A temperature control device for PCR instrument well plate partition

Abstract

The invention relates to a temperature control device for partitioning a well plate of a PCR instrument. The device comprises a temperature-averaging plate, a semiconductor thermoelectric module, a temperature control module and a detection module. The temperature-averaging plate is laid directly below a reaction plate and is in close contact with the bottom of each partition of the reaction plate respectively; the semiconductor thermoelectric module is used for directly heating the temperature-averaging plate under the control of the temperature control module; the temperature-averaging plate comprises a plurality of temperature-averaging sub-plates; the detection module is used for detecting the current temperature of each partition of the reaction plate; the temperature control module comprises a correction unit, the correction unit is used for calculating the heating power of each partition of the reaction plate according to the current temperature and set temperature of each partition of the reaction plate; and the heating power of each partition of the reaction plate is corrected according to a power correction function to obtain a heating correction power; the temperature control module controls the semiconductor thermoelectric module to heat each corresponding partition of the reaction plate according to the calculated heating correction power.

Description

Technical Field

[0001] The invention relates to the technical field of PCR instruments, and in particular to a temperature control device for partitioning a well plate of a PCR instrument. Background Art

[0002] Fluorescence quantitative PCR (Real-time qPCR) technology refers to a method that adds fluorescent genes to the PCR reaction system, uses the accumulation of fluorescent signals to monitor the entire PCR process in real time, and finally uses a standard curve to quantitatively analyze the unknown template. The number of DNA copies generated during the PCR reaction increases exponentially. As the number of reaction cycles increases, the final PCR reaction no longer generates templates exponentially, and enters a plateau phase. In traditional PCR, gel electrophoresis is often used to separate and fluorescent staining is used to detect the final amplification product of the PCR reaction. Therefore, there is unreliability in using this endpoint method to quantify PCR products. In Real-time qPCR, the entire PCR reaction amplification process is monitored in real time and the amplification-related fluorescent signals are continuously analyzed. As the reaction time progresses, the changes in the monitored fluorescent signals can be plotted into a curve. At this stage, Real-time qPCR is mainly achieved through fluorescent quantitative PCR instruments.

[0003] Fluorescence quantitative PCR instruments have high requirements for temperature control accuracy, especially the temperature uniformity between different partitions, which directly affects the efficiency of DNA fragment amplification. Summary of the invention

[0004] The present invention aims to provide a temperature control device and a control method for partitioning a well plate of a PCR instrument. The technical problems to be solved at least include how to improve the temperature control uniformity of the well plate partitions of the PCR instrument.

[0005] In order to achieve the above-mentioned purpose, the present invention provides a temperature control device for partitioning a well plate of a PCR instrument, comprising a temperature-averaging plate, a semiconductor thermoelectric module, a temperature control module and a detection module, wherein the temperature-averaging plate is laid directly below a reaction plate and is in close contact with the bottom of each partition of the reaction plate; the semiconductor thermoelectric module is used to directly heat the temperature-averaging plate under the control of the temperature control module; the temperature-averaging plate comprises a plurality of temperature-averaging sub-plates, the number of which is the same as the number of partitions of the reaction plate, so that a temperature-averaging sub-plate is arranged under each partition of the reaction plate; the detection module is used to detect the current temperature of each partition of the reaction plate; the temperature control module comprises a correction unit, and the correction unit is used to calculate the heating power of each partition of the reaction plate according to the current temperature and set temperature of each partition of the reaction plate; and the heating power of each partition of the reaction plate is corrected according to the power correction function to obtain the heating correction power; the power correction function is:

[0006]

[0007] Wherein, P(x,y) is the heating power of the partition with coordinates (x,y) in each partition of the reaction plate; P(x+1,y) is the heating power of the partition with coordinates (x+1,y) in each partition of the reaction plate; P(x-1,y) is the heating power of the partition with coordinates (x-1,y) in each partition of the reaction plate; P(x,y+1) is the heating power of the partition with coordinates (x,y+1) in each partition of the reaction plate; P(x,y-1) is the heating power of the partition with coordinates (x,y-1) in each partition of the reaction plate; δ is the empirical correction coefficient;

[0008] The temperature control module controls the semiconductor thermoelectric module to heat each corresponding partition of the reaction plate according to the calculated heating correction power.

[0009] Preferably, the value of the empirical correction coefficient δ is between 1.025 and 1.038.

