PCR thermal cycler and control method

Abstract

The application relates to the technical field of rapid molecular diagnosis, in particular to a PCR (Polymerase Chain Reaction) thermal cycle device and a control method. The PCR thermal cycle device comprises a first constant-temperature tank with a constant first temperature, a moving mechanism and a control module. The moving mechanism is used for synchronously moving or taking out a capillary tube and a temperature measurement module from or in the first constant-temperature tank, so that the temperature of a sample in the capillary tube is controlled by the first constant-temperature tank. The temperature measurement module is used for detecting the temperature. The control module is configured as follows: the control module is provided with a first set temperature. When the temperature detected by the temperature measurement module in the first constant-temperature tank reaches the first set temperature, the control module immediately controls the moving mechanism to take out the capillary tube from the first constant-temperature tank, so that the temperature of the sample is indirectly monitored by the temperature measurement module, and the sample in the capillary tube can be taken out from the first constant-temperature tank in time when the temperature of the sample reaches a first temperature threshold.

Description

[0001] This application is a divisional application of the invention patent filed on December 6, 2021, with application number 2021114807285, entitled "PCR Thermal Cycling Device and Control Method". Technical Field

[0002] This application relates to the field of rapid molecular diagnostic technology, and in particular to a PCR thermal cycling device and control method. Background Technology

[0003] In molecular diagnostics, the amplification process of samples is usually completed in a PCR thermal cycling device. Current PCR thermal cycling devices are generally equipped with multiple thermostatic baths, such as high temperature baths, low temperature baths and optical baths, so that the capillary tubes are circulated multiple times in multiple thermostatic baths according to a predetermined cycle to complete the amplification process.

[0004] During high-speed capillary transfer, sample temperature needs to be monitored to determine the optimal time to remove the sample from the thermostat to complete the denaturation, annealing, and extension thermal cycles. However, due to the extremely small size of the capillary, the sample experiences very rapid temperature changes in the high-temperature and low-temperature baths, making it impossible to accurately monitor the real-time sample temperature and thus difficult to determine the optimal time to remove the capillary from the thermostat. Related techniques employ fixed-time control of capillary transfer or the use of rapid-response temperature detection devices to ensure the sample in the capillary meets the temperature requirements of each cycle. However, because the sample experiences very rapid temperature changes in the thermostat, the sensitivity and material requirements of the rapid-response temperature sensor and control device are extremely high. While these methods address the sample temperature control issue to some extent, they still result in low reliability and high cost in PCR thermal cycling. Summary of the Invention

[0005] The purpose of this invention is to provide a PCR thermal cycling device and control method to improve the detection efficiency and reliability of PCR thermal cycling.

[0006] This invention provides a PCR thermal cycling device, including a moving mechanism, a first thermostat, and a control module; the first thermostat can be configured to have a first controlled temperature; a capillary tube containing a sample and a temperature measuring module are placed on the moving mechanism, and the moving mechanism can drive the capillary tube and the temperature measuring module to move synchronously; the control module is configured to:

[0007] The moving mechanism is controlled to transfer the capillary tube and the temperature measuring module into the first constant temperature bath; when the temperature measured by the temperature measuring module reaches the first set temperature, the moving mechanism is controlled to move the capillary tube out of the first constant temperature bath, so that the temperature of the sample is within the first temperature threshold range after the capillary tube is moved out of the first constant temperature bath; the first control temperature is greater than the first set temperature.

[0008] Furthermore, the first set temperature is greater than the first temperature threshold.

[0009] Furthermore, the first temperature threshold is (94±2)℃;

[0010] The first set temperature ranges from 90℃ to 105℃.

[0011] Furthermore, the PCR thermal cycling device also includes a second thermostat, which can be configured to have a second controlled temperature.

[0012] The control module is also configured to:

[0013] The moving mechanism is controlled to transfer the capillary tube from the first constant temperature bath to the second constant temperature bath;

[0014] When the temperature measured by the temperature measuring module reaches the second set temperature, the moving mechanism is controlled to move the capillary tube out of the second constant temperature bath, so that the temperature of the sample is within the second temperature threshold range after the capillary tube is moved out of the second constant temperature bath.

[0015] The second control temperature is lower than the second set temperature.

[0016] Furthermore, the second set temperature is greater than the second temperature threshold.

[0017] Furthermore, the second temperature threshold is (57.5±2)℃;

[0018] The second set temperature range is 50℃-70℃.

[0019] Furthermore, the PCR thermal cycling device also includes a third thermostat, which can be configured to have a third controlled temperature.

[0020] The control module is also configured to:

[0021] The moving mechanism is controlled to transfer the capillary tube from the second constant temperature bath to the third constant temperature bath;

[0022] After the sample in the capillary completes fluorescence signal acquisition in the third constant temperature bath, the moving mechanism is controlled to remove the capillary from the third constant temperature bath.

[0023] Furthermore, the PCR thermal cycling device also includes a fourth thermostat, which can be configured to have a fourth controlled temperature.

[0024] The control module is also configured to:

[0025] The moving mechanism is controlled to sequentially transfer the capillary tube to the first constant temperature bath, the fourth constant temperature bath, the second constant temperature bath, and the third constant temperature bath, and the capillary tube stays in the fourth constant temperature bath with the fourth controlled temperature for a first predetermined time.

[0026] Furthermore, the fourth control temperature is the target temperature for sample denaturation.

[0027] Furthermore, the fourth constant temperature bath can also be configured to control the fifth temperature;

[0028] The control module can also be configured to:

[0029] The moving mechanism is controlled to sequentially transfer the capillary to a fourth thermostat bath with a fifth controlled temperature, a first thermostat bath, a fourth thermostat bath with a fourth controlled temperature, a second thermostat bath, and a third thermostat bath, and the capillary is kept in the fourth thermostat bath with a fifth controlled temperature for a second predetermined time to complete the reverse transcription of the sample.

[0030] Furthermore, the fifth control temperature is the target temperature for reverse transcription of the sample.

[0031] Furthermore, the first constant temperature bath, the fourth constant temperature bath, the second constant temperature bath, and the third constant temperature bath are arranged side by side with intervals.

