CN107739710B - Rapid nucleic acid amplification system - Google Patents
Rapid nucleic acid amplification system Download PDFInfo
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- CN107739710B CN107739710B CN201711091962.2A CN201711091962A CN107739710B CN 107739710 B CN107739710 B CN 107739710B CN 201711091962 A CN201711091962 A CN 201711091962A CN 107739710 B CN107739710 B CN 107739710B
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Classifications
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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
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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
- 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
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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
- B01L2200/00—Solutions for specific problems relating to chemical or physical laboratory apparatus
- B01L2200/10—Integrating sample preparation and analysis in single entity, e.g. lab-on-a-chip concept
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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
- B01L2300/00—Additional constructional details
- B01L2300/18—Means for temperature control
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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
- B01L2300/00—Additional constructional details
- B01L2300/18—Means for temperature control
- B01L2300/1805—Conductive heating, heat from thermostatted solids is conducted to receptacles, e.g. heating plates, blocks
Abstract
The invention provides a rapid nucleic acid amplification system, which transfers nucleic acid amplification reaction solution in reaction consumable materials in different temperature ranges in a rotating mode to realize rapid nucleic acid amplification, and solves the technical problems that the existing nucleic acid amplification system is long in time consumption or difficult to realize reaction consumable materials. The system of the invention comprises a disc-shaped or fan-shaped reaction consumable, a temperature control component and a rotary support component. The reaction consumable is provided with a PCR reaction area close to the edge, and the temperature control assembly comprises two or more groups of PCR temperature area control units which are circumferentially distributed and used for controlling the temperature of the PCR reaction area on the reaction consumable. The reaction consumable is placed on the support component of the rotary support component, and the PCR reaction area is controlled by the rotary motor to be transferred between different temperature zone control units so as to realize rapid nucleic acid amplification.
Description
Technical Field
The invention relates to the fields of molecular biology and molecular diagnosis, in particular to a system for rapidly realizing nucleic acid amplification.
Background
Nucleic acid detection is a fundamental experimental method of molecular biology. Molecular diagnosis is a technology for diagnosing exogenous or self genetic material 'nucleic acid' and expression level thereof in a patient body or body fluid by using a molecular biological method, and is one of important front fields of modern medical development. The molecular diagnosis is based on detection and analysis of target nucleic acid, and the molecular diagnosis technology mainly comprises real-time fluorescence quantitative PCR, fluorescence In Situ Hybridization (FISH), gene sequencing, gene chip and the like. Molecular diagnostic techniques essentially all include three steps: nucleic acid extraction and purification, nucleic acid amplification and detection. Nucleic acid amplification is a process of amplifying a target nucleic acid or a target gene in vitro, and has been developed to include Polymerase Chain Reaction (PCR), loop-mediated isothermal amplification reaction (LAMP), rolling circle amplification Reaction (RCA), ligation amplification reaction (LCR), recombinase Polymerase Amplification (RPA), and the like. PCR is the most commonly used method for amplifying nucleic acid in vitro, and is an essential link for real-time fluorescent quantitative PCR, gene sequencing, gene chip and other molecular diagnostics.
Most of the most commonly used gene amplification instruments and real-time fluorescent quantitative PCR in molecular biology research and clinical molecular diagnosis are based on porous PCR plates or PCR tubes and other reaction consumables, and the thermal cycle process of PCR denaturation, annealing and extension is realized by controlling the temperature rise and fall of a metal temperature block through a semiconductor device. However, such methods require a long time, and PCR processes involving reverse transcription and pre-denaturation are typically performed for 1.5 to 3 hours; by improving the new materials and the new processes, the method can shorten the time to be within 1 hour. The light cycler 2.0 from Roche and the router-Gene Q from Qiagen use centrifugal air-heated thermal systems with very fast temperature change rates and good temperature uniformity, which control the shortening of the PCR process containing reverse transcription and pre-denaturation to within 1.5 hours, with the light cycler using capillary glass as the reaction consumable and requiring only 0.5-1 hour. But the temperature control accuracy and precision of such methods are not good. The third kind of PCR amplification method is to transfer the PCR reaction system between fixed temperature areas and to transfer the PCR tube between three water baths. Some existing systems transfer the PCR reaction chamber between different temperature zones of a metal bath or water bath, oil bath, or flow the PCR reaction solution between different temperature zones by microfluidic technology. This method does not involve a temperature-increasing or decreasing process, and it is even possible to shorten the PCR process including reverse transcription and pre-denaturation to within 15 minutes. However, the method has a plurality of problems in realization, firstly, the reaction consumable material is difficult to design, the mechanical transfer and the full contact of the temperature zone require that the reaction consumable material has certain strength, and meanwhile, the heat conduction performance is good; secondly, the transfer of reaction consumables among multiple temperature areas needs complicated mechanical motion control, and the flux, the automation degree and the size of an instrument are difficult to be compatible, so that a rapid nucleic acid amplification system based on a temperature area transfer mode is not clinically applied at present.
