CN118813376A

CN118813376A - Reaction module, nucleic acid amplification device and nucleic acid amplification control method

Info

Publication number: CN118813376A
Application number: CN202310420702.4A
Authority: CN
Inventors: 韦嘉; 徐强; 赵蒙; 徐涛; 冼志科
Original assignee: Guangzhou National Laboratory
Current assignee: Guangzhou National Laboratory
Priority date: 2023-04-18
Filing date: 2023-04-18
Publication date: 2024-10-22
Also published as: WO2024217049A1

Abstract

The present invention relates to a reaction module, a nucleic acid amplification apparatus, and a nucleic acid amplification control method. And the reaction assembly comprises a containing cavity for containing a reaction sample and a heater for heating the reaction sample. The nucleic acid amplification apparatus includes a reaction component. The nucleic acid amplification control method comprises: and controlling the reaction assembly through a resistance temperature measurement method and a double temperature measurement method for calibrating the temperature of the reaction assembly. The heater and the accommodating cavity are of an integral structure, the heater is tightly connected with the accommodating cavity, and an air layer does not exist, so that the heat transfer speed of the heater and the reaction sample in the accommodating cavity is accelerated, the nucleic acid amplification process is accelerated, and the detection efficiency is improved. The nucleic acid amplification control method can realize rapid and accurate control of the temperature of the reaction component, and achieves the aim of accurate temperature control.

Description

Reaction module, nucleic acid amplification device and nucleic acid amplification control method

Technical Field

The invention relates to the technical field of in-vitro diagnosis, in particular to a reaction component, a nucleic acid amplification device and a nucleic acid amplification control method.

Background

PCR (polymerase chain reaction) is a molecular biological experimental method for in vitro enzymatic synthesis of specific DNA fragments, and PCR amplification, namely nucleic acid amplification, mainly comprises three repeated thermal cycles of high-temperature denaturation, low-temperature annealing and temperature-adaptive extension.

In the prior art, a housing or a tube (such as a PCR tube) having a housing cavity is placed on a heater to facilitate the later removal of the housing or tube, the housing cavity is used for housing a reaction sample, the heater can heat the housing or tube, and the housing or tube transfers heat to the reaction sample, thereby realizing the amplification of the reaction sample. However, as the shell or the tube body is only placed on the heater for heating, an air layer exists between the shell or the tube body and the heater, the heat transfer speed between the heater and the shell or the tube body is reduced, the reaction sample is slow in temperature rise and reduction, and the nucleic acid amplification process and the detection efficiency are further affected. Meanwhile, the thickness of the reaction sample in the shell or the tube body is large, the time required for the uniform temperature of the reaction sample is long, and the reaction sample is slow in temperature rising and reducing speed.

In the prior art, a multi-layer structure layer is arranged between the heater and the reaction sample in the accommodating cavity, and the multi-layer structure layer can further obstruct heat transfer between the heater and the reaction sample, so that the reaction sample is slow in temperature rise and reduction.

In order to facilitate the control of the heater, there are two main methods for detecting the temperature of the reaction sample, which are commonly used in the prior art, the first method is to detect the temperature of the heater or detect the temperature of the reaction sample by a temperature sensor, but since a certain time is required for transferring heat to the temperature sensor, the detection result measured by the temperature sensor has a delay of 1-2 s, which results in inaccurate control of the heater and greatly affects the detection result. The other method is to detect the resistance value of the heater and then combine the resistance temperature coefficient with the nominal resistance value to obtain the corresponding temperature value, but for the same type of resistance, such as copper wire resistance, the nominal resistance value and the resistance temperature coefficient between the resistances are slightly different, so that the temperature measurement error is large and the detection result is affected.

Disclosure of Invention

It is an object of the present invention to provide a reaction module which solves at least one of the above problems.

To achieve the above object, a first aspect of the present invention provides a reaction module including a receiving chamber for receiving a reaction sample and a heater for heating the reaction sample.

Optionally, the heater is in direct contact with the reaction sample within the receiving chamber.

Optionally, at least part of the upper surface of the heater is in direct contact with the reaction sample in the holding chamber, or

The surface of the heater is provided with a groove, and at least part of the wall surface of the groove is in direct contact with the reaction sample in the accommodating cavity.

Optionally, the heater comprises a soaking layer in direct contact with the reaction sample within the containment chamber.

Optionally, the heater further comprises a heating element and a temperature calibration part for the temperature detection unit to detect temperature.

Optionally, the heater further comprises an upper conductive component and a lower conductive component, the heating element is sandwiched between the upper conductive component and the lower conductive component, and the upper conductive component comprises the soaking layer.

Optionally, the temperature calibration part is connected to the upper conductive assembly or the lower conductive assembly.

Optionally, the temperature calibration part is connected to one side of the upper conductive component, which is close to the lower conductive component, and the lower conductive component is provided with a first through hole opposite to the temperature calibration part.

Optionally, the temperature calibration portion is part of the upper conductive assembly or the lower conductive assembly.

Optionally, a second through hole is formed in the lower conductive component along the thickness direction of the reaction component, and the surface, opposite to the second through hole, of the upper conductive component is the temperature calibration part.

Optionally, the reaction assembly further comprises a fast conducting part for conducting heat of the heating element to the temperature calibration part.

Optionally, one side of the rapid conduction part is connected to one side of the upper conduction assembly close to the heating element or connected to one side of the lower conduction assembly close to the heating element, and the other side is connected to the temperature calibration part.

Optionally, the temperature calibration portion is located on a side of the lower conductive assembly remote from the heating element.

Optionally, a receiving groove is formed in a side, far away from the heating element, of the lower conduction assembly, and the temperature calibration part is located in the receiving groove and is connected with the bottom of the receiving groove.

Optionally, the quick-conduction part comprises one or more first guide posts, one ends of the one or more first guide posts are attached to one side of the upper conduction assembly close to the heating element or one side of the lower conduction assembly close to the heating element, and the other ends of the one or more first guide posts are connected with the temperature calibration part.

Optionally, the quick conduction portion includes paster and one or more second guide pillar, the paster with go up the conduction subassembly is close to one side laminating of heating element or with down the conduction subassembly is close to one side laminating of heating element, one end of one or more second guide pillar is connected in the paster, the other end wears to locate down the conduction subassembly and with the temperature calibration portion is connected.

Optionally, a third through hole is formed in the lower conductive component, and the guide post is disposed in the third through hole.

Optionally, the lower conductive assembly further comprises an insulating thermal resistance layer.

Optionally, the lower conductive assembly further comprises a thermally conductive layer located on a side of the insulating thermal resistance layer remote from the heating element.

Optionally, the soaking layer is made of a conductive material or an insulating material.

Optionally, when the soaking layer is made of an electrically conductive material, the upper conductive assembly further comprises an insulating layer, the insulating layer being located between the heating element and the soaking layer;

When the soaking layer is made of an insulating material, the soaking layer is adjacent to the heating member.

Optionally, the receiving cavity includes a bottom wall;

the bottom wall is closely contacted with at least part of the upper surface of the heater, or

The surface of the heater is provided with a groove, and at least part of the wall surface of the groove is tightly contacted with the bottom wall.

Optionally, a flexible heat conducting member is arranged between the bottom wall and the heater.

Optionally, the accommodating cavity is of a flat structure.

Optionally, the cross section of the accommodating cavity is polygonal, circular or elliptical.

Optionally, the reaction assembly comprises a first contact temperature detection unit connected to the temperature calibration part and configured to measure a temperature at the temperature calibration part.

Another object of the present invention is to provide a nucleic acid amplification apparatus for amplifying a nucleic acid.

To achieve the object, the second aspect of the present invention adopts the following technical scheme:

a nucleic acid amplification apparatus comprising the reaction assembly.

To achieve the object, a third aspect of the present invention adopts the following technical scheme:

A nucleic acid amplification device,

The reaction component comprises a second contact type temperature detection unit and a reaction component, wherein the second contact type temperature detection unit can be separated from or contacted with the temperature calibration part, and the temperature of the temperature calibration part can be measured when the second contact type temperature detection unit is contacted with the temperature calibration part.

To achieve the object, a fourth aspect of the present invention adopts the following technical scheme:

A nucleic acid amplification apparatus comprising a non-contact temperature detection unit for measuring a temperature at the temperature calibration part and a reaction assembly as described above.

Optionally, the nucleic acid amplification apparatus further comprises a cooling mechanism for cooling the reaction sample in the accommodation chamber.

Optionally, an avoidance portion is formed on a side, close to the reaction component, of the cooling mechanism.

Optionally, the cooling mechanism cools the reaction assembly by a fluid.

Optionally, the cooling mechanism cools the reaction assembly by spraying a fluid or fluid flow.

Optionally, when the cooling mechanism cools the reaction assembly by fluid flow, the cooling mechanism includes a cooling body having a cooling flow passage provided therein to flow a cooling medium.

Optionally, the cooling mechanism adopts a solid cooling mode to cool the reaction component.

Optionally, the nucleic acid amplification apparatus further comprises a resistance detection unit for detecting the temperature of the reaction component.

Optionally, the resistance detection unit obtains the temperature value of the reaction component by detecting the resistance of the reaction component.

It is still another object of the present invention to provide a nucleic acid amplification control method for performing nucleic acid amplification.

To achieve the object, a fifth aspect of the present invention adopts the following technical scheme:

a nucleic acid amplification control method, which is performed using the reaction module as described above, or a nucleic acid amplification apparatus;

The nucleic acid amplification control method comprises: and controlling the reaction assembly through a resistance temperature measurement method and a double temperature measurement method for calibrating the temperature of the reaction assembly.

Optionally, controlling the reaction assembly by a resistance thermometry method and a dual thermometry method for calibrating the temperature of the reaction assembly includes:

And measuring a temperature calibration value of the reaction component through a temperature detection unit, and calibrating a temperature value obtained through the resistance value of the reaction component through the temperature calibration value.

