Nucleic acid extraction device and extraction method

Abstract

The present invention discloses a nucleic acid extraction device and an extraction method, which relate to the technical field of nucleic acid extraction devices. The device comprises a nucleic acid storage table, wherein an isolation cover is fixedly connected to the middle part of the upper surface of the nucleic acid storage table, a preheating dispersion cylinder is arranged on the circumference of the upper surface, a multifunctional sampling tube is arranged in the preheating dispersion cylinder, a nucleic acid detection rod is arranged on the upper end of the multifunctional sampling tube, and further comprises a uniform dispersion preheating adjustment mechanism and an auxiliary sampling mechanism. The uniform dispersion preheating adjustment mechanism is arranged to not only realize the control of the internal shaking of the specimen, but also enable the turntable to push the preheating dispersion cylinder to slide back and forth in the isolation cover through the arc surface arrangement of the inclined convex corner block, and the frequency of the reciprocating sliding of the preheating dispersion cylinder in the isolation cover can be directly controlled by controlling the rotation speed of the motor, which can not only reduce the damage of the sample caused by excessive physical force generated during the shaking process, especially for those fragile cells or tissue structures, but also increase the integrity and extraction efficiency of the nucleic acid extraction.

Description

Technical Field

[0001] The present invention relates to the technical field of nucleic acid extraction devices, and in particular to a nucleic acid extraction device and an extraction method. Background Art

[0002] Nucleic acid testing of pigs is an efficient and accurate detection method that plays an important role in the diagnosis, monitoring and prevention of pig diseases. Nucleic acid testing can be used to diagnose a variety of pig diseases such as swine fever, African swine fever, pseudorabies, etc. By detecting the DNA or RNA of pathogens in pigs, it can be accurately determined whether the pigs are infected with related viruses. During the peak period of the epidemic or in the area where the epidemic is spreading, by regularly collecting pig samples for nucleic acid testing, the epidemic dynamics can be discovered in a timely manner, providing important data support for epidemic prevention and control.

[0003] However, when the existing pig nucleic acid is tested, when the sampled nucleic acid test stick is stored, the nucleic acid test stick needs to be placed in a multifunctional sampling tube, the handle needs to be broken, and it needs to be clearly marked. When the operator breaks the handle, he must ensure that the position of the sample tube nucleic acid test stick in the container multifunctional sampling tube is neither offset nor unstable. This is because any slight shaking may affect the smooth breaking of the handle and may even damage the sample tube, resulting in sample contamination or failure to successfully break the handle. However, the existing collection tube design has not fully taken this into consideration. The existing collection tube has a relatively simple design and lacks sufficient stability and support, so that the sample tube nucleic acid test stick is easily shaken during the process of breaking the handle. This shaking not only increases the difficulty of breaking the handle, but also greatly increases the risk of sample contamination.

[0004] Existing pig nucleic acid specimens need to be manually shaken by operators after collection. During the shaking process, the cell walls or tissue structures in the sample will be broken by physical force, making the nucleic acid in the cells easier to extract. Moderate shaking can also help remove impurities in the sample and improve the purity of the nucleic acid. However, if the physical force generated during the shaking process is too large, it may cause damage to the sample, especially for those fragile cells or tissue structures, which will affect the integrity and extraction efficiency of the nucleic acid. If a separate instrument is used for shaking, additional operating steps and time are required, which may increase the workload of the experimenter. Especially when processing a large number of samples, this cumbersome operating process may reduce work efficiency.

[0005] Therefore, a nucleic acid extraction device and an extraction method are proposed to solve the above problems. Summary of the Invention

[0006] In view of this, the technical problem to be solved by the present invention is to propose a nucleic acid extraction device and extraction method to solve the problems of easy contamination during collection of pig nucleic acid samples and low detection efficiency caused by the single mixing and shaking after sample collection in the existing technology.

[0007] To achieve the above objectives, the present invention provides the following technical solutions: a nucleic acid extraction device and extraction method, comprising a nucleic acid storage table, an isolation cover fixedly connected to the middle portion of the upper surface of the nucleic acid storage table, a preheating dispersion cylinder provided around the circumference of the upper surface, a multifunctional sampling tube provided in the preheating dispersion cylinder, a nucleic acid detection stick provided at the upper end of the multifunctional sampling tube, and a uniform dispersion preheating adjustment mechanism and an auxiliary sampling mechanism, the uniform dispersion preheating adjustment mechanism being provided in the middle portion of the isolation cover, and the auxiliary sampling mechanism being provided in the multifunctional sampling tube;

[0008] The uniform dispersion preheating regulating mechanism is used for uniform mixing and heating of nucleic acid with reagents after extraction;

[0009] The auxiliary sampling mechanism is used to assist in the extraction of nucleic acids.

[0010] Preferably, the uniformly dispersed preheating adjustment mechanism includes a protective cover, the bottom of the protective cover is fixedly connected to the upper surface of the isolation cover, a motor is installed in the middle of the upper surface of the protective cover, and the motor drive shaft passes through the middle of the protective cover, and a rotating drum is rotatably connected to the middle of the upper end of the isolation cover, and the middle of the upper surface of the rotating drum is fixedly connected to the motor drive shaft.

[0011] Preferably, an eccentric hole is provided at an eccentric position of one end of the rotating drum away from the motor, a conical sliding column is slidably connected in the eccentric hole, and a turntable is fixedly connected to the end of the conical sliding column away from the eccentric hole, the turntable is arranged on the inner wall, and an inclined convex corner block is provided on the side of the turntable away from the eccentric hole, and the inclined convex corner block is fixedly connected to the bottom of the isolation cover on the side away from the turntable.