Disclosure of Invention
The invention aims to provide a rapid nucleic acid amplification system, which enables nucleic acid amplification reaction solutions in reaction consumables to be transferred between different temperature ranges to realize rapid nucleic acid amplification so as to solve the technical problems of the existing nucleic acid amplification system or time consumption or difficulty in realizing.
In order to achieve the above purpose, the invention provides a rapid nucleic acid amplification system, which comprises a reaction consumable and a temperature control component, wherein a PCR reaction area is arranged on the reaction consumable, and the rapid nucleic acid amplification system is characterized in that:
the PCR reaction area is arranged in the area of the reaction consumable close to the edge;
the temperature control assembly comprises two or more groups of PCR temperature zone control units which are circumferentially distributed, the PCR temperature zone control units control the temperature of a PCR reaction zone on a reaction consumable, and the temperatures controlled by different PCR temperature zone control units can be the same or different; the nucleic acid amplification system further comprises a rotary support assembly comprising a support member and a rotary motor; the supporting component is horizontally arranged and used for supporting and fixing reaction consumable materials; the rotating motor is used for controlling the rotation of the supporting component, and the PCR reaction area of the reaction consumable fixed on the supporting component is subjected to rapid nucleic acid amplification by rotating and changing temperature areas between two or more groups of circumferentially arranged PCR temperature area control units.
Further, the supporting component is a turntable; the reaction consumable is a disc-shaped reaction consumable or a plurality of fan-shaped reaction consumable, and the fan-shaped reaction consumable is circumferentially distributed on the supporting component.
In some embodiments, the disc-shaped reaction consumable comprises one or more PCR units uniformly distributed around the circumference, each PCR unit comprises a closable sample inlet and a PCR reaction area, the sample inlet and the PCR reaction area are communicated through a flow channel, and each PCR reaction area comprises one or more PCR reaction chambers uniformly distributed around the circumference; the PCR reaction chambers are all arranged on the circumference of the outermost ring of the reaction consumable.
In some embodiments, the PCR unit of the disc-shaped reaction consumable further comprises a PCR premix chamber or a plurality of PCR premix chambers in one-to-one correspondence with the plurality of PCR reaction chambers, the PCR premix chambers being in communication with the sample addition port and the PCR reaction chambers through the flow channels.
In some embodiments, the sector reaction consumable has a closable loading port and a PCR reaction region coupled to the loading port, each PCR reaction region comprising one or more circumferentially distributed PCR reaction chambers; the PCR reaction chamber is arranged on the circumference of the outermost ring of the reaction consumable.
In some embodiments, a PCR premix chamber or a plurality of PCR premix chambers corresponding to the PCR reaction chambers one to one are arranged between the sample inlet of the sector reaction consumable and the PCR reaction chambers, and the sample inlet, the PCR premix chambers and the PCR reaction chambers are communicated.
In some embodiments, the PCR temperature zone control unit may be a temperature control module having a notch at a front end thereof, where a local air heating zone is formed by a constant temperature air flow, and may provide a temperature required for a PCR reaction to each PCR reaction chamber in the PCR reaction zone rotationally embedded therein.