Optionally, the temperature coefficient of resistance and the nominal resistance value of the reaction component are obtained according to the temperature calibration value so as to calibrate the temperature value.

Optionally, the temperature value is calibrated by measuring the temperature calibration value before the nucleic acid amplification process, during a first temperature increase of the nucleic acid amplification process and/or during a first amplification cycle of the nucleic acid amplification process.

Optionally, the temperature values are calibrated using at least two different temperature calibration values.

Optionally, controlling the reaction assembly by a resistance thermometry method and a dual thermometry method for calibrating the temperature of the reaction assembly includes the steps of:

Obtaining at least two different temperature calibration values; detecting a first voltage and a first current of the reaction component under the temperature calibration value, and obtaining a first resistance value of the reaction component according to the first voltage and the first current; detecting a second voltage and a second current of the reaction component under the other temperature calibration value, and obtaining a second resistance value of the reaction component according to the second voltage and the second current;

Obtaining a resistance temperature coefficient of the reaction component and a resistance at a nominal temperature according to at least the first resistance value, the second resistance value and the corresponding temperature calibration value;

and continuously detecting the current and the voltage of the reaction component, and controlling the reaction component according to the temperature coefficient of resistance and the resistance at the nominal temperature.

Optionally, before calibrating the temperature value, the temperature of the reaction component measured by the temperature detection unit controls the reaction component, or controls the temperature of the reaction component according to an RT temperature curve preset for the reaction component.

Alternatively, according to the formula: and (2) calibrating the temperature coefficient of resistance and the nominal resistance value of the reaction component by R=R0 (1+alpha delta T), and obtaining the temperature curve of the reaction component according to the formula by continuously measuring the voltage and the current of the reaction component, wherein R0 is the nominal resistance value and alpha is the temperature coefficient of resistance of the material.

In view of the above, the reaction assembly according to the present invention includes a receiving chamber for receiving a reaction sample and a heater for heating the reaction sample. The heater and the accommodating cavity are of an integral structure, the heater is tightly connected with the accommodating cavity, and an air layer does not exist, so that the heat transfer speed of the heater and the reaction sample in the accommodating cavity is accelerated, the nucleic acid amplification process is accelerated, and the detection efficiency is improved. The heater is in direct contact with the reaction sample in the accommodating cavity, the accommodating cavity is directly formed on the upper surface of the heater, and no other conduction interface exists between the reaction sample and the heater, so that the conduction interface between the heater and the accommodating cavity is reduced, and the conduction efficiency is further improved.

Drawings

FIG. 1 is a schematic structural view of a first reaction module according to an embodiment of the present invention;

FIG. 2 is a schematic structural view of a second reaction module according to an embodiment of the present invention;

FIG. 3 is a schematic view of a third reaction assembly and a fluid flow cooling mechanism according to an embodiment of the present invention;

FIG. 4 is a schematic structural view of a fourth reaction module according to an embodiment of the present invention;

FIG. 5 is a schematic structural view of a fifth reaction module according to an embodiment of the present invention;

FIG. 6 is a schematic structural view of a sixth reaction module according to an embodiment of the present invention;

FIG. 7 is a schematic structural view of a seventh reaction module according to an embodiment of the present invention;

FIG. 8 is a schematic structural view of an eighth reaction module according to an embodiment of the present invention;

FIG. 9 is a schematic structural view of a ninth reaction module according to an embodiment of the present invention;

FIG. 10 is a schematic structural view of a tenth reaction module according to an embodiment of the present invention;

FIG. 11 is a schematic structural view of a reaction assembly and cooling mechanism for the injected fluid provided by an embodiment of the present invention;

FIG. 12 is a schematic diagram showing a structure of a nucleic acid amplification apparatus according to an embodiment of the present invention when a first contact temperature detecting unit is used for detecting temperature;

FIG. 13 is a partial cross-sectional view showing a nucleic acid amplification apparatus according to an embodiment of the present invention when a second contact temperature detection unit is used for detecting temperature;

FIG. 14 is a schematic diagram showing a structure of a nucleic acid amplification apparatus according to an embodiment of the present invention when a second contact temperature detection unit is used for detecting temperature;

FIG. 15 is a schematic diagram showing a structure of a nucleic acid amplification apparatus according to an embodiment of the present invention when a non-contact temperature detection unit is used for detecting temperature;

FIG. 16 is a flowchart of a method for controlling nucleic acid amplification according to an embodiment of the present invention;

FIG. 17 is a temperature versus time curve of a reaction assembly provided by an embodiment of the present invention.

In the figure:

1. A cooling mechanism; 11. a cooling flow passage; 12. cooling the body; 13. an avoidance unit;

2. A reaction assembly;

21. a receiving chamber; 22. a bottom wall; 23. a heating member; 24. an upper conductive assembly; 241. a soaking layer; 242. an insulating layer; 25. a temperature calibration unit; 251. an external electrical connection contact; 252. an electrical connection lead; 26. a fast conduction part; 261. a patch; 262. a second guide post; 263. a first guide post; 27. a lower conductive assembly; 271. an insulating thermal resistance layer; 272. a heat conducting layer; 273. a first through hole; 274. a second through hole; 275. a receiving groove; 28. a cavity sidewall; 29. a cover plate; 20. a second contact; 201. a groove; 202. a wall surface; 2021. a sidewall; 2022. a groove bottom; 203. a flexible heat conducting member;

200. A heater;

3. a first contact temperature detection unit; 4. a second contact temperature detection unit; 5. a non-contact temperature detection unit; 6. and a resistance detection unit.

Detailed Description

The technical scheme of the invention is further described below by the specific embodiments with reference to the accompanying drawings. It is to be understood that the specific embodiments described herein are merely illustrative of the invention and are not limiting thereof. It should be further noted that, for convenience of description, only some, but not all of the drawings related to the present invention are shown.

In the present invention, directional terms such as "upper", "lower", "left", "right", "inner" and "outer" are used for convenience of understanding, and thus do not limit the scope of the present invention unless otherwise specified.

In the present invention, unless expressly stated or limited otherwise, a first feature "above" or "below" a second feature may include both the first and second features being in direct contact, as well as the first and second features not being in direct contact but being in contact with each other through additional features therebetween. Moreover, a first feature being "above," "over" and "on" a second feature includes the first feature being directly above and obliquely above the second feature, or simply indicating that the first feature is higher in level than the second feature. The first feature being "under", "below" and "beneath" the second feature includes the first feature being directly under and obliquely below the second feature, or simply means that the first feature is less level than the second feature.

In the description of the present invention, unless explicitly stated and limited otherwise, the terms "connected," "connected," and "fixed" are to be construed broadly, and may be, for example, fixedly connected, detachably connected, or integrally formed; can be mechanically or electrically connected; can be directly connected or indirectly connected through an intermediate medium, and can be communicated with the inside of two elements or the interaction relationship of the two elements. The specific meaning of the above terms in the present invention will be understood in specific cases by those of ordinary skill in the art.

Example 1

The present embodiment provides a reaction module 2 for amplifying a reaction sample, but is not limited thereto, and can be used in other occasions where heating of the reaction sample is required to improve the detection efficiency.

As shown in fig. 1 to 10, the reaction module 2 provided in the present embodiment includes a housing chamber 21 for housing a reaction sample and a heater 200 for heating the reaction sample. The heater 200 and the accommodating cavity 21 are of an integral structure, the heater 200 and the accommodating cavity 21 are tightly connected, and an air layer does not exist, so that the heat transfer speed of the reaction sample in the heater 200 and the accommodating cavity 21 is increased, the nucleic acid amplification process is accelerated, and the detection efficiency is improved.

As shown in fig. 1 and 2, the heater 200 is in direct contact with the reaction sample in the accommodation chamber 21. There is no other conductive interface between the reaction sample and the heater 200, thereby reducing the conductive interface between the heater 200 and the receiving chamber 21 and further improving the conduction efficiency. Meanwhile, no interface exists between the heater 200 and the reaction sample, so that the thermal resistance is small, and the rapid heat conduction and the higher-speed temperature control can be realized.

At least a portion of the upper surface of the heater 200 is in direct contact with the reaction sample in the receiving chamber 21, i.e., the receiving chamber 21 is directly formed on the upper surface of the heater 200, and no other conductive interface exists between the reaction sample and the heater 200, thereby reducing the conductive interface between the heater 200 and the receiving chamber 21 and further improving the conductive efficiency.

As shown in fig. 1, an annular chamber sidewall 28 may be optionally attached to the upper surface of the heater 200, and the top end of the chamber sidewall 28 is attached to a cover plate 29. The chamber side wall 28, the cover plate 29 and at least part of the upper surface of the heater 200 enclose the above-mentioned accommodation chamber 21, thereby accommodating and confining the reaction sample. The cover plate 29 may be made of glass or transparent plastic to allow optical detection of the sample liquid from above.

In a specific embodiment, the cover plate 29 is made of a glass material, and is exemplified by a thickness of 0.2mm, and the chamber side wall 28 supports the cover plate 29, and is exemplified by a height of 0.5mm of the chamber side wall 28, that is, a thickness of 0.5mm of the receiving chamber 21, and a thickness of 0.5mm of the reaction sample when the reaction sample fills the receiving chamber 21. The cover plate 29 is provided with a sample inlet hole for loading the reaction sample, and the reaction sample can be sealed by a glue film after being put in.