[0012] Preferably, the preheating dispersion cylinder is evenly arranged on the upper surface of the turntable, and a buffer spring is provided on the outer surface of the preheating dispersion cylinder near the inner wall of the isolation cover, one end of the buffer spring is fixedly connected to the preheating dispersion cylinder, and the other end of the buffer spring is fixedly connected to the inner wall of the isolation cover. A copper power ring is provided on the outer surface of the preheating dispersion cylinder near the top of the isolation cover, a shock-absorbing block is slidably connected to the inner wall of the preheating dispersion cylinder, and a first reset spring is fixedly connected to the bottom of the preheating dispersion cylinder, and one end of the first reset spring away from the shock-absorbing block is fixedly connected to the bottom of the preheating dispersion cylinder.

[0013] Preferably, the upper end of the preheating and dispersion cylinder is also provided with an anti-falling mechanism, which is used to prevent the multi-functional sampling tube from detaching. The anti-falling mechanism includes a multi-functional clamping and prying plate, and the middle part of the multi-functional clamping and prying plate is rotatably connected to a support plate, and the support plate is rotatably connected to the outer surface of the upper end of the preheating and dispersion cylinder at one end away from the multi-functional clamping and prying plate. One end of the multi-functional clamping and prying plate is fixedly connected to a pressing plate, and the other end of the multi-functional clamping and prying plate is fixedly connected to a splint. A card slot is provided at the upper end of the preheating and dispersion cylinder, and a second return spring is evenly and fixedly connected in the card slot, and the second return spring is fixedly connected to the splint of the multi-functional clamping and prying plate at one end away from the card slot.

[0014] Preferably, the auxiliary sampling mechanism includes a retaining ring, which is fixedly connected to the inner wall of the upper end of the multifunctional sampling tube. A limiting hole is provided on the retaining ring. A sealing cover is rotatably connected to the outer surface of the upper end of the multifunctional sampling tube. The sealing cover is fixedly connected to an anti-slip limiting plate on the side away from the connection with the multifunctional sampling tube.

[0015] Preferably, the nucleic acid detection stick is arranged below the retaining ring near the nucleic acid extraction end, an auxiliary groove is provided on the nucleic acid detection stick, and the nucleic acid detection stick is arranged at the upper end of the multifunctional sampling tube.

[0016] The nucleic acid extraction device and the extraction method include the following steps:

[0017] Step 1: Sampling of clinical specimens: Use a nucleic acid detection stick to scrape the inside of the pig's mouth, cheeks, and nasal cavity. If necessary, such as anal swabs or vaginal swabs, they must also be collected according to the corresponding methods. Then, place the nucleic acid detection stick in a multi-functional sampling tube, break off the handle, and mark it clearly;

[0018] Step 2: Extraction of nucleic acid: Extract viral RNA or DNA from the sample. RNA can be reverse transcribed (converting RNA into cDNA) to obtain a certain number of DNA or cDNA molecules;

[0019] Step 3: Multiplex quantitative PCR detection and nanopore sequencing library construction. After 1.5 hours of multiplex quantitative PCR detection, a graphical curve display can be obtained on the terminal. After 10 minutes of nanopore sequencing library construction, the ends of these DNA or cDNA molecules are repaired and connected to sequencing adapters to complete the library construction. Then, they are placed in the sequencing chip. The process of molecules entering and passing through the nanopore is monitored in real time by an electrical signal feedback system, and the sequence information of each molecule is recorded.

[0020] Step 4, MinION sequencing: Basic preprocessing is performed on the data obtained through sequencing, such as filtering low-quality data, removing the sequencing adapter sequences, splicing and assembling the DNA or cDNA sequences, etc.; finally, the data results are compared with the known gene library, and the viral infection status and abnormal genes are identified through Centrifuge software analysis (R language analysis).

[0021] Compared with the prior art, the present invention provides a nucleic acid extraction device and method, which have the following beneficial effects:

[0022] 1. Through the setting of the retaining ring and the anti-slip limiting plate in the present invention, the nucleic acid detection end of the nucleic acid detection stick is placed at the bottom of the retaining ring and in the limiting hole, and the limiting action of the limiting hole and the anti-slip limiting plate on the detection rod of the nucleic acid detection stick makes the detection rod stably tilted at the upper end of the multi-functional sampling tube. At this time, when the sealing cover is closed, the auxiliary groove of the nucleic acid detection stick is pressed and engaged by the anti-slip limiting plate. At this time, the operator can easily break the handle of the nucleic acid detection stick. Compared with the prior art, through the setting of the present invention, the operator only needs to tilt the nucleic acid detection stick and the limiting hole and then press the sealing cover to easily break the handle. This can not only avoid the sample falling and contamination phenomenon caused by the sliding of the nucleic acid detection stick when breaking in the prior art, but also reduce the operator's control over the breaking of the nucleic acid detection stick, enhance the operator's stability and anti-falling when breaking the handle of the nucleic acid detection stick, and thus reduce the contamination phenomenon during sample collection.

[0023] 2. The uniform dispersion preheating adjustment mechanism can not only control the shaking inside the specimen, but also enable the turntable to push the preheating dispersion cylinder to slide back and forth in the isolation cover through the arc surface setting of the inclined convex corner block. By controlling the rotation speed of the motor, the frequency of the reciprocating sliding of the preheating dispersion cylinder in the isolation cover can be directly controlled. This can not only reduce the damage to the sample caused by excessive physical force generated during the shaking process, especially for those fragile cells or tissue structures, but also increase the integrity and efficiency of nucleic acid extraction.

[0024] Moreover, by placing the multifunctional sampling tube in a preheated dispersion cylinder for storage, not only can the contamination from external pollution be reduced, but the rotation of the turntable can also drive