be formed as a protrusion that protrudes like a sawtooth shape on the inner surface that meets the end of the curved channel (560) toward the nucleic acid separation channel (530). The protrusions may be formed at regular intervals. The washing liquid flows toward the nucleic acid separation channel (530) that extends downward by gravity, and at this time, the washing liquid collides with the resistance portion (580) while passing through the curved channel (560), so that the washing liquid can be more easily guided and flow toward the lower waste chamber (510).

[0109] In addition, according to the present embodiment, as illustrated in FIG. 8, the width (W2) of the washing liquid collection channel (520) is formed wider than the width (W1) of the nucleic acid separation channel (530). The washing liquid flows into the inflow channel (570), passes through the curved channel (560), and can flow more smoothly toward the wider washing liquid collection channel (520) at the end of the curved channel (560). In addition, a washing liquid guide channel (524) is formed in the washing liquid collection channel (520) that has a narrower width than the width of the washing liquid collection channel (520) and is formed by being sunken deeper than the bottom of the washing liquid collection channel (520). The above washing liquid induction path (524) allows the washing liquid to easily flow downward from the end of the curved path (560) toward the waste chamber (510) by capillary phenomenon.

[0110] The waste chamber (510) is provided at an end of the washing liquid collection channel (520) to provide a space in which the washing liquid is filled. According to the present embodiment, the washing liquid flows in from the bottom of the waste chamber (510) and fills the space of the waste chamber (510). According to the present embodiment, as illustrated in FIG. 9, a support rib (501) protruding vertically from the bottom surface is formed on the inside of the waste chamber (510). According to the present embodiment, a plurality of the support ribs (501) are provided. According to the present embodiment, one side and the other side of the body part (10) are sealed by adhering a film. The support ribs (501) prevent the surface of the film from sinking into the space of the waste chamber (510) when sealing the other side of the body part (10) with the film. When the film is adhered to the other surface of the body part (10), if the area of ​​the space secured by sinking into the space of the waste chamber (510) is reduced, the storage space for the washing liquid is reduced accordingly, which may cause a problem in that the washing liquid is discharged together with the nucleic acid through the nucleic acid separation path (530). Therefore, the support rib (501) ensures that the space of the waste chamber (510) can secure a space in which the washing liquid can be stored.

[0111] According to the present embodiment, a pressure-providing portion (540) is provided on the upper side of the waste chamber (510) to prevent nucleic acids from flowing further into the waste chamber (510) by the pressure of the washing liquid by filling the washing liquid. The pressure-providing portion (540) includes a microchannel (550) formed on the upper side of the waste chamber (510). The width of the microchannel (550) is narrower than that of the inflow channel (570), the curved channel (560), the waste collection channel (520), and the nucleic acid separation channel (530). A plurality of micro-spaces (541) are included on the microchannel (550).

[0112] When the washing liquid is filled in the waste chamber (510), the internal pressure of the waste chamber (510) increases by the microchannel (550) and the microspace (541). Some of the washing liquid may penetrate into the microchannel (550) and the microspace (541). The microspace (541) is provided to prevent sufficient pressure from being applied to a portion of the microchannel (550) connected to the waste chamber (510), and even if the washing liquid penetrates through the microchannel (550) near the waste chamber (510), pressure is applied again by the remaining microchannel (550) and the microspace (541).

[0113] In addition, the microchannel (550) and microspace (541) prevent the pressure of the waste chamber (510) from increasing rapidly. When the pressure increases by the pressure providing unit (540), the washing liquid is no longer transported toward the waste chamber (510), so that the nucleic acid subsequently introduced through the inflow channel (570) can flow upward through the nucleic acid separation channel (530) due to the repulsive force. Through this process, the washing liquid that washes away the absorbent of the absorption chamber (200) is collected and separated in the waste chamber (510), and only the nucleic acid can move to the nucleic acid chamber (300) through the nucleic acid separation channel (530).

[0114]

[0115] Meanwhile, according to another aspect of the present invention, the double-sided chip (1000) may be manufactured and provided in the form of a cartridge (2000).

[0116] As illustrated in FIGS. 11 and 12, a cartridge (2000) according to the present embodiment includes a body portion (10), and a front cover (2100) and a rear cover (2200) which are respectively coupled to one side and the other side of the body portion (10). The body portion (100) is positioned between the front cover (2100) and the rear cover (2200), and separates nucleic acids from a sample as the sample passes through the absorption chamber (200).