[0032] Furthermore, the control module is also configured to:

[0033] When the capillary tube remains in the first constant temperature bath for a first set time, and the temperature measured by the temperature measuring module does not reach the first set temperature, the moving mechanism is controlled to remove the capillary tube from the first constant temperature bath.

[0034] Furthermore, the control module is also configured to:

[0035] When the capillary tube remains in the second constant temperature bath for a second set time, and the temperature measured by the temperature measuring module does not reach the second set temperature, the moving mechanism is controlled to remove the capillary tube from the second constant temperature bath.

[0036] Furthermore, the temperature measurement module is a temperature sensor with a metal casing or a ceramic casing.

[0037] Furthermore, the control module is also configured to: in the first cycle, when the temperature measured by the temperature measuring module in the first constant temperature bath reaches the third set temperature, control the moving mechanism to remove the capillary tube from the first constant temperature bath; the third set temperature is higher than the first set temperature.

[0038] This application also provides a method for controlling a PCR thermal cycling device, including the following steps:

[0039] Step 100: Place the capillary tube into the first constant temperature bath, and remove it when the temperature measured by the temperature measuring module reaches the first set temperature, so that the sample temperature in the capillary tube is within the first temperature threshold range.

[0040] Step 200: Transfer the capillary tube to the second constant temperature bath. When the temperature measured by the temperature measuring module reaches the second set temperature, remove the capillary tube so that the sample temperature in the capillary tube is within the second temperature threshold range.

[0041] Step 300: Transfer the capillary tube to the third constant temperature bath, and remove it after completing the fluorescence signal acquisition of the sample;

[0042] Step 400: Repeat steps 100 to 300 a predetermined number of times.

[0043] Further, in step 100:

[0044] In the first cycle, when the temperature measured by the temperature measuring module in the first constant temperature bath reaches the third set temperature, the capillary is removed from the first constant temperature bath, and the third set temperature is greater than the first set temperature.

[0045] Furthermore, step 101 is included between step 100 and step 200:

[0046] The capillary tube is transferred to a fourth thermostatic bath with a fourth controlled temperature, and removed after a first predetermined time.

[0047] Furthermore, the fourth thermostat of the PCR thermal cycling device can be configured to control a sixth temperature.

[0048] Before step 100, step 100c is also included: the capillary tube is placed in a fourth constant temperature bath with a sixth controlled temperature for a third predetermined time and then removed.

[0049] Preferably, the sixth control temperature is the target temperature at the start of the cycle.

[0050] Furthermore, step 101 is included between step 100 and step 200:

[0051] The capillary tube is transferred to a fourth thermostatic bath with a fourth controlled temperature, and removed after a first predetermined time.

[0052] Furthermore, step 100a is included before step 100:

[0053] The capillary tube is placed in the fourth constant temperature bath with the fifth controlled temperature, and after staying for a second predetermined time, it is transferred to the first constant temperature bath.

[0054] Furthermore, step 100 also includes step 100b:

[0055] If the temperature measured by the temperature measuring module in the first constant temperature bath does not reach the first set temperature within the first set time, the control module controls the moving mechanism to remove the capillary tube from the first constant temperature bath.

[0056] And / or,

[0057] Step 200 further includes step 200a:

[0058] If the temperature measured by the temperature measuring module in the second constant temperature bath does not reach the second set temperature within the second set time, the control module controls the moving mechanism to remove the capillary tube from the second constant temperature bath.

[0059] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0060] The PCR thermal cycling device provided by this invention includes a first thermostatic bath, a fourth thermostatic bath, a moving mechanism, and a control module. The fourth thermostatic bath can be configured to have a sixth control temperature, and the first thermostatic bath has a constant first control temperature. The moving end of the moving mechanism is spaced apart with a capillary tube containing a sample and an independent temperature measuring module. The moving mechanism can drive the capillary tube and the temperature measuring module to move synchronously. The moving mechanism first drives the capillary tube and the temperature measuring module into the fourth thermostatic bath to reach the target temperature for the start of the cycle, avoiding the influence of ambient temperature causing the starting temperature to be too low, which would prevent the first temperature threshold requirement from being met quickly during the cycle, thus improving the detection efficiency and reliability of the PCR thermal cycling. Then, the moving mechanism puts the capillary tube and the temperature measuring module into the first thermostatic bath together or removes them together from the first thermostatic bath. The first thermostatic bath can control the temperature of the sample placed inside the capillary tube, such as by heating or cooling. The control module is configured such that: the control module has a first set temperature, and when the temperature measured by the temperature measuring module in the first thermostatic bath reaches the first set temperature, the control module immediately controls the moving mechanism to remove the capillary tube from the first thermostatic bath, thereby indirectly monitoring the temperature of the sample through the temperature measuring module, ensuring that the temperature of the sample in the capillary tube remains within the first temperature threshold after it is removed from the first thermostatic bath.

[0061] The present invention also provides a control method for a PCR thermal cycling device, comprising the following steps: First, placing a capillary tube into a fourth constant temperature bath, and removing it when the sample or temperature measuring module in the capillary tube reaches the target temperature for the start of the cycle; Next, placing the capillary tube into a first constant temperature bath, and removing it when the temperature measured by the temperature measuring module reaches a first set temperature; Next, transferring the capillary tube to a second constant temperature bath, and removing it when the temperature measured by the temperature measuring module reaches a second set temperature; Next, transferring the capillary tube to a third constant temperature bath, and removing it after completing the fluorescence signal acquisition of the sample; Next, repeating steps 100 to 300 a predetermined number of times; thereby completing the sample amplification process. Attached Figure Description

[0062] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0063] Figure 1 This is a schematic diagram of the PCR thermal cycling device provided in an embodiment of the present invention;

[0064] Figure 2 A schematic diagram of the capillary and temperature measuring module of the PCR thermal cycling device provided in an embodiment of the present invention;

[0065] Figure 3 A sample temperature curve in a capillary tube provided in an embodiment of the present invention;

[0066] Figure 4 The temperature curve is provided by the temperature measurement module in an embodiment of the present invention.