Alternatively, the accommodating chamber 21 is of a flat structure, and it is understood that the flat structure may mean that the dimension in the thickness direction of the accommodating chamber 21 is much smaller than the dimension in the width or length direction, and as an example, the accommodating chamber 21 is a rectangular parallelepiped, and the ratio of the length to the thickness of the rectangular parallelepiped may be greater than 5:1, such as 90:1, for example, the dimension of the accommodating chamber 21 in the thickness direction may be 0.3-1.0mm, and the width and length of the accommodating chamber 21 may be about 10mm and 20mm, respectively, wherein the dimension in the thickness direction is the arrangement direction of the heating member 23 and the accommodating chamber 21. As an example, the housing chamber 21 may also have a cylindrical structure with a diameter to thickness ratio greater than 5:1, for example, 0.3-1.0mm in thickness and 5-20mm in diameter. Of course, the cross section of the accommodating chamber 21 may be polygonal or elliptical, etc. Of course, the cross section of the accommodating chamber 21 may be polygonal or elliptical, etc.

The reaction sample in the accommodating cavity 21 of the flat structure is thin, the distance between the center of the reaction sample and the surface of the liquid is small, the temperature of the reaction sample can be consistent in a short time, and the flat structure can ensure that the contact area between the reaction sample and the heater 200 is large, the heat transfer efficiency is high, and the temperature rising and reducing speed and the detection efficiency of the reaction sample are greatly improved. The inner diameter of the PCR tube is larger than that of the flat accommodating cavity 21, the distance between the center of the reaction sample and the surface of the liquid is large, the temperature of the reaction sample needs a long time to be consistent, the temperature rising and reducing speed of the reaction sample is low, and the detection efficiency is low.

In other alternative embodiments, as shown in fig. 2, the surface of the heater 200 is formed with a groove 201, and at least part of the wall 202 of the groove 201 is in direct contact with the reaction sample in the accommodating chamber 21, i.e., the reaction sample is in direct contact with the wall 202 of the groove 201, and no other conductive interface exists between the reaction sample and the heater 200, so that the conductive interface between the heater 200 and the accommodating chamber 21 is reduced, and the conductive efficiency is further improved. Meanwhile, the side wall 2021 of the wall surface 202 of the groove 201 is also in contact with the reaction sample, so that the contact area between the heater 200 and the reaction sample can be increased, and the heat transfer efficiency can be further increased, compared with the case that the heater 200 is not provided with the groove 201.

As shown in fig. 2, it is preferable that the receiving chamber 21 coincides with the recess 201, that is, the space in the recess 201 forms the receiving chamber 21, and the reaction sample directly contacts with the side wall 2021 and the groove bottom 2022 of the wall 202 of the recess 201, so that heat exchange between the heater 200 and the reaction sample proceeds from between the side wall 2021 and the groove bottom 2022 of the recess 201, increasing the contact area of the heater 200 and the reaction sample, and increasing the heat transfer efficiency.

In this embodiment, the depth of the recess 201 is 0.5mm, i.e., the thickness of the reaction sample is 0.5mm when the reaction sample fills the accommodating chamber 21. The top of the recess 201 may be connected to a cover plate 29 to confine the reaction sample within the receiving chamber 21.

For the scheme that the reaction sample is in direct contact with the heater 200, no detachable interface exists between the heater 200 and the reaction sample, so that the reaction sample has smaller thermal resistance, and higher-speed temperature control can be realized while rapid heat conduction is realized.

In other alternative embodiments, as shown in fig. 3, the accommodating chamber 21 includes a bottom wall 22, the surface of the heater 200 is formed with a groove 201, and at least part of the wall 202 of the groove 201 is in close contact with the bottom wall 22. Or the bottom wall 22 is in close contact with at least part of the upper surface of the heater 200, the air layer is not formed between the bottom wall 22 and the heater 200 due to the close contact between the bottom wall 22 and the heater 200, and thus heat can be rapidly transferred. However, the heat transfer efficiency is reduced by adding a heat transfer layer such as the bottom wall 22 to the case where the reaction sample is in direct contact with the heater 200.

In order to make the bottom wall 22 closely contact with the heater 200, alternatively, the bottom wall 22 and the heater 200 may be formed by injection molding, or of course, the close contact may be achieved by other methods, which will not be described herein. Specifically, the bottom wall 22 is in close contact with the soaking layer 241 of the heater 200.

Of course, in other alternative embodiments, the flexible heat conducting member 203 may be disposed between the bottom wall 22 and the heater 200, and the flexible heat conducting member 203 may ensure a more compliant bond with the bottom wall 22 and the heater 200, so that both the bottom wall 22 and the heater 200 closely contact the flexible heat conducting member 203, thereby avoiding the occurrence of an air layer. The flexible heat conductive member 203 may be a heat conductive silicone or the like.

As shown in fig. 1-5, alternatively, the heater 200 includes a heating element 23, and a power source is connected to the heating element 23, where the heating element 23 is a controllable heating source inside the heater 200, and may be a resistor, for example, a copper material may be made into a resistor fine wire structure, and the heating power is controlled by controlling the magnitude of the current flowing through the resistor, so as to achieve temperature control. In other alternative embodiments, the heating element 23 may be configured as a coil or heated by electromagnetic induction through ferromagnetic materials or the like.

As shown in fig. 4, the heater 200 may further include an upper conductive member 24 and a lower conductive member 27, with the heating member 23 interposed between the upper conductive member 24 and the lower conductive member 27. The upper and lower conductive assemblies 24 and 27 have a conductive heat and insulating effect.

As shown in fig. 1 and 4-5, the heater 200 includes a soaking layer 241, and in particular, the upper conductive assembly 24 may also include a soaking layer 241. The soaking layer 241 is in direct contact with the reaction sample in the accommodating cavity 21 or in close contact with the bottom wall 22, and the soaking layer 241 can ensure uniform conduction of heat in the longitudinal direction and the transverse direction (namely the thickness direction of the reaction sample and the surface perpendicular to the thickness direction), so that the temperature uniformity of the sample liquid is ensured. Alternatively, the soaking layer 241 is made of an electrically conductive material or an insulating material, for example, the soaking layer 241 is made of an electrically conductive material such as aluminum, copper, or an insulating material such as high thermal conductivity ceramic.

As shown in fig. 1, when the soaking layer 241 is made of an insulating material, the soaking layer 241 is adjacent to the heating member 23, and at this time, the number of layers of the reaction module 2 can be reduced, the time for transferring heat from the heating member 23 to the reaction module 2 can be shortened, and the time required for heat dissipation from the reaction module 2 can be shortened.

As shown in fig. 4, when the soaking layer 241 is made of a conductive material, the upper conductive member 24 further includes an insulating layer 242, and the insulating layer 242 is located between the heating member 23 and the soaking layer 241. The insulating layer 242 is made of a material having a high thermal conductivity for electrically isolating the heating member 23 from the electrically conductive soaking layer 241. Alternatively, the insulating layer 242 is made of a material having a thermal conductivity >1W/mK, and the thermal conductivity of the insulating layer 242 is 3W/mK in this embodiment.

As shown in fig. 1 and 4-5, the lower conductive assembly 27 further includes an insulating thermal resistance layer 271. The insulating thermal resistance layer 271 has a certain thermal resistance characteristic and insulating characteristic. The insulating thermal resistance layer 271 may form a longitudinal thermal resistance in addition to insulating the heating element 23. The magnitude of the thermal resistance can be designed by material selection and thickness selection. Typically, the thermal resistance of this layer is much greater than the thermal resistance of the other layers of the structure, so that the thermal edge resistance layer 271 is the primary thermal resistance source for the reaction component 2 to dissipate heat to and cool down the cooling mechanism 1 (cooling mechanism 1 will be described in detail below). The insulating thermal resistance layer 271 is one of the main influencing factors for the thermal performance of the reaction module 2.

Optionally, the lower conductive assembly 27 further includes a thermally conductive layer 272, the thermally conductive layer 272 being located on a side of the insulating thermal resistance layer 271 remote from the heating element 23. Further, the heat conductive layer 272 is the outermost layer of the lower conductive assembly 27, which is in direct contact with the cooling mechanism 1. The heat conductive layer 272 is made of metal such as copper or other material with high thermal conductivity. The surface of the lower conductive assembly 27 that contacts the cooling mechanism 1 is difficult to avoid point contact due to cost control or tooling limitations. When the outermost layer of the lower conductive component 27 is the heat conductive layer 272, even if the heat conductive layer 272 is in point contact with the cooling mechanism 1, the heat conductive layer 272 can uniformly distribute heat over the entire heat conductive layer 272 due to its good conductivity, so that heat of other layers of the lower conductive component 27 can be uniformly distributed.

Preferably, the heating element 23 of the present embodiment is a resistor, and there is a specific relationship between the resistor and the temperature thereof, so that the real-time resistance change of the heating element 23 is measured while heating, and the average temperature of the heating element 23 is deduced through the temperature coefficient of resistance and the nominal resistance value. The temperature shows the current temperature of the reaction component 2 in real time without time delay, so that the temperature of the reaction component 2 and the reaction sample can be quickly controlled in a feedback manner, and compared with the prior art, the temperature of the sample can be accurately controlled, and the overall reaction speed of a temperature control system is improved.

The disadvantage of this method is that for the same type of resistor, such as copper wire resistor, the nominal resistance value and the temperature coefficient of resistance are slightly different, which results in that the temperature coefficient of resistance between the single heating elements 23 is slightly different from the nominal resistance value, which may cause temperature measurement errors, so that, preferably, as shown in fig. 1 and fig. 4-8, the heater 200 provided in this embodiment may further include a temperature calibration part 25 for allowing the temperature detection unit to detect the temperature, so that the reaction assembly 2 can be controlled by a resistance thermometry method and a dual temperature measurement method for calibrating the temperature of the reaction assembly 2.

Alternatively, the temperature detection unit may be a contact temperature detection unit or a non-contact temperature detection unit 5.

As shown in fig. 1, 4-6, 8 and 10, when the temperature of the temperature calibration part 25 is detected in a contact or non-contact manner, for example, when the temperature detection unit is a contact type temperature detection unit or a non-contact type temperature detection unit 5, the temperature calibration part 25 is connected to the upper conductive member 24 or the lower conductive member 27 so that the temperature of the heating element 23 is conducted to the temperature calibration part 25.