[0117] The above body part (10) is formed in a plate shape with a predetermined thickness and includes an injection chamber (100), an absorption chamber (200), a nucleic acid chamber (300), a washing liquid storage chamber (400), and a waste collection and nucleic acid separation chamber (500). A repetitive description of each chamber formed in the body part (10) is omitted as described above.

[0118] The body part (10) further includes a tubular flow path (700) in addition to the flow path formed on the surface of the body part (10) so that a sample can be transferred from the injection chamber (100) to the nucleic acid chamber (300). According to the present embodiment, a valve (600) for opening and closing the tubular flow path (700) is provided. As illustrated in Fig. 12, the tubular flow path (700) is connected by having both ends inserted into the tube flow path connecting portion (13) provided in the body part (10).

[0119] The above tubular flow path (700) includes a first tube flow path provided between the injection chamber (100) and the washing liquid storage chamber (400), a second tube flow path provided between the washing liquid storage chamber (400) and the absorption chamber (200), and a third tube flow path provided between the absorption chamber (200) and the waste waste collection and nucleic acid separation chamber (500). In addition, according to the present embodiment, the valve (600) simultaneously opens or closes the first, second, and third tube flow paths.

[0120] As illustrated in FIGS. 14 and 15, the valve (600) is coupled to the body part (10) so as to be movable in the vertical direction. As illustrated in FIG. 14(c), when the valve (600) is in the first position, the pressure rib (610) formed in the valve (600) presses the tubular flow path (700) to keep the flow path closed. That is, the tubular flow path (700) is in a state where the valve (600) is pressed in the first position as illustrated in FIGS. 14(a) and 14(b). Then, as illustrated in FIG. 15, when the valve (600) moves to a second position higher than the first position, the pressure rib (610) pressing the tubular flow path (700) moves upward, opening the tubular flow path (700) so that the sample can flow in one direction.

[0121] A groove is formed in the rear cover (2200) coupled to one side of the body portion (10), and a valve exposure hole (2210) is formed on the upper side of the groove to expose the valve (600) so as to transmit a pressing force to move the valve (600) upward. As illustrated in FIG. 16, when a cartridge (2000) is inserted into the nucleic acid extraction device (3000) and a nucleic acid extraction operation is performed, the pressing force transmission means (3600) provided in the nucleic acid extraction device (3000) transmits a force to move the valve (600) upward through the valve exposure hole (2210). The valve (600) moves upward by the pressing force, and the tubular flow path (700) is opened.

[0122] In addition, a cap (2300) for opening and closing the inlet (101) of the injection chamber (100) is coupled to the upper side of the body part (10). As illustrated in FIG. 17, the gap (2300) is slidably coupled to the upper surface of the body part (10). The cap (2300) includes a guide protrusion (2370) that is inserted into and guided by a sliding groove (14) formed in the body part (10), and a sealing member (2330) that seals the edge of the inlet (101) at a position where the inlet (101) is closed.

[0123] As illustrated in FIGS. 17 and 18, the cap (2300) is formed with a cut groove (2310) that extends and faces each other in the longitudinal direction. A catch (2320) is formed to protrude in the cut groove (2310). A protrusion (15) is formed in the width direction on the upper surface of the body (10), and when the cap (2300) is slid to close the inlet (101), the cut groove (2310) is elastically deformed so that the catch (2320) can pass over the protrusion (15). When the cap (2320) moves to the upper side of the inlet (101), a sealing member (2330), such as an O-ring, which is coupled to the lower surface of the cap (2300) is placed on the edge of the inlet (101) to seal the inlet (101), and the protrusion (15) is maintained in a state of being caught by the catch (2320), so that the cap (2300) closes the inlet (101).

[0124] As illustrated in FIG. 18, the cap (2300) includes a handle (2340), a guide piece (2360) on which the guide protrusion (2370) is formed, and a position control protrusion (2350) that regulates the sliding range of the cap (2300). As illustrated in FIG. 19, the guide piece (2360) extends downward from both ends of the cap (2300) and is coupled to the upper side of the body part (10), and the guide protrusion (2370) formed on the inside of the guide piece (2360) is inserted into the sliding groove (14) of the body part (10). The position control protrusion (2350) is formed longitudinally on the upper surface of the cap (2300) so that its end can contact the front cover (2100) or the rear cover (2200) to limit the movement range of the cap (2300).