In some embodiments, the PCR temperature zone control unit may also include two temperature control modules installed at the upper and lower parts of the reaction consumable material, and the two temperature control modules control the same temperature. The two temperature control modules are driven by the driving structure to clamp the PCR reaction area, and provide one temperature required by the PCR reaction for each PCR reaction cavity in the PCR reaction area.
In some embodiments, the PCR temperature zone control unit includes a temperature control module for controlling annealing extension, and the upper part or the lower part of the temperature control module, or the upper temperature control module, or the lower temperature control module is provided with a fluorescence detection unit, when the PCR reaction zone rotates to the temperature control module, the fluorescence detection unit performs real-time fluorescence detection, so as to realize a nucleic acid quantitative detection function.
In some embodiments, the turntable is provided with a clamping mechanism or a fixing clamping groove, so that the disc-shaped or fan-shaped reaction consumable material can be clamped and fixed on the turntable for rotary positioning or high-speed centrifugation.
In some embodiments, the rotating electrical machine is provided with a coded disc, and the absolute position and the rotating speed of the reaction consumable material can be subjected to negative feedback control. The speed curve of the rotating speed control can be triangular wave, sawtooth wave, sine wave or parabola, and the frequency and amplitude of the rotating speed curve can be dynamically adjusted, so that the rapid and efficient mixing of different volumes of liquid in different cavities in the reaction box is realized.
In some embodiments, the rotating motor can drive the reaction consumable to rotate unidirectionally at a high speed, and the rotating speed can reach 2000-10000 revolutions per minute. The liquid such as the sample, the template, the reagent or the amplification reaction solution can be driven to flow from the cavity with the reaction consumable close to the center to the cavity with the reaction consumable far from the center at a proper rotating speed.
Compared with the prior art, the invention has the beneficial effects that:
1. the invention discloses a rapid nucleic acid amplification system, which provides a disc-shaped or fan-shaped reaction consumable, wherein nucleic acid amplification reaction solution finally enters a PCR reaction cavity distributed on the periphery of the consumable, and rapid transfer is realized between PCR temperature zone control units under the control of the system. Since there is no temperature rise and drop process, it is theoretically possible to complete the PCR process including reverse transcription and pre-denaturation even within 15 minutes.
2. The design of the disc-shaped or fan-shaped reaction consumable material can enable the nucleic acid amplification reaction solution to be spread, and is more beneficial to temperature conversion of the nucleic acid amplification reaction solution.
3. The rapid nucleic acid amplification system of the invention performs switching between different temperature areas through the rotation of the reaction consumable in the circumferential direction, and has the characteristics of high speed, low noise, accurate positioning, good repeatability and the like compared with the existing reaction consumable temperature area switching mode; in addition, centrifugal fluid driving and reaction solution mixing functions can be realized, and the system integration level is improved.
4. Compared with the temperature control modes of water bath, oil bath and metal bath in the background technology, the temperature control mode of the local air bath of the PCR temperature zone control unit of the rapid nucleic acid amplification system has the characteristics of high heat exchange efficiency, high heat balance speed, non-contact, good uniformity, no pollution, no loss and the like, and can reduce the non-specificity in the PCR amplification process.
5. The reaction consumable disclosed by the invention is flexible in design, can realize multiple detection of one sample and multiple detection of a plurality of samples, has a reagent premixing function, and is wider in application range.
Drawings
FIG. 1 is a schematic plan view of a disc-shaped reaction consumable material according to an embodiment of the present invention;
FIG. 2 is a schematic diagram of a rapid nucleic acid amplification system according to an embodiment of the present invention;
FIG. 3 shows a top view of a rapid nucleic acid amplification system in accordance with one embodiment of the present invention;
FIG. 4 is a schematic plan view of a second exemplary embodiment of a reactive consumable material;
FIG. 5 is a schematic diagram of a second rapid nucleic acid amplification system according to an embodiment of the present invention;
FIG. 6 shows a top view of a second rapid nucleic acid amplification system in accordance with an embodiment of the present invention;
FIGS. 7 and 8 are schematic diagrams showing the switching of temperature control modules of a third rapid nucleic acid amplification system according to an embodiment of the present invention;
FIG. 9 is a schematic plan view of a second embodiment of a disc-shaped reaction consumable of the present invention;
FIG. 10 is a schematic plan view of a third exemplary disc-shaped reaction consumable material of the present invention;
FIG. 11 is a schematic plan view of a second exemplary embodiment of a fan-shaped reactive consumable of the present invention.
