CN212955133U - Reaction tube and reaction tube set for two-step nucleic acid amplification and detection - Google Patents
Reaction tube and reaction tube set for two-step nucleic acid amplification and detection Download PDFInfo
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- CN212955133U CN212955133U CN202021464690.3U CN202021464690U CN212955133U CN 212955133 U CN212955133 U CN 212955133U CN 202021464690 U CN202021464690 U CN 202021464690U CN 212955133 U CN212955133 U CN 212955133U
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Abstract
The utility model discloses a reaction tube and a reaction tube set for two-step nucleic acid amplification and detection. The reaction tube comprises a tube body and a cover body, wherein the cover body comprises a liquid box for temporarily storing the second reaction liquid during the first-step reaction; the reaction tube also comprises an upper cover, the upper cover comprises a sleeve joint part and a sealing plate, and the bottom surface of the sealing plate is provided with a puncture needle; and a protection ring surrounding the sealing plate and used for protecting the sealing plate is also arranged above the sleeving part. Through sealing the second reaction liquid in the liquid box, and because the closing plate receives the protection of protection ring, thereby be difficult to take place the mistake and bump the condition that the closing plate punctures the seal membrane, the guarantee adds the safety of reaction tube at in-process such as transportation, sale behind the second reaction liquid in advance, maloperation when also can avoiding using. It is necessary to pierce the sealing membrane and to operate it with a specially configured pressing tool.
Description
Technical Field
The utility model relates to a biological detection technical field especially relates to a reaction tube and reaction tube suit that two-step method nucleic acid amplification, detection used.
Background
The nucleic acid amplification detection technology is the basis for developing molecular biology research, can be used for qualitatively and quantitatively analyzing and detecting trace nucleic acid, plays an important role in various related fields of clinical medicine, inspection medicine, molecular biology, genomics, food safety and the like, and is an important inspection method indispensable to the development of life science. Existing nucleic acid amplification techniques can be divided into two categories depending on whether temperature cycling is required: the first type is a temperature-variable amplification system, which includes Polymerase Chain Reaction (PCR), ligase chain reaction (LRC), and the like. Among them, the PCR technique is the most widely used technique for in vitro amplification of nucleic acids at present because of its advantages such as strong specificity, high sensitivity and low cost. However, this technique requires special equipment and also has certain professional requirements on the operator and the experimental conditions. The second type is an isothermal Amplification system, which includes Loop-mediated isothermal Amplification (LAMP), Rolling Circle Amplification (RCA), Recombinase Polymerase Amplification (RPA), etc., and the technology does not require a thermal cycling process or a special instrument, but the Amplification is prone to false positive.
In recent years, the advent of CRISPR technology has brought about a more convenient and sensitive detection format. The main principle is that after the Cas enzyme is combined with a target sequence under the guidance of guide RNA in a system, the Cas enzyme is switched into an activated state, and other single-stranded DNA in the system is efficiently cut. After a single-stranded DNA probe substrate containing a reporter group is added into the system, the Cas protein recognizes the existence of a target sequence, and then cuts the single-stranded DNA probe substrate to release a fluorescent reporter group to generate a fluorescent signal. In the existing system for detecting CRISPR, the CRISPR system cannot be added into the reaction system at the initial moment due to the influence of the reaction temperature, and the subsequent addition is carried out after the reaction is finished and secondary uncapping is carried out, so that serious aerosol pollution and false positive problems are easily caused, and the whole detection operation process becomes complicated and fussy.
The invention patent application with the publication number of CN111334425A, which was previously filed by the applicant, discloses a reaction tube for coupling two systems for nucleic acid amplification and CRISPR detection, and a method for coupling two systems, wherein the reaction tube for coupling two systems comprises a tube body and a cover body which is matched and sealed with the tube body, and the inner cavity of the tube body comprises a reaction area at the lower part and a storage area at the upper part; the storage region is provided with a storage chamber for temporarily storing the CRISPR detection reagent during nucleic acid amplification, and a release mechanism for releasing the CRISPR detection reagent from the storage chamber to the reaction region after the nucleic acid amplification is completed. In the prior art, either the opening of the storage chamber is arranged, the CRISPR detection reagent is released from the storage chamber to the reaction region in an inclined and inverted manner after the first-step reaction is completed, and due to the arrangement of the opening of the storage chamber, the liquids of the two-step reaction may be accidentally mixed, which affects the result; or the storage chamber is sealed, but the CRISPR detection reagent is released from the storage chamber to the reaction region by directly pressing the cover, and the corresponding region on the cover is easily touched accidentally, so that the CRISPR detection reagent is released from the storage chamber to the reaction region, and the result is influenced.
