A thermostated nucleic acid amplification reaction tube

Abstract

The utility model relates to medical detection tool technical field, proposes a constant temperature nucleic acid amplification reaction tube, including first pipe body, the pipe mouth end threaded connection of first pipe body has the second pipe body, the inboard fixed connection of second pipe body has the seal, the pipe mouth end sliding fit of second pipe body has the pipe cover, and the sealed cabin is formed between the pipe cover and the seal membrane, one end fixed connection of first pipe body inboard has the conical puncture cover, is set up a plurality of leak holes on the conical puncture cover, the inboard rotatory connection of first pipe body has the rotating rod, the rotating rod's outboard fixed connection has a plurality of magnetic stripe along the rotating rod length direction equidistance distribution, the outboard fixed connection of first pipe body has two parallelly arranged limit ring, set up the driving element for urging the rotating rod rotation between two limit rings. The utility model discloses through the innovative one -tube method design, high -performance sealing material, built -in magnetic force stirring system and magnetic force drive mechanism, and the accuracy, reliability and operation convenience of constant temperature nucleic acid amplification reaction are comprehensively promoted.

Description

Technical Field

[0001] This utility model relates to the field of medical testing tools, specifically to an isothermal nucleic acid amplification reaction tube. Background Technology

[0002] Recombinase polymerase amplification (RPA) is a technique based on the recombinase polymerase-mediated amplification principle. It mimics the enzymatic reaction process of DNA replication in vivo and relies on a specific combination of enzymes and proteins (recombinase, single-strand binding protein, and DNA polymerase) to specifically amplify the DNA template. It is an isothermal nucleic acid amplification technique that can achieve rapid and specific amplification at near-body temperature (25℃-42℃, 5-30 min). As an isothermal technique, it reduces the dependence on high-precision and expensive instruments, stable power supply facilities, and high-level laboratories. It only requires a thermostat and can be completed with a fluorescence analysis device to determine the presence of amplification products. It has the characteristics of relatively simple facilities and operation requirements and has been widely used in many fields such as life science research, medical testing, agriculture, food safety, and transgenic detection.

[0003] To further improve the detection sensitivity of RPA, researchers have proposed a nested RPA detection method: first, the first fragment of the target gene is amplified using outer primers, and then the second fragment of the target gene, located entirely within the first amplified fragment, is amplified using inner primers. To eliminate the influence of the enzyme's fluorescence signal, the first RPA reaction system does not contain a fluorescent probe. A fluorescent probe is added in the second RPA reaction. Experiments have confirmed that nestRPA has extremely high sensitivity, providing strong technical support for the early detection of various viruses such as SARS-CoV-2, influenza, and swine fever. However, nestRPA requires two RPA reactions. After the first RPA reaction, the reaction tube cap must be opened, the inner primers and probes required for the second RPA reaction added, and then the reaction tube cap closed for the second RPA reaction, continuing until the reaction is complete and the detection result is obtained.

[0004] One issue is that opening the reaction tube cap after the first RPA reaction can easily lead to nucleic acid aerosol contamination, resulting in false positive results. Therefore, there is an urgent need to develop a reaction tube that can complete nestRPA in a "one-tube" manner, allowing for a second RPA reaction without opening the tube cap after the first RPA reaction. Utility Model Content

[0005] This invention proposes an isothermal nucleic acid amplification reaction tube, which solves the problems of easy contamination and reduced accuracy of detection results in related technologies.

[0006] The technical solution of this utility model is as follows: A constant temperature nucleic acid amplification reaction tube includes a first tube body for pre-storing a second reactant, a second tube body threadedly connected to the end of the first tube body, a sealing film fixedly connected to the inner side of the second tube body, a tube cap slidably fitted to the end of the second tube body, a sealed chamber for placing the first reactant formed between the tube cap and the sealing film, a conical puncture sleeve fixedly connected to one end of the inner side of the first tube body, the conical puncture sleeve having a plurality of leakage holes, a rotating rod rotatably connected to the inner side of the first tube body, a plurality of magnetic strips equidistantly distributed along the length direction of the rotating rod fixedly connected to the outer side of the rotating rod, two parallel limiting rings fixedly connected to the outer side of the first tube body, and a driving component for driving the rotating rod to rotate between the two limiting rings.

