Portable convection PCR (Polymerase Chain Reaction) amplification instrument

Abstract

The utility model discloses a portable convection PCR (Polymerase Chain Reaction) amplification instrument, relates to a PCR technology, and solves the technical problem that an existing PCR amplification device needs a relatively long time to repeatedly heat and cool. The device comprises a shell, an internal control circuit is arranged in the shell, a cooling fan is installed on the shell, a convection heating module is installed on the shell, and the cooling fan and the convection heating module are both electrically connected with the internal control circuit; the convection heating module comprises an upper constant-temperature control unit, a middle unit and a lower constant-temperature control unit, the upper constant-temperature control unit is installed above the middle unit, the middle unit is installed above the lower constant-temperature control unit, and an air heat insulation layer is arranged between the middle unit and the lower constant-temperature control unit; and a plurality of convection heating chambers are arranged among the upper constant temperature control unit, the middle unit and the lower constant temperature control unit. Compared with the traditional PCR instrument, the PCR instrument disclosed by the utility model does not need to be repeatedly heated and cooled and is short in reaction time.

Description

Technical Field

[0001] The utility model relates to PCR technology, and more specifically, to a convenient convection PCR amplification instrument. Background Art

[0002] Polymerase chain reaction (PCR) is a technology for enzymatically amplifying specific DNA fragments in vitro. Under the catalysis of thermostable DNA polymerase, the nucleic acid fragment defined by a pair of oligonucleotide primers is amplified. One cycle consists of three reaction stages: denaturation, annealing, and extension. These three stages can be repeated for several cycles and have the advantages of strong specificity, high sensitivity, and simple operation. PCR technology can be used not only for basic research, but also for disease diagnosis. Since Manz, Widmer and others proposed the concept of microfluidic total analysis system (Miniaturized Total Analysia Systems, μTAS) in 1990, biochemical analysis technology based on microfluidic chips has developed very rapidly. Biochemical analysis technology based on microfluidic chips has truly moved from the theoretical level to the practical application level.

[0003] Traditional PCR requires repeated heating and cooling of samples in a precise control circuit system for temperature cycling, which usually requires 30-40 cycles. The entire reaction takes 2-3 hours, most of which is taken up by repeated heating and cooling. The energy consumption is relatively high, and the traditional PCR device is large in size, making it difficult to achieve high throughput and rapid detection at low cost. Utility Model Content

[0004] The technical problem to be solved by the utility model is to provide a portable convection PCR amplification instrument with low cost, convenient maintenance, simple structural design and high integration, in order to solve the technical problem that the existing PCR amplification device needs a long time to repeatedly heat up and cool down.

[0005] The utility model discloses a portable convection PCR amplification instrument, which comprises a housing, an internal control circuit is arranged in the housing, a heat dissipation fan is installed on the housing, a convection heating module is installed on the housing, wherein the convection heating module is provided with a plurality of reaction holes, reaction containers are inserted into the reaction holes, the reaction containers contain corresponding PCR reagents, and the heat dissipation fan and the convection heating module are both electrically connected to the internal control circuit;

[0006] The convection heating module includes an upper thermostatic control unit, an intermediate unit and a lower thermostatic control unit to maintain the temperature gradient required for convection PCR, so that the sample DNA undergoes thermal cycling under the dual effects of temperature difference and gravity field. The upper thermostatic control unit is installed above the intermediate unit, the intermediate unit is installed above the lower thermostatic control unit, an air insulation layer is provided between the intermediate unit and the lower thermostatic control unit, and a plurality of convection heating chambers are provided between the upper thermostatic control unit, the intermediate unit and the lower thermostatic control unit, and the convection heating chambers are connected to the air insulation layer.

[0007] As a further improvement, the upper constant temperature control unit is provided with a plurality of first reaction holes, the middle unit is provided with a plurality of second reaction holes corresponding one-to-one to the first reaction holes, and the lower constant temperature control unit is provided with a plurality of third reaction holes corresponding one-to-one to the second reaction holes. The first reaction holes and the third reaction holes are respectively in close contact with the reaction container for effective contact and heat transfer. The aperture of the second reaction hole is larger than that of the first reaction hole and the third reaction hole. The first reaction hole, the second reaction hole and the third reaction hole constitute a stepped convection heating chamber with a large middle portion and small ends.

[0008] Furthermore, the upper constant temperature control unit is provided with a first positioning block and a second positioning block which are integrally formed therewith, the first positioning block being provided with a first temperature acquisition sensor element interface, the first temperature acquisition sensor element being installed in the first temperature acquisition sensor element interface, the second positioning block being provided with a first heating element interface, the first heating element being installed in the first heating element interface, the first temperature acquisition sensor element and the first heating element being both electrically connected to the internal control circuit.