[0125]

[0126] Meanwhile, according to another aspect of the present invention, a nucleic acid extraction device (3000) is proposed that receives the cartridge and extracts nucleic acids. The extraction device (3000) includes a cartridge (2000) including the double-sided chip described above, and a mounting portion (3100) on which the cartridge (2000) is mounted. Since the configuration of the cartridge (2000) has been described above, a repetitive description thereof will be omitted.

[0127] The above-mentioned mounting portion (3100) may include an insertion portion (3200) into which the cartridge (2000) is inserted and mounted, a door (3300) for opening and closing the insertion portion (3200), a select bar (3400) for controlling the nucleic acid extraction device (3000), and a display portion (3500) for externally displaying whether the nucleic acid extraction device (3000) is operating.

[0128] As illustrated in FIG. 20, according to the present embodiment, the insertion portion (3200) is provided so that two cartridges (2000) can be inserted. Of course, the number of cartridges (2000) that can be accommodated is not limited thereto.

[0129] The above select bar (3400) can be implemented to enable selection of the on / off operation and operation time of the nucleic acid extraction device (3000). The display unit (3500) is provided on the upper surface of the mounting unit (3100) to visually check the operation progress status of the nucleic acid extraction device (3000). According to the present embodiment, the select bar (3400) is provided in a knob type, and by rotating the select bar (3400), the nucleic acid extraction device (3000) programmed under predetermined conditions can be operated.

[0130] As illustrated in Fig. 21, the mounting portion (3100) is provided with a pump connection portion (3700) for supplying air to the cartridge (2000). A docking portion (11) for receiving air pressure from an air pump is provided in the body portion (10) of the cartridge (2000), and a docking hole (2230) is provided to expose the docking portion (11) at the lower side when the front cover (2100) and the rear cover (2200) are coupled to each other. When the cartridge (200) is inserted into and mounted in the insertion portion (3200), as illustrated in Fig. 21, the upper portion of the pump connection portion (3700) is inserted into the docking portion (11), and a pump is connected to the lower portion.

[0131]

[0132] Hereinafter, the function and effect of the double-sided chip for nucleic acid extraction for PCR testing according to the above configuration are described based on experimental results.

[0133]

[0134] The present invention relates to a double-sided chip (1000), a cartridge (2000), and a nucleic acid extraction device (3000) for extracting nucleic acids. In contrast to the conventional concept of capturing and concentrating nucleic acids using a column method or magnetic beads and then re-extracting nucleic acids from the beads, the present invention improves the nucleic acid extraction process to be simpler by removing inhibitory substances using an absorbent (resin type) in an absorption chamber (200), thereby dramatically speeding up and simplifying the nucleic acid extraction process. In addition, the nucleic acids can be automatically separated from a buffer (washing solution) in a waste collection and nucleic acid separation chamber (500) by utilizing the difference in pressure.

[0135]

[0136] - Flow of sample in double-sided chips

[0137]

[0138] After a sample is introduced into the injection chamber (100), the cartridge (2000) is mounted on the nucleic acid extraction device (3000) to perform the nucleic acid extraction process. Air is supplied into the interior of the injection chamber (100) through the docking portion (11) of the cartridge (2000) from a pump connected to the nucleic acid extraction device (3000).

[0139] As shown in FIGS. 5 and 6, air is supplied to No. 0, exits to No. 1, and flows to No. 2. No. 2 is connected to No. 3. The air flowing to No. 2 moves to No. 3 on the opposite side. The air moves from No. 3 to No. 4, moves to the opposite side again through No. 5, which is connected to No. 4, and rises to No. 6 before being supplied to the injection chamber (100). The sample received in the injection chamber (100) is discharged to No. 7 by air pressure. The sample moves to No. 8, exits to No. 9 on the opposite side, and moves from No. 9 to No. 10 along the tube-shaped path. The sample introduced to No. 10 exits to No. 11 on the opposite side and flows into the washing liquid storage chamber (400). At this time, the washing liquid stored in the washing liquid storage chamber (400) moves to No. 12 due to the movement pressure of the sample and is discharged to No. 13 on the opposite side. Nos. 13 and 14 are connected by a tubular path. The washing liquid passing through No. 14 flows again into No. 15 on the opposite side and washes the resin contained in the first, second, and third absorption chambers (210, 220, 230) while passing through the absorption chamber (200). The washing liquid passing through the absorption chamber (200) is discharged to No. 16 and passes through Nos. 17 and 18. Subsequently, the washing liquid passes through Nos. 19 and 20 and flows into No. 21. At this time, Nos. 17 and 18 are connected by a tubular path.