[0067] Figure label:

[0068] 1-First constant temperature bath, 2-Second constant temperature bath, 3-Third constant temperature bath, 4-Fourth constant temperature bath, 5-Moving mechanism, 6-Capillary tube, 7-Temperature measuring module, 8-Support. Detailed Implementation

[0069] The technical solution of the present invention will now be clearly and completely described with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments.

[0070] The components of the embodiments of the invention described and shown in the accompanying drawings can typically be arranged and designed in a variety of different configurations. Therefore, the following detailed description of the embodiments of the invention provided in the drawings is not intended to limit the scope of the claimed invention, but merely to illustrate selected embodiments of the invention.

[0071] Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0072] In the description of this invention, it should be noted that the terms "center," "upper," "lower," "left," "right," "vertical," "horizontal," "inner," and "outer," etc., indicate the orientation or positional relationship based on the orientation or positional relationship shown in the accompanying drawings. They are used only for the convenience of describing the invention and for simplifying the description, and do not indicate or imply that the device or element referred to must have a specific orientation, or be constructed and operated in a specific orientation. Therefore, they should not be construed as limitations on the invention. Furthermore, the terms "first," "second," and "third" are used for descriptive purposes only and should not be construed as indicating or implying relative importance.

[0073] In the description of this invention, it should be noted that, unless otherwise explicitly specified and limited, the terms "installation," "connection," and "linking" should be interpreted broadly. For example, they can refer to a fixed connection, a detachable connection, or an integral connection; they can refer to a mechanical connection or an electrical connection; they can refer to a direct connection or an indirect connection through an intermediate medium; and they can refer to the internal connection of two components. Those skilled in the art can understand the specific meaning of the above terms in this invention based on the specific circumstances.

[0074] The following reference Figures 1 to 4 This application describes a PCR thermal cycling apparatus and control method according to some embodiments.

[0075] Example 1

[0076] Example 1 provides a PCR thermal cycling device, such as Figure 1 and Figure 2 As shown, it includes a first constant temperature bath 1 and a moving mechanism 5. The first constant temperature bath 1 has a constant first control temperature. The moving end of the moving mechanism 5 is provided with a support 8. The capillary tube 6 containing the sample and the temperature measuring module 7 are installed on the support 8 at intervals. The moving mechanism 5 can drive the capillary tube 6 and the temperature measuring module 7 to move synchronously, so as to put the capillary tube 6 and the temperature measuring module 7 into the first constant temperature bath 1 together or take them out of the first constant temperature bath 1 together.

[0077] The first thermostat 1 can control the temperature of the sample placed inside the capillary tube 6, such as by heating or cooling. In this embodiment, the first thermostat 1 is used as a high-temperature bath of the PCR thermal cycling device to heat the sample so that the temperature of the sample is heated to a first temperature threshold range to complete the DNA denaturation reaction.

[0078] When the sample is heated in the first constant temperature bath 1, the temperature measuring module 7 can monitor the temperature of the sample to ensure that the temperature of the sample inside the capillary tube 6 is within the first temperature threshold range after the capillary tube 6 is removed from the first constant temperature bath 1. Because the capillary tube 6 is very small, the temperature measuring module 7 cannot be placed inside the capillary tube 6 to directly monitor the sample temperature, resulting in a temperature difference between the temperature measured by the temperature measuring module 7 and the temperature of the sample inside the capillary tube 6. To accurately determine the timing of removing the capillary tube 6 from the first constant temperature bath 1 based on the temperature measured by the temperature measuring module 7, the PCR thermal cycling device is also equipped with a control module. This control module is configured to have a first set temperature. When the temperature measured by the temperature measuring module 7 in the first constant temperature bath 1 reaches the first set temperature, the control module immediately controls the moving mechanism 5 to remove the capillary tube 6 from the first constant temperature bath 1. This allows the temperature measuring module 7 to indirectly monitor the sample temperature, ensuring that the temperature of the sample inside the capillary tube 6 is within the first temperature threshold after being removed from the first constant temperature bath 1, thus ensuring rapid and smooth DNA denaturation of the sample.

[0079] In this embodiment, preferably, there are two temperature measuring modules 7, which are arranged side by side on both sides of the capillary tube. When the temperature of any one of the temperature measuring modules 7 reaches the first set temperature, the control module immediately controls the moving mechanism 5 to remove the capillary tube 6 from the first constant temperature bath 1, so as to further reduce the risk caused by temperature uniformity.

[0080] In this embodiment, the temperature measuring module 7 can be a tubular structure, with the temperature sensor inserted into the water or oil inside the tube and sealed. Preferably, the temperature measuring module 7 is a temperature sensor with a metal or ceramic shell, which makes the temperature measuring module 7 less prone to breakage or damage during the movement of the capillary tube 6, thereby ensuring the smooth completion of the entire detection process.

[0081] However, because the temperature measuring module 7 is made of a different material than the capillary tube 6, their transient thermal response characteristics in the constant temperature bath differ. This leads to a difference between the temperature measured by the temperature measuring module 7 and the temperature of the sample in the capillary tube 6, even when both are under the same ambient temperature. Therefore, a stable temperature correspondence needs to be established between them. In this embodiment, the first temperature threshold is the target temperature for the sample to complete DNA denaturation, such as 95°C. Preferably, the first control temperature range of the first constant temperature bath 1 is 100°C-130°C. By setting the first control temperature much higher than the target temperature for DNA denaturation, the sample can be rapidly heated to the required temperature and denatured within the first constant temperature bath 1. Preferably, the first control temperature is 120°C.

[0082] In this embodiment, due to individual differences between temperature measuring modules 7 made of different materials or with different structures, they exhibit different transient thermal response deviations with the capillary tube 6. Therefore, the first set temperature range of the temperature measuring module 7 is 90℃-105℃ to meet the DNA denaturation temperature requirement of 95℃ for the sample. Simultaneously, to improve the reliability of system control and reduce control difficulty, the first temperature threshold is further set to (94±2)℃, so that temperature measuring modules 7 made of different materials or with different structures can establish a stable temperature correspondence with the capillary tube 6. Preferably, when the temperature measuring module 7 is an NTC steel tube, the first set temperature is 102℃; that is, when the temperature measured by the NTC steel tube reaches 102℃, the control module immediately controls the moving mechanism 5 to move the capillary tube 6 out of the first constant temperature bath 1. At this time, it can be ensured that the sample temperature is always at (94±2)℃ to meet the temperature requirements for completing DNA denaturation in multiple high-speed cycles.