Wherein the reference numerals are as follows: 21. 31, 41, 51, 61, 71-reactive consumables; 22. 32, 52, 62, 72-spindle detents; 23. 33, 53, 63, 73-PCR units; 24. 34, 54, 64, 74-sample addition port; 25. 35, 65, 75-PCR premix chamber; 26. 36, 56, 66, 76-PCR reaction chambers; 27. 37, 67, 77-PCR reaction region; 28. 38, 58, 68-loading channels; 29. 39, 69, 79-split flow paths; 210. 310-a turntable; 211. 311-rotating electric machine; 212. 312-PCR temperature zone control unit; 12a 1 -a temperature control module on the first PCR temperature zone control unit; 12a 2 -a first PCR temperature zone control unit lower temperature control module; 12b 1 -a temperature control module on the second PCR temperature zone control unit; 12b 2 -a second PCR temperature zone control unit lower temperature control module; 213. 313-rotating a stationary shaft; 314414-fiber optic probe.
Detailed Description
Embodiment one:
in the following, referring to fig. 1, 2 and 3, a specific embodiment of the present invention will be described, and the rapid nucleic acid amplification system shown in this example can load a disc-shaped reaction consumable and achieve rapid amplification of a plurality of nucleic acid samples simultaneously by using a local air bath.
As shown in fig. 1, the disc-shaped reaction consumable 21 in this embodiment includes a spindle slot 22 and six PCR units 23, each PCR unit 23 includes a sample inlet 24, a PCR pre-mixing chamber 25 pre-packaged with dry powder reagent, and three PCR reaction chambers 26, and the three PCR reaction chambers 26 form a PCR reaction area 27; the sample adding port 24 is communicated with the PCR premixing cavity 25 through a sample adding flow channel 28; the PCR premixing chamber 25 is communicated with three PCR reaction chambers 26 through a liquid separation flow channel 29.
As shown in fig. 2 and 3, the rotary amplifying device in this embodiment includes a turntable 210, a rotary motor 11, and six PCR temperature zone control units 212 (two of which are not shown in fig. 2 for convenience of explanation). The turntable 210 is provided with a rotation fixing shaft 213 for loading and fixing the reaction consumable 21. The rotating motor 211 is connected to the turntable 210, and can drive the turntable 210 and the loaded reaction consumable 21 to rotate or centrifuge at a high speed so as to drive the liquid in the reaction consumable 21 to flow. The front ends of the six PCR temperature zone control units 212 are provided with notches into which the six PCR reaction areas 27 on the reaction consumable 21 can be inserted. Meanwhile, six PCR temperature zone control units 212 provide constant temperature air flow through the notch to form a local air bath, wherein three PCR temperature zone control units 212 provide the same temperature (higher temperature), the other three PCR temperature zone control units 212 provide the same temperature (lower temperature), the high and low temperature PCR temperature zone control units 212 are alternately distributed to form different temperature zones required by the PCR reaction, and the PCR temperature zone control units 212 can adopt heating wires or heating films.