As shown in fig. 7, when the temperature detecting means is the noncontact temperature detecting means 5, the number of calibration portions may be one or two, as long as the temperature of the temperature calibration portion 25 can be detected by the noncontact temperature detecting means 5.

When the temperature detecting unit is a contact temperature detecting unit as shown in fig. 4 and 6, the two first contacts of the contact temperature detecting unit are respectively contacted with the two temperature calibrating parts 25, and the two temperature calibrating parts 25 are not electrically conductive, at this time, as shown in fig. 6, optionally, the nucleic acid amplifying device may further include an external electrical connecting contact 251 and an electrical connecting lead 252, the number of the external electrical connecting contact 251 and the electrical connecting lead 252 may be two, the two external electrical connecting contacts 251 are respectively located at the sides of the two temperature calibrating parts 25 apart from each other, one external connecting contact is electrically connected with one temperature calibrating part 25 through one electrical connecting lead 252, and the other external connecting contact is electrically connected with the other temperature calibrating part 25 through the other electrical connecting lead 252.

Referring to fig. 5 and 10, heat of the upper conductive member 24 (e.g., the soaking layer 241 of the upper conductive member 24 contacting the patch 261 of the rapid conductive portion 26, which will be described later in detail) is conducted to the temperature calibration portion 25 through the patch 261 and the guide post 262, the temperature calibration portion 25 is electrically connected to the outside through the electrical connection lead 252 at the external electrical connection contact 251, wherein the diameter of the electrical connection lead 252 is smaller than that of the temperature calibration portion 25 and the external electrical connection contact 251, thereby reducing heat loss generated by the temperature calibration portion 25 through the electrical connection lead 252, and thus the temperature calibration portion 25 can better embody the temperature of the upper conductive member 24 (e.g., the soaking layer 241 of the upper conductive member 24 contacting the patch 261), and the temperature detection unit realizes good electrical and thermal contact with the temperature calibration portion 25 through the solder joint, and when the temperature of the upper conductive member 24 (e.g., the soaking layer 241 of the upper conductive member 24 contacting the patch 261) is changed, the temperature detection unit senses a temperature change rapidly and precisely, and the temperature change causes a resistance change of the temperature detection unit, and the temperature detection can be realized by detecting the resistance change of the temperature detection unit in real time at the external electrical connection contact 251.

Specifically, as shown in fig. 4 and 5, the temperature calibration part 25 is located at a side of the lower conductive member 27 away from the heating member 23 so as to fix the temperature calibration part 25. In connection with fig. 6 and 7, further, a receiving groove 275 is formed in a side of the lower conductive member 27 remote from the heating member 23, and the temperature calibrating portion 25 is disposed in the receiving groove 275 and connected to a bottom of the receiving groove 275. Optionally, the accommodating groove 275 penetrates through the heat conducting layer 272, the bottom of the accommodating groove 275 is an insulating thermal resistance layer 271, and the temperature calibration part 25 is connected to the insulating thermal resistance layer 271. In other alternative embodiments, the receiving groove 275 may not extend through the heat conductive layer 272, and the temperature calibration portion 25 is connected to the heat conductive layer 272. The receiving groove 275 may prevent the temperature calibrating portion 25 from protruding from the lower conductive member 27, thereby maintaining the flatness of the lower surface of the reaction module 2 and facilitating the smooth placement of the reaction module 2. Of course, in still another embodiment, the lower conductive member 27 may not be provided with the receiving groove 275, and the temperature calibrating portion 25 is connected to the lower surface of the lower conductive member 27.

In yet another alternative embodiment, as shown in fig. 8, the temperature calibration portion 25 is connected to the side of the upper conductive member 24 near the lower conductive member 27, e.g., the temperature calibration portion 25 is connected to the lower surface of the insulating soaking layer 241 (as shown in fig. 8, the insulating layer 242 may not be provided when the soaking layer 241 is insulated, and thus the temperature calibration portion 25 may be directly connected to the lower surface of the soaking layer 241), or connected to the lower surface of the insulating layer 242, so that the temperature of the heating element 23 is rapidly transferred to the temperature calibration portion 25.

The lower conductive member 27 may further be provided with a first through hole 273 disposed opposite to the temperature calibration portion 25 so that the temperature detecting member can detect the temperature of the temperature calibration portion 25. The lower surface of the upper conductive member 24 and the upper surface of the lower conductive member 27 are closest to the heating member 23, and the temperature thereof is closest to the temperature of the heating member 23 first, so that the temperature closest to the heating member 23 can be detected more quickly by the temperature detecting unit by connecting the temperature calibrating portion 25 to the side of the upper conductive member 24 closest to the lower conductive member 27.

As shown in fig. 1 and 4, alternatively, the temperature detecting unit may be a first contact temperature detecting unit 3, and the reaction assembly 2 may include the first contact temperature detecting unit 3 connected to the temperature calibrating portion 25 and configured to measure the temperature at the temperature calibrating portion 25. The first contact temperature detecting unit 3 may be a temperature sensor or the like. The first contact temperature detecting unit 3 may be connected to the temperature calibrating portion 25 by welding or the like, and the first contact temperature detecting unit 3 may be discarded together with the reaction module 2 after the reaction module 2 is used.

When the temperature of the temperature calibration portion 25 is detected in a non-contact manner, for example, when the temperature detection unit is a non-contact temperature detection unit 5 such as an infrared temperature measurement unit, the temperature calibration portion 25 may be provided in a manner other than the above-mentioned manner, as shown in fig. 9, and the temperature calibration portion 25 may be a part of the upper conductive member 24 or the lower conductive member 27, so that no additional connection of the temperature calibration portion 25 is required, and only the position of the temperature calibration portion 25 needs to be reserved for the upper conductive member 24 or the lower conductive member 27, so that the non-contact temperature detection unit 5 can be aligned with the position and the temperature of the position can be detected. Preferably, the lower conductive member 27 is provided with a second through hole 274 along the thickness direction of the reaction member 2, and the surface of the upper conductive member 24 facing the second through hole 274 is the temperature calibration portion 25. The lower surface of the upper conductive member 24 and the upper surface of the lower conductive member 27 are closest to the heating member 23, and the temperature thereof is first close to the temperature of the heating member 23, so that the temperature detecting unit can detect the temperature closest to the heating member 23 more quickly by detecting the temperature of the lower surface of the upper conductive member 24.

Of course, in other alternative embodiments, the temperature calibration portion 25 may be a lower surface of the heat conducting layer 272 to simplify the structure of the reaction module 2.

As shown in fig. 4 and 5, in order to shorten the time when the temperature of the temperature calibration part 25 coincides with the temperature of the heating member 23, the reaction assembly 2 may optionally further include a rapid conduction part 26, and the rapid conduction part 26 is used to conduct heat of the heating member 23 to the temperature calibration part 25. Specifically, in the present embodiment, the heat of the heating element 23 is indirectly transferred to the temperature calibration portion 25, for example, the heating element 23 heats the soaking layer 241, and the heat of the soaking layer 241 is transferred to the temperature calibration portion 25 through the rapid conduction portion 26, so that the temperature calibration portion 25 accurately reflects the temperature of the soaking layer 241, and the temperature detection unit can accurately measure the temperature of the soaking layer 241. Since the reaction sample has a small thickness, the temperature of the reaction sample substantially coincides with the temperature of the soaking layer 241, and the temperature of the reaction sample can be obtained by detecting the temperature of the temperature calibration section 25.

Preferably, one side of the rapid conduction part 26 is connected to one side of the upper conduction assembly 24 near the heating member 23 or to one side of the lower conduction assembly 27 near the heating member 23, and the other side is connected to the temperature calibration part 25. The lower surface of the upper conductive member 24 and the upper surface of the lower conductive member 27 are closest to the heating member 23 and have temperatures closest to the temperature of the heating member 23 first, so that the rapid conduction portion 26 is disposed in such a manner that the temperature of the rapid conduction portion 26 and the temperature of the heating member 23 are consistent in the shortest time. Alternatively, the rapid conduction portion 26 is made of a material having a high thermal conductivity, such as a metal material of copper or aluminum, or a thermally conductive ceramic, or the like. The thermal conductivity of the fast conducting portion 26 is particularly superior to that of the lower conducting assembly 27 to rapidly transfer heat to the temperature calibrating portion 25.

The quick-speed conduction part 26 comprises a patch 261 and one or more second guide posts 262, wherein the patch 261 is attached to one side of the upper conduction assembly 24 close to the heating element 23 or one side of the lower conduction assembly 27 close to the heating element 23, one end of the one or more second guide posts 262 is connected to the patch 261, and the other end of the one or more second guide posts 262 penetrates through the lower conduction assembly 27 and is connected with the temperature calibration part 25. The lower surface of the upper conductive member 24 and the upper surface of the lower conductive member 27 are closest to the heating member 23, and the temperature thereof is closest to the temperature of the heating member 23 first, so that the patches 261 are arranged in such a manner that the temperature of the rapid conductive portion 26 is most rapidly coincident with the temperature of the heating member 23. The patch 261 can increase the contact area of the rapid conduction portion 26 with the upper conduction member 24 or the lower conduction member 27, improving the conduction efficiency. The cross-sectional area of the second guide post 262 can be smaller than that of the patch 261, so that the temperature of the patch 261 can be quickly conducted to the temperature calibration part 25, and meanwhile, the volume of the second guide post 262 can be reduced as much as possible, so that the influence of the quick conduction part 26 on the lower conduction assembly 27 is reduced, and the thermal resistance required by the thermal edge resistance layer 271 is ensured to be generated according to design. Alternatively, the patch 261 and the second guide post 262 are made of a material with high thermal conductivity such as copper, and when the patch 261 and the second guide post 262 are required to be made of an insulating material to avoid the reaction assembly 2 from being shorted, the patch 261 or the second guide post 262 may be made of a material with high thermal conductivity such as ceramic.

