Variable-temperature PCR temperature adjusting device based on MEMS thermopile technology

Abstract

The invention discloses a variable-temperature PCR temperature adjusting device based on an MEMS thermopile technology, and relates to the technical field of microelectronics, the variable-temperature PCR temperature adjusting device comprises a bottom lining base and a PCR reaction plate, the upper end face of the bottom lining base is provided with a thermopile assembly used for temperature detection and control, the thermopile assembly comprises a heat conduction layer deposited on the upper end face of the bottom lining base, and the heat conduction layer is arranged on the upper end face of the bottom lining base. According to the invention, the temperature of the PCR reaction is adjusted through the thermopile, and the switching from heating to cooling can be realized by changing the direction of the current, so that the temperature change requirement can be quickly responded, and the operation is simple and convenient. According to the temperature-variable PCR reaction device, the requirements of different steps in the temperature-variable PCR process are met, meanwhile, the temperature can be monitored, a sensor does not need to be additionally arranged, the temperature rising speed is higher than that of a traditional device, accurate temperature circulation can be achieved in a short time, and the PCR reaction efficiency and accuracy are improved.

Description

Technical Field

[0001] The invention relates to the field of microelectronic technology, in particular to a variable temperature PCR temperature regulating device based on MEMS thermopile technology. Background Art

[0002] PCR stands for polymerase chain reaction, and its principle is the in vitro amplification of DNA. The standard PCR process is divided into three steps: DNA denaturation: under the action of heat, the hydrogen bonds of the double-stranded DNA template are broken to form single-stranded DNA; annealing: the system temperature is lowered, the primers bind to the DNA template, and a local double strand is formed; extension: under the action of Taq enzyme (the activity is best at around 72°C), dNTP is used as the raw material, starting from the 3′ end of the primer and extending in the direction from 5′ to 3′ to synthesize a DNA chain complementary to the template. After each cycle of denaturation, annealing and extension, the DNA content doubles;

[0003] Now, some PCR can be changed to a two-step method because the amplification region is very short, so even if the Taq enzyme activity is not optimal, the replication can be completed in a very short time. That is, annealing and extension are carried out simultaneously at 60℃-65℃ to reduce the temperature rise and fall process once and increase the reaction speed.

[0004] At present, variable temperature PCR generally adopts the mode of TEC cooling sheet. TEC cooling sheet, also known as thermoelectric cooler, works based on the Peltier effect, which is a kind of heat transfer caused by one conductor absorbing heat and the other conductor releasing heat when current passes through a circuit composed of two different conductors. In TEC cooling sheet, current is introduced into semiconductor materials, so that one material absorbs heat and the other releases heat, thereby achieving cooling effect.

[0005] The TEC cooling sheet is composed of a pair of units consisting of a P-type semiconductor and an N-type semiconductor. When powered on, these units will generate electron-hole pairs at one end, resulting in a decrease in internal energy, a decrease in temperature, and a cold end; at the other end, the recombination of the electron-hole pairs leads to an increase in internal energy and an increase in temperature, forming a hot end. Through the series and parallel connection of multiple pairs of PN units, and the use of ceramic plates for upper and lower packaging, the TEC cooling sheet forms a structure with one end as the cold side and the other end as the hot side. This structure enables the TEC to transfer heat from the cold end to the hot end. During the heat transfer process, the TEC itself needs to pass current and voltage, which will also generate heat. The temperature difference between the cold and hot ends of a single TEC can reach 60 to 70°C, and the cold end temperature can reach -20 to -10°C. If a larger temperature difference or a lower cold end temperature is required, it can be achieved by stacking multiple TECs. Depending on the usage scenario and method, TECs of various shapes can be purchased on the market to meet specific cooling needs;

[0006] The main disadvantages of TEC cooling chips include high price, professional maintenance, low efficiency and high energy consumption;

[0007] Higher price: The price of TEC cooling chips is higher than that of traditional refrigeration equipment, which requires users to invest more money;

[0008] Requires professional maintenance: TEC cooling chips require professional maintenance, and it is difficult for ordinary users to maintain them by themselves;

[0009] Low efficiency: Compared with vapor compression refrigeration, TEC refrigeration efficiency is low at large capacities;

[0010] High energy consumption: Due to the lack of better semiconductor materials, TEC cooling consumes relatively high energy;

[0011] In addition, TEC cooling chips may also experience failure mechanisms such as overload, electrochemical corrosion, material migration (diffusion), semiconductor crystal damage and thermal stress during use. These problems may cause the cooling chip to fail to work properly.