[0140] The washing liquid introduced through No. 21 flows downward and enters the waste collection and nucleic acid separation chamber (500). As described above, the washing liquid first flows downward through the washing liquid collection channel (520) to fill the waste chamber (510), and when the waste chamber (510) is filled with the washing liquid, pressure is applied by the narrow microchannel (550) of No. 22, and the nucleic acid flowing following the washing liquid (the nucleic acid to which the inhibitor is adsorbed in the absorption chamber (200) and discharged from the absorption chamber (200)) flows toward No. 23. The nucleic acid discharged through No. 23 exits through No. 24 on the opposite side and flows into the nucleic acid chamber (300) through No. 25 to be extracted. The user can capture the nucleic acid extracted through No. 26 using a pipette (330), etc.

[0141]

[0142] - Experimental verification of the function and effectiveness of the absorption chamber

[0143]

[0144] When the sample does not contain an inhibitor, and the nucleic acid is passed through a polar resin layer and then PCR is performed, it was confirmed that the effect on the number of repetitions of the PCR cycle for detecting a specific base sequence is minimal when performing PCR using Direct Lysis Buffer (DLB) prepared using a conventional lysis buffer and when performing PCR after passing through at least one absorbent (an absorption chamber including at least one of the first, second, and third chambers can be configured). The PCR cycle is defined as the following three steps as one cycle. The above three steps refer to: 1) a denaturing step in which a sample solution containing double-stranded DNA is heated to a specific temperature, for example, about 95°C, to separate the double-stranded DNA into single-stranded DNA; 2) an annealing step in which an oligonucleotide primer having a sequence complementary to a specific base sequence to be amplified is provided to the sample solution after the denaturing step, and the sample solution is cooled to a specific temperature, for example, 55°C, together with the separated single-stranded DNA, to bind the primer to the specific base sequence of the single-stranded DNA to form a partial DNA-primer complex; and 3) an extension (or amplification) step in which the sample solution after the annealing step is maintained at an active temperature of a DNA polymerase, for example, 72°C, to form double-stranded DNA based on the primer of the partial DNA-primer complex by a DNA polymerase. PCR exponentially amplifies target nucleic acids with a specific base sequence by repeating the above three steps multiple times, enabling the detection of a specific base sequence. In Table 1 below, the CT value of 29.56 indicates that the average number of repetitions is 29.56.From these experimental results, it was confirmed that although nucleic acids have a weak cathode, when PCR was performed by passing them through an anion exchange resin, a chelating resin, and a cation exchange resin, respectively, and when these absorbents were configured in three stages and passed through a three-stage chamber, the number of repetitions of PCR cycles that enabled the detection of a specific gene was only slightly different from the conventional method, and thus the nucleic acids were not affected by the resin layer.

[0145] CT values ​​of clinical samples without inhibitors: DLB 29.56, cation exchange resin 29.84, chelate resin 29.77, anion exchange resin 30.15, three-stage resin of anion, chelate, and cation 30.63

[0146] And, when the sample contains an inhibitor, as shown in Table 2 below, the sample prepared using the conventional dissolution buffer contained an inhibitor, so even if the three steps for PCR were performed, amplification of a specific base sequence was not achieved due to the inhibitor. However, as shown in Table 2, when the sample was passed through an anion exchange resin, a chelating resin, and a cation exchange resin and the three steps for PCR were repeated, it was confirmed that a specific base sequence was amplified to a detectable level. In particular, it was confirmed that the number of PCR cycle repetitions for detecting a specific base sequence was the shortest when the absorption chamber was configured as a three-stage chamber with resin layers including an anion exchange resin, a chelating resin, and a cation exchange resin from the top to the bottom. In other words, when the absorption chamber is configured to sequentially pass through the three-stage resin layers, amplification of a specific base sequence can be completed quickly.

[0147] CT values ​​of clinical samples with inhibitors DLB (control) 0 Cation exchange resin layer (123) 36.68 Chelate resin layer 41.34 Anion exchange resin layer (121) 39.56 Three-layer resin layer of anion, chelate, and cation 35.13

[0148] According to the present embodiment, the absorption chamber (200) is sequentially arranged with first, second, and third absorption chambers (210, 220, 230) including a cathode-bearing anion exchange resin, a chelating resin, and a cathode-bearing cation exchange resin. In another embodiment, the absorption chamber (200) may be formed by selecting at least one of the first, second, and third absorption chambers (210, 220, 230). That is, the absorption chamber may be formed by a combination of at least one, two, or three layers.