[0009] Furthermore, the aperture depth of the first reaction hole is 1-4 mm, the aperture depth of the second reaction hole is 4-8 mm, the aperture depth of the third reaction hole is 1-4 mm, and the thickness of the air insulation layer is 4-8 mm.

[0010] Furthermore, the intermediate unit is provided with a first groove and a second groove, the first positioning block is plugged into the first groove, and the second positioning block is plugged into the second groove.

[0011] Furthermore, a second temperature acquisition sensor element interface is provided in the lower constant temperature control unit, a second temperature acquisition sensor element is installed in the second temperature acquisition sensor element interface, a second heating element is provided in the lower constant temperature control unit, a second heating element is installed in the second heating element interface, and the second temperature acquisition sensor element and the second heating element are both electrically connected to the internal control circuit.

[0012] Furthermore, a display module is fixedly installed on the top of the shell, a first switch and a second switch are provided on one side of the shell, and a power switch and a power socket are provided on the other side of the shell, and the display module, the first switch and the second switch are all connected to the internal control circuit.

[0013] Furthermore, the upper constant temperature control unit, the middle unit and the lower constant temperature control unit are all made of 6061 aluminum alloy plates.

[0014] Furthermore, the thickness of the upper constant temperature control unit is 1-4 mm, the thickness of the middle unit is 1-10 mm, and the thickness of the lower constant temperature control unit is 1-20 mm.

[0015] Furthermore, the heat dissipation fan is located on one side of the convection heating module.

[0016] The above-mentioned portable convection PCR amplifier is used for nucleic acid amplification, and the steps are as follows:

[0017] Step 1: Extract nucleic acid and prepare samples to be amplified;

[0018] Step 2: Start the portable convection PCR amplification instrument, preheat the convection heating module, and then insert the reaction vessel into it;

[0019] Step 3: The reaction vessel is heated in the thermal cycler for several minutes;

[0020] Step 4: After the reaction is completed, take out the reaction container and place it in a cold environment for cooling;

[0021] Step 5: Analyze the amplified products in the reaction vessel by agarose gel electrophoresis.

[0022] Beneficial Effects

[0023] The advantages of the utility model are:

[0024] The utility model sets a convection heating module so that the bottom of the reaction container containing the sample to be amplified reaches the denaturation temperature required for the PCR amplification reaction, the top of the reaction container containing the sample to be amplified reaches the annealing temperature required for the PCR amplification reaction, and the middle part of the reaction container containing the sample to be amplified reaches the extension temperature required for the PCR amplification reaction for a longer time. Compared with the traditional PCR instrument, it does not need to repeatedly heat up and cool down, the reaction time is short, and the work efficiency can be improved. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 It is a schematic diagram of the overall structure of the amplifier of the utility model;

[0026] Figure 2a This is a schematic diagram of the overall structure of the convection heating module of the utility model;

[0027] Figure 2b This is the installation diagram of the temperature collection sensor element and the heating element of the utility model;

[0028] Figure 3 This is a disassembled diagram of the convection heating module of the utility model;

[0029] Figure 4 This is a schematic diagram of the convection heating chamber structure of the utility model;

[0030] Figure 5 It is a cross-sectional view of the convection heating module of the utility model;

[0031] Figure 6 It is a top view of the convection heating module of the utility model;

[0032] Figure 7 It is a schematic diagram of the structure of an existing reaction vessel;

[0033] Figure 8 This is the gel electrophoresis diagram showing the relationship between the convective PCR amplification effect and the amplification reaction time.

[0034] Wherein: 1-housing, 2-display module, 3-cooling fan, 4-convection heating module, 5-first switch, 6-second switch, 7-power switch, 8-power socket, 9-reaction container, 10-air insulation layer, 11-convection heating chamber, 12-placement port, 13-upper constant temperature control unit, 131-first temperature acquisition sensor element interface, 132-first heating element interface, 133-first reaction hole, 134-second positioning block, 135-first positioning block, 14-middle unit, 141-second reaction hole, 142-first groove, 143-second groove, 15-lower constant temperature control unit, 151-second temperature acquisition sensor element interface, 152-second heating element interface, 153-third reaction hole, 16-cylindrical gasket, 17-first temperature acquisition sensor element, 18-first heating element, 19-second temperature acquisition sensor element, 20-second heating element. DETAILED DESCRIPTION

[0035] The utility model is further described below in conjunction with the embodiments, but does not constitute any limitation to the utility model. Any limited modifications made by anyone within the scope of the claims of the utility model are still within the scope of the claims of the utility model.