[0012] At present, most of the TECs on the market, whether from ABI or well-known domestic brands, can only achieve an average heating and cooling speed of 4 to 5 degrees per second. Due to the unevenness of TEC power, in order to meet the demand for PCR temperature uniformity, the traditional PCR using TEC refrigeration plates needs to add multiple temperature sensors on the heating plate near the TEC refrigeration plates, which requires a high response speed of the temperature sensors. Summary of the invention

[0013] The purpose of the present invention is to provide a variable temperature PCR temperature control device based on MEMS thermopile technology, which has the advantages of low heating power, fast response time, small heat loss, compatibility with CMOS process, and easy integration with other microelectronic devices, so as to solve the problems raised in the above background technology.

[0014] To achieve the above-mentioned purpose, the present invention provides the following technical solutions: a variable temperature PCR temperature control device based on MEMS thermopile technology, comprising a substrate base and a PCR reaction plate, wherein the upper end surface of the substrate base is provided with a thermopile assembly for temperature detection and control, wherein the thermopile assembly comprises a heat conductive layer deposited on the upper end surface of the substrate base, wherein the upper end surface of the heat conductive layer is provided with a thermocouple layer, wherein the thermocouple layer is formed by a plurality of thermocouples connected in series, wherein the thermocouple comprises a first thermocouple material on the bottom side and a second thermocouple material on the upper side, wherein the connection between the first thermocouple material and the second thermocouple material forms a cold end and a hot end, wherein the connection at the bottom is the cold end, and the connection at the top is the hot end, wherein the upper end of the thermocouple layer is connected with a heat absorbing layer, wherein the bottom of the heat absorbing layer abuts against the hot end, and the PCR reaction plate is mounted on the upper end surface of the heat absorbing layer.

[0015] Preferably, the heat conductive layer is provided with sinking areas at the left and right edges respectively, a thermal isolation layer is embedded in the sinking areas, and the cold end connection between the first thermocouple material and the second thermocouple material is located above the thermal isolation layer.

[0016] Preferably, both ends of the second thermocouple material are respectively provided with thickened portions at the connection positions with the first thermocouple material, and the thickened portions abut against the bottom of the heat absorption layer.

[0017] Preferably, the first thermocouple material and the second thermocouple material are connected via a vertical portion, and the second thermocouple material is located at the top and is inclined.

[0018] Preferably, the thermal isolation layer is a silicon nitride film with a thickness controlled at 5-15 microns.

[0019] Preferably, a heat sink is fixedly mounted on the bottom of the substrate base, and heat sinks are distributed on the bottom of the heat sink.

[0020] Preferably, a stable connection layer is provided between the bottom of the substrate base and the heat sink.

[0021] Preferably, a cavity is provided at the bottom of the substrate base, and a film layer is formed on the top of the cavity.

[0022] Preferably, the stable connection layer is a graphene layer.

[0023] In summary, the beneficial effects of the present invention are:

[0024] 1. The present invention uses a thermopile to adjust the temperature of the PCR reaction. By changing the direction of the current, it can achieve switching from heating to cooling, so that it can quickly respond to temperature changes and meet the requirements of different steps in the variable temperature PCR process. At the same time, it can also monitor the temperature. It has both temperature conduction and temperature rise and fall functions and can be used as a temperature sensor. There is no need to add additional sensors, and the heating speed is faster than the traditional one. It can achieve accurate temperature cycles in a short time, thereby improving the efficiency and accuracy of the PCR reaction.

[0025] 2. Compared with the traditional thermocouple arrangement, the thermocouple arrangement can effectively improve the heat conduction efficiency, making the temperature response in the heat absorption layer faster and the data of the entire temperature change more accurate. At the same time, it can enhance the mechanical connection between the hot end of the thermocouple and the heat absorption layer, and improve the stability of the overall structure. This is especially important in high temperature or vibration environments. It can prevent the hot end from breaking or shifting due to mechanical stress and provide better thermal stability. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] In order to more clearly illustrate the embodiments of the invention or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without paying any creative work.