[0007] Preferably, the outer diameter of the first tube is the same as the inner diameter of the second tube, and the length of the first tube is greater than that of the second tube.

[0008] Preferably, the pipe end of the first pipe body is fixedly connected with an external thread, and the inner side of the second pipe body is provided with an internal thread groove that mates with the external thread.

[0009] Preferably, the sealing film is a brittle metal foil or a polymer diaphragm.

[0010] Preferably, a first rotating sleeve is fixedly connected to the outer side of the second tube, and the outer edge of the first rotating sleeve is provided with a plurality of annularly distributed anti-slip grooves.

[0011] Preferably, the driving component includes a sleeve fitted on the outside of the first tube body, the sleeve being rotatably connected to two limiting rings, an arc-shaped plate being fixedly connected to one end of the inner side of the sleeve, and an arc-shaped magnetic block being fixedly connected to the other end of the inner side of the sleeve, the arc-shaped magnetic block and the arc-shaped plate being spliced ​​together to form a ring structure.

[0012] Preferably, the length of the arc-shaped magnetic block is greater than the length of the arrangement of several magnetic strips, and the arc-shaped magnetic block is concentrically arranged with the first tube body.

[0013] Preferably, a second rotating cap is fixedly connected to the outer side of the sleeve, and the outer edge of the second rotating cap is provided with a plurality of annularly distributed anti-slip grooves.

[0014] The working principle and beneficial effects of this utility model are as follows:

[0015] 1. After the first reaction is completed in the sealed chamber of the second tube, simply remove the tube cap, connect the second tube to the first tube containing the second reactant via thread, and puncture the sealing film with a conical puncture sleeve to allow the first reaction product to flow into the first tube, thus avoiding the risk of contamination from opening the cap. The entire process is carried out in a sealed environment, which significantly reduces the probability of false positives and improves the accuracy of medical testing.

[0016] 2. The sealing film is made of brittle metal foil or polymer membrane, which has high brittleness and temperature resistance, ensuring that the first reactant will not leak or evaporate during storage and reaction; the sealing film completely isolates the first reactant and the second reactant until the mixing is triggered, avoiding cross-contamination or degradation of pre-stored reagents.

[0017] 3. The magnetic strips on the rotating rod are driven by the driving component to rotate, which magnetically stirs the mixture, so that the nucleic acid template and the second reactant are quickly and evenly mixed, which is beneficial to the efficiency and repeatability of the second reaction. The equidistantly distributed magnetic strips and the optimized driving component provide continuous and stable stirring force, reducing reaction errors. Attached Figure Description

[0018] The present invention will now be described in further detail with reference to the accompanying drawings and specific embodiments.

[0019] Figure 1 This is a schematic diagram of the structure of an isothermal nucleic acid amplification reaction tube proposed in this utility model. Figure 1 ;

[0020] Figure 2 This is a schematic diagram of the structure of an isothermal nucleic acid amplification reaction tube proposed in this utility model. Figure 2 ;

[0021] Figure 3 This is a schematic diagram of the structure of the second tube body proposed in this utility model;

[0022] Figure 4 This is a schematic diagram of the internal structure of the first tube body proposed in this utility model;

[0023] Figure 5 This is a schematic diagram of the structure of the driving component proposed in this utility model.

[0024] In the figure: 1. First tube body; 11. External thread; 12. Conical puncture sleeve; 13. Leakage hole; 14. Rotating rod; 15. Magnetic strip; 16. Limiting ring; 17. Driving component; 171. Tube sleeve; 172. Second rotating cap; 173. Arc plate; 174. Arc magnetic block; 2. Second tube body; 21. Internal thread groove; 22. First rotating sleeve; 23. Sealing film; 3. Tube cap. Detailed Implementation

[0025] The technical solutions of this utility model will be clearly and completely described below with reference to the embodiments of this utility model. Obviously, the described embodiments are only some embodiments of this utility model, and not all embodiments. Based on the embodiments of this utility model, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of this utility model.

[0026] Please see Figure 1 and Figure 5 This application discloses an isothermal nucleic acid amplification reaction tube, including a first tube body 1 for pre-storing a second reactant, a second tube body 2 being threadedly connected to the end of the first tube body 1, an external thread 11 being fixedly connected to the end of the first tube body 1, and an internal thread groove 21 being formed on the inner side of the second tube body 2 that is threadedly engaged with the external thread 11.