[0036] See also Figure 1-Figure 8The utility model discloses a portable convection PCR amplification instrument, which comprises a shell 1, an internal control circuit is installed in the shell 1, a bracket is fixedly installed in the shell 1, a placement opening 12 is opened on the shell 1, a display module 2 and a cooling fan 3 are fixedly installed on the shell 1, a convection heating module 4 is fixedly installed on one side of the bracket close to the placement opening 12, and the display module 2 and the convection heating module 4 are both electrically connected to the internal control circuit.

[0037] The convection heating module 4 includes an upper thermostatic control unit 13, an intermediate unit 14 and a lower thermostatic control unit 15. The upper thermostatic control unit 13 is installed above the intermediate unit 14. The intermediate unit 14 is fixedly connected to the lower thermostatic control unit 15 through a cylindrical gasket 16. An air insulation layer 10 for heat insulation is provided between the intermediate unit 14 and the lower thermostatic control unit 15. The air insulation layer 10 can be used for heat insulation. The upper thermostatic control unit 13 and the lower thermostatic control unit 15 are used for heating.

[0038] The upper constant temperature control unit 13 is provided with a plurality of first reaction holes 133 for placing reaction containers 9. In the present embodiment, 6 (3X2) first reaction holes 133 are provided. The middle unit 14 is provided with 6 second reaction holes 141 corresponding one-to-one to the first reaction holes 133. The lower constant temperature control unit 15 is provided with 6 third reaction holes 153 corresponding one-to-one to the second reaction holes 141, so as to facilitate the insertion of the reaction container 9 containing the sample to be amplified for the convection PCR amplification reaction.

[0039] These reaction wells are Figure 6 The 2 rows and 3 columns shown are arranged in an array. The spacing between adjacent reaction wells is 18 mm. The material of the upper constant temperature control unit 13, the middle unit 14, and the lower constant temperature control unit 15 is 6061 aluminum alloy with good ductility and strong ductility. A polytetrafluoroethylene plate is placed at the bottom for heat insulation.

[0040] In this solution, the size of the convection heating module 4 is 60 mm*60 mm, the thickness of the upper constant temperature control unit 13 is 2 mm, the thickness of the middle unit 14 is 5 mm, and the thickness of the lower constant temperature control unit 15 is 10 mm.

[0041] The first reaction hole 133, the second reaction hole 141 and the third reaction hole 153 constitute a convection heating chamber 11. The convection heating chamber 11 can improve the extension effect of thermal convection PCR. The convection heating chamber 11 is connected to the air insulation layer 10.

[0042] The aperture depth of the third reaction hole 153 is 2 mm. The insulation thickness of the air insulation layer 10 is 4 mm, and the diameter of the cross section of the convection heating chamber 11 is 7.96 mm. The convection heating chamber 11 and the upper constant temperature control unit 13 and the lower constant temperature control unit 15 can conduct step-by-step temperature changes to enhance the extension effect of the thermal convection PCR.

[0043] like Figure 2b As shown, the upper thermostatic control unit 13 is installed with a first temperature acquisition sensor element 17 and a first heating element 18, and the lower thermostatic control unit 15 is installed with a second temperature acquisition sensor element 19 and a second heating element 20. The first temperature acquisition sensor element 17, the first heating element 18, the second temperature acquisition sensor element 19 and the second heating element 20 are all electrically connected to the internal control circuit. The upper thermostatic control unit 13 is provided with a first positioning block 135 and a second positioning block 134 which are an integral structure therewith. The first positioning block 135 is provided with a first temperature acquisition sensor element interface 131, and the first temperature acquisition sensor element 17 is installed in the first temperature acquisition sensor element interface 131. The second positioning block 134 is provided with a first heating element interface 132, and the first heating element 18 is installed in the first heating element interface 132.

[0044] The middle unit 14 is provided with a first groove 142 and a second groove 143 . The first positioning block 135 is inserted into the first groove 142 , and the second positioning block 134 is inserted into the second groove 143 .

[0045] The lower constant temperature control unit 15 has a second temperature acquisition sensor element interface 151 , in which the second temperature acquisition sensor element 19 is installed. The lower constant temperature control unit 15 has a second heating element interface 152 , in which the second heating element 20 is installed.