[0083] In this embodiment, preferably, the PCR thermal cycling device further includes a second constant temperature bath 2 used as a low temperature bath and a third constant temperature bath 3 used as an optical bath, and the second constant temperature bath 2 is set to have a constant second control temperature, and the third constant temperature bath 3 is set to have a constant third control temperature; the control module can control the moving mechanism 5 to sequentially transfer the capillary tube 6 taken out from the first constant temperature bath 1 to the second constant temperature bath 2 and the third constant temperature bath 3, so that the sample can complete cooling and annealing in the second constant temperature bath 2 and complete fluorescence signal acquisition in the third constant temperature bath 3 (completed by an external fluorescence signal acquisition device).

[0084] Preferably, the sample in the capillary 6 needs to be cooled in the second constant temperature bath 2 so that the temperature of the sample is within the second temperature threshold before the capillary 6 reaches the third constant temperature bath 3, so as to meet the temperature requirements for completing DAN regeneration in multiple high-speed cycles. Preferably, the second temperature threshold is (57.5±2)℃.

[0085] Since the temperature of the sample in the capillary tube 6 will continue to decrease during the process of removing the capillary tube 6 from the second constant temperature bath 2 and transferring it to the third constant temperature bath 3, in order to ensure that the temperature of the sample can be rapidly reduced to the second temperature threshold range before the capillary tube 6 enters the third constant temperature bath 3, so that the temperature of the second constant temperature bath 2, i.e. the second control temperature, is much lower than the second temperature threshold; preferably, the second control temperature range of the second constant temperature bath 2 is 10℃-25℃; further, the second control temperature is 15℃.

[0086] Meanwhile, during the cooling process of capillary tube 6 in the second constant temperature bath 2, the temperature of the sample inside capillary tube 6 is still indirectly monitored by temperature measuring module 7 to determine the timing for removing capillary tube 6 from the second constant temperature bath 2 based on the temperature measured by temperature measuring module 7. Preferably, the control module is configured to set a second set temperature, preferably ranging from 50℃ to 70℃. When the temperature measured by temperature measuring module 7 in the second constant temperature bath 2 reaches the second set temperature, the control module immediately controls the moving mechanism 5 to remove capillary tube 6 from the second constant temperature bath 2, thereby ensuring that the temperature of the sample inside capillary tube 6 is within the second temperature threshold range before being transferred to the third constant temperature bath 3. Preferably, the second set temperature is 65℃.

[0087] It should be noted that the specific values ​​of the first set temperature and the second set temperature can be set according to the actual situation. For example, when the temperature of the first constant temperature bath 1 and the second constant temperature bath 2 changes, or the material and structure of the temperature measuring module changes, or the first temperature threshold and the second temperature threshold that the sample needs to reach change, the values ​​of the first set temperature and the second set temperature can also be adjusted accordingly.

[0088] Specifically, the specific values ​​of the first and second set temperatures are obtained through temperature calibration and compensation of the constant temperature bath and temperature curve calibration. For example, a temperature measuring module and a capillary tube containing a predetermined amount of water are placed at the moving end of the moving mechanism to simulate the state of a sample in the capillary tube. A thermocouple is inserted into the capillary tube to collect the sample temperature, and the thermocouple probe is connected to a temperature acquisition instrument. The moving mechanism drives the capillary tube and temperature measuring module to circulate between the first and second constant temperature baths. A first extraction temperature and a second extraction temperature are given; that is, when the temperature of the temperature measuring module reaches the first extraction temperature, the capillary tube is removed from the high-temperature bath, and when the temperature of the temperature measuring module reaches the second extraction temperature, the capillary tube is removed from the low-temperature bath. The temperature acquisition instrument will then provide the following data: Figure 3 The temperature curve of the sample inside the capillary tube measured by the thermocouple is shown below. Simultaneously, based on the temperature measured by the temperature sensing module, the following can be obtained: Figure 4 The temperature curves shown are both regular curves with peaks and troughs. When the first extraction temperature is the first set temperature of 102℃ mentioned above and the second extraction temperature is the second set temperature of 65℃ mentioned above, the peak of the temperature curve of the sample in the capillary can be stabilized at (94±2)℃, which is the first temperature threshold, and the trough can be stabilized at (57.5±2)℃, which is the second temperature threshold, thus determining the first set temperature and the second set temperature.

[0089] In this embodiment, preferentially, since a stable correspondence has not yet been established between the temperature measuring module 7 and the sample tube when the temperature measuring module 7 is first inserted into the high-temperature bath at the beginning of the cycle, the first extraction temperature of the temperature measuring module is set differently in multiple cycles. In the first cycle, the first extraction temperature of the temperature measuring module 7 is higher than the first extraction temperature in subsequent cycles. For example, in 40 cycles, the extraction temperature of the temperature measuring module in the high-temperature bath in the first cycle will be 1-4°C higher than the extraction temperature in the following 39 cycles, preferably 2°C. Figure 4 As shown, the first extraction temperature for the first cycle is 104°C, while the first extraction temperature for the second to 39th cycles is 102°C, in order to meet the first temperature threshold requirement.

[0090] In this embodiment, preferably, the third control temperature of the third constant temperature bath 3 is 75°C, so as to extend the temperature of the sample placed in the capillary tube 6 through the third constant temperature bath 3, ensuring that the sample can successfully complete the fluorescence signal acquisition. Preferably, the control module is communicatively connected to the fluorescence signal acquisition device, and the control module is also configured to: after the fluorescence signal acquisition device completes the fluorescence signal acquisition of the sample, the control module controls the moving mechanism 5 to move the capillary tube 6 out of the third constant temperature bath 3.

