WO2014035124A1 - Dispositif de pcr rotatif et puce dédiée à la pcr - Google Patents

Dispositif de pcr rotatif et puce dédiée à la pcr Download PDF

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Publication number
WO2014035124A1
WO2014035124A1 PCT/KR2013/007699 KR2013007699W WO2014035124A1 WO 2014035124 A1 WO2014035124 A1 WO 2014035124A1 KR 2013007699 W KR2013007699 W KR 2013007699W WO 2014035124 A1 WO2014035124 A1 WO 2014035124A1
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WO
WIPO (PCT)
Prior art keywords
pcr
film
sample
chip
forming means
Prior art date
Application number
PCT/KR2013/007699
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English (en)
Korean (ko)
Inventor
김성재
이종철
김원정
황병갑
정재안
Original Assignee
(주) 메디센서
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from KR1020130061468A external-priority patent/KR101513273B1/ko
Application filed by (주) 메디센서 filed Critical (주) 메디센서
Publication of WO2014035124A1 publication Critical patent/WO2014035124A1/fr

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    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L7/00—Heating or cooling apparatus; Heat insulating devices
    • B01L7/52—Heating or cooling apparatus; Heat insulating devices with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples
    • B01L7/525—Heating or cooling apparatus; Heat insulating devices with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples with physical movement of samples between temperature zones
    • B01L7/5255—Heating or cooling apparatus; Heat insulating devices with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples with physical movement of samples between temperature zones by moving sample containers
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0803—Disc shape
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L2300/00—Additional constructional details
    • B01L2300/08—Geometry, shape and general structure
    • B01L2300/0809—Geometry, shape and general structure rectangular shaped
    • B01L2300/0816—Cards, e.g. flat sample carriers usually with flow in two horizontal directions
    • B—PERFORMING OPERATIONS; TRANSPORTING
    • B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
    • B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
    • B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
    • B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
    • B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
    • B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip

Definitions

  • the present invention relates to a PCR device and a PCR chip, and more particularly, to a rotating PCR device having a plurality of temperature ranges and an insertable strip type PCR chip.
  • DNA amplification is widely used for research and development and diagnostic purposes in the life sciences, genetic engineering, and medicine fields, and DNA amplification by polymerase chain reaction (PCR) is widely used.
  • PCR polymerase chain reaction
  • Polymerase chain reaction is a technique for amplifying exponentially a nucleic acid by serially replicating a region having a specific base sequence of a nucleic acid by repeatedly heating and cooling a sample solution containing a nucleic acid.
  • the most important factor when synthesizing DNA using a PCR device is firstly to keep the surroundings of the PCR device containing the DNA sample uniformly at a specific temperature, and secondly, to maintain the specific temperature. As a result, DNA is not synthesized if the temperature range required for DNA synthesis is not correct.
  • a method of maintaining a specific temperature for a predetermined time by increasing or decreasing the temperature from one zone to a specific temperature is adopted. Therefore, when one cycle (cycle) it takes a lot of time about 90 seconds to rise and fall the temperature to reach the temperature required for DNA synthesis.
  • the degree of amplification of the DNA sample is confirmed by electrophoresis. Therefore, the amplification degree of the DNA sample could not be confirmed in real time during the amplification process.
  • the technology for amplifying genes has evolved from using plastic tubes to chips with microchannels.
  • gene amplification on a microchip was made of silicon or glass substrate using a semiconductor manufacturing process.
  • chips made of materials such as polydimethylsiloxane (PDMS), polycarbonate (PC), and acrylic resin (Poly methyl methacrylate, PMMA).
  • PDMS polydimethylsiloxane
  • PC polycarbonate
  • acrylic resin Poly methyl methacrylate
  • microfabrication technology using fluid technology and MEMS (Microelectromechanical System) is a trend to combine the existing analysis technology. At this time, all components necessary for sample analysis can be miniaturized and integrated so that a small amount of liquid sample can be handled in a unit chip and on-chip to be lab-on-achip.
  • the problem to be solved by the present invention is to provide a rotary PCR device that can reduce the PCR process time and can measure a large number of samples.
  • Another object of the present invention is to provide a PCR chip with a simplified manufacturing process.
  • a PCR device includes at least one module including an instrument portion and a measurement portion disposed on the instrument portion, and a support column passing through the center of the instrument portion, wherein the instrument portion has different temperature regions. And a rotary part having chip insertion holes disposed on or above the heating block, the temperature area forming means including heating blocks and contacting the temperature area forming means.
  • the support pillar is a PCR device for rotating the rotating unit so that one or more PCR chips provided in the chip insertion holes pass through the heating blocks.
  • PCR chips may be inserted into the chip insertion holes.
  • the measurement unit may include a short wavelength uniform irradiator for irradiating light to the target material provided on the PCR chip and a detection sensor for detecting light of a specific wavelength emitted from the target material.
  • the temperature region forming means may further include an insulating block interposed between the heating blocks.
  • the insulating block may be formed of an empty space formed by insulating material or the heating blocks.
  • the rotating part may further include a pair of one or more propellers formed on both sides of the support pillar and protruding from the surface of the rotating part.
  • the propellers may have convex curved wings.
  • the rotating part may further include holes adjacent to the propellers and passing through the rotating part.
  • PCR chip includes a support film, a sample reaction film adhered on the support film, a sample injection film adhered on the sample reaction film, and a coating film adhered on the sample injection film
  • the sample reaction film, the inlet and exhaust ports spaced apart from each other by the short axis of the sample reaction film, the sample movement passages extending the inlet and the exhaust port to the long axis of the sample reaction film, the sample movement passage And a sample reaction area portion connected to each other and disposed to face the inlet and the exhaust port.
  • the sample injection film may include holes formed at the same position as the inlet and the exhaust port.
  • the coating film may include covering the holes.
  • the films may include a polymer material having good light transmittance and thermal conductivity.
  • the films may include a polyethylene terephthalate (PET) film or a polycarbonate film.
  • PET polyethylene terephthalate
  • Each of the sample movement passages may include a holding layer application region in which a thermoplastic material that is deformable by heat is disposed.
  • the polymer material may include paraffin or wax.
  • a rotating part including a plurality of PCR chip insertion holes may rotate on a temperature region forming means having different temperature regions. Since a plurality of PCR chips may be inserted into the PCR chip insertion holes, target materials included in the PCR chips may be amplified at the same time by a single PCR process.
  • the PCR device may include one or more modules.
  • the module may include a temperature range forming unit, a rotating unit, and a measuring unit.
  • the PCR device may additionally mount a plurality of modules according to the number of samples to be measured. Thus, multiple samples may be measurable in one PCR device.
  • PCR chip according to an embodiment of the present invention can be formed by bonding a plurality of plastic films with high permeability and high thermal conductivity.
  • the PCR chip can be provided with a simplified manufacturing process.
  • FIG. 1 is a perspective view showing a PCR device equipped with one module according to an embodiment of the present invention.