[0010] Preferably, thermally conductive silicone grease is provided between the upper surface of the temperature averaging plate and the bottom of each partition of the reaction plate to ensure that there is no air thermal resistance between the temperature averaging plate and the reaction plate.

[0011] Preferably, the temperature homogenizing plate is made of copper having a thermal conductivity greater than or equal to 386.4 W / (m·K) at room temperature.

[0012] Preferably, the material of the temperature homogenizing plate is silver having a thermal conductivity greater than or equal to 426.2 W / (m·K) at room temperature.

[0013] Preferably, a heat-insulating isolation plate is provided between adjacent said temperature-averaging sub-plates; said heat-insulating isolation plate is used to prevent temperature transfer between adjacent said temperature-averaging sub-plates.

[0014] Preferably, after a heat insulating isolation plate is provided between adjacent temperature-averaging sub-plates, the value of the empirical correction coefficient δ is fixed between 1.002 and 1.005.

[0015] Preferably, thermal insulation pads are provided between adjacent different partitions of the reaction plate; the thermal insulation pads are used to prevent temperature transfer between adjacent different partitions of the reaction plate.

[0016] Preferably, after heat insulating spacers are arranged between adjacent different partitions of the reaction plate, the value of the empirical correction coefficient δ is fixed between 0.993 and 0.997.

[0017] Further preferably, when insulating isolation plates are provided between adjacent temperature-averaging sub-plates and insulating isolation pads are provided between adjacent different sub-zones of the reaction plate, the value of the empirical correction coefficient δ is fixed between 0.985 and 0.991.

[0018] Compared with the prior art, the present invention has the following beneficial effects:

[0019] The temperature control device for PCR instrument well plate partitions of the present invention corrects the heating power of each partition of the reaction plate through a power correction function, which can eliminate the mutual influence between adjacent partitions during the heating process and improve the temperature uniformity between different partitions. After correction by the power correction function, the value of the empirical correction coefficient δ is fixed between 1.025 and 1.038, and the temperature uniformity of each partition of the entire reaction plate can be stabilized below ±0.3°C.

[0020] The present invention can ensure good heat conduction between the upper surface of the temperature equalizing plate and the bottom of each partition of the reaction plate by eliminating the air thermal resistance, which is beneficial to improving the temperature uniformity of each partition of the entire reaction plate. The experimental results show that the temperature uniformity of each partition of the entire reaction plate is improved by at least 0.06°C between the experimental group using thermal grease and the control group without thermal grease. When other conditions are the same, the temperature uniformity of each partition of the entire reaction plate is improved by at least 0.03°C between the experimental group using thermal insulation isolation board and the control group without thermal insulation isolation board. The temperature uniformity of each partition of the entire reaction plate is improved by at least 0.05°C between the experimental group using thermal insulation isolation board and the control group without thermal insulation isolation board. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] The accompanying drawings are used to provide a further understanding of the technical solution of the present invention and constitute a part of the specification. Together with the specific implementation methods of the present application, they are used to explain the technical solution of the present invention and do not constitute a limitation on the technical solution of the present invention.

[0022] Figure 1 It is a structural schematic diagram of the temperature control device for the PCR instrument well plate partitions of the present invention.

[0023] Figure 2 It is a top view of a position layout of partitions with coordinates (x, y) in the reaction plate. DETAILED DESCRIPTION

[0024] The present invention is described in more detail hereinafter to facilitate understanding of the present invention.

[0025] The temperature control device for the partition of the PCR instrument well plate of the present invention comprises a temperature averaging plate 1, a semiconductor thermoelectric module 2, a temperature control module 3 and a detection module 4, wherein the temperature averaging plate 1 is laid directly below the reaction plate 5 and is in close contact with the bottom of each partition of the reaction plate; the semiconductor thermoelectric module 2 is used to directly heat the temperature averaging plate 1 under the control of the temperature control module 3; the temperature averaging plate 1 comprises a plurality of temperature averaging sub-plates, the number of which is the same as the number of partitions of the reaction plate, so that a temperature averaging sub-plate is arranged under each partition of the reaction plate; the detection module 4 is used to detect the current temperature of each partition of the reaction plate; the temperature control module 3 comprises a correction unit, which is used to calculate the heating power of each partition of the reaction plate according to the current temperature and set temperature of each partition of the reaction plate; and the heating power of each partition of the reaction plate is corrected according to the power correction function to obtain the heating correction power; the power correction function is:

[0026]

[0027] Wherein, P(x,y) is the heating power of the partition with coordinates (x,y) in each partition of the reaction plate; P(x+1,y) is the heating power of the partition with coordinates (x+1,y) in each partition of the reaction plate; P(x-1,y) is the heating power of the partition with coordinates (x-1,y) in each partition of the reaction plate; P(x,y+1) is the heating power of the partition with coordinates (x,y+1) in each partition of the reaction plate; P(x,y-1) is the heating power of the partition with coordinates (x,y-1) in each partition of the reaction plate; δ is the empirical correction coefficient;

[0028] The temperature control module controls the semiconductor thermoelectric module 2 to heat each corresponding partition of the reaction plate according to the calculated heating correction power.