The nucleic acid amplification process of this example is: six identical or different liquid nucleic acid samples are added from the sample addition port 24 to the reaction consumable 21, the added samples flow into the PCR premix chamber 25 via the sample addition channel 28, then the sample addition port 24 will be sealed, and the reaction consumable 21 will become a closed system. The closed reaction consumable 21 is loaded onto the turntable 210 of the rotary amplification device, and specifically is operated to fix the reaction consumable on the rotary fixing shaft 213 of the turntable through the rotary shaft clamping groove 22 thereof. After the reaction consumable 21 is loaded, the rotating motor 211 drives the reaction consumable 21 to rotate reciprocally, so that the nucleic acid sample added from the sample adding port flows into the PCR premixing cavity 25 and is fully and uniformly mixed with the PCR dry powder reagent in the PCR premixing cavity 25, then the rotating motor 211 drives the reaction consumable 21 to centrifugally rotate at a high speed, so that the mixed solution in the PCR premixing cavity 25 enters the three PCR reaction cavities 26 through the liquid separating flow channels 29, the rotary amplification device controls the six PCR temperature zone control units 212 to move to positions corresponding to the six PCR reaction zones 27 of the reaction consumable 21, and each PCR reaction zone 27 is embedded into a front end notch of the corresponding PCR temperature zone control unit 212. Finally, the rotating motor 211 drives the reaction consumable 21 to reciprocally rotate among the three groups of high and low temperature PCR temperature zone control units 212 to provide different temperatures required by the PCR reaction so as to realize rapid amplification of the nucleic acid sample.
Embodiment two:
in the following, referring to fig. 4, 5 and 6, a specific embodiment of the present invention will be described, in which the rapid nucleic acid amplification system shown in this embodiment can load six fan-shaped reaction consumables and achieve rapid amplification of a plurality of nucleic acid samples simultaneously by using a local air bath.
As shown in fig. 4, the fan-shaped reaction consumable 31 in this embodiment includes two spindle clamping grooves 32 and a PCR unit 33, the PCR unit 33 includes a sample inlet 34, a PCR pre-mixing chamber 35 pre-packaged with dry powder reagent, and three PCR reaction chambers 36, and the three PCR reaction chambers 36 form a PCR reaction area 37; the sample inlet 34 is communicated with the PCR premixing cavity 35 through a sample inlet channel 38, and the PCR premixing cavity 35 is communicated with the PCR reaction cavity 36 through a liquid separation channel 39.
As shown in fig. 5 and 6, the rotary amplifying device in this embodiment includes a turntable 310, a rotary motor 311 and six PCR temperature zone control units 312 (two of which are not shown in fig. 5 for convenience of explanation). Twelve rotation fixing shafts 313 are provided on the turntable 310, and two rotation fixing shafts 313 are provided as a group for loading and fixing one reaction consumable 31. The rotating motor 311 is connected to the turntable 310, and can drive the turntable 310 and the loaded reaction consumable 31 to rotate or centrifuge at a high speed so as to drive the liquid in the reaction consumable 31 to flow. The front ends of the six PCR temperature zone control units 312 are provided with notches into which the PCR reaction zones 37 on the reaction consumable 31 can be inserted. Meanwhile, six PCR temperature zone control units 312 provide constant temperature air flow through the notch to form a local air bath, wherein three PCR temperature zone control units 312 provide the same temperature (higher temperature), the other three PCR temperature zone control units 312 provide the same temperature (lower temperature), the high and low temperature PCR temperature zone control units 312 are alternately distributed to form different temperature zones required by the PCR reaction, and the PCR temperature zone control units 312 can adopt heating wires or heating films. Meanwhile, three optical fiber probes 314 are installed on the three low-temperature PCR temperature zone control units 312, and are used for performing multipath fluorescence detection on the PCR reaction.