[0027] Figure 1 This is a schematic diagram of the overall structure of a variable temperature PCR temperature control device based on MEMS thermopile technology of the present invention;

[0028] Figure 2 This is a schematic diagram of the main structure of a variable temperature PCR temperature control device based on MEMS thermopile technology of the present invention;

[0029] Figure 3 It is a schematic diagram of the thermocouple structure of the thermocouple layer 21 in a variable temperature PCR temperature regulating device based on MEMS thermopile technology of the present invention;

[0030] Figure 4 The figure is a schematic diagram of the top view of the thermocouple structure of the thermocouple layer 21 in the variable temperature PCR temperature control device based on the MEMS thermopile technology of the present invention.

[0031] The symbols in the accompanying drawings are described as follows: substrate base 10; heat sink 11; heat sink 12; stable connection layer 13; cavity 14; film layer 15; PCR reaction plate 16; thermopile assembly 20; thermocouple layer 21; heat conductive layer 22; heat absorbing layer 23; thermal isolation layer 24; first thermocouple material 25; second thermocouple material 26; thickened portion 27; power terminal one 28; power terminal two 29. DETAILED DESCRIPTION

[0032] The present invention will now be further described in detail in conjunction with the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, rather than all of the embodiments. Based on the embodiments in the present invention, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention. These drawings are all simplified schematic diagrams, which only illustrate the basic structure of the present invention in a schematic way, and therefore they only show the structures related to the present invention.

[0033] To facilitate understanding of the present invention, the present invention will be described more comprehensively below with reference to the relevant drawings. Several embodiments of the present invention are shown in the drawings. However, the present invention can be implemented in many different forms and is not limited to the embodiments described herein. On the contrary, the purpose of providing these embodiments is to make the disclosure of the present invention more thorough and comprehensive.

[0034] All features disclosed in this specification, or steps in all methods or processes disclosed, except mutually exclusive features and / or steps, can be combined in any manner.

[0035] Any feature disclosed in this specification (including any additional claims, abstract and drawings), unless otherwise stated, may be replaced by other equivalent or alternative features with similar purposes. That is, unless otherwise stated, each feature is only an example of a series of equivalent or similar features.

[0036] In the present invention, unless otherwise clearly defined and specified, the terms "installed", "connected", "connected", "fixed" and the like should be understood in a broad sense, for example, it can be a fixed connection, a detachable connection, or an integral connection; it can be a mechanical connection, a direct connection, or an indirect connection through an intermediate medium, it can be the internal connection of at least two elements or the interaction relationship between at least two elements, unless otherwise clearly defined. For those of ordinary skill in the art, the specific meanings of the above terms in the present invention can be understood according to specific circumstances.

[0037] Combine the following Figure 1-4 The present invention is described in detail. For the convenience of description, the directions mentioned below are defined as follows: the up, down, left, right, front, and back directions mentioned below are the same as Figure 1 The view direction is consistent in front, back, left, right, up and down. Figure 1 The directions shown are consistent with the front, back, left, right, up and down directions of the device of the present invention when viewed from the front.

[0038] See also Figure 1-4An embodiment of the present invention is a variable temperature PCR temperature control device based on MEMS thermopile technology, comprising a substrate base 10, a PCR reaction plate 16 and a thermopile assembly 20, wherein the substrate base 10 can be silicon, germanium or germanium silicon, and the best is silicon. Then, etching is performed in the substrate base 10 to form a cavity 14, and a thin film layer 15 is formed on the top of the cavity 14. The thermopile assembly 20 includes a thermocouple layer 21, a heat conductive layer 22 and a heat absorbing layer 23. The heat conductive layer 22 is deposited on the upper end surface of the substrate base 10, wherein the heat conductive layer 22 can be boron nitride, and the thermocouple layer 21 is formed by connecting a plurality of thermocouples in series, wherein the thermocouple includes a first thermocouple material 25 on the bottom side and a second thermocouple material 26 on the top side, and the first The raw material of the thermocouple material 25 is deposited on the thermal conductive layer 22 and patterned into a horizontal row distribution arrangement to form the first thermocouple material 25. The thermocouple vertical parts are respectively made at both ends of the first thermocouple material 25 through a single damascene process. Similarly, the second thermocouple material 26 is patterned, and the two ends of the second thermocouple material 26 are respectively connected to the thermocouple vertical parts, so that the connection between the first thermocouple material 25 and the second thermocouple material 26 forms a cold end and a hot end, wherein the connection at the bottom is the cold end A, and the connection at the top is the hot end B. Finally, a heat absorption layer 23 is set on the upper end surface of the second thermocouple material 26, and the bottom of the heat absorption layer 23 is in contact with the hot end, and the PCR reaction plate 16 is installed on the upper end surface of the heat absorption layer 23;