[0027] The outer diameter of the first tube 1 is the same as the inner diameter of the second tube 2, and the length of the first tube 1 is greater than that of the second tube 2. A sealing film 23 is fixedly connected to the inner side of the second tube 2. The sealing film 23 is a brittle metal foil film. A tube cap 3 is slidably fitted to the tube end of the second tube 2. A sealed chamber for placing the first reactant is formed between the tube cap 3 and the sealing film 23. A conical puncture sleeve 12 is fixedly connected to one end of the inner side of the first tube 1. Several leakage holes 13 are opened on the conical puncture sleeve 12. A rotating rod 14 is rotatably connected to the inner side of the first tube 1. Several magnetic strips 15 are fixedly connected to the outer side of the rotating rod 14 and are evenly distributed along the length direction of the rotating rod 14. Two parallel limiting rings 16 are fixedly connected to the outer side of the first tube 1. A driving member 17 for driving the rotating rod 14 to rotate is provided between the two limiting rings 16.

[0028] In this embodiment, the overall structure's workflow is as follows:

[0029] Preparation stage: The second reaction reagent is added to the bottom of the first tube 1 beforehand. The first reaction reagent is added to the sealed compartment of the second tube 2, and the tube cap 3 is closed for sealing. At this time, the two reaction systems are physically isolated by the sealing film 23, and can be stored and transported independently;

[0030] First reaction: Place only the second tube portion of the assembled reaction tube into the isothermal amplification instrument to carry out the first nucleic acid amplification reaction. The reaction is completed in a sealed chamber.

[0031] Mixing triggering stage: After the first reaction is completed, remove the tube cap 3 and screw the second tube 2 with the first tube 1 which has been pre-stored with the second reactant through the thread. During the screwing process, the conical puncture sleeve 12 at the top of the first tube 1 will puncture and completely destroy the sealing film 23 at the bottom of the second tube 2.

[0032] Liquid transfer: After the sealing film 23 is punctured, the first reaction product containing the amplified nucleic acid template flows into the bottom of the first tube 1 through the leak hole 13 on the conical puncture sleeve 12 under the action of gravity, and comes into contact with the second reaction reagent.

[0033] Mixing and Second Reaction: After the liquid is mixed, the internal magnetic strip 15 is rotated by the external drive 17, which drives the rotating rod 14 to rotate, and the mixture is magnetically stirred to ensure that it is fully mixed. Then the entire reaction tube is placed in the constant temperature amplification instrument for the second amplification reaction.

[0034] The entire process does not require opening the main reaction chamber. The product of the first reaction flows into the second reaction chamber through the punctured sealing film, enabling two reactions to be completed within a closed system. This fundamentally eliminates aerosol contamination caused by opening the lid and significantly improves detection accuracy. The use of brittle metal foil or polymer diaphragms replaces the ordinary sealing film previously used in patents, greatly improving sealing strength, temperature resistance, and reliability, ensuring that the reaction solution does not leak or evaporate during storage and the first reaction. The built-in magnetic stirring system provides active mixing capabilities, ensuring that the two reaction solutions are quickly and thoroughly mixed uniformly, overcoming the problem of insufficient mixing that may result from relying solely on centrifugation or diffusion. This improves the efficiency and repeatability of the second reaction.

[0035] In a further preferred embodiment of this utility model, the driving component 17 includes a sleeve 171 sleeved on the outside of the first tube body 1. The sleeve 171 is rotatably connected to two limiting rings 16. An arc-shaped plate 173 is fixedly connected to one end of the inner side of the sleeve 171, and an arc-shaped magnetic block 174 is fixedly connected to the other end of the inner side of the sleeve 171. The arc-shaped magnetic block 174 and the arc-shaped plate 173 are spliced ​​to form a ring structure. The length of the arc-shaped magnetic block 174 is greater than the arrangement length of a number of magnetic strips 15. The arc-shaped magnetic block 174 is concentrically arranged with the first tube body 1.

[0036] In this embodiment, when the operator manually rotates the second rotating cap 172, it will cause the sleeve 171 to rotate. The arc-shaped magnetic block 174 fixed inside the sleeve 171 will rotate accordingly. Due to the magnetic force, this rotating external magnetic field will drive the magnetic strip 15 inside the first tube 1 to rotate as well, thereby driving the rotating rod 14 to rotate and realize the stirring function. Power transmission is realized through magnetic coupling. The inside of the reaction tube remains completely sealed, and there is no need to open a hole in the tube wall to introduce a drive shaft, which further ensures the sealing integrity of the system and prevents potential contamination.