[0091] Preferably, the control module is further configured to control the moving mechanism 5 to drive the capillary tube 6 to move cyclically multiple times between the first constant temperature bath 1, the second constant temperature bath 2 and the third constant temperature bath 3, so that the sample completes the amplification process in the PCR thermal cycling device.

[0092] Example 2

[0093] This second embodiment is an improvement based on the above embodiments. The technical content disclosed in the above embodiments will not be described again, and the content disclosed in the above embodiments also belongs to the content disclosed in this embodiment.

[0094] In Example 2, preferably, the PCR thermal cycling device may also be equipped with a fourth thermostat 4, which can be used as a reverse transcription tank. When the fourth thermostat 4 is used as a reverse transcription tank, it is set to a constant fifth control temperature, which is the target temperature for sample reverse transcription. Preferably, the fifth control temperature is 55°C. When the fourth thermostat 4 is used as a reverse transcription tank, the four thermostats are arranged side by side in the order of the fourth thermostat 4, the first thermostat 1, the second thermostat 2, and the third thermostat 3.

[0095] The control module is configured to: control the moving mechanism 5 to sequentially transfer the capillary 6 to the fourth thermostat 4, the first thermostat 1, the second thermostat 2, and the third thermostat 3 and cycle them multiple times; and to keep the sample in the fourth thermostat 4 for a second predetermined time to complete reverse transcription. The timing of removing the capillary 6 from the first thermostat 1, the second thermostat 2, and the third thermostat 3 is as described in Example 1 above, so that the sample can complete DNA denaturation in the first thermostat 1, complete cooling and annealing in the second thermostat 2, and complete fluorescence signal acquisition in the third thermostat 3.

[0096] Example 3

[0097] This third embodiment is an improvement based on the above embodiments. The technical content disclosed in the above embodiments will not be described again, and the content disclosed in the above embodiments also belongs to the content disclosed in this embodiment.

[0098] In Embodiment 3, preferably, the fourth constant temperature bath 4 can be used as an auxiliary denaturation bath, and when the fourth constant temperature bath 4 is used as an auxiliary denaturation bath, the fourth constant temperature bath 4 is set to a constant fourth control temperature, which is the target temperature for sample denaturation; preferably, the fourth control temperature is 95°C; the four constant temperature baths are arranged side by side in the order of the first constant temperature bath 1, the fourth constant temperature bath 4, the second constant temperature bath 2 and the third constant temperature bath 3.

[0099] At this time, the control module is configured to: control the moving mechanism 5 to sequentially transfer the capillary tube 6 to the first constant temperature bath 1, the fourth constant temperature bath 4, the second constant temperature bath 2 and the third constant temperature bath 3 and cycle them multiple times, and make the sample stay in the fourth constant temperature bath 4 for a first predetermined time so that the sample continues to denature in the fourth constant temperature bath 4 for a period of time. The timing of removing the capillary tube 6 from the first constant temperature bath 1, the second constant temperature bath 2 and the third constant temperature bath 3 is the same as in the above embodiment 1.

[0100] In Example 3, preferably, when the fourth isothermal bath 4 is used as an auxiliary denaturation bath, the fourth isothermal bath 4 can also be set to a constant sixth control temperature, which is the target temperature for the start of the cycle. Before starting the first cycle, the moving mechanism 5 inserts the capillary tube 6 and the temperature measuring module 7 into the fourth isothermal bath 4 with the sixth control temperature, so that the sample in the capillary tube 6 or the temperature measuring module 7 reaches the target temperature for the start of the cycle, to avoid the sample starting at too low a temperature due to the influence of the ambient temperature, which would prevent the first temperature threshold requirement from being met quickly during the cycle. The value range of the sixth control temperature is 50℃-57℃, preferably 55℃. Further, the moving mechanism 5 inserts the capillary tube 6 and the temperature measuring module 7 into the fourth isothermal bath 4 with the sixth control temperature for a predetermined time, preferably 1 minute, to ensure that the sample in the capillary tube 6 can reach the target temperature for the start of the cycle, thereby improving the detection efficiency and reliability of the PCR thermal cycling.

[0101] Example 4

[0102] This fourth embodiment is an improvement based on the above embodiments. The technical content disclosed in the above embodiments will not be described again, and the content disclosed in the above embodiments also belongs to the content disclosed in this embodiment.

[0103] In embodiment four, preferably, the fourth thermostat 4 is used as both a reverse transcription tank and an auxiliary denaturation tank. In this case, the four thermostats are arranged side by side in the order of the first thermostat 1, the fourth thermostat 4, the second thermostat 2 and the third thermostat 3.

[0104] At this time, the control module is configured to control the capillary tube 6 to transfer and cycle multiple times in the order of the fourth constant temperature bath 4, the first constant temperature bath 1, the fourth constant temperature bath 4, the second constant temperature bath 2, and the third constant temperature bath 3.

[0105] In each cycle, the temperature of the fourth thermostat 4 is first set to the fifth control temperature, and the capillary 6 is first placed in the fourth thermostat 4 with the fifth control temperature for reverse transcription. After the capillary 6 is removed from the fourth thermostat 4 and transferred to the first thermostat 1, the temperature of the fourth thermostat 4 is increased to the fourth control temperature, so that the sample can be transferred to the fourth thermostat 4 with the fourth control temperature after DNA denaturation in the first thermostat 1 for a period of time to continue denaturation, and then transferred to the subsequent thermostats.

[0106] For the timing of removing the capillary 6 from the fourth thermostat 4 used for reverse transcription, the first thermostat 1, the fourth thermostat 4 used as an auxiliary denaturation tank, the second thermostat 2, and the third thermostat 3, please refer to the above embodiments one to three.

[0107] Example 5

[0108] This fifth embodiment is an improvement based on the above embodiments. The technical content disclosed in the above embodiments will not be described again, and the content disclosed in the above embodiments also belongs to the content disclosed in this embodiment.