It is understood that the temperature calibration portions 25 may be disposed in one-to-one correspondence with the patches 261, and that two temperature calibration portions 25 may be connected to one patch 261. One temperature calibration part 25 may be connected to one second guide post 262, and in order to improve temperature uniformity of the temperature calibration part 25, the temperature calibration part 25 may be connected to a plurality of second guide posts 262.

In other alternative embodiments, as shown in fig. 10, the fast-conducting portion 26 may include one or more first guide posts 263, where the fast-conducting portion 26 is not provided with a patch 261, and one end of the one or more first guide posts 263 is attached to a side of the upper conductive assembly 24 near the heating element 23, and the other end of the one or more first guide posts 263 is connected to the temperature calibration portion 25. The first guide post 263 does not affect other structures of the reaction module 2, and can rapidly transfer heat to the temperature calibration part 25. Optionally, the first guide post 263 is a metal post with high thermal conductivity such as a copper post. One temperature calibration part 25 may be connected to one first guide post 263, and in order to improve temperature uniformity of the temperature calibration part 25, the temperature calibration part 25 may be connected to a plurality of first guide posts 263.

Optionally, the lower conductive component 27 is provided with a third through hole, and the first guide post 263 or the second guide post 262 is disposed in the third through hole. The third through hole can facilitate the placement of the first guide post 263 or the second guide post 262, and meanwhile, the continuity of the lower conductive assembly 27 is not affected, and the performance of the lower conductive assembly 27 is ensured. Optionally, the first guide post 263 or the second guide post 262 is not in contact with the inner wall of the third through hole, so as to avoid heat conduction between the first guide post 263 or the second guide post 262 and the lower conductive component 27, thereby affecting the temperature of the temperature calibration portion 25.

As shown in fig. 7, in order to obtain the resistance of the heating element 23 and to supply power to the heating element 23, the reaction assembly 2 is optionally provided with a plurality of second contacts 20 on the side facing away from the receiving chamber 21, the second contacts 20 being electrically connected to the heating element 23. The current and voltage of the heating element 23 can be obtained through the second contact 20, and thus the resistance value of the heating element 23 can be obtained.

In this embodiment, the second contact 20 enables the reaction component 2 to realize its own temperature measurement function, compared with the conventional structure that can only measure the temperature through the external temperature measurement unit, the embodiment can directly measure the temperature of the reaction component 2, so that the temperature measurement is more accurate and rapid, and the accuracy and control speed of the temperature control system can be improved.

In the prior art, the temperature of the reaction component 2 is detected by the isothermal detection unit of the temperature sensor, but because a certain time is required for heat transfer from the reaction component 2 to the isothermal detection unit, the detection result measured by the isothermal detection unit has a temperature measurement delay of 1-2s under normal conditions, and the temperature change of the reaction component 2 can reach more than 30 ℃ in the rapid temperature rise and fall process, so that the temperature of the reaction component 2 is relatively difficult to control by the isothermal detection unit in the rapid temperature rise and fall process. The temperature of the reaction component 2 is not completely dependent on the temperature value measured by an uncalibrated resistance temperature measurement method, and the reaction component 2 is not completely dependent on the temperature detected by a temperature detection unit, but the temperature of the temperature calibration part 25 of the reaction component 2 and the temperature of the heating element 23 are measured by a resistance temperature measurement method by combining the two methods, so that the temperature of the reaction component 2 can be rapidly and accurately controlled, the aim of accurately controlling the temperature is fulfilled, and the problems of temperature detection delay and large temperature measurement error caused by a common temperature detection method in the prior art are solved.

The embodiment also provides a nucleic acid amplification device, which comprises the reaction component 2, and the nucleic acid amplification device provided by the embodiment can improve the amplification efficiency and shorten the detection time.

As shown in FIG. 3, the nucleic acid amplification apparatus further includes a cooling mechanism 1, the cooling mechanism 1 being for cooling the reaction sample in the accommodation chamber 21 so that the reaction sample can complete a thermal cycle.

Alternatively, the cooling mechanism 1 is located on the side of the reaction module 2 remote from the accommodation chamber 21, and the cooling mechanism 1 cools the portion of the reaction module 2 located between the reaction sample and the cooling mechanism 1 first when cooling the reaction sample.

Alternatively, the cooling mechanism 1 cools the reaction assembly 2 by a fluid flow, which may be a gas or a liquid. Specifically, as shown in fig. 3, the cooling mechanism 1 cools the reaction module 2 by fluid flow. Alternatively, the cooling mechanism 1 includes a cooling body 12, and a cooling flow passage 11 is provided in the cooling body 12 to flow a cooling medium. The cooling medium may be water or other liquid.

As shown in fig. 11, the cooling mechanism 1 may also cool the reaction module 2 by spraying a fluid, alternatively the cooling medium may be water or gas or the like (the arrow in fig. 11 shows a partial flow direction of the cooling medium). For example, the cooling mechanism 1 may comprise a pump and a spray assembly, the spray assembly being in communication with the pump, the pump pumping a high pressure cooling medium into the spray assembly, the spray assembly spraying the cooling medium towards the reaction assembly 2.

In other alternative embodiments, the cooling mechanism 1 may not use a flowable or sprayed medium to cool the reaction assembly 2, but use a solid cooling mode to cool the reaction assembly 2, for example, the cooling mechanism 1 includes a semiconductor refrigerator.

As shown in fig. 12 and 13, optionally, a dodging portion 13 is formed on a side, close to the reaction module 2, of the cooling mechanism 1, and specifically, the dodging portion 13 is used for dodging the temperature calibration portion 25. The escape portion 13 may be a groove, a hole, or the like provided in the cooling mechanism 1. The avoiding portion 13 can prevent the cooling mechanism 1 from affecting the temperature of the temperature calibration portion 25, and ensure that the temperature calibration portion 25 accurately reflects the temperature of the heating element 23.

When the temperature detecting unit is the first contact temperature detecting unit 3, the avoidance portion 13 may also avoid the first contact temperature detecting unit 3.

As shown in fig. 13 and 14, the temperature detection unit in the present embodiment may be not the first contact temperature detection unit 3 but the second contact temperature detection unit 4. Specifically, the nucleic acid amplification apparatus includes the second contact temperature detection unit 4, and the second contact temperature detection unit 4 is capable of being separated from or in contact with the temperature calibration part 25, and is capable of measuring the temperature of the temperature calibration part 25 when it is in contact with the temperature calibration part 25.

Specifically, the second contact type temperature detecting unit 4 may be provided at the escape portion 13 to make full use of space and to ensure that the temperature detecting unit can be brought into contact with the temperature calibrating portion 25. The second contact temperature detecting unit 4 is in elastic contact with the temperature calibrating portion 25, for example, the second contact temperature detecting unit 4 is connected with the cooling mechanism 1 through a spring, so as to achieve elastic contact. The second contact temperature detecting unit 4 is not discarded with the reaction component 2, so that the cost of the reaction component 2 and the detecting cost can be reduced.

The nucleic acid amplification apparatus further includes a resistance detection unit 6, the resistance detection unit 6 being configured to detect a temperature of the heating element 23. Specifically, the resistance detection unit 6 measures the resistance of the heating element 23 by a four-wire method, and the resistance detection unit 6 may be electrically connected to the second contact 20 to detect the current and voltage of the heating element 23. Alternatively, the resistance detection unit 6 is located at the side of the cooling mechanism 1. The second contact 20 is provided at a position that facilitates stable contact with the resistance detection unit 6, and the resistance detection unit 6 can be provided at the side of the cooling mechanism 1, so that the nucleic acid amplification apparatus is compact in structure.

Alternatively, the cooling mechanism 1 is in continuous contact with the reaction module 2, generating heat by the heating member 22, and heating the reaction sample by the soaking layer 241; at the same time, heat is transferred downward through the insulating thermal resistance layer 271 and dissipated through the interface of the heat conduction layer 272 and the cooling mechanism 1. When the heat dissipation power is the same as the heat generation power, the system realizes heat balance, and the reaction sample can be maintained at a specific temperature. By changing the heating power of the inner heating member 22, the temperature of the reaction sample at the time of reaching the thermal equilibrium can be adjusted, thereby realizing the dynamic adjustment of the temperature of the reaction sample.

The reaction module 2 in this embodiment adopts a heating-integrated thin-layer structure to achieve rapid temperature change and heat balance of the reaction sample.

The temperature rising speed of the reaction sample is proportional to the product of the heat capacity and the heating thermal resistance of the reaction sample. Wherein, the heat capacity of the reaction sample comprises the heat capacity of the liquid, and the auxiliary structure such as the heat capacity of the conduction assembly 24 and the cover plate 29 on the side of the heating element 22 close to the reaction sample; the thermal resistance of heating includes the thermal resistance of the various interfaces and materials from the heating element 22 to the reaction sample, as well as the equivalent thermal resistance of the heat transfer within the reaction sample. At the same temperature rise amplitude (same temperature rise variation value), the smaller the product of the heat capacity and the heating resistance of the reaction sample, the faster the temperature rise speed. In this embodiment, since the heating element 22 has a thin layer structure (without conventional heating metal blocks) bonded to the soaking layer 241, and the soaking layer 241 is tightly bonded to the reaction sample and the cover plate 29, the heat capacity of the reaction sample is much smaller than that of the conventional structure; meanwhile, as the reaction component is of an integral structure and has no contact thermal resistance, the structure of the heating element 22 close to the reaction sample adopts high heat conduction materials such as metal, and the thickness of the reaction sample layer is thinner (the longitudinal heat conduction thermal resistance of the reaction sample is small), so that the overall heating thermal resistance is also small. The reaction assembly provided by this embodiment can therefore achieve a much higher rate of temperature rise than conventional pcr heating schemes.