[0039] It is worth mentioning that in order to make the thermocouple structure more stable, in this embodiment, the heat conductive layer 22 is provided with sinking areas at the left and right edges, and a thermal isolation layer 24 is embedded in the sinking area. The cold end connection between the first thermocouple material 25 and the second thermocouple material 26 is located above the thermal isolation layer 24. The thermal isolation layer 24 is a silicon nitride film with a thickness controlled at 5-15 microns, preferably 10 microns.

[0040] Silicon nitride has high hardness and wear resistance, and can provide good mechanical support for the cold end A of the thermocouple, especially in applications that need to withstand high mechanical stress. It can reduce the performance degradation of the cold end due to wear during use. At the same time, it serves as an insulating layer between the cold end A and other electronic components to prevent short circuits and other problems. At the same time, it maximizes the temperature stability of the cold end and extends the service life of the thermocouple.

[0041] It should be noted that, in order to further improve the heat conduction performance of the hot end, in this embodiment, thickened portions 27 are respectively provided at both ends of the second thermocouple material 26 at the connection positions with the first thermocouple material 25, and the thickened portions 27 abut against the bottom of the heat absorption layer 23, and the second thermocouple material 26 is arranged in an inclined distribution above the second thermocouple material 26;

[0042] By utilizing the thickened portion 27 and the obliquely distributed second thermocouple material 26, the contact area between the hot end of the thermocouple and the heat absorption layer 23 can be effectively increased, thereby improving the heat conduction efficiency, making the temperature response in the heat absorption layer 23 more rapid, and the data of the entire temperature change more accurate. At the same time, the mechanical connection between the hot end of the thermocouple and the heat absorption layer 23 can be enhanced, and the stability of the overall structure can be improved, which is particularly important in high temperature or vibration environments, and can prevent the hot end from breaking or shifting due to mechanical stress, thereby providing better thermal stability.

[0043] In addition, in one embodiment, a heat sink 11 is fixedly installed at the bottom of the substrate base 10, and a stable connection layer 13 is arranged between the substrate base 10 and the heat sink 11. The stable connection layer 13 can be a graphene layer to improve the strength of the connection. Heat sinks 12 are distributed at the bottom of the heat sink 11. The heat sink 11 is installed at the bottom to improve the overall heat dissipation, and the stable connection layer 13 can further improve the conduction of heat and accelerate the heat dissipation of the device.

[0044] When variable temperature PCR is required, the required reaction mixture is first added to the PCR reaction plate 16, wherein the reaction mixture includes: target DNA sample to be amplified, primers, dNTPs (deoxyribonucleotide triphosphates), buffer, DNA polymerase and water. During the initial denaturation, the reaction mixture is heated to 94-98° C. for about 2-5 minutes. This high temperature will cause the DNA double strand to unwind into a single strand because the high temperature destroys the hydrogen bonds that maintain the double-stranded structure. Then a cyclic reaction is performed. Each cycle generally includes the following three steps. The whole process may be repeated 25 to 35 times:

[0045] (a) Denaturation

[0046] At this stage, the reaction mixture is heated to approximately 94-98°C for 20-30 seconds. This causes any re-formed double-stranded DNA to unwind back into a single-stranded state.

[0047] (b) Annealing

[0048] The temperature is rapidly dropped to between 50-65°C for 20-60 seconds. This temperature range allows the primers to bind to the complementary sequences on the single strand of the template DNA. The design of the primers determines their optimal annealing temperature.

[0049] (c) Extension

[0050] The temperature rises to about 72°C, and the duration depends on the length of the target fragment to be amplified, which takes about 1 minute per 1kb (kilobase). At this temperature, DNA polymerase begins to synthesize a new DNA chain, extending from the primer to the other end of the template DNA.

[0051] After all cycles are completed, there is usually a final extension step where the temperature is maintained at 72°C for 5-10 minutes to ensure that all DNA fragments are fully extended. After the reaction is complete, the samples are usually cooled to 4°C to prevent non-specific reactions and can be stored for a period of time until further processing or analysis.