The nucleic acid amplification process of this example is: liquid nucleic acid sample is added from the sample addition port 34 to the reaction consumable 31, the added sample flows into the PCR premix chamber 35 through the sample addition flow channel 38, then the sample addition port 34 is sealed, and the reaction consumable 31 becomes a closed system. The six reaction consumables 31 to which the same or different nucleic acid samples are added and which are sealed are then loaded onto the turntable 310 of the rotary amplification device, specifically operating to fix the six reaction consumables 31 on the six sets of rotation fixing shafts 312 of the turntable through their respective two rotation shaft clamping grooves 32, respectively. After the six reaction consumable materials 31 are loaded, the rotating motor 311 drives the six reaction consumable materials 31 to rotate reciprocally, so that the nucleic acid sample in the PCR premixing cavity 35 and the PCR dry powder reagent are fully and uniformly mixed, and then the rotating motor 311 drives the six reaction consumable materials 31 to rotate centrifugally at a high speed, so that the mixed solution in the PCR premixing cavity 35 enters the three PCR reaction cavities 36 through the liquid separating flow channels 39. The rotary amplification device controls the six PCR temperature zone control units 312 to move to positions corresponding to the PCR reaction regions 37 of the six reaction consumables 31, so that each PCR reaction region 37 is embedded in a front end notch of the PCR temperature zone control unit 312 opposite thereto. Finally, the rotating motor 311 drives the reaction consumable 31 to reciprocally rotate among the three groups of high-temperature PCR temperature zone control units 312 to provide different temperatures required by PCR reaction so as to realize rapid nucleic acid amplification; every time the PCR reaction area 37 is embedded into the front end notch of the three low-temperature PCR temperature area control units 312, the three optical fiber probes 314 on the low-temperature PCR temperature area control units 312 can perform multiplex fluorescence detection on the PCR reaction, so that the nucleic acid fluorescence detection is realized.
Embodiment III:
in the following, a specific embodiment of the rotary amplification apparatus according to the present invention will be described with reference to fig. 7 and 8, in which the rapid nucleic acid amplification system shown in this embodiment employs a temperature control module to achieve rapid amplification of a single or multiple nucleic acid samples.
As shown in fig. 7 to 8, the rotary amplification apparatus in this embodiment includes a turntable (not shown), a rotary motor 411 and two PCR temperature zone control units: the first PCR temperature zone control unit comprises an upper temperature control module 12a 1 And a lower temperature control module 12a 2 The second PCR temperature zone control unit comprises an upper temperature control module 12b 1 And a lower temperature control module 12b 2 The two groups of upper and lower temperature control modules are oppositely arranged at the upper part and the lower part of the reaction consumable, and the same temperature is controlled. The two groups of temperature control modules can be driven by the driving structure to clamp the PCR reaction area, and provide one temperature required by the PCR reaction for each PCR reaction cavity in the PCR reaction area; meanwhile, the upper temperature control module 12b of the second PCR temperature zone control unit 1 Five fiber probes 414 are mounted for multiplexed fluorescence detection of the PCR reactions.
The specific implementation manner of the embodiment is as follows: after the reaction consumable 41 is loaded into the rotary amplification device, the rotary motor 411 drives the reaction consumable 41 to reciprocally rotate to complete the uniform mixing of the sample in the PCR premixing cavity and the packaged dry powder reagent; after mixing, the rotating motor 411 drives the reaction consumable 41 to move back and forth between the first PCR temperature zone control unit and the second PCR temperature zone control unit, so as to perform PCR amplification, wherein the first PCR temperature zone control unit and the second PCR temperature zone control unit can adopt heating wires or heating films. As shown in fig. 7, when the PCR reaction region of the reaction consumable 41 is moved to the first PCR temperature zone control unit under the driving of the rotating motor 411, the upper temperature control module 12a of the first PCR temperature zone control unit 1 Temperature control module12a 2 The driving structure drives and clamps the PCR reaction area and controls the temperature; as shown in fig. 8, when the PCR reaction region of the reaction consumable 41 is moved to the second PCR temperature zone control unit under the driving of the rotating motor 411, the upper temperature control module 12b of the second PCR temperature zone control unit 1 Lower temperature control module 12b 2 The driving structure drives the PCR reaction region to be clamped and the temperature is controlled, so that different temperatures required by the PCR reaction are provided, and the rapid amplification of nucleic acid is realized; upper temperature control module 12b of each second PCR temperature zone control unit 1 Lower temperature control module 12b 2 When the temperature control is performed by clamping the PCR reaction area, the upper temperature control module 12b 1 The five optical fiber probes 414 can perform multiplex fluorescence detection on the PCR reaction, so that the nucleic acid fluorescence detection is realized.