[0037] The length of the arc-shaped magnetic block 174 is greater than the length of all the internal magnetic strips 15), and it is concentrically set with the first tube 1. This design ensures that the arc-shaped magnetic block 174 can effectively couple to the magnetic force regardless of the angle of the rotating rod 14 on the circumference, providing a stable and continuous torque and avoiding jamming.

[0038] In a further preferred embodiment of the present invention, a first rotating sleeve 22 is fixedly connected to the outer side of the second tube body 2, and a plurality of annularly distributed anti-slip grooves are provided on the outer edge of the first rotating sleeve 22; a second rotating cap 172 is fixedly connected to the outer side of the tube sleeve 171, and a plurality of annularly distributed anti-slip grooves are provided on the outer edge of the second rotating cap 172.

[0039] In this embodiment, the second rotating cap 172 and its anti-slip groove, as well as the first rotating sleeve 22 on the second tube body 2, provide convenience for manual operation, making the action of rotating the tube body and driving the stirring more effortless and stable.

[0040] The above are merely preferred embodiments of the present utility model and are not intended to limit the present utility model. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present utility model shall be included within the protection scope of the present utility model.

Claims

1. A isothermal nucleic acid amplification reaction tube, characterized in that, The device includes a first tube (1) for pre-storing a second reactant, a second tube (2) threadedly connected to the end of the first tube (1), a sealing film (23) fixedly connected to the inner side of the second tube (2), a tube cap (3) slidably fitted to the end of the second tube (2), a sealed chamber for inserting the first reactant formed between the tube cap (3) and the sealing film (23), a conical puncture sleeve (12) fixedly connected to one end of the inner side of the first tube (1), a plurality of leakage holes (13) provided on the conical puncture sleeve (12), a rotating rod (14) rotatably connected to the inner side of the first tube (1), a plurality of magnetic strips (15) equidistantly distributed along the length direction of the rotating rod (14) fixedly connected to the outer side of the rotating rod (14), two parallel limiting rings (16) fixedly connected to the outer side of the first tube (1), and a driving component (17) for driving the rotating rod (14) to rotate is provided between the two limiting rings (16).

2. The isothermal nucleic acid amplification reaction tube according to claim 1, characterized in that, The outer diameter of the first tube (1) is the same as the inner diameter of the second tube (2), and the length of the first tube (1) is greater than that of the second tube (2).

3. The isothermal nucleic acid amplification reaction tube according to claim 1, characterized in that, The first tube (1) has an external thread (11) fixedly connected to its opening end, and the second tube (2) has an internal thread groove (21) that is threaded to the external thread (11).

4. The isothermal nucleic acid amplification reaction tube according to claim 1, characterized in that, The sealing film (23) is a brittle metal foil or a polymer diaphragm.

5. The isothermal nucleic acid amplification reaction tube according to claim 1, characterized in that, The outer side of the second tube (2) is fixedly connected to a first rotating sleeve (22), and the outer edge of the first rotating sleeve (22) is provided with several annularly distributed anti-slip grooves.

6. The isothermal nucleic acid amplification reaction tube according to claim 1, characterized in that, The driving component (17) includes a sleeve (171) sleeved on the outside of the first tube body (1). The sleeve (171) is rotatably connected to two limiting rings (16). An arc plate (173) is fixedly connected to one end of the inner side of the sleeve (171), and an arc magnetic block (174) is fixedly connected to the other end of the inner side of the sleeve (171). The arc magnetic block (174) and the arc plate (173) are spliced ​​together to form a ring structure.

7. The isothermal nucleic acid amplification reaction tube according to claim 6, characterized in that, The length of the arc-shaped magnetic block (174) is greater than the length of the arrangement of several magnetic strips (15), and the arc-shaped magnetic block (174) is concentrically arranged with the first tube body (1).

8. The isothermal nucleic acid amplification reaction tube according to claim 6, characterized in that, The outer side of the sleeve (171) is fixedly connected to a second rotating cap (172), and the outer edge of the second rotating cap (172) is provided with several annularly distributed anti-slip grooves.

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