[0029] Preferably, the value of the empirical correction coefficient δ is between 1.025 and 1.038.

[0030] At present, the temperature uniformity of the temperature control system of the first-line fluorescent quantitative PCR instrument abroad is above ±0.2℃, and the temperature uniformity of the temperature control system of the domestic fluorescent quantitative PCR instrument is about ±0.5℃. During the development process, the applicant found that one of the reasons for the poor temperature uniformity between different partitions is that adjacent partitions will have mutual influence during the heating process. In order to eliminate the mutual influence between adjacent partitions during the heating process, the applicant obtained the above-mentioned power correction function through a large number of experiments and combined with Octave for function fitting.

[0031] In actual use, a rectangular coordinate system can be established with any partition of the reaction plate as the origin, for example, with the partition at the edge of the lower left corner as the origin.

[0032] The test results show that after the power correction function is used, the value of the empirical correction coefficient δ is fixed between 1.025 and 1.038, and the temperature uniformity of each partition of the entire reaction plate can be stabilized below ±0.3°C.

[0033] Preferably, thermal conductive silicone grease is provided between the upper surface of the temperature averaging plate 1 and the bottom of each partition of the reaction plate to ensure that there is no air thermal resistance between the temperature averaging plate and the reaction plate.

[0034] By eliminating the air thermal resistance, it is possible to ensure good heat conduction between the upper surface of the temperature plate 1 and the bottom of each partition of the reaction plate, which is conducive to improving the temperature uniformity of each partition of the entire reaction plate. The experimental results show that the temperature uniformity of each partition of the entire reaction plate is improved by at least 0.06°C between the experimental group using thermal conductive silicone grease and the control group without thermal conductive silicone grease.

[0035] Preferably, the temperature homogenizing plate is made of copper having a thermal conductivity greater than or equal to 386.4 W / (m·K) at room temperature.

[0036] During the research and development process, the applicant found that the material of the temperature plate has a significant impact on the temperature uniformity of each partition of the entire reaction plate. The experimental results show that the temperature uniformity of each partition of the entire reaction plate is at least 0.04°C higher in the experimental group using copper with a thermal conductivity greater than or equal to 386.4W / (m·K) at room temperature than in the control group without a temperature plate.

[0037] Preferably, the material of the temperature homogenizing plate is silver having a thermal conductivity greater than or equal to 426.2 W / (m·K) at room temperature.

[0038] The experimental results show that the temperature uniformity of each partition of the entire reaction plate is improved by at least 0.09°C between the experimental group using silver with a thermal conductivity greater than or equal to 426.2W / (m·K) at room temperature as the temperature equalizing plate and the control group without the temperature equalizing plate.

[0039] Preferably, a heat insulating isolation plate is provided between adjacent temperature-averaging sub-plates, and the heat insulating isolation plate is used to prevent temperature transfer between adjacent temperature-averaging sub-plates.

[0040] As mentioned above, one of the reasons for the poor temperature uniformity between different partitions is the mutual influence between adjacent partitions during the heating process. In order to eliminate the mutual influence between adjacent partitions during the heating process, the previous method is to use a power correction function to correct the heating power of each partition of the reaction plate. Another idea is to reduce the mutual influence between adjacent partitions during the heating process from the hardware aspect. The specific method is to set an insulating isolation plate between adjacent temperature-averaging panels. The experimental results show that when other conditions are the same, the temperature uniformity of each partition of the entire reaction plate is improved by at least 0.05°C between the experimental group using an insulating isolation plate and the control group without an insulating isolation plate.

[0041] It should be noted that the use of a power correction function to correct the heating power of each partition of the reaction plate and the provision of an insulating isolation plate between adjacent said temperature-averaging sub-plates do not conflict with each other, and the two can exist at the same time. However, after the insulating isolation plate is provided between adjacent said temperature-averaging sub-plates, the value of the empirical correction coefficient δ needs to be corrected, and preferably, the value of the empirical correction coefficient δ is fixed between 1.002 and 1.005.