[0046] The first temperature collection sensor element 17 and the second temperature collection sensor element 19 are both electrically connected to the internal control circuit, and the first heating element 18 and the second heating element 20 are both electrically connected to the internal control circuit. The first heating element 18 and the second heating element 20 are both 24V, 48W heating rods.

[0047] The first temperature collection sensor element 17 and the second temperature collection sensor element 19 are both DS18B20, with a power supply voltage of 3.3V, and are used to collect temperature.

[0048] A first switch 5 and a second switch 6 are provided on one side of the housing 1, and a power switch 7 and a power socket 8 are provided on the other side of the housing 1. The power switch 7 and the power socket 8 are electrically connected to an internal power supply, and the internal power supply supplies power to the electrical equipment. The first switch 5 is a standby switch for maintaining the temperature of the heating rod, and the second switch 6 is a heat dissipation switch for starting the heat dissipation fan 3. Both the first switch 5 and the second switch 6 are self-locking switches. Both the first switch 5 and the second switch 6 are electrically connected to the internal control circuit.

[0049] The display module 2 is a capacitive touch screen, which is convenient for viewing the controlled temperature results. The rated power of the cooling fan 3 is 2.4W. The central axis of the cooling fan 3 and the central axis of the convection heating module 4 are located on the same straight line. The horizontal distance between the central axis of the cooling fan 3 and the central axis of the convection heating module 4 is 70mm.

[0050] The internal control circuit in the utility model is prior art, and the utility model does not improve it.

[0051] With respect to the above-mentioned portable convection PCR amplification instrument, this article illustrates the technical effect of using the convection heating module 4 on the experiment through the following application method:

[0052] Step 1: Extract the target DNA and prepare the sample to be amplified. The preferred composition of the convection PCR mixture is as follows: according to the experimental requirements, add 1.6 μL of target DNA (4 ng / μL), 1.6 μL of upstream and downstream primers (2.5 μmol / L), 2 μL of 10× PCR buffer (containing Mg 2+ ), 0.8U of Ex Taq DNA polymerase, mixed with pure distilled water to form a total solution of 20 μL, added to the reaction container 9, the reaction container 9 was centrifuged for uniform mixing, and the solution was sealed with 2 μL of silicone oil to prevent the solution from volatilizing, thereby completing the preparation of the reaction solution in the reaction container;

[0053] The above upstream and downstream primer sequences are as follows:

[0054] Upstream primer: 5′-CAGTTTACCAACCGTCAT-3′;

[0055] Downstream primer: 5′-GAGCAACCGTTCCATTAC-3′.

[0056] Step 2: Turn on the first switch 5 and the second switch 6 to start the portable convection PCR amplifier, so that the first temperature acquisition sensor element 17, the second temperature acquisition sensor element 19, the first heating element 18 and the second heating element 20 in the convection heating module 4 are all powered on to increase their temperature, and the temperature of the reaction container 9 containing the sample to be amplified is maintained within a constant range through the convection heating container module 4 to complete the preheating of the convection heating module 4, and then the reaction container 9 is inserted into the reaction hole of the convection heating module 4.

[0057] Step 3: Set up several experimental groups: set the amplification reaction time to several different values ​​(10min, 15min, 20min, 25min, 30min, 35min). When the heating time of the reaction container 9 in the amplification instrument reaches the set amplification reaction time, turn off the convection heating module 4.

[0058] Step 4: After the desired convection PCR reaction time has passed, the reaction container 9 is taken out and placed in a refrigeration device for cooling, so that the convection PCR amplification reaction can be fully realized.

[0059] Step 5: Take out the amplified product from the reaction container 9 and perform electrophoresis analysis on the amplified product using 2% agarose gel. The analysis results are as follows: Figure 8 shown.

[0060] Figure 8 The experimental conditions for lanes 1 to 6 are: target DNA fragment length of 168 bp; target DNA concentration of 4 ng / μL; the temperatures of the upper and lower thermostatic control units are set to 53°C and 98°C, respectively; and the amplification reaction times are 10 min (lane 1), 15 min (lane 2), 20 min (lane 3), 25 min (lane 4), 30 min (lane 5), and 35 min (lane 6).

[0061] from Figure 8 It can be seen that the brightness of the band of the amplified product gradually becomes brighter as the amplification reaction time increases, and then reaches a plateau. Obviously, a relatively obvious amplified product band will appear when the reaction is 15 minutes, and the plateau is basically reached at 25-35 minutes. Therefore, the set amplification reaction time is preferably 25 minutes, and the acceptable range is 15-35 minutes. Compared with the existing PCR instrument that takes 2 hours to complete the complete amplification process, the amplification reaction time required for the PCR amplification instrument using the convection heating module 4 is greatly shortened.