The cooling speed of the reaction sample is proportional to: the product of the heat capacity and the heat dissipation resistance of the reaction sample. Wherein the heat capacity of the reaction sample includes the heat capacity of the liquid, and the auxiliary structure such as the heat capacity of the conduction assembly 24 and the cover plate 29 on the side of the heating element 22 close to the reaction sample; the thermal resistance to dissipate heat includes the thermal resistance from the heating element 22 to the cooling mechanism interface and materials, as well as the equivalent thermal resistance to reflect heat transfer within the sample. Under the same cooling amplitude (same cooling temperature rise change value), the smaller the product of the heat capacity and the heat dissipation thermal resistance of the reaction sample is, the faster the cooling speed is. In this embodiment, since the heating element 22 has a thin layer structure (without conventional heating metal blocks) bonded to the soaking layer 241, and the soaking layer 241 is tightly bonded to the reaction sample and the cover plate 29, the heat capacity of the reaction sample is much smaller than that of the conventional structure; meanwhile, the only contact surface is the reaction component 2 and the cooling mechanism 1, so that the thermal resistance of the insulating thermal resistance layer 271 can be designed to be smaller, the reaction component is of an integral structure, the soaking layer 241 is made of high-heat-conductivity materials such as metal, and the thickness of a reaction sample is thinner (the longitudinal heat conduction thermal resistance of liquid is small), so that the overall heat dissipation thermal resistance is also small. Therefore, the nucleic acid amplification device of the embodiment can realize a cooling speed far higher than that of the conventional pcr heating scheme. Specifically, the nucleic acid amplification apparatus of the present embodiment can achieve a temperature rise and fall rate of 20℃per second or higher.

Selection of the insulating thermal resistance layer 271: in the same size of the reaction module 1 design, the temperature control performance of the sample can be optimized by the design of the insulating thermal resistance layer 271. When the thermal resistance of the insulating thermal resistance layer 271 is large, the heat dissipation is small, and when the same heat balance temperature is realized, the heating power of the heating element 22 is low, which is favorable for reducing the total power consumption of the nucleic acid amplification device, but because the thermal resistance of the insulating thermal resistance layer 271 is large, the heat dissipation thermal resistance is improved, and the cooling speed of the reaction sample is reduced. Similarly, when the thermal resistance of the insulating thermal resistance layer 271 is smaller, the heat dissipation is faster, and when the same heat balance temperature is realized, the heating power of the heating element 22 is higher, and the total power consumption of the nucleic acid amplification device is larger, but at the same time, the temperature reduction speed of the reaction sample is increased due to the reduction of the thermal resistance of the insulating thermal resistance layer 271, which is favorable for shortening the overall time of the pcr flow. The insulating thermal resistance layer 271 can be adjusted in both material and thickness to meet different design requirements. In the general design, a thin layer with the thickness of 0.1-0.3mm can be adopted, the thermal conductivity of the material is selected from the range of 0.2-0.5W/mK, the temperature reduction speed of 20 ℃/sec can be realized corresponding to the reaction sample with the thickness of 0.5mm and the corresponding auxiliary structural design, and the average power of pc temperature control is about 30-50W.

Example two

As shown in FIG. 15, the nucleic acid amplification apparatus of the second embodiment is substantially the same as that of the first embodiment, and differs therefrom in that the temperature detecting unit in the second embodiment is not the first contact temperature detecting unit 3, and the temperature detecting unit is not the non-contact temperature detecting unit 5. In the second embodiment, the nucleic acid amplification apparatus includes a noncontact temperature detection unit 5 for measuring the temperature at the temperature calibration portion 25.

Specifically, the noncontact temperature detecting unit 5 may be provided at the escape portion 13 to make full use of space and to ensure that the noncontact temperature detecting unit 5 detects the temperature of the temperature correcting portion 25. The non-contact temperature detection unit 5 is not discarded with the reaction component 2, so that the cost of the reaction component 2 and the detection cost can be reduced.

Example III

In the third embodiment, a nucleic acid amplification control method is provided, which can be performed by using the reaction module 2 or the nucleic acid amplification apparatus of the first to second embodiments.

In the prior art, the temperature of the reaction component is detected by the isothermal detection unit of the temperature sensor, but because heat is transferred from the reaction component 2 to the isothermal detection unit for a certain time, the detection result measured by the isothermal detection unit has a temperature measurement delay of 1-2s under normal conditions, and the temperature change of the reaction component 2 can reach more than 30 ℃ in the rapid temperature rise and fall process, so that the temperature of the reaction component 2 is relatively difficult to control by the isothermal detection unit in the rapid temperature rise and fall process.

There is a specific relation between the resistance of the heating element 23 and its temperature, so that the real-time resistance change of the heating element 23 of the reaction assembly 2 is measured while heating, and the average temperature of the heating element 23 is deduced by the temperature coefficient of resistance and the resistance value at the nominal temperature (the resistance value at the nominal temperature is simply referred to as the nominal resistance value, and the nominal resistance means that the asserted (or noted) resistance value is true at this temperature, wherein this temperature is the nominal temperature, and the nominal temperature can be arbitrarily selected according to the requirement). The temperature shows the current temperature of the reaction component 2 in real time without delay, so that the temperature can be used for quickly feedback-controlling the temperature of the reaction component 2 and a reaction sample. The disadvantage of this method is that for the same type of resistance, such as copper wire resistance, the nominal resistance and the temperature coefficient of resistance differ slightly between the resistances, resulting in a slight difference between the temperature coefficient of resistance and the nominal resistance between the individual heating elements 23, which may cause temperature measurement errors.

In this embodiment, in order to avoid the defects of the two temperature measurement methods described above and to improve the accuracy of temperature control, as shown in fig. 16 and 17, the nucleic acid amplification control method includes: the reaction assembly 2 is controlled by a resistance temperature measurement method and a double temperature measurement method for calibrating the temperature of the reaction assembly 2.

The temperature value measured by the uncalibrated resistance temperature measurement method is not completely relied on, and the temperature of the reaction component 2 is not completely controlled by the temperature detection unit, but the temperature of the reaction component 2 is calibrated by combining the two, so that the defects are overcome, the temperature of the reaction component 2 is rapidly and accurately controlled, and the aim of accurately controlling the temperature is fulfilled.

The dual temperature measurement mode control reaction assembly 2 for calibrating the temperature of the reaction assembly 2 by a resistance temperature measurement method comprises:

and measuring the temperature calibration value of the reaction component 2 by a temperature detection unit, and acquiring the temperature value corresponding to the resistance value of the reaction component 2 by the temperature calibration value. That is, the temperature detecting unit detects the temperature calibration value of the reaction component 2, and the temperature measurement by the resistance temperature measurement method is calibrated by combining the temperature calibration value with the voltage and the current of the heating element 23 of the reaction component 2 detected by the resistance temperature measurement method. Preferably, the temperature detecting unit detects the temperature at the temperature calibrating portion 25 of the reaction component 2, thereby obtaining a temperature calibration value. It will be appreciated that the temperature calibration value is an artificially selected temperature value that is the current actual temperature of the reaction block 2.

Specifically, the temperature calibration value is combined with the resistance value of the heating element 23 under the temperature calibration value to obtain the resistance temperature coefficient and the nominal resistance value corresponding to the specific heating element 23, and the actual temperature of the reaction component 2 can be obtained through the resistance temperature coefficient and the nominal resistance value when the temperature is measured by using a resistance temperature measurement method.

The temperature measurement is realized by a resistance temperature measurement method and a double temperature measurement mode for calibrating the temperature of the reaction component 2, and comprises the following steps:

Obtaining at least two different temperature calibration values; detecting a first voltage and a first current of the reaction component 2 under a temperature calibration value, and obtaining a first resistance value of the reaction component 2 according to the first voltage and the first current; detecting a second voltage and a second current of the reaction component 2 at another temperature calibration value, and obtaining a second resistance value of the reaction component 2 according to the second voltage and the second current;

Obtaining a resistance temperature coefficient and a nominal resistance value of the reaction component 2 at least according to the first resistance value, the second resistance value, a temperature calibration value corresponding to the first resistance value and a temperature calibration value corresponding to the second resistance value;

the current and voltage of the reaction module 2 are continuously detected, and the reaction module 2 is controlled according to the temperature coefficient of resistance and the nominal resistance value.

It will be appreciated that the at least two different temperature calibrations obtained are read when the temperature detection unit is substantially in agreement with the temperature of the reaction assembly 2, which temperature is considered to be the actual temperature of the reaction assembly 2. At this time, the corresponding current and voltage of the reaction component 2 are read, and the temperature coefficient of resistance and the nominal resistance value are reversely deduced, that is, calibration of the temperature coefficient of resistance and the nominal resistance value is achieved, so that the temperature of the reaction component 2 can be considered to be the accurate temperature of the reaction component 2 when the temperature of the reaction component 2 is obtained according to the current and voltage of the reaction component 2 later.

Still further, according to the formula r=r ₀ (1+αΔt) (where Δt=t-T ₀, R is the corresponding resistance value of the heating element 23 at the temperature T, T ₀ is the nominal temperature, R ₀ is the nominal resistance value, α is the temperature coefficient of resistance of the material) to calibrate the temperature coefficient of resistance and nominal resistance of the reaction block 2. that is, the temperature detecting unit detects a first temperature calibration value T ₁, the resistance detecting unit 6 detects a first voltage U ₁ and a first current I ₁ of the heating element 23 at a temperature T ₁, From r=u/I, the resistance R ₁ of the heating element 23 at the temperature T ₁ can be obtained; The temperature detection unit then detects a second temperature calibration value T ₂, the resistance detection unit 6 detects a second voltage U ₂ and a second current I ₂ of the heating element 23 at a temperature T ₂, The resistance R ₂ of the heating element 23 at the temperature T ₂ can be obtained from r=u/I, finally from two sets of binary once equations: r ₁＝R₀(1+αΔT₁) and R ₂＝R₀(1+αΔT₂), (wherein, Δt _1＝T₁-T_0;ΔT_2＝T₂-T_0;) yields specific values for α and R ₀. The temperature profile of the reaction component 2 is then obtained according to the formula r=r ₀ (1+αΔt) by continuously measuring the voltage and current of the reaction component 2. Since the formula is the R ₀ and the α value corresponding to the specific heating element 23, the temperature value can be accurately obtained.