[0042] Preferably, a heat-insulating spacer is provided between adjacent different partitions of the reaction plate, and the heat-insulating spacer is used to prevent temperature transfer between adjacent different partitions of the reaction plate.

[0043] Similar to the measure of setting up an insulating isolation plate between adjacent temperature-averaging sub-plates, setting up an insulating isolation pad between adjacent different partitions of the reaction plate can also reduce the mutual influence between adjacent partitions during the heating process. The experimental results show that when other conditions are the same, the temperature uniformity of each partition of the entire reaction plate is improved by at least 0.03°C between the experimental group using the insulating isolation plate and the control group without the insulating isolation plate.

[0044] There is no conflict between using a power correction function to correct the heating power of each partition of the reaction plate and providing thermal insulation pads between adjacent different partitions of the reaction plate, and the two can exist at the same time.

[0045] After the heat insulating spacers are arranged between the adjacent different partitions of the reaction plate, the value of the empirical correction coefficient δ also needs to be corrected. Preferably, the value of the empirical correction coefficient δ is fixed between 0.993 and 0.997.

[0046] Further preferably, when insulating isolation plates are provided between adjacent said temperature-averaging plates and insulating isolation pads are provided between adjacent different partitions of the reaction plate, the value of the empirical correction coefficient δ needs to be further corrected. Preferably, the value of the empirical correction coefficient δ is fixed between 0.985 and 0.991.

[0047] Further preferably, in actual calculation, the empirical correction coefficient δ may be an intermediate value, such as a numerical range between 0.985 and 0.991, and preferably δ=0.988.

[0048] The present invention also provides a temperature control method for partitioning a PCR instrument plate, which uses the aforementioned temperature control device to perform temperature control, and specifically includes the following steps:

[0049] S1, detecting the current temperature of each partition of the reaction plate;

[0050] S2, calculating the heating power of each partition of the reaction plate according to the current temperature and the set temperature of each partition of the reaction plate;

[0051] S3, correcting the heating power of each partition of the reaction plate according to the power correction function to obtain the heating correction power; the power correction function is:

[0052]

[0053] Wherein, P(x,y) is the heating power of the partition with coordinates (x,y) in each partition of the reaction plate; P(x+1,y) is the heating power of the partition with coordinates (x+1,y) in each partition of the reaction plate; P(x-1,y) is the heating power of the partition with coordinates (x-1,y) in each partition of the reaction plate; P(x,y+1) is the heating power of the partition with coordinates (x,y+1) in each partition of the reaction plate; P(x,y-1) is the heating power of the partition with coordinates (x,y-1) in each partition of the reaction plate; δ is the empirical correction coefficient;

[0054] S4. Heat each corresponding partition of the reaction plate according to the calculated heating correction power.

[0055] Preferably, the specific method for calculating the heating power of each partition of the reaction plate according to the current temperature and the set temperature of each partition of the reaction plate is: obtaining a preset heating time; calculating the heating amount of each partition of the reaction plate according to the set temperature and the current temperature and the specific heat capacity of each partition of the reaction plate; and dividing the heating amount of each partition of the reaction plate by the heating time to obtain the heating power of each partition of the reaction plate.

[0056] Specifically, taking one of the partitions of the reaction plate (denoted as partition A) as an example, the heating power of partition A is calculated using the following formula:

[0057] P A =Q A / t;

[0058] Among them, P A Indicates the heating power of zone A, Q A represents the heating amount of zone A, Q A =C·M·(T 1 -T 2 ), where C is the specific heat capacity of partition A, which can refer to the average specific heat capacity, M represents the volume of partition A, T 1 is the set temperature, T 2 is the current temperature, and t is the heating time, for example 1 minute.

[0059] Preferably, in order to further improve the synchronization of temperature control, after heating each corresponding partition of the reaction plate for a preset time according to the calculated heating correction power, it also includes: obtaining the post-heating temperature of each partition of the reaction plate, wherein the preset time is less than the heating time; obtaining the second heating power of each partition of the well plate according to the set temperature and the post-heating temperature; and continuing to control the heating of each partition of the reaction plate according to the second heating power.

[0060] The determination process of the second heating power is similar to the determination process of the heating power of the A partition, except that the heating time is different. The heating time in the heating power of the A partition is the heating time, and the heating time of the second heating power is the remaining time obtained by subtracting the preset time from the heating time. The specific calculation process will not be repeated here.