[0109] In Embodiment 5, the control module is also configured with an early warning function. Preferably, the control module is further configured to: set a first set time and a second set time; when the capillary tube 6 stays in the first constant temperature bath 1 for the first set time, and the temperature measured by the temperature measuring module 7 still has not reached the first set temperature, the control module determines that the first constant temperature bath 1 or the temperature measuring module 7 is faulty and issues a fault warning, while controlling the moving mechanism 5 to remove the capillary tube 6 from the first constant temperature bath 1; or when the capillary tube 6 stays in the second constant temperature bath 2 for the second set time, and the temperature measured by the temperature measuring module 7 still has not reached the second set temperature, the control module determines that the second constant temperature bath 2 or the temperature measuring module 7 is faulty and issues a fault warning, while controlling the moving mechanism 5 to remove the capillary tube 6 from the second constant temperature bath 2.

[0110] Examples 6 to 9

[0111] Examples 6 to 9 provide a control method for the PCR thermal cycling device in the above examples, so that the sample completes the amplification process in the PCR thermal cycling device.

[0112] Example 6

[0113] In Example 6, preferably, the control method for the PCR thermal cycling device includes the following steps:

[0114] Step 100: Set the temperature of the first constant temperature bath to the first control temperature, and the moving mechanism puts the capillary tube and the temperature measuring module into the first constant temperature bath together so as to heat the sample in the capillary tube through the first constant temperature bath.

[0115] When the temperature measured by the temperature measuring module reaches the first set temperature within the first set time, the capillary is removed from the first constant temperature bath to bring the temperature of the sample to the first temperature threshold range to complete DNA denaturation, and then step 200 is executed.

[0116] When the capillary tube remains in the first constant temperature bath for a first set time, and the temperature measured by the temperature measuring module still has not reached the first set temperature, the control module determines that the first constant temperature bath or the temperature measuring module is faulty and issues a fault warning. At the same time, it controls the moving mechanism to remove the capillary tube from the first constant temperature bath and stops operation. The following steps are not performed. The device is restarted after the fault is cleared.

[0117] Step 200: Set the temperature of the second constant temperature bath to the second control temperature, and the moving mechanism will transfer from the capillary tube to the second constant temperature bath so that the sample can be cooled and annealed in the second constant temperature bath.

[0118] When the temperature measured by the temperature measuring module reaches the second set temperature within the second set time, the capillary is removed from the second constant temperature bath so that the temperature of the sample can reach the second temperature threshold range when it enters the third constant temperature bath, so as to complete the cooling annealing, and then continue to execute step 300.

[0119] When the capillary tube remains in the second constant temperature bath for a second set time, and the temperature measured by the temperature measuring module still has not reached the second set temperature, the control module determines that the second constant temperature bath or the temperature measuring module is faulty and issues a fault warning. At the same time, it controls the moving mechanism to remove the capillary tube from the second constant temperature bath and stops operation. The following steps are not performed. The device is restarted after the fault is cleared.

[0120] Step 300: Set the temperature of the third constant temperature bath to the third control temperature, and the moving mechanism transfers the capillary to the third constant temperature bath so that the sample can complete the fluorescence signal acquisition in the third constant temperature bath. After the fluorescence signal acquisition of the sample is completed, the capillary is removed from the third constant temperature bath.

[0121] Step 400: Repeat steps 100-300, using a moving mechanism to drive the capillary to circulate multiple times in a predetermined order between multiple constant temperature baths to complete the sample amplification process.

[0122] Example 7

[0123] In Example 7, preferably, the control method for the PCR thermal cycling device includes the following steps:

[0124] Step 100c: Place the capillary tube into the fourth constant temperature bath with the sixth controlled temperature for a third predetermined time, and then remove it so that the sample in the capillary tube reaches the target temperature for the start of the cycle.

[0125] Step 100: Set the temperature of the first constant temperature bath to the first control temperature, and the moving mechanism puts the capillary tube and the temperature measuring module into the first constant temperature bath together so as to heat the sample in the capillary tube through the first constant temperature bath.

[0126] When the temperature measured by the temperature measuring module reaches the first set temperature within the first set time, the capillary is removed from the first constant temperature bath to bring the temperature of the sample to the first temperature threshold range to complete DNA denaturation, and then step 200 is executed.

[0127] When the capillary tube remains in the first constant temperature bath for a first set time, and the temperature measured by the temperature measuring module still has not reached the first set temperature, the control module determines that the first constant temperature bath or the temperature measuring module is faulty and issues a fault warning. At the same time, it controls the moving mechanism to remove the capillary tube from the first constant temperature bath and stops operation. The following steps are not performed. The device is restarted after the fault is cleared.

[0128] Step 101: Set the temperature of the fourth constant temperature bath to the fourth control temperature. The moving mechanism transfers the capillary tube removed from the first constant temperature bath to the fourth constant temperature bath with the fourth control temperature, and keeps the capillary tube in the fourth constant temperature bath for a first predetermined time, so that the sample continues to denature in the fourth constant temperature bath for a period of time.

[0129] Step 200: Set the temperature of the second constant temperature bath to the second control temperature, and the moving mechanism will transfer from the capillary tube to the second constant temperature bath so that the sample can be cooled and annealed in the second constant temperature bath.

[0130] When the temperature measured by the temperature measuring module reaches the second set temperature within the second set time, the capillary is removed from the second constant temperature bath so that the temperature of the sample can reach the second temperature threshold range when it enters the third constant temperature bath, so as to complete the cooling annealing, and then continue to execute step 300.

[0131] When the capillary tube remains in the second constant temperature bath for a second set time, and the temperature measured by the temperature measuring module still has not reached the second set temperature, the control module determines that the second constant temperature bath or the temperature measuring module is faulty and issues a fault warning. At the same time, it controls the moving mechanism to remove the capillary tube from the second constant temperature bath and stops operation. The following steps are not performed. The device is restarted after the fault is cleared.

[0132] Step 300: Set the temperature of the third constant temperature bath to the third control temperature, and the moving mechanism transfers the capillary to the third constant temperature bath so that the sample can complete the fluorescence signal acquisition in the third constant temperature bath. After the fluorescence signal acquisition of the sample is completed, the capillary is removed from the third constant temperature bath.

[0133] Step 400: Repeat steps 100-300, using a moving mechanism to drive the capillary to circulate multiple times in a predetermined order between multiple constant temperature baths to complete the sample amplification process.