OTHER EMBODIMENTS
FIG. 9 is a schematic plan view of a second embodiment of a disc-shaped reaction consumable of the present invention. In this embodiment, the disc-shaped reaction consumable 51 includes a spindle clamping groove 52 and six PCR units 53, and each PCR unit 53 includes a sample inlet 54 and a PCR reaction chamber 56; the sample inlet 54 communicates with the PCR reaction chamber 56 via a sample inlet channel 58. Six identical or different prepared PCR reaction liquids can be added into the disc-shaped reaction consumable to carry out rapid amplification of nucleic acid.
FIG. 10 is a schematic plan view of a third exemplary disc-shaped reaction consumable material according to the present invention. In this embodiment, the disc-shaped reaction consumable 61 includes a spindle slot 62 and four PCR units 63, each PCR unit 63 includes a sample inlet 64, three PCR pre-chambers 65 pre-packaged with the same or different dry powder reagents, and three PCR reaction chambers 66, and the three PCR reaction chambers 66 form a PCR reaction area 67. The sample inlet 64 communicates with three PCR premixing chambers 65 through sample addition flow passages 68, and the three PCR premixing chambers 65 communicate with three PCR reaction chambers 66 through three flow passages 69. Four identical or different nucleic acid samples can be added to the four PCR units 63 of the disc-shaped reaction consumable, and three identical or different PCR amplification reactions can be performed in the three PCR reaction chambers 66 of each PCR unit 63.
FIG. 11 is a schematic plan view of a second exemplary embodiment of a fan-shaped reactive consumable of the present invention. In this embodiment, the sector reaction consumable 71 includes two spindle clamping grooves 72 and one PCR unit 73. The PCR unit 73 comprises a sample inlet 74, three PCR premix chambers 75 pre-packed with the same or different PCR dry powder reagents, and three PCR reaction chambers 76, the three PCR reaction chambers 76 forming a PCR reaction area 77. The sample inlet 74 communicates with three PCR premixing chambers 75 through sample addition flow passages 78, and the three PCR premixing chambers 75 communicate with three PCR reaction chambers 76 through three flow passages 79. Four sector reaction consumables 71 can be loaded in the rotary amplification device, each sector reaction consumable 71 can be added with a nucleic acid sample, and each nucleic acid sample can carry out three identical or different PCR amplification reactions.
The nucleic acid sample added to the sample addition port in the first embodiment, the second embodiment, the third embodiment, and the fig. 10 and 11 of the present invention may be an extracted and purified nucleic acid sample, or may be an original nucleic acid sample, and when the nucleic acid sample is the original nucleic acid sample, the nucleic acid sample may be a purified nucleic acid sample when flowing into the premix chamber by designing the structure of the flow channel between the reaction consumable and the premix chamber.
It is to be understood that the above examples of the present invention are provided by way of illustration only and not by way of limitation of the embodiments of the present invention. Other variations or modifications may be made in the light of the above description, and need not be, nor should it be exhaustive of all embodiments. While remaining within the scope of the invention, obvious variations or modifications are incorporated by reference herein.
Claims (8)
1. The utility model provides a quick nucleic acid amplification system, includes reaction consumable and temperature control subassembly, be provided with PCR reaction area, its characterized in that on the reaction consumable:
the PCR reaction area is arranged in the area of the reaction consumable close to the edge;
the temperature control assembly comprises two or more groups of PCR temperature zone control units which are circumferentially distributed, and the PCR temperature zone control units control the temperature of a PCR reaction zone on the reaction consumable;
the front ends of the plurality of groups of circumferentially arranged PCR temperature zone control units are provided with notches, reaction areas on reaction consumable materials can be embedded into the notches, meanwhile, the temperature zone control units provide constant-temperature air flow through the notches to form a local air bath, part of the PCR temperature zone control units provide higher temperature, and the other part of the PCR temperature zone control units provide lower temperature, and the high-temperature and low-temperature PCR temperature zone control units are alternately distributed to form different temperature zones required by PCR reaction;
the nucleic acid amplification system further comprises a rotary support assembly comprising a support member and a rotary motor; the supporting component is horizontally arranged and is used for supporting and fixing reaction consumables; the rotating motor is used for controlling the rotation of the supporting component, one or more PCR reaction areas of the reaction consumable fixed on the supporting component change temperature areas in a rotating mode between two or more groups of circumferentially arranged PCR temperature area control units, and nucleic acid amplification is achieved.