[0061] Preferably, the second heating power of each partition of the reaction plate is corrected according to the power correction function to obtain the second heating correction power; and each corresponding partition of the reaction plate is heated according to the calculated second heating correction power, thereby improving the synchronization of temperature control.

[0062] Preferably, the preset duration can be half of the heating duration, or 2 / 3 thereof. Of course, after heating for a period of time according to the second heating correction power and before each partition reaches the set temperature, the heating power can be determined again for heating control, and this can be repeated until the heated temperature approaches the set temperature (when the difference is the preset temperature difference), and then the heating correction power determined last time is directly used to heat to the set temperature. In this way, the synchronization of the temperature control of each partition is higher.

[0063] The above describes the preferred embodiments of the present invention, but it is not intended to limit the present invention. Those skilled in the art may make improvements and changes to the embodiments disclosed herein without departing from the scope and spirit of the present invention.

Claims

1. A temperature control device for PCR instrument well plate partitions, characterized in that: The temperature control device for the partition of the PCR instrument plate comprises a temperature averaging plate, a semiconductor thermoelectric module, a temperature control module and a detection module. The temperature averaging plate is laid directly below the reaction plate and is in close contact with the bottom of each partition of the reaction plate respectively; the semiconductor thermoelectric module is used to directly heat the temperature averaging plate under the control of the temperature control module; the temperature averaging plate comprises a plurality of temperature averaging sub-plates, the number of which is the same as the number of partitions of the reaction plate, so that a temperature averaging sub-plate is arranged under each partition of the reaction plate; the detection module is used to detect the current temperature of each partition of the reaction plate; the temperature control module comprises a correction unit, which is used to calculate the heating power of each partition of the reaction plate according to the current temperature and set temperature of each partition of the reaction plate; and the heating power of each partition of the reaction plate is corrected according to the power correction function to obtain the heating correction power; the power correction function is: ； Wherein, P(x,y) is the heating power of the partition with coordinates (x,y) in each partition of the reaction plate; P(x+1,y) is the heating power of the partition with coordinates (x+1,y) in each partition of the reaction plate; P(x-1,y) is the heating power of the partition with coordinates (x-1,y) in each partition of the reaction plate; P(x,y+1) is the heating power of the partition with coordinates (x,y+1) in each partition of the reaction plate; P(x,y-1) is the heating power of the partition with coordinates (x,y-1) in each partition of the reaction plate; δ is the empirical correction coefficient; The temperature control module controls the semiconductor thermoelectric module to heat each corresponding partition of the reaction plate according to the calculated heating correction power; The value of the empirical correction coefficient δ is between 1.025 and 1.

038.

2. The temperature control device for PCR instrument well plate partition according to claim 1, characterized in that: Thermally conductive silicone grease is provided between the upper surface of the temperature averaging plate and the bottom of each partition of the reaction plate to ensure that there is no air thermal resistance between the temperature averaging plate and the reaction plate.

3. The temperature control device for PCR instrument well plate partition according to claim 1, characterized in that: The material of the temperature homogenizing plate is copper with a thermal conductivity greater than or equal to 386.4 W / (m·K) at room temperature.

4. The temperature control device for zoning well plates of a PCR instrument according to claim 1, characterized in that: The material of the temperature homogenizing plate is silver with a thermal conductivity greater than or equal to 426.2 W / (m·K) at room temperature.

5. The temperature control device for zoning well plates of a PCR instrument according to claim 1, characterized in that: An insulating isolation plate is arranged between adjacent temperature-averaging sub-plates; the insulating isolation plate is used to prevent temperature transfer between adjacent temperature-averaging sub-plates.

6. The temperature control device for zoning well plates of a PCR instrument according to claim 5, characterized in that: After the heat insulating isolation plates are arranged between the adjacent temperature-averaging sub-plates, the value of the empirical correction coefficient δ is fixed between 1.002 and 1.

005.

7. The temperature control device for zoning well plates of a PCR instrument according to claim 1, characterized in that: A heat-insulating isolation pad is arranged between adjacent different partitions of the reaction plate; the heat-insulating isolation pad is used to prevent temperature transfer between adjacent different partitions of the reaction plate.

8. The temperature control device for zoning well plates of a PCR instrument according to claim 7, characterized in that: After the heat insulating spacers are arranged between the adjacent different partitions of the reaction plate, the value of the empirical correction coefficient δ is fixed between 0.993 and 0.997.

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