  • Figure 2 is a plan view showing a PCR device equipped with one module according to an embodiment of the present invention.
  • Figure 3 is a cross-sectional view taken in the direction of AA 'of Figure 2 showing a PCR device equipped with a module according to an embodiment of the present invention.
  • FIG. 4 is a plan view showing a temperature region forming means according to an embodiment of the present invention.
  • FIG. 5 is a schematic view showing a measuring unit according to an embodiment of the present invention.
  • FIG. 6 is a perspective view showing a PCR device equipped with a plurality of modules according to an embodiment of the present invention.
  • FIG. 7 is a perspective view showing a PCR chip according to an embodiment of the present invention.
  • FIG. 8 is an exploded perspective view of a PCR chip according to an embodiment of the present invention.
  • FIG. 1 is a perspective view showing a PCR device equipped with one module according to an embodiment of the present invention.
  • Figure 2 is a plan view showing a PCR device equipped with one module according to an embodiment of the present invention.
  • Figure 3 is a cross-sectional view taken in the direction of AA 'of Figure 2 showing a PCR device equipped with a module according to an embodiment of the present invention.
  • 4 is a plan view showing a temperature region forming means according to an embodiment of the present invention.
  • 5 is a schematic view showing a measuring unit according to an embodiment of the present invention.
  • the PCR apparatus 1000 includes an instrument unit 100, a measurement unit 300, and a support column 305 penetrating the center of the instrument unit 100.
  • the mechanism part 100 may include a temperature region forming means 110 and a rotating part 115.
  • the rotating part 115 may be disposed on the upper or lower portion of the temperature region forming means 110.
  • the temperature region forming means 110 and the rotating part 115 may be in contact with each other.
  • the measuring unit 300 may be disposed on the rotating unit 115.
  • the instrument unit 100 and the measurement unit 300 may be a module 10 of the PCR device 1000.
  • the temperature region forming means 110 may include heating blocks 110a, 110b and 110c and insulating blocks 110d disposed between the heating blocks 110a, 110b and 110c. have.
  • the temperature region forming means 110 may have three unit heating blocks 110a, 110b, 110c to have at least three temperature regions. This is because one cycle of the PCR process generally goes through three temperature stages.
  • Each of the heating blocks 110a, 100b, and 100c may be regions in which a temperature of a predetermined level is maintained, and may be independently temperature controlled. The region where the temperature is maintained may be all or part of the heating blocks 110a, 110b and 110c.
  • the denaturation step is a step of thermally denaturing the DNA to be amplified, the DNA can be separated into two strands. The DNA may function as a template.
  • the binding step lowers the temperature so that primers can hydrogen bond with the DNA strand.
  • the stretching step is a heat-resistant DNA polymerase can synthesize a copy of the target DNA using the deoxynucleotide triphosphate.
  • the insulating blocks 110d may include an insulating material to prevent heat conduction between the heating blocks 110a, 110b, and 110c.
  • the insulating blocks 110d may be empty spaces formed by separating the heating blocks 110a, 110b, and 110c from each other.
  • the rotation unit 115 may rotate using the support pillar 305.
  • the support pillar 305 may further include a bearing structure (not shown) in an area in contact with the rotating part 115.
  • the rotating part 115 may rotate on the support pillar 305 by the bearing structure.
  • the rotating part 115 may include four propellers 225 extending from the rotating part 115 and protruding from the surface of the rotating part 115.
  • the four propellers 225 may be spaced apart from each other by a predetermined interval.
  • the two propellers 225 may be formed in a pair to correspond to each other about the support pillar 305.
  • the propellers 225 may have wings of convex curves.
  • the rotating part 115 may include holes 227 adjacent to the propellers 225 and having the same shape as the propellers.
  • One of the two holes 227 corresponding to each other as the center of the support pillar 305 may be an area in which air is introduced, and the other hole may be an area in which resistance is generated.
  • air is introduced into and / or discharged from the holes 227, so that a pressure difference between the temperature range forming means 110 and the rotary part 115 is increased. May be generated. Accordingly, the buoyancy may be generated in the rotating part 115 to be spaced apart from the temperature area forming means 110 and the rotating part 115 in contact with each other.
  • the rotating unit 115 may have chip insertion holes 220 into which at least one chip is inserted.
  • the chip insertion holes 220 may be formed to be disposed on one heating block.
  • the surface of the PCR chip is in contact with the surface of one heating block to apply the temperature applied to the heating block to the PCR chip.
  • the chip insertion holes 220 are disposed on the first heating block 110a, the PCR chips disposed at the chip insertion holes 220 may be generated at the first heating block 110a. Temperature may be applied.
  • the PCR chip contacts the surfaces of the heating blocks 110a, 110b, and 110c having different temperatures, and is applied to the heating blocks 110a, 110b, and 110c.
  • the PCR process may be performed while going through.
  • the plurality of the PCR chips inserted into the plurality of chip insertion holes 220 are subjected to a PCR process at the same time. Therefore, the target material included in the PCR chips can be amplified at the same time.
  • the measurement unit 300 may be disposed on the third heating block 110c. Alternatively, the measurement unit 300 may be disposed on the lower side and the side of the third heating block (110c). Referring to FIG. 5, the measurement unit 300 may include a multi-wavelength uniform irradiator 300a and a detection sensor 300b. The fluorescent signal of the PCR chip 400 may be measured through the measuring unit 300.
  • the short wavelength uniform irradiator 300a irradiates light onto a target material provided to a PCR chip