It will be appreciated that the greater the difference between the different temperature calibration values, the more accurate the resulting temperature coefficient of resistance and nominal resistance value, and therefore, optionally, the difference between adjacent two temperature calibration values is not less than 20 ℃.

It will be appreciated that the temperature calibration may be detected throughout the nucleic acid amplification process, but that only a few of the temperature calibration values may be selected to calibrate the temperature. Optionally, the temperature value is calibrated using at least two unequal temperature calibration values. Such as measuring two, three, four or more temperature calibration values. The temperature calibration values can be used for obtaining a resistance temperature coefficient and a nominal resistance value, the temperature calibration values can be used for obtaining the resistance temperature coefficient and the nominal resistance value, and the temperature coefficient and the nominal resistance value can be used for obtaining the value which is closer to the actual resistance temperature coefficient and the nominal resistance value, so that the detection precision is further improved.

In order to overcome the problem of inaccurate temperature measurement caused by the temperature measurement delay of 1-2s of the temperature detection unit, the current actual temperature of the reaction component 2 is ensured to be reflected by the temperature calibration value, in this embodiment, after the temperature of the temperature detection unit is consistent with the temperature of the reaction component 2, at least two unequal temperature calibration values for calibrating the temperature value are read. For example, after stopping for 1-2s at a certain temperature, the temperature calibration value used for calibrating the temperature value is read to ensure the accuracy of temperature calibration. For another example, if the temperature calibration value is obtained in the temperature increasing and decreasing process, the speed of increasing and decreasing is slowed down so that the temperature of the temperature detecting unit is consistent with the temperature of the reaction component 2, or the temperature change is suspended in the temperature increasing and decreasing process, and the temperature increasing and decreasing process is executed again after the temperature calibration value is obtained; if the temperature calibration value is obtained in the heat preservation stage, the temperature detected by the temperature detection unit is read after the heat preservation is carried out for a certain period of time, for example, the temperature detected by the temperature detection unit is read after the heat preservation is carried out for 2s, 3s or 8s, etc.

It will be appreciated that the first warming up of the nucleic acid amplification process is the warming up to the pretreatment stage. The temperature rising process, the high-temperature denaturation stage, the cooling process, the low-temperature annealing stage, the temperature rising process and the temperature-adaptive extension stage are used as a primary amplification cycle.

Optionally, the timing for acquiring the temperature calibration value for calibrating the temperature value is: before the nucleic acid amplification process, during the first temperature increase of the nucleic acid amplification process and/or during the first amplification cycle of the nucleic acid amplification process. That is, the temperature calibration value is preferably calibrated using the calibration temperature value measured in the above-described period of time. The temperature calibration value used for calibrating the temperature value is measured during or before the first amplification cycle, and the calibration of the temperature value is completed during the first amplification cycle, so that the accuracy of the subsequent temperature control can be ensured.

It will be appreciated that the temperature detection unit detects the room temperature of the reaction component 2 prior to the progress of nucleic acid amplification, the room temperature being used as a temperature calibration value for one of the calibrated temperature values. It is also possible to make the reaction component 2 have a certain temperature change before the nucleic acid amplification process, and the temperature calibration value required for calibrating the temperature value is obtained by the temperature detection unit, but in this case, since it takes a certain time for the temperature of the temperature detection unit to coincide with the temperature of the reaction component 2, the detection time is increased, and therefore, it is preferable to measure the temperature calibration value in combination with the room temperature of the reaction component 2 detected by the temperature detection unit and during the first temperature increase of the nucleic acid amplification process and/or during the first amplification cycle of the nucleic acid amplification process.

As shown in fig. 16 and 17, alternatively, before the temperature value is calibrated, the temperature of the reaction module 2 is controlled according to the RT temperature curve preset for the reaction module 2, so that an excessive deviation of the temperature of the reaction module 2 due to rapid temperature change of the reaction module 2 can be avoided. After calibrating the temperature values, the temperature values detected by the resistance thermometry are continued to control the reaction module 2. Of course, in other alternative embodiments, the temperature of the reaction module 2 measured by the temperature detecting unit may control the reaction module 2 before calibrating the temperature value, and at this time, the temperature rise and fall rate is reduced during the temperature rise and fall phases, so as to avoid the temperature being too high or too low.

In order to more clearly describe the nucleic acid amplification control method in this embodiment, a procedure of calibrating a resistance thermometry by a temperature detection unit in an actual detection is shown in conjunction with FIG. 17. Before calibrating the temperature values, an initial RT temperature profile, i.e. a temperature preset profile, is preset, and then a small current, e.g. less than 1 ma, is applied to the heating element 23 of the reaction assembly 2. Wherein a small current is applied in order to read the resistance of the heating element 23 without heating the heating element 23.

First calibration: the temperature detecting unit detects a first temperature calibration value T ₁, and the resistance detecting unit detects a first voltage U ₁ and a first current I ₁ of the heating element 23 at the temperature of T ₁, and can obtain a resistance R ₁ of the heating element 23 at the temperature of T ₁ according to r=u/I.

Second calibration: then the temperature detecting unit detects a second temperature calibration value T ₂, and the resistance detecting unit 5 detects a second voltage U ₂ and a second current I ₂ of the heating element 23 at the temperature T ₂, and obtains a resistance R ₂ of the heating element 23 at the temperature T ₂ according to r=u/I.

Finally, according to two sets of binary once equations: r ₁＝R₀(1+αΔT₁) and R ₂＝R₀(1+αΔT₂) to obtain specific values of R ₀ and alpha, namely an accurate R-T curve is obtained, and the temperature of the heating element 23 measured by a resistance temperature measurement method can be used as feedback for accurately controlling the temperature.

The temperature calibration value can be detected in the whole process of nucleic acid amplification, so that the temperature can be calibrated for multiple times in the subsequent process, and the detection precision is further improved.

With continued reference to fig. 17, when the first temperature calibration value is obtained, the temperature calibration value may be regarded as a nominal resistance value, and thus, according to the temperature calibration value, the R ₀ in the formula r=r ₀ (1+αΔt) may be corrected for the first time, so as to implement the first calibration of the temperature value measured by the resistance temperature measurement method (such as the temperature value curve measured by the resistance temperature measurement method in fig. 17 fluctuates once after the first calibration is performed), then the temperature value obtained by the resistance temperature measurement method or the temperature value measured by the temperature detection unit controls the reaction component 2 to be raised to the first stage of pretreatment, and after the reaction component 2 is raised to the first stage of pretreatment for 8 seconds (not limited to 8 seconds, but may be any duration exceeding 2 seconds and less than the first stage of pretreatment), the second temperature calibration value for calibrating the temperature value is obtained, the temperature value detected by the resistance temperature measurement method is calibrated for the second time according to the two temperature calibration values, and then the temperature value obtained by the resistance temperature measurement method is controlled to continue. It will be appreciated that when the first temperature calibration value is room temperature, the temperature value measured by the resistance temperature measurement method may be calibrated for the first time, then a second temperature calibration value may be obtained at any temperature higher than room temperature, and the temperature value detected by the resistance temperature measurement method may be calibrated for the second time according to the two temperature calibration values, thereby completing the calibration of the temperature value.

Because heat is transferred from the reaction component 2 to the temperature detection unit for a certain time, the temperature measurement delay of 1-2s exists in the detection result measured by the temperature detection unit under normal conditions, and the temperature change of the reaction component 2 can reach more than 30 ℃ in the rapid temperature rise and fall process for 1-2s, so that the control of the reaction component 2 by the temperature detection unit is relatively difficult in the rapid temperature rise and fall process. The resistance temperature measurement method can measure the real-time resistance change of the resistor while heating, and deduce the average temperature of the resistor through the temperature coefficient of the resistor and the nominal resistance. The temperature shows the current temperature of the reaction component 2 in real time without delay, so that the temperature can be used for rapidly controlling the temperature of a sample. Since the temperature measurement method has the disadvantage that the temperature coefficient of resistance of a single resistor is slightly different from the nominal resistance value, temperature measurement errors may be caused. The temperature value measured by the uncalibrated resistance temperature measurement method is not completely relied on, and the temperature of the reaction component 2 is not completely controlled by the temperature detection unit, but the temperature of the reaction component 2 is calibrated by combining the two, so that the defects are overcome, the temperature of the reaction component 2 is rapidly and accurately controlled, and the aim of accurately controlling the temperature is fulfilled.

While the invention has been described in detail in the foregoing general description, embodiments and experiments, it will be apparent to those skilled in the art that modifications and improvements can be made thereto. Accordingly, such modifications or improvements may be made without departing from the spirit of the invention and are intended to be within the scope of the invention as claimed.

Claims

1. A reaction module comprising a receiving chamber (21) for receiving a reaction sample and a heater (200) for heating the reaction sample.

2. The reaction assembly according to claim 1, wherein the heater (200) is in direct contact with the reaction sample within the receiving chamber (21).

3. The reaction module according to claim 2, characterized in that at least part of the upper surface of the heater (200) is in direct contact with the reaction sample in the receiving chamber (21), or

A groove (201) is formed on the surface of the heater (200), and at least part of the wall surface (202) of the groove (201) is in direct contact with the reaction sample in the accommodating cavity (21).