[0062] The above is only a preferred embodiment of the present invention. It should be pointed out that for those skilled in the art, several modifications and improvements can be made without departing from the structure of the present invention, which will not affect the effect of the implementation of the present invention and the practicality of the patent.

Claims

1. A portable convection PCR amplification instrument, comprising a housing (1), wherein an internal control circuit is arranged in the housing (1), characterized in that: A heat dissipation fan (3) is installed on the housing (1), and a convection heating module (4) is installed on the housing (1), and both the heat dissipation fan (3) and the convection heating module (4) are electrically connected to an internal control circuit; The convection heating module (4) comprises an upper constant temperature control unit (13), an intermediate unit (14) and a lower constant temperature control unit (15); the upper constant temperature control unit (13) is installed above the intermediate unit (14); the intermediate unit (14) is installed above the lower constant temperature control unit (15); an air insulation layer (10) is provided between the intermediate unit (14) and the lower constant temperature control unit (15); a plurality of convection heating chambers (11) are provided between the upper constant temperature control unit (13), the intermediate unit (14) and the lower constant temperature control unit (15); and the convection heating chambers (11) are in communication with the air insulation layer (10).

2. A portable convection PCR amplification instrument according to claim 1, characterized in that: The upper constant temperature control unit (13) is provided with a plurality of first reaction holes (133), the middle unit (14) is provided with a plurality of second reaction holes (141) corresponding one-to-one to the first reaction holes (133), the lower constant temperature control unit (15) is provided with a plurality of third reaction holes (153) corresponding one-to-one to the second reaction holes (141), the aperture of the second reaction hole (141) is larger than the aperture of the first reaction hole (133) and the third reaction hole (153), and the first reaction hole (133), the second reaction hole (141) and the third reaction hole (153) constitute a stepped convection heating chamber (11) with a large middle portion and small ends.

3. A portable convection PCR amplification instrument according to claim 1, characterized in that: The upper constant temperature control unit (13) is provided with a first positioning block (135) and a second positioning block (134) which are integrally structured therewith; a first temperature acquisition sensor element interface (131) is provided in the first positioning block (135); a first temperature acquisition sensor element (17) is installed in the first temperature acquisition sensor element interface (131); a first heating element interface (132) is provided in the second positioning block (134); a first heating element (18) is installed in the first heating element interface (132); and both the first temperature acquisition sensor element (17) and the first heating element (18) are electrically connected to an internal control circuit.

4. A portable convection PCR amplification instrument according to claim 2, characterized in that: The aperture depth of the first reaction hole (133) is 1-4 mm, the aperture depth of the second reaction hole (141) is 4-8 mm, the aperture depth of the third reaction hole (153) is 1-3 mm, and the thickness of the air insulation layer (10) is 4-8 mm.

5. A portable convection PCR amplification instrument according to claim 3, characterized in that: The intermediate unit (14) is provided with a first groove (142) and a second groove (143); the first positioning block (135) is plugged into the first groove (142); and the second positioning block (134) is plugged into the second groove (143).

6. A portable convection PCR amplification instrument according to claim 1, characterized in that: The lower constant temperature control unit (15) is provided with a second temperature acquisition sensor element interface (151), and a second temperature acquisition sensor element (19) is installed in the second temperature acquisition sensor element interface (151). The lower constant temperature control unit (15) is provided with a second heating element interface (152), and a second heating element (20) is installed in the second heating element interface (152). Both the second temperature acquisition sensor element (19) and the second heating element (20) are electrically connected to an internal control circuit.

7. A portable convection PCR amplification instrument according to claim 1, characterized in that: A display module (2) is fixedly mounted above the housing (1); a first switch (5) and a second switch (6) are provided on one side of the housing (1); a power switch (7) and a power socket (8) are provided on the other side of the housing (1); and the display module (2), the first switch (5) and the second switch (6) are all connected to an internal control circuit.

8. A portable convection PCR amplification instrument according to claim 1, characterized in that: The upper constant temperature control unit (13), the middle unit (14) and the lower constant temperature control unit (15) are all made of 6061 aluminum alloy plates.

9. A portable convection PCR amplification instrument according to claim 1, characterized in that: The thickness of the upper constant temperature control unit (13) is 1-4 mm, the thickness of the middle unit (14) is 1-10 mm, and the thickness of the lower constant temperature control unit (15) is 1-20 mm.

10. A portable convection PCR amplification instrument according to claim 1, characterized in that: The heat dissipation fan (3) is located on one side of the convection heating module (4).

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