2. The rapid nucleic acid amplification system of claim 1, wherein:
the supporting component is a turntable; the reaction consumable is a disc-shaped reaction consumable or a plurality of fan-shaped reaction consumable, and the fan-shaped reaction consumable is circumferentially distributed on the supporting component.
3. The rapid nucleic acid amplification system of claim 2, wherein:
the disc-shaped reaction consumable comprises one or more PCR units uniformly distributed on the circumference, each PCR unit comprises a closable sample adding port and a PCR reaction area, and the sample adding ports are communicated with the PCR reaction areas through flow channels;
each PCR reaction area comprises one or more PCR reaction chambers uniformly distributed on the circumference.
4. The rapid nucleic acid amplification system of claim 3, wherein:
each PCR unit of the disc-shaped reaction consumable further comprises a PCR premix chamber or a plurality of PCR premix chambers corresponding to the plurality of PCR reaction chambers one by one, and the PCR premix chambers are communicated with the sample adding port and the PCR reaction chambers through flow channels.
5. The rapid nucleic acid amplification system of claim 2, wherein:
the sector reaction consumable comprises a sample adding port which can be closed and a PCR reaction area connected with the sample adding port, wherein the PCR reaction area comprises one or more circumferentially distributed PCR reaction cavities.
6. The rapid nucleic acid amplification system of claim 5, wherein:
a PCR premix cavity or a plurality of PCR premix cavities corresponding to the PCR reaction cavities one by one are arranged between the sample adding port of the sector reaction consumable and the PCR reaction cavities, and the sample adding port, the PCR premix cavities and the PCR reaction cavities are communicated.
7. The rapid nucleic acid amplification system of claim 2, wherein:
the rotary table is provided with a clamping mechanism or a fixing clamping groove for clamping and fixing the disc-shaped or fan-shaped reaction consumable clamp on the rotary table.
8. The rapid nucleic acid amplification system of claim 1, wherein:
the rotary motor is provided with a coding disc, the coding disc carries out negative feedback control on the absolute position and the rotating speed of the reaction consumable, and the speed curve is triangular wave, sawtooth wave, sine wave or parabola.
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| CN108342301B (en) * | 2018-05-18 | 2023-11-28 | 西安天隆科技有限公司 | Nucleic acid extraction and purification device |
| CN109046477B (en) * | 2018-07-23 | 2023-12-19 | 深圳市呈晖医疗科技有限公司 | Centrifugal Microfluidic Device |
| WO2020124510A1 (en) * | 2018-12-20 | 2020-06-25 | 广州市华南医学研究中心 | Extremly rapid nucleic acid amplification method, apparatus and use thereof |
| CN111607509B (en) * | 2019-02-22 | 2023-04-07 | 西安天隆科技有限公司 | Automatic amplification and detection device for digital PCR |
| CN110804540B (en) * | 2019-10-22 | 2023-03-31 | 深圳市刚竹医疗科技有限公司 | Microfluidic temperature control system and nucleic acid analysis system |
| CN110747294B (en) * | 2019-11-08 | 2021-06-04 | 宁波胤瑞生物医学仪器有限责任公司 | Method for regulating and controlling digital nucleic acid amplification instrument |
| CN110699243B (en) * | 2019-11-11 | 2022-09-23 | 西南医科大学附属医院 | Multifunctional nucleic acid detection device |
| CN111004708B (en) * | 2019-12-24 | 2023-05-30 | 中国科学院长春光学精密机械与物理研究所 | PCR temperature cycle control method and rotary driving type PCR temperature cycle control device |
| CN112094740A (en) * | 2020-09-25 | 2020-12-18 | 壹宏(深圳)基因有限公司 | PCR amplification method and PCR amplification device |
| EP4328291A1 (en) * | 2021-05-28 | 2024-02-28 | Chin Hung Wang | Polymerase chain reaction device |
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