[0052] The entire reaction above requires temperature control and regulation, and the PCR reaction plate 16 is located at the upper end of the heat absorption layer 23, and the positive and negative electrodes of the external power supply are respectively connected to the power connection terminal 1 28 and the power connection terminal 2 29. The current passes through the thermocouple, so that the top is heated and heated, and the heat absorption layer 23 is used to conduct the heat to the PCR reaction plate 16 for heating the PCR reaction plate 16. By changing the direction of the current, the switching from heating to cooling can be achieved, so that the temperature change requirements can be quickly responded to, and the temperature can be quickly increased or decreased by adjusting the current to meet the requirements of different steps in the variable temperature PCR process. At the same time, the temperature can be monitored and the signal can be fed back to the control system. The control system adjusts the current size or direction according to the feedback temperature signal to achieve the required temperature. It has both the temperature conduction and lifting effect and can be used as a temperature sensor. No additional sensor is required, and the heating speed is also faster than the traditional one. It can achieve accurate temperature cycle in a short time, improving the efficiency and accuracy of the PCR reaction.

[0053] The above is only a specific implementation of the invention, but the protection scope of the invention is not limited to it. Any changes or substitutions that are not conceived through creative work should be included in the protection scope of the invention. Therefore, the protection scope of the invention should be based on the protection scope defined in the claims.

Claims

1. A variable temperature PCR temperature control device based on MEMS thermopile technology, comprising a substrate base (10) and a PCR reaction plate (16), characterized in that: The upper end surface of the substrate base (10) is provided with a thermopile assembly (20) for temperature detection and control. The thermopile assembly (20) comprises a heat-conducting layer (22) deposited on the upper end surface of the substrate base (10). The upper end surface of the heat-conducting layer (22) is provided with a thermocouple layer (21). The thermocouple layer (21) is formed by a plurality of thermocouples connected in series. The thermocouple comprises a first thermocouple material (25) on the bottom side and a second thermocouple material (26) on the top side. The connection between the first thermocouple material (25) and the second thermocouple material (26) forms a cold end and a hot end, wherein the connection at the bottom is the cold end and the connection at the top is the hot end. The upper end of the thermocouple layer (21) is connected with a heat-absorbing layer (23). The bottom of the heat-absorbing layer (23) is in contact with the hot end. The PCR reaction plate (16) is mounted on the upper end surface of the heat-absorbing layer (23).

2. The variable temperature PCR temperature control device based on MEMS thermopile technology according to claim 1, characterized in that: The heat-conducting layer (22) is provided with sinking areas at the left and right edges respectively, a thermal isolation layer (24) is embedded in the sinking areas, and the cold end connection point of the first thermocouple material (25) and the second thermocouple material (26) is located above the thermal isolation layer (24).

3. The variable temperature PCR temperature control device based on MEMS thermopile technology according to claim 2, characterized in that: Both ends of the second thermocouple material (26) are respectively provided with thickened portions (27) at the connection positions with the first thermocouple material (25), and the thickened portions (27) are in contact with the bottom of the heat absorption layer (23).

4. The variable temperature PCR temperature control device based on MEMS thermopile technology according to claim 3, characterized in that: The first thermocouple material (25) and the second thermocouple material (26) are connected via a vertical portion, and the second thermocouple material (26) is located at the top and is inclined.

5. The variable temperature PCR temperature control device based on MEMS thermopile technology according to claim 4, characterized in that: The thermal isolation layer (24) is a silicon nitride film with a thickness controlled at 5-15 microns.

6. The variable temperature PCR temperature control device based on MEMS thermopile technology according to claim 5, characterized in that: A heat sink (11) is fixedly mounted on the bottom of the substrate base (10), and heat sink grooves (12) are distributed on the bottom of the heat sink (11).

7. The variable temperature PCR temperature control device based on MEMS thermopile technology according to claim 6, characterized in that: A stable connection layer (13) is provided between the bottom of the substrate base (10) and the heat dissipation plate (11).

8. The variable temperature PCR temperature control device based on MEMS thermopile technology according to claim 7, characterized in that: A cavity (14) is provided at the bottom of the substrate base (10), and a thin film layer (15) is formed on the top of the cavity (14).

9. The variable temperature PCR temperature control device based on MEMS thermopile technology according to claim 8, characterized in that: The stable connection layer (13) is a graphene layer.

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