  • the detection sensor 300b detects light of a specific wavelength emitted from the target material to perform real-time measurement of a fluorescent material. Can be.
  • the measurement unit 300 may measure in real time a sample included in a plurality of PCR chips 400 that have undergone a PCR process at the same time.
  • the light source used in the short wavelength uniform irradiator 300a may be any one of a white light source such as a tungsten halogen lamp and a xenon discharge lamp, or a single color light source such as an LED and a laser.
  • the measuring method of the measuring unit 300 includes, but is not limited to, a photo analysis method by a camera or a quantitative analysis method by a detector.
  • FIG. 6 is a perspective view showing a PCR device equipped with a plurality of modules according to an embodiment of the present invention.
  • the PCR device 2000 may include a plurality of the modules 10.
  • the modules 10 may be additionally installed and used in the support pillar 305 according to the number of samples.
  • the mechanism parts 100 may be spaced apart from the support pillar 305 by a predetermined interval.
  • the measurement unit 300 may be disposed on each of the instrument units 100.
  • the modules 10 may be driven independently. That is, each of the rotating parts 115 included in the mechanism parts 100 may be driven independently of each other. Therefore, the PCR device can adjust the number of mounting of the modules 10, it is possible to implement a high yield PCR process in an economical manner.
  • FIG. 7 is a perspective view showing a PCR chip according to an embodiment of the present invention.
  • 8 is an exploded perspective view of a PCR chip according to an embodiment of the present invention.
  • the PCR chip 400 may include four films.
  • the PCR chip 400 may include a supporting film 410, a sample reaction film 420, a sample injection film 430, and a coating film 440, which are sequentially stacked.
  • the films 410, 420, 430, and 440 may include a material having high light transmittance and high thermal conductivity.
  • the films 410, 420, 430, and 440 may include, for example, polyethylene terephalate (PET) or polycarbonate (PC).
  • the sample reaction film 420 may be attached onto the support film 410.
  • the support film 410 may have a flat plane.
  • the sample reaction film 420 may include a sample inlet 400a, a sample movement passage 400b, a sample reaction region 400d, and an exhaust port 400e.
  • the sample reaction film 420 has a through hole 423 penetrated in the shape of the sample inlet 400a, the sample movement passage 400b, the sample reaction region 400d and the exhaust port 400e. Can have Therefore, when the sample reaction film 420 is bonded to the support film 410, the through hole 423 is covered on the surface of the support film 410, the sample inlet 400a, the sample movement passage 400b ), The sample reaction region 400d and the exhaust port 400e may be formed.
  • the sample inlet 400a and the exhaust port 400e may be spaced apart from each other by a short axis of the PCR chip at one edge of the sample reaction film 420.
  • the sample movement passages 400b may extend from the sample inlet 400a and the exhaust port 400e with a long axis.
  • the sample reaction region 400d may be disposed at the other edge of the sample reaction film 420 and may be connected to the sample movement passages 400b.
  • the sample containing the target material of the DNA to be replicated and amplified is provided to the sample inlet 400a and moved to the sample reaction region 400d through the sample movement passage 400b.
  • the exhaust port 400e may be an area where air is discharged.
  • the sample moved to the sample reaction region 400d may be subjected to a real time polymerase chain reaction through the heating blocks 110a, 110b, and 110c (see FIG. 5) having different temperatures gradientd. .
  • the sample movement passages 400b may include a holding layer coating area portion 400c.
  • the holding layer coating area 400c is disposed between the sample inlet 400a and the sample reaction area 400d in the sample moving passage 400b and the exhaust port 400e. And the sample reaction region portion 400d.
  • the retaining layer application region 400c may be provided with a thermoplastic material that is deformable by heat.
  • the polymer material may be, for example, paraffin or wax.
  • the polymer material may serve as a valve of the PCR chip 400. In detail, when heat is not applied to the polymer material, the sample injected into the sample inlet 400a may be moved to the sample reaction region 400d through the sample moving passage 400b.
  • the polymer material when heat is applied to the polymer material, the polymer material is expanded to block the sample movement passage 400b. That is, the polymer material blocks the movement of the sample from the sample reaction region 400d to the sample inlet 400a or from the sample reaction region 400d to the exhaust port 400e. Therefore, the problem of the backflow into the sample inlet 400a and the exhaust port 400e can be prevented during the PCR process.
  • the sample injection film 430 may be attached onto the sample reaction film 420.
  • the sample injection film 430 may cover the sample movement passage 400b, the holding layer application region 400c, and the sample reaction region 400d.
  • the sample injection film 430 may include holes 433 exposing the sample injection port 400a and the exhaust port 400e. Therefore, the sample may be injected into the sample inlet 400a through one of the holes 433, and the air discharged to the exhaust unit 400e may be discharged through the other one of the holes 433. have.
  • the coating film 440 may be adhered to the sample injection film 430.
  • the coating film 400 may cover the sample inlet 400a and the exhaust port 400e.
  • a film coated with an adhesive material may be used, or a double-sided tape may be used between the films.
  • the PCR chip 400 is formed by adhering a plurality of plastic films having high permeability and high thermal conductivity to form the PCR chip 400 without additional processes such as microfluid technology or etching used to manufacture the conventional PCR chip 400. Can simplify the manufacturing process. In addition, the manufacturing process of the PCR chip 400 may be simplified to provide an economic advantage of the PCR chip used for single use.