[0134] Example 8

[0135] In Example 8, preferably, the control method for the PCR thermal cycling device includes the following steps:

[0136] Step 100a: Set the temperature of the fourth thermostat to the fifth control temperature. The moving mechanism places the capillary and temperature measuring module into the fourth thermostat and removes the capillary after a second predetermined time, so that the sample can complete reverse transcription in the fourth thermostat with the fifth control temperature.

[0137] Step 100: Set the temperature of the first constant temperature bath to the first control temperature, and the moving mechanism puts the capillary tube and the temperature measuring module into the first constant temperature bath together so as to heat the sample in the capillary tube through the first constant temperature bath.

[0138] When the temperature measured by the temperature measuring module reaches the first set temperature within the first set time, the capillary is removed from the first constant temperature bath to bring the temperature of the sample to the first temperature threshold range to complete DNA denaturation, and then step 200 is executed.

[0139] When the capillary tube remains in the first constant temperature bath for a first set time, and the temperature measured by the temperature measuring module still has not reached the first set temperature, the control module determines that the first constant temperature bath or the temperature measuring module is faulty and issues a fault warning. At the same time, it controls the moving mechanism to remove the capillary tube from the first constant temperature bath and stops operation. The following steps are not performed. The device is restarted after the fault is cleared.

[0140] Step 200: Set the temperature of the second constant temperature bath to the second control temperature, and the moving mechanism will transfer from the capillary tube to the second constant temperature bath so that the sample can be cooled and annealed in the second constant temperature bath.

[0141] When the temperature measured by the temperature measuring module reaches the second set temperature within the second set time, the capillary is removed from the second constant temperature bath so that the temperature of the sample can reach the second temperature threshold range when it enters the third constant temperature bath, so as to complete the cooling annealing, and then continue to execute step 300.

[0142] When the capillary tube remains in the second constant temperature bath for a second set time, and the temperature measured by the temperature measuring module still has not reached the second set temperature, the control module determines that the second constant temperature bath or the temperature measuring module is faulty and issues a fault warning. At the same time, it controls the moving mechanism to remove the capillary tube from the second constant temperature bath and stops operation. The following steps are not performed. The device is restarted after the fault is cleared.

[0143] Step 300: Set the temperature of the third constant temperature bath to the third control temperature, and the moving mechanism transfers the capillary to the third constant temperature bath so that the sample can complete the fluorescence signal acquisition in the third constant temperature bath. After the fluorescence signal acquisition of the sample is completed, the capillary is removed from the third constant temperature bath.

[0144] Step 400: Repeat steps 100a-300, using a moving mechanism to drive the capillary to circulate multiple times in a predetermined order between multiple constant temperature baths to complete the sample amplification process.

[0145] Example 9

[0146] In Example 9, preferably, the control method for the PCR thermal cycling device includes the following steps:

[0147] Step 100a: Set the temperature of the fourth thermostat to the fifth control temperature. The moving mechanism places the capillary and temperature measuring module into the fourth thermostat and removes the capillary after a second predetermined time, so that the sample can complete reverse transcription in the fourth thermostat with the fifth control temperature.

[0148] Step 100: Set the temperature of the first constant temperature bath to the first control temperature, and the moving mechanism puts the capillary tube and the temperature measuring module into the first constant temperature bath together so as to heat the sample in the capillary tube through the first constant temperature bath.

[0149] When the temperature measured by the temperature measuring module reaches the first set temperature within the first set time, the capillary is removed from the first constant temperature bath to bring the temperature of the sample to the first temperature threshold range to complete DNA denaturation, and then step 200 is executed.

[0150] When the capillary tube remains in the first constant temperature bath for a first set time, and the temperature measured by the temperature measuring module still has not reached the first set temperature, the control module determines that the first constant temperature bath or the temperature measuring module is faulty and issues a fault warning. At the same time, it controls the moving mechanism to remove the capillary tube from the first constant temperature bath and stops operation. The following steps are not performed. The device is restarted after the fault is cleared.

[0151] Step 101: Set the temperature of the fourth constant temperature bath to the fourth control temperature. The moving mechanism transfers the capillary tube removed from the first constant temperature bath to the fourth constant temperature bath with the fourth control temperature, and keeps the capillary tube in the fourth constant temperature bath for a first predetermined time, so that the sample continues to denature in the fourth constant temperature bath for a period of time.

[0152] Step 200: Set the temperature of the second constant temperature bath to the second control temperature, and the moving mechanism will transfer from the capillary tube to the second constant temperature bath so that the sample can be cooled and annealed in the second constant temperature bath.

[0153] When the temperature measured by the temperature measuring module reaches the second set temperature within the second set time, the capillary is removed from the second constant temperature bath so that the temperature of the sample can reach the second temperature threshold range when it enters the third constant temperature bath, so as to complete the cooling annealing, and then continue to execute step 300.

[0154] When the capillary tube remains in the second constant temperature bath for a second set time, and the temperature measured by the temperature measuring module still has not reached the second set temperature, the control module determines that the second constant temperature bath or the temperature measuring module is faulty and issues a fault warning. At the same time, it controls the moving mechanism to remove the capillary tube from the second constant temperature bath and stops operation. The following steps are not performed. The device is restarted after the fault is cleared.

[0155] Step 300: Set the temperature of the third constant temperature bath to the third control temperature, and the moving mechanism transfers the capillary to the third constant temperature bath so that the sample can complete the fluorescence signal acquisition in the third constant temperature bath. After the fluorescence signal acquisition of the sample is completed, the capillary is removed from the third constant temperature bath.

[0156] Step 400: Repeat steps 100a-300, using a moving mechanism to drive the capillary to circulate multiple times in a predetermined order between multiple constant temperature baths to complete the sample amplification process.