[0149] According to an embodiment of the present invention, the volume ratio of the anion exchange resin, the chelate resin, and the cation exchange resin, which are made of spherical resin particles included in the first, second, and third absorption chambers (210, 220, and 230), is substantially 1:1:1. In addition, the flow rate of the sample passing through the interior of the absorption chamber is adjusted to 15 μL / sec or less. The flow rate of the sample passing through the interior of the absorption chamber is adjusted to 15 μL / sec or less, so that the flow rate of nucleic acid discharged to the lower part of the absorption chamber is adjusted to 15 μL per second or less, and the inhibitor can be sufficiently absorbed into the absorbent of the spherical particles as the sample passes through the first, second, and third absorption chambers (210, 220, and 230). When the flow rate of the nucleic acid discharged exceeds 15 μL per second, the movement speed of the sample becomes fast, so that the inhibitor cannot be sufficiently absorbed.

[0150] According to an embodiment of the present invention, the amount of nucleic acid collected in the nucleic acid chamber (300) can be secured to be substantially the same as the amount of nucleic acid injected into the injection chamber (100). For example, when 500 μl of the biological sample is injected, the inhibitor contained in the sample is removed, and the flow rate discharged to the lower portion of the absorption chamber (200) is maintained at approximately 500 μl, and the inside is obtained in a state in which the inhibitor is removed and only the nucleic acid is contained. This significantly improves the secured flow rate of the sample containing only the nucleic acid compared to the prior art.

[0151] In this way, the double-sided chip for nucleic acid extraction for PCR testing, the cartridge including the same, and the nucleic acid extraction device according to the embodiment of the present invention provide the effect of effectively removing foreign substances from the biological sample while moving the biological sample within a single chip without using additional equipment, easily separating the nucleic acid and buffer, and quickly and easily extracting the nucleic acid. In addition, the present invention eliminates the use of additional equipment such as a centrifuge for nucleic acid separation, thereby making it easy to carry and move to the field and improving user convenience.

[0152]

[0153] Above, the present invention has been described in detail with reference to preferred embodiments, but the present invention is not limited to the above embodiments, and many modifications can be provided within a range that does not depart from the scope of the present invention.

Claims

1. A plate-shaped body including an absorption chamber that absorbs and removes an inhibitor that inhibits PCR performance from a sample; A front cover coupled to one side of the above body part; and Including a rear cover that is connected to the front cover with the body part in between; A cartridge including a nucleic acid extraction chip, characterized in that nucleic acids are separated from the sample as the sample passes through the absorption chamber.

2. In paragraph 1, The above body part, An injection chamber into which a biological sample to be tested is injected; A washing liquid storage chamber that receives a washing liquid for washing the absorbent filled in the above absorption chamber; and It includes a waste collection and nucleic acid separation chamber in which the washing liquid that washes the above absorption chamber is collected, passes through the above absorption chamber, and separates and discharges the nucleic acid from which the inhibitor has been removed. A cartridge including a nucleic acid extraction chip, characterized in that the sample passes through the injection chamber, the washing liquid storage chamber, the absorption chamber, and the waste collection and nucleic acid separation chamber in sequence.

3. In paragraph 2, At least one of the washing liquid storage chamber, the absorption chamber, and the waste collection and nucleic acid separation chamber is formed on one surface of the body portion having a predetermined thickness, A cartridge including a chip for nucleic acid extraction, characterized in that the remainder of the washing liquid storage chamber, the absorption chamber, and the waste collection and nucleic acid separation chamber that are not formed on the one side are formed on the other side opposite the one side.

4. In paragraph 2, A cartridge including a nucleic acid extraction chip, characterized in that the body part has a nucleic acid chamber in which the nucleic acid separated through the waste collection and nucleic acid separation chamber is stored.

5. In paragraph 1, The above body part includes an injection chamber having an injection port into which a biological sample to be tested is injected, The above body part includes a cap that opens and closes the inlet by sliding, A cartridge including a nucleic acid extraction chip, characterized in that the cap includes a guide projection that is inserted into a sliding groove formed in the body portion and guided, and a sealing member that seals the edge of the inlet at a position where the inlet is closed.