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  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Biochemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Molecular Biology (AREA)
  • Clinical Laboratory Science (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)

Abstract

Un mode de réalisation de la présente invention concerne un dispositif de PCR comprenant : au moins un module comprenant une unité mécanique et une unité de mesure disposée sur l'unité mécanique ; et une colonne de soutien traversant le centre de l'unité mécanique. L'unité mécanique comprend : un moyen formant une région de température comprenant des blocs thermiques présentant différentes régions de températures ; et une partie rotative en contact avec le moyen formant des régions de températures et disposée au-dessus ou en dessous du moyen formant des régions de températures, la partie rotative présentant des ouvertures pour l'insertion de puces formées sur l'un des blocs thermique.
PCT/KR2013/007699 2012-08-30 2013-08-28 Dispositif de pcr rotatif et puce dédiée à la pcr WO2014035124A1 (fr)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
KR20120095600 2012-08-30
KR10-2012-0095600 2012-08-30
KR10-2013-0061468 2013-05-30
KR1020130061468A KR101513273B1 (ko) 2012-08-30 2013-05-30 회전형 pcr 장치 및 pcr 칩

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WO2014035124A1 true WO2014035124A1 (fr) 2014-03-06

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Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2008200006A (ja) * 2007-02-22 2008-09-04 Toyobo Co Ltd 核酸増幅装置、核酸増幅容器及び核酸増幅方法
JP2009136250A (ja) * 2007-12-10 2009-06-25 Seiko Epson Corp 生体試料反応用チップ、生体試料反応装置、および生体試料反応方法
KR20090133079A (ko) * 2008-06-23 2009-12-31 (주)바이오니아 중합효소 연쇄반응 블록 및 이를 이용한 연속형 실시간 모니터링 장치
KR20100070977A (ko) * 2008-12-18 2010-06-28 유니버시티 세인즈 말레이시아 일회용 멀티플렉스 중합효소 연쇄 반응(ｐｃｒ)용 칩 및 장치
KR20120016934A (ko) * 2010-08-17 2012-02-27 한국과학기술원 회전 ｐｃｒ 장치, 이를 위한 ｐｃｒ 칩 및 이를 이용한 회전 ｐｃｒ 방법

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2008200006A (ja) * 2007-02-22 2008-09-04 Toyobo Co Ltd 核酸増幅装置、核酸増幅容器及び核酸増幅方法
JP2009136250A (ja) * 2007-12-10 2009-06-25 Seiko Epson Corp 生体試料反応用チップ、生体試料反応装置、および生体試料反応方法
KR20090133079A (ko) * 2008-06-23 2009-12-31 (주)바이오니아 중합효소 연쇄반응 블록 및 이를 이용한 연속형 실시간 모니터링 장치
KR20100070977A (ko) * 2008-12-18 2010-06-28 유니버시티 세인즈 말레이시아 일회용 멀티플렉스 중합효소 연쇄 반응(ｐｃｒ)용 칩 및 장치
KR20120016934A (ko) * 2010-08-17 2012-02-27 한국과학기술원 회전 ｐｃｒ 장치, 이를 위한 ｐｃｒ 칩 및 이를 이용한 회전 ｐｃｒ 방법

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