4. The reaction assembly according to claim 1, wherein the heater (200) comprises a soaking layer (241), the soaking layer (241) being in direct contact with the reaction sample within the receiving cavity (21).

5. The reaction assembly according to claim 4, wherein the heater (200) further comprises a heating member (23) and a temperature calibration part (25) for the temperature detection unit to detect the temperature.

6. The reaction assembly of claim 5, wherein the heater (200) further comprises an upper conductive assembly (24) and a lower conductive assembly (27), the heating element (23) being sandwiched between the upper conductive assembly (24) and the lower conductive assembly (27), the upper conductive assembly (24) comprising the soaking layer (241).

7. The reaction assembly according to claim 6, wherein the temperature calibration part (25) is connected to the upper conductive assembly (24) or the lower conductive assembly (27).

8. The reaction module according to claim 7, wherein the temperature calibration part (25) is connected to a side of the upper conductive module (24) close to the lower conductive module (27), and the lower conductive module (27) is provided with a first through hole (273) disposed opposite to the temperature calibration part (25).

9. The reaction assembly of claim 6, wherein the temperature calibration part (25) is part of the upper conductive assembly (24) or the lower conductive assembly (27).

10. The reaction module according to claim 9, wherein the lower conductive member (27) is provided with a second through hole (274) in a thickness direction of the reaction module, and a surface of the upper conductive member (24) facing the second through hole (274) is the temperature calibration portion (25).

11. The reaction assembly according to claim 6, further comprising a fast conducting portion (26) for conducting heat of the heating element (23) to the temperature calibration portion (25).

12. Reaction module according to claim 11, wherein one side of the fast conducting part (26) is connected to one side of the upper conducting component (24) close to the heating element (23) or to one side of the lower conducting component (27) close to the heating element (23) and the other side is connected to the temperature calibration part (25).

13. A reaction assembly according to claim 11, wherein the temperature calibration part (25) is located at a side of the lower conduction assembly (27) remote from the heating element (23).

14. The reaction module according to claim 11, wherein a receiving groove (275) is provided on a side of the lower conduction module (27) remote from the heating member, and the temperature calibration part (25) is located in the receiving groove (275) and is connected to a groove bottom of the receiving groove (275).

15. The reaction assembly according to claim 13, wherein the fast conducting part (26) comprises one or more first guide posts (263), one end of the one or more first guide posts (263) is attached to a side of the upper conducting assembly (24) close to the heating element (23) or to a side of the lower conducting assembly (27) close to the heating element (23), and the other end of the one or more first guide posts (263) is connected to the temperature calibrating part (25).

16. The reaction assembly according to claim 13, wherein the fast conducting part (26) comprises a patch (261) and one or more second guide posts (262), the patch (261) is attached to one side of the upper conducting assembly (24) close to the heating element (23) or one side of the lower conducting assembly (27) close to the heating element (23), one end of the one or more second guide posts (262) is connected to the patch (261), and the other end of the one or more second guide posts (262) is penetrated through the lower conducting assembly (27) and is connected to the temperature calibration part (25).

17. Reaction module according to claim 15 or 16, wherein the lower conductive component (27) is provided with a third through hole, the guide post being arranged in the third through hole.

18. The reaction assembly of claim 6, wherein the lower conductive assembly (27) further comprises an insulating thermal resistance layer (271).

19. The reaction assembly of claim 18, wherein the lower conductive assembly (27) further comprises a thermally conductive layer (272), the thermally conductive layer (272) being located on a side of the insulating thermal resistance layer (271) remote from the heating element (23).

20. The reaction assembly of claim 6, wherein the soaking layer (241) is made of a conductive or insulating material.

21. The reaction assembly according to claim 20, wherein when the soaking layer (241) is made of an electrically conductive material, the upper conductive assembly (24) further comprises an insulating layer (242), the insulating layer (242) being located between the heating element (23) and the soaking layer (241);

when the soaking layer (241) is made of an insulating material, the soaking layer (241) is adjacent to the heating member (23).

22. A reaction assembly according to claim 1, wherein the containing chamber (21) comprises a bottom wall (22);

The bottom wall (22) is in close contact with at least part of the upper surface of the heater (200), or

A groove (201) is formed on the surface of the heater (200), and at least part of the wall surface (202) of the groove (201) is in close contact with the bottom wall (22).

23. A reaction assembly according to claim 22, wherein a flexible heat conducting member (203) is arranged between the bottom wall (22) and the heater (200).

24. Reaction module according to claim 1, wherein the receiving chamber (21) is of flat construction.

25. A reaction assembly according to claim 1, wherein the receiving chamber (21) is polygonal, circular or elliptical in cross-section.

26. A reaction assembly according to any one of claims 5-8, 11-16, 18, 19, characterized in that the reaction assembly comprises a first contact temperature detection unit (3) connected to the temperature calibration part (25) and adapted to measure the temperature at the temperature calibration part (25).

27. A nucleic acid amplification apparatus comprising the reaction module (2) according to any one of claims 1 to 26.

28. A nucleic acid amplification device characterized in that,

A reaction assembly (2) comprising a second contact temperature detection unit (4) and any of claims 5-8, 11-19, said second contact temperature detection unit (4) being capable of being separated from or in contact with said temperature calibration portion (25), and being capable of measuring the temperature of said temperature calibration portion (25) when it is in contact with said temperature calibration portion (25).

29. A nucleic acid amplification device, characterized by comprising a non-contact temperature detection unit (5) and a reaction assembly (2) according to any one of claims 5-19, for measuring the temperature at the temperature calibration part (25).

30. The nucleic acid amplification apparatus as set forth in any one of claims 27 to 29, further comprising a cooling mechanism (1), the cooling mechanism (1) being for cooling the reaction sample in the accommodating chamber (21).

31. The nucleic acid amplification apparatus according to claim 30, wherein a side of the cooling mechanism (1) adjacent to the reaction module (2) is provided with a relief portion (13).

32. The nucleic acid amplification apparatus as set forth in claim 30, wherein the cooling mechanism (1) cools the reaction component (2) by a fluid.

33. The nucleic acid amplification apparatus as set forth in claim 32, wherein the cooling mechanism (1) cools the reaction component (2) by spraying a fluid or a fluid flow.

34. The nucleic acid amplification apparatus as set forth in claim 33, wherein when the cooling mechanism (1) cools the reaction component (2) by a fluid flow, the cooling mechanism (1) includes a cooling body (12), and a cooling flow passage (11) is provided in the cooling body (12) to flow a cooling medium.

35. The nucleic acid amplification apparatus of claim 30, wherein the cooling mechanism (1) cools the reaction component (2) by means of solid cooling.

36. The nucleic acid amplification apparatus as set forth in any one of claims 27 to 29, further comprising a resistance detection unit (6) for detecting a temperature of the reaction component (2).

37. The nucleic acid amplification apparatus as set forth in claim 36, wherein the resistance detection unit (6) obtains the temperature value of the reaction component (2) by detecting the resistance of the reaction component (2).

38. A nucleic acid amplification control method, characterized in that the nucleic acid amplification control method is performed using the reaction module (2) according to any one of claims 1 to 26, or the nucleic acid amplification apparatus according to claims 26 to 37;

The nucleic acid amplification control method comprises: the reaction assembly (2) is controlled by a resistance temperature measurement method and a double temperature measurement method for calibrating the temperature of the reaction assembly (2).

39. The method according to claim 38, wherein controlling the reaction component (2) by a resistance thermometry method and a dual thermometry method of calibrating a temperature of the reaction component (2) comprises:

and measuring a temperature calibration value of the reaction component (2) through a temperature detection unit, and calibrating the temperature value acquired through the resistance value of the reaction component (2) through the temperature calibration value.

40. The method according to claim 39, wherein a temperature coefficient of resistance and a nominal resistance value of the reaction component (2) are obtained from the temperature calibration value to calibrate the temperature value.

41. The method according to claim 39, wherein the temperature value is calibrated by measuring the temperature calibration value before the nucleic acid amplification process, during the first temperature increase of the nucleic acid amplification process, and/or during the first amplification cycle of the nucleic acid amplification process.

42. The method according to claim 39, wherein the temperature values are calibrated using at least two different temperature calibration values.

43. The method according to any one of claims 39 to 42, wherein controlling the reaction component (2) by a resistance thermometry method and a dual thermometry method for calibrating the temperature of the reaction component (2) comprises the steps of:

obtaining at least two different temperature calibration values; detecting a first voltage and a first current of the reaction component (2) at the temperature calibration value, and obtaining a first resistance value of the reaction component (2) according to the first voltage and the first current; detecting a second voltage and a second current of the reaction component (2) at the other temperature calibration value, and obtaining a second resistance value of the reaction component (2) according to the second voltage and the second current;

Obtaining a temperature coefficient of resistance of the reaction component (2) and a resistance at a nominal temperature according to at least the first resistance value, the second resistance value and the corresponding temperature calibration value;

The current and voltage of the reaction component (2) are continuously detected, and the reaction component (2) is controlled according to the temperature coefficient of resistance and the resistance at the nominal temperature.

44. The method according to any one of claims 39 to 42, wherein the temperature of the reaction component (2) measured by the temperature detecting unit controls the reaction component (2) before the temperature value is calibrated, or the temperature of the reaction component (2) is controlled according to an RT temperature profile preset for the reaction component (2).

45. The method of nucleic acid amplification control of any one of claims 39 to 42, wherein, according to the formula: -r=r ₀ (1+αΔt) calibrating the temperature coefficient of resistance and the nominal resistance value of the reaction component (2) and-by continuously measuring the voltage and current of the reaction component (2), the temperature profile of the reaction component (2) is obtained according to the formula, wherein R ₀ is the nominal resistance value and α is the temperature coefficient of resistance of the material.