[0157] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A PCR thermal cycling device, characterized in that, Includes a moving mechanism, a first constant temperature bath, a fourth constant temperature bath, and a control module; The first thermostatic bath can be configured to have a first control temperature; The moving mechanism is equipped with a capillary tube containing a sample and a temperature measuring module, and the moving mechanism can drive the capillary tube and the temperature measuring module to move synchronously. The fourth thermostatic bath can be configured to have a sixth controlled temperature; The temperature measuring module is a temperature sensor with a metal shell or a ceramic shell. The temperature measuring module and the capillary have different transient thermal response characteristics, and a stable temperature correspondence is established between the temperature measuring module and the capillary: in the first constant temperature bath with a first control temperature, when the temperature measured by the temperature measuring module reaches the first set temperature, the temperature of the sample in the capillary is within the first temperature threshold range. The control module is configured to: The moving mechanism is controlled to transfer the capillary tube and the temperature measuring module into the fourth constant temperature bath with the sixth control temperature, and the moving mechanism is controlled to move the capillary tube out of the fourth constant temperature bath so that the sample or the temperature measuring module reaches the cycle start target temperature after the capillary tube is moved out of the fourth constant temperature bath. The moving mechanism is controlled to transfer the capillary tube and the temperature measuring module from the fourth constant temperature bath to the first constant temperature bath; When the temperature measured by the temperature measuring module reaches the first set temperature, the moving mechanism is controlled to move the capillary tube out of the first constant temperature bath, so that the temperature of the sample is within the first temperature threshold range after the capillary tube is moved out of the first constant temperature bath; the first control temperature is greater than the first set temperature. The sixth control temperature is the target temperature at the start of the cycle, and the sixth control temperature is lower than the first set temperature.

2. The PCR thermal cycling apparatus according to claim 1, characterized in that, The moving mechanism is controlled to transfer the capillary tube and the temperature measuring module into the fourth constant temperature bath with the sixth control temperature for a third predetermined time, and then remove them so that the sample or the temperature measuring module reaches the target temperature for the start of the cycle.

3. The PCR thermal cycling apparatus according to claim 1, characterized in that, The first set temperature is greater than the first temperature threshold.

4. The PCR thermal cycling apparatus according to claim 1, characterized in that, The PCR thermal cycling device further includes a second thermostat, which can be configured to have a second controlled temperature. The control module is also configured to: The moving mechanism is controlled to transfer the capillary tube from the first constant temperature bath to the second constant temperature bath; When the temperature measured by the temperature measuring module reaches the second set temperature, the moving mechanism is controlled to move the capillary tube out of the second constant temperature bath, so that the temperature of the sample is within the second temperature threshold range after the capillary tube is moved out of the second constant temperature bath. The second control temperature is lower than the second set temperature.

5. The PCR thermal cycling apparatus according to claim 4, characterized in that, The PCR thermal cycling device also includes a third thermostat, which can be configured to have a third controlled temperature. The control module is also configured to: The moving mechanism is controlled to transfer the capillary tube from the second constant temperature bath to the third constant temperature bath; After the sample in the capillary completes fluorescence signal acquisition in the third constant temperature bath, the moving mechanism is controlled to remove the capillary from the third constant temperature bath.

6. The PCR thermal cycling apparatus according to claim 5, characterized in that, The fourth thermostatic bath can also be configured to have a fourth controlled temperature; The control module is also configured to: The moving mechanism is controlled to sequentially transfer the capillary to the fourth constant temperature bath with the sixth control temperature, the first constant temperature bath, the fourth constant temperature bath with the fourth control temperature, the second constant temperature bath, and the third constant temperature bath, and to keep the capillary in the fourth constant temperature bath with the fourth control temperature for a first predetermined time. The fourth control temperature is the target temperature for sample denaturation.

7. The PCR thermal cycling apparatus according to claim 6, characterized in that, The first constant temperature bath, the fourth constant temperature bath, the second constant temperature bath, and the third constant temperature bath are arranged side by side with intervals.

8. The PCR thermal cycling apparatus according to claim 1, characterized in that, The control module is also configured to: When the capillary tube remains in the first constant temperature bath for a first set time, and the temperature measured by the temperature measuring module does not reach the first set temperature, the moving mechanism is controlled to remove the capillary tube from the first constant temperature bath.

9. A method for controlling a PCR thermal cycling apparatus, used in any one of claims 5 to 8; characterized in that, Includes the following steps: Step 100c: Place the capillary tube into the fourth thermostat bath with the sixth temperature control. Remove the sample or temperature measuring module inside the capillary tube after it reaches the target temperature for the start of the cycle. Step 100: Place the capillary tube into the first constant temperature bath, and remove it when the temperature measured by the temperature measuring module reaches the first set temperature, so that the sample temperature in the capillary tube is within the first temperature threshold range. Step 200: Transfer the capillary tube to the second constant temperature bath. When the temperature measured by the temperature measuring module reaches the second set temperature, remove the capillary tube so that the sample temperature in the capillary tube is within the second temperature threshold range. Step 300: Transfer the capillary tube to the third constant temperature bath, and remove it after completing the fluorescence signal acquisition of the sample; Step 400: Repeat steps 100 to 300 a predetermined number of times to complete multiple loops; Step 200 also includes step 200a: when the temperature measured by the temperature measuring module in the second constant temperature bath within the second set time does not reach the second set temperature, the control module determines that the second constant temperature bath or the temperature measuring module is faulty and issues a fault warning. The control module controls the moving mechanism to remove the capillary tube from the second constant temperature bath.

10. The control method for the PCR thermal cycling apparatus according to claim 9, characterized in that, In step 100c, the capillary is placed in the fourth constant temperature bath with the sixth control temperature for a third predetermined time and then removed so that the sample in the capillary reaches the target temperature for the start of the cycle.

11. The control method for the PCR thermal cycling apparatus according to claim 9, characterized in that, Step 101 is further included between step 100 and step 200: The capillary tube is transferred to a fourth thermostatic bath with a fourth controlled temperature, and removed after a first predetermined time.

12. The control method for the PCR thermal cycling apparatus according to claim 9, characterized in that, Step 100 further includes step 100b: If the temperature measured by the temperature measuring module in the first constant temperature bath does not reach the first set temperature within a first set time, the control module controls the moving mechanism to remove the capillary tube from the first constant temperature bath.

13. The control method for the PCR thermal cycling apparatus according to claim 9, characterized in that, In step 400, during the first cycle, the first extraction temperature of the temperature measuring module is higher than the first extraction temperature in subsequent cycles.

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