6. In paragraph 1, The above absorption chamber includes a first absorption chamber, a second absorption chamber, and a third absorption chamber that are connected in the vertical direction, The above first, second, and third absorption chambers contain a resin to which the inhibitor is adsorbed, A cartridge including a chip for nucleic acid extraction, characterized in that a mesh-structured barrier is provided between the first absorption chamber and the second absorption chamber, and between the second absorption chamber and the third absorption chamber, so that the sample passes through the first, second, and third absorption chambers sequentially, and the resin is prevented from moving by the barrier.

7. In paragraph 2, The body part is provided with a first tube passage connecting the injection chamber and the washing liquid storage chamber; a second tube passage connecting the washing liquid storage chamber and the absorption chamber; and a third tube passage connecting the absorption chamber and the waste collection and nucleic acid separation chamber. It includes a valve that moves up and down with respect to the body part to simultaneously open or close the first tube channel, the second tube channel, and the third tube channel, A cartridge including a nucleic acid extraction chip, characterized in that the rear cover includes a valve exposure hole exposing the valve so as to transmit a pressure force causing the valve to move upward.

8. In paragraph 2, An air injection hole is formed in the above injection chamber through which air is injected by an air pump, and the sample is pushed and moved toward the absorption chamber by the air pressure provided through the air injection hole. A cartridge including a nucleic acid extraction chip, characterized in that the body part is provided with a docking part connected to the air pump.

9. A cartridge comprising a plate-shaped body part including an absorption chamber for absorbing and removing an inhibitor that inhibits PCR performance from a sample; a front cover coupled to one side of the body part; and a rear cover coupled to the front cover with the body part in between; wherein the cartridge separates nucleic acid from the sample as the sample passes through the absorption chamber; and A nucleic acid extraction device using a cartridge, characterized by comprising: an insertion portion into which the cartridge is inserted and mounted; and a mounting portion including a door provided on the insertion portion for opening and closing the insertion portion.

10. In paragraph 9, The above body part, An injection chamber into which a biological sample to be tested is injected; A washing liquid storage chamber that receives a washing liquid for washing the absorbent filled in the above absorption chamber; and It includes a waste collection and nucleic acid separation chamber in which the washing liquid that washes the above absorption chamber is collected, passes through the above absorption chamber, and separates and discharges the nucleic acid from which the inhibitor has been removed. A nucleic acid extraction device using a cartridge, characterized in that the sample passes through the injection chamber, the washing liquid storage chamber, the absorption chamber, and the waste collection and nucleic acid separation chamber in sequence.

11. In paragraph 10, At least one of the washing liquid storage chamber, the absorption chamber, and the waste collection and nucleic acid separation chamber is formed on one surface of the body portion having a predetermined thickness, A nucleic acid extraction device using a cartridge, characterized in that the remainder of the washing liquid storage chamber, the absorption chamber, and the waste collection and nucleic acid separation chamber that are not formed on the one side are formed on the other side opposite the one side.

12. In paragraph 10, A nucleic acid extraction device using a cartridge, characterized in that the body part is provided with a nucleic acid chamber in which the nucleic acid separated through the waste collection and nucleic acid separation chamber is stored.

13. In paragraph 9, The above body part includes an injection chamber having an injection port into which a biological sample to be tested is injected, The above body part includes a cap that opens and closes the inlet by sliding, A nucleic acid extraction device using a cartridge, characterized in that the cap includes a guide projection that is inserted into and guided by a sliding groove formed in the body portion, and a sealing member that seals the edge of the inlet at a position where the inlet is closed.

14. In paragraph 10, The body part is provided with a first tube passage connecting the injection chamber and the washing liquid storage chamber; a second tube passage connecting the washing liquid storage chamber and the absorption chamber; and a third tube passage connecting the absorption chamber and the waste collection and nucleic acid separation chamber. It includes a valve that moves up and down with respect to the body part to simultaneously open or close the first tube channel, the second tube channel, and the third tube channel, The above mounting part is provided with a pressure transmission means that transmits a pressure force that moves the valve upward, A nucleic acid extraction device using a cartridge, characterized in that the rear cover includes a valve exposure hole that exposes the valve so that the pressure transmission means pressurizes the valve.

15. In paragraph 10, An air injection hole is formed in the above injection chamber through which air is injected by an air pump, and the sample is pushed and moved toward the absorption chamber by the air pressure provided through the air injection hole. The above body part is provided with a docking part connected to the air pump, A nucleic acid extraction device using a cartridge, characterized in that the air pump is connected to the above mounting portion and a pump connection portion inserted into the above docking portion is provided.

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