SUMMERY OF THE UTILITY MODEL
The utility model discloses to the above-mentioned not enough that exists among the prior art, provide a space of storing second reaction liquid and set up for sealing, and be used for the structure of second reaction liquid release to be difficult to by the two-step method nucleic acid amplification of maloperation, the reaction tube of detection usefulness.
A reaction tube for nucleic acid amplification and detection in a two-step method comprises a tube body for adding a first reaction liquid and a cover body matched and sealed with the tube body, wherein the cover body comprises a liquid box for temporarily storing a second reaction liquid in a first-step reaction, the bottom surface of the liquid box is provided with a first opening for allowing the second reaction liquid to flow into the tube body, the top surface of the liquid box is provided with a second opening for adding the second reaction liquid into the liquid box, and a sealing film is used for sealing the first opening before adding the second reaction liquid;
the liquid box is provided with a plug part extending into the tube body and matched with the opening part of the tube body in an inserted manner, and a sealing part positioned outside the tube body, the first opening is positioned on the bottom surface of the plug part, and the second opening is positioned on the top surface of the sealing part;
the reaction tube also comprises an upper cover used for sealing the second opening, the upper cover comprises a sleeve joint part matched with the sealing part in a sleeve joint way, and a sealing plate which is arranged on the top surface of the sleeve joint part and can be bent towards one side of the tube body, and the bottom surface of the sealing plate is provided with a puncture needle used for puncturing the sealing membrane when the sealing plate is bent towards one side of the tube body; and a protection ring surrounding the sealing plate and used for protecting the sealing plate is also arranged above the sleeving part.
Preferably, the sealing plate is an arc protruding towards the outer side of the reaction tube, and a plurality of thinning grooves for thinning the sealing plate are arranged on the outer side surface and/or the inner side surface of the sealing plate at intervals. This application reaction tube generally uses plastics preparation, and the closing plate of plastics makes the closing plate conveniently buckle through setting up some attenuate grooves, buckles to become to the inboard protrusion from the protrusion of outside to pierce the seal membrane, release second reaction liquid.
More preferably, all the thinning grooves are arranged in concentric circles centering on the center of the sealing plate, so that the sealing plate can be bent by pressing the middle region of the sealing plate.
Further preferably, at least one side of the cross section of the thinning groove close to the bottom surface is an arc groove; one thinning groove at the outermost ring is arranged on the inner side surface of the sealing plate, and the other thinning grooves are arranged on the outer side surface of the sealing plate. Compared with the design with a sharp angle, the arc design is not easy to break in the bending process. The design of the thinning groove can achieve the purpose of being convenient to bend on the inner side face or the outer side face of the sealing plate, when the sealing plate is bent, the connection part of the outermost side region and the sleeve joint part of the sealing plate is not easy to bend, and the middle region is bent towards the inner side, so that the thinning groove on the outermost ring is arranged on the inner side face of the sealing plate, and the rest thinning grooves are arranged on the outer side face of the sealing plate, so that the thinning groove can be adapted to the bending condition of the sealing plate, and the thinning groove is in a furled state when the sealing plate is bent, and the sealing.
More preferably, the central region of the inner side surface of the sealing plate protrudes outwards to form a mounting groove for fixedly mounting the puncture needle, and one end of the puncture needle is provided with a mounting part in interference fit with the mounting groove. The puncture needle is clamped into the mounting groove through the mounting part and fixed.
Preferably, the outer periphery of the puncture needle is provided with a flow guide groove arranged along the axial direction. The design of the diversion trench is convenient for the second reaction liquid to flow into the pipe body along the diversion trench after the sealing film is punctured.
Preferably, a first annular groove and a first convex ring which are matched with each other are arranged between the inner side wall of the tube body and the outer side wall of the plug part; and a second annular groove and a second convex ring which are matched with each other are arranged between the inner side wall of the sleeving part and the outer side wall of the sealing part. The design of the annular groove and the convex ring can enhance the sealing effect.
Preferably, a baffle ring is arranged on the outer side wall of the liquid box and between the plug part and the sealing part, during assembly, the top surface of the pipe body is abutted against the lower surface of the baffle ring, and the bottom surface of the sleeve part is abutted against the upper surface of the baffle ring. The design of retaining ring can be spacing to the cooperation between liquid box and the body and the cooperation between upper cover and the liquid box.
Preferably, the inner cavity of the liquid cartridge is tapered from top to bottom. The gradual reduction in this application can be the reduction of continuous variation, also can be the reduction of sectional type, as long as there is the trend that reduces, then the second reaction liquid of being convenient for assembles the bottom of liquid box inner chamber, punctures the seal membrane after, makes things convenient for the second reaction liquid to flow to the body in.
The application also provides a reaction tube set for two-step nucleic acid amplification and detection, which comprises the reaction tube and an extrusion tool for extruding the sealing plate to bend the sealing plate towards one side of the tube body, wherein the extrusion tool comprises a finger sleeve for a finger to pass through during operation and an extrusion head arranged on the periphery of the finger sleeve; the extrusion head comprises an extrusion part and a limiting part, wherein the extrusion part extends into the protective ring to extrude the sealing plate when in use, and the limiting part limits the depth of the extrusion part extending into the protective ring by abutting against the top surface of the protective ring. The design of protective ring in this application upper cover for the closing plate is difficult to touch to the finger, thereby can avoid the maloperation. When the second reaction liquid needs to be released, the sealing plate is pressed down by using the extrusion tool, and the pressing stroke is limited by the limiting part, so that overpressure is prevented.
The utility model discloses a seal the second reaction liquid in the liquid box, the reaction tube slope, can not influence the second reaction liquid under the circumstances such as rock yet to can be added the second reaction liquid in advance to the reaction tube by the producer, then transport, sell etc.. And because the closing plate receives the protection of protection ring, thereby be difficult to take place the mistake and touch the condition that the closing plate thereby punctures the seal membrane, the safety of reaction tube in transportation, selling etc. in-process after further the guarantee adds second reaction liquid in advance also can avoid the maloperation when using.
Drawings
FIG. 1 is a schematic perspective view of a reaction tube according to the present invention.
FIG. 2 is a schematic diagram of an exploded structure of a reaction tube according to the present invention.
FIG. 3 is a schematic diagram of a side view of a reaction tube according to the present invention.
Fig. 4 is a sectional view taken along a-a in fig. 3.
FIG. 5 is a partial cross-sectional view of a reaction tube according to the present application.
Fig. 6 is a sectional view of the upper cover.
Fig. 7 is a sectional view of the liquid cartridge.
FIG. 8 is a perspective view of the lancet.
FIG. 9 is a side view of the lancet.
Fig. 10 is a sectional view taken along the direction B-B in fig. 9.
Fig. 11 is a schematic structural view of the pressing tool according to the present invention before the sealing plate is bent downward.
Fig. 12 is a schematic structural view of the sealing plate press-bending process by the pressing tool according to the present invention.
Fig. 13 is a schematic structural view of the sealing plate after the pressing tool of the present application presses and bends the sealing plate.
Detailed Description
As shown in FIGS. 1 to 13, a two-step nucleic acid amplification and detection reaction tube comprises a tube body 1 for adding a first reaction solution and a cover body 2 sealed with the tube body 1, wherein the cover body 2 comprises a liquid box 3 for temporarily storing a second reaction solution during a first reaction step, the bottom surface of the liquid box 3 is provided with a first opening 31 for allowing the second reaction solution to flow into the tube body 1, the top surface of the liquid box 3 is provided with a second opening 32 for adding the second reaction solution into the liquid box 3, and the first opening 31 is sealed with a sealing film before adding the second reaction solution. The whole structure of the tube body 1 is similar to that of a conventional PCR reaction tube, and the bottom of the tube body is used for adding reaction liquid for reaction.
As shown in fig. 5 and 7, the liquid cartridge 3 has a plug portion 33 inserted into the tube 1 to be fitted to the mouth of the tube 1, and a sealing portion 34 located outside the tube 1, the first opening 31 being located on the bottom surface of the plug portion 33, and the second opening 32 being located on the top surface of the sealing portion 34.
As shown in fig. 5 and 6, the reaction tube of the present application further includes an upper cap 4 for sealing the second opening 32, the upper cap 4 includes a sleeve portion 41 in sleeve fit with the sealing portion 34, and a sealing plate 42 disposed on the top surface of the sleeve portion 41 and capable of bending toward one side of the tube body 1, and the bottom surface of the sealing plate 42 is provided with a puncture needle 5 for puncturing the sealing membrane 42 when the sealing plate 42 bends toward one side of the tube body 1; a protective ring 43 surrounding the sealing plate 42 and protecting the sealing plate 42 is provided above the socket 41.
The sealing plate 42 is an arc shape protruding to the outside of the reaction tube, and a plurality of thinning grooves 44 for thinning the sealing plate 42 are provided at intervals on the outer side and/or inner side of the sealing plate 42. The reaction tube is generally made of plastic, and the sealing plate 42 of the plastic is provided with a plurality of thinning grooves 44, so that the sealing plate 42 is convenient to bend and protrudes outwards to be bent inwards, and a sealing film is punctured to release the second reaction liquid. All the thinning grooves 44 are arranged in concentric circles centering on the center of the sealing plate 42, so that the sealing plate 42 can be bent by pressing the middle region of the sealing plate 42. At least one side of the cross section of the thinning groove 44 close to the bottom surface is an arc groove, and compared with a design with a sharp angle, the arc groove is not easy to break in the bending process. One of the outermost reducing grooves 44 is provided on the inner side surface of the sealing plate 42, and the remaining reducing grooves 44 are provided on the outer side surface of the sealing plate 42. The thinning groove 44 is designed on the inner side surface or the outer side surface of the sealing plate 42 to achieve the purpose of being convenient to bend, when the sealing plate is bent, the joint of the outermost region of the sealing plate 42 and the sleeve joint part 41 is not easy to bend, and the middle region is bent inwards, so that the outermost thinning groove 44 is arranged on the inner side surface of the sealing plate 42, and the rest thinning grooves 44 are arranged on the outer side surface of the sealing plate 42, so that the thinning grooves 44 can be adapted to the bending condition of the sealing plate 42, and when the sealing plate is bent, the thinning grooves 44 are in a furled state, and the sealing plate 42 is not easy to break.
The central region of the inner side surface of the sealing plate 42 protrudes outwards to form a mounting groove 45 for fixedly mounting the puncture needle 5, one end of the puncture needle 5 is provided with a mounting part 51 which is in interference fit with the mounting groove 45, the puncture needle 5 is clamped into the mounting groove 45 through the mounting part 51 for fixing, and the other end of the puncture needle 5 is a needle tip for puncturing a sealing membrane. The outer periphery of the puncture needle 5 is provided with a guide groove 52 arranged along the axial direction, the guide groove 52 is designed such that the second reaction liquid can flow into the pipe body 1 along the guide groove 52 after the sealing film is punctured, as shown in fig. 8 to 10, and two opposite guide grooves 52 are provided on the outer periphery of the puncture needle 5.
A first annular groove 11 and a first convex ring 35 which are matched with each other are arranged between the inner side wall of the tube body 1 and the outer side wall of the plug part 33. A second annular groove 46 and a second convex ring 36 which are matched with each other are arranged between the inner side wall of the sleeve part 41 and the outer side wall of the sealing part 34. The design of the annular groove and the convex ring can enhance the sealing effect.
A stopper ring 37 is provided on the outer side wall of the liquid cartridge 3 between the stopper portion 33 and the sealing portion 34, and when assembled, the top surface of the tube body 1 abuts against the lower surface of the stopper ring 37, and the bottom surface of the sleeve portion 41 abuts against the upper surface of the stopper ring 37. The design of the baffle ring 37 can limit the matching between the liquid box 3 and the tube body 1 and the matching between the upper cover 4 and the liquid box 3.
The inner cavity of the liquid cartridge 3 is gradually reduced from the top to the bottom. The gradual reduction in this application can be the reduction of continuous change, also can be the reduction of sectional type, as long as there is the trend that reduces, then the second reaction liquid of being convenient for assembles the bottom of 3 inner chambers of liquid box, punctures the seal membrane after, makes things convenient for the second reaction liquid to flow to body 1 in.
The application also provides a reaction tube set for two-step nucleic acid amplification and detection, which comprises the reaction tube with the structure and an extrusion tool 6 for extruding the sealing plate 42 to bend the sealing plate 42 towards one side of the tube body 1, wherein the extrusion tool 6 comprises a finger sleeve 61 for a finger to pass through during operation and an extrusion head arranged on the periphery of the finger sleeve 61, as shown in figures 11-13; the extrusion head includes an extrusion portion 62 that, in use, extends into the protective ring 43 to extrude the seal plate 42, and a stop portion 63 that limits the depth of extension of the extrusion portion 62 into the protective ring 43 by abutting against the top surface of the protective ring 43. The design of the protection ring 43 in the upper cover 4 of the present application makes the sealing plate 42 less likely to be touched by a finger, thereby avoiding misoperation. When the second reaction solution needs to be discharged, the sealing plate 42 is pressed down by using the pressing tool 6, and the stroke of pressing down is limited by the stopper 63, thereby preventing overpressure.
When the device is used, the puncture needle 5 and the upper cover 4 are fixedly inserted, then the first reaction liquid for the first-step reaction is added into the tube body 1, the second reaction liquid which needs to be added into the reaction product of the first step during the second-step reaction is added into the liquid box 3, the first opening 31 of the liquid box 3 needs to be sealed by a sealing film before liquid is added, then the upper cover 4 and the liquid box 3 are assembled and sealed, and then the device is assembled with the tube body 1; when the first-step reaction is completed, the sealing plate 42 is pressed down by the pressing tool 6 to make the puncture needle 5 puncture the sealing film, the second reaction liquid flows down into the tube body 1 to be mixed with the product of the first-step reaction, and then the second-step reaction is performed. For the LAMP-CRISPR coupled two-step detection, the first step LAMP reaction comprises a reaction reagent and an object to be detected, the object to be detected is a nucleic acid sample serving as a template, and the second step CRISPR detection is used for detecting the result of the LAMP reaction. The reaction tube can be pre-installed in the liquid box 3 before leaving factory and selling, the first reaction liquid of the first step reaction can be pre-installed in the tube body 1 by other reagents except for being used as a detection object of the template, and then the reaction tube is conveyed and sold, and during detection, the cover body 2 only needs to be pulled out of the tube body 1, and then the template is added.
Claims (10)
1. A reaction tube for nucleic acid amplification and detection in a two-step method comprises a tube body for adding a first reaction solution and a cover body matched and sealed with the tube body, and is characterized in that the cover body comprises a liquid box for temporarily storing a second reaction solution in a first-step reaction, the bottom surface of the liquid box is provided with a first opening for allowing the second reaction solution to flow into the tube body, the top surface of the liquid box is provided with a second opening for adding the second reaction solution into the liquid box, and a sealing film is used for sealing the first opening before adding the second reaction solution;
the liquid box is provided with a plug part extending into the tube body and matched with the opening part of the tube body in an inserted manner, and a sealing part positioned outside the tube body, the first opening is positioned on the bottom surface of the plug part, and the second opening is positioned on the top surface of the sealing part;
the reaction tube also comprises an upper cover used for sealing the second opening, the upper cover comprises a sleeve joint part matched with the sealing part in a sleeve joint way, and a sealing plate which is arranged on the top surface of the sleeve joint part and can be bent towards one side of the tube body, and the bottom surface of the sealing plate is provided with a puncture needle used for puncturing the sealing membrane when the sealing plate is bent towards one side of the tube body; and a protection ring surrounding the sealing plate and used for protecting the sealing plate is also arranged above the sleeving part.
2. The reaction tube according to claim 1, wherein the sealing plate has an arc shape protruding to the outside of the reaction tube, and a plurality of thinning grooves for thinning the sealing plate are provided at intervals on the outer side and/or the inner side of the sealing plate.
3. The reactor tube as set forth in claim 2, wherein all the thinning grooves are arranged in concentric circles centered on the center of the sealing plate.
4. A reaction tube as set forth in claim 3, wherein the cross-section of said thinning groove is a circular arc groove at least on the side near the bottom surface; one thinning groove at the outermost ring is arranged on the inner side surface of the sealing plate, and the other thinning grooves are arranged on the outer side surface of the sealing plate.
5. The reaction tube of claim 2, wherein the sealing plate has an inner surface with a central region protruding outward to form a mounting groove for fixedly mounting the lancet, and a mounting portion is provided at one end of the lancet and is in interference fit with the mounting groove.
6. The reaction tube of claim 1, wherein the puncture needle is provided at its outer circumference with guide grooves arranged in an axial direction.
7. The reaction tube of claim 1, wherein a first annular groove and a first convex ring which are matched with each other are formed between the inner side wall of the tube body and the outer side wall of the plug portion; and a second annular groove and a second convex ring which are matched with each other are arranged between the inner side wall of the sleeving part and the outer side wall of the sealing part.
8. The reaction tube according to claim 1, wherein a stopper ring is provided on an outer side wall of the liquid cartridge between the stopper portion and the sealing portion, and when the cartridge is assembled, a top surface of the tube body abuts against a lower surface of the stopper ring, and a bottom surface of the sleeve portion abuts against an upper surface of the stopper ring.
9. The reaction tube of claim 1, wherein the inner cavity of the liquid cartridge is tapered from top to bottom.
10. A reaction tube set for two-step nucleic acid amplification and detection, which is characterized by comprising the reaction tube according to any one of claims 1 to 9, and an extrusion tool for extruding the sealing plate to bend the sealing plate towards one side of the tube body, wherein the extrusion tool comprises a finger sleeve for a finger to pass through during operation, and an extrusion head arranged on the periphery of the finger sleeve; the extrusion head comprises an extrusion part and a limiting part, wherein the extrusion part extends into the protective ring to extrude the sealing plate when in use, and the limiting part limits the depth of the extrusion part extending into the protective ring by abutting against the top surface of the protective ring.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202021464690.3U CN212955133U (en) | 2020-07-22 | 2020-07-22 | Reaction tube and reaction tube set for two-step nucleic acid amplification and detection |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202021464690.3U CN212955133U (en) | 2020-07-22 | 2020-07-22 | Reaction tube and reaction tube set for two-step nucleic acid amplification and detection |
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| CN212955133U true CN212955133U (en) | 2021-04-13 |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113234591A (en) * | 2021-05-28 | 2021-08-10 | 宁波康程德诺生物医药有限公司 | Integrated nucleic acid rapid-extraction test tube, rapid-extraction detection device and method |
| CN113980797A (en) * | 2021-10-29 | 2022-01-28 | 广东粤港澳大湾区国家纳米科技创新研究院 | Detection device and detection method for nucleic acid amplification product |
| WO2023138148A1 (en) * | 2022-01-19 | 2023-07-27 | 江苏为真生物医药技术股份有限公司 | Sealing device for rapid detection, use method thereof, and application thereof |
-
2020
- 2020-07-22 CN CN202021464690.3U patent/CN212955133U/en active Active
Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113234591A (en) * | 2021-05-28 | 2021-08-10 | 宁波康程德诺生物医药有限公司 | Integrated nucleic acid rapid-extraction test tube, rapid-extraction detection device and method |
| CN113234591B (en) * | 2021-05-28 | 2024-03-22 | 宁波康程德诺生物医药有限公司 | Integrated nucleic acid quick-lifting test tube, quick-lifting detection device and method |
| CN113980797A (en) * | 2021-10-29 | 2022-01-28 | 广东粤港澳大湾区国家纳米科技创新研究院 | Detection device and detection method for nucleic acid amplification product |
| CN113980797B (en) * | 2021-10-29 | 2024-05-24 | 武汉纳达康生物科技有限公司 | Detection device and detection method for nucleic acid amplification product |
| WO2023138148A1 (en) * | 2022-01-19 | 2023-07-27 | 江苏为真生物医药技术股份有限公司 | Sealing device for rapid detection, use method thereof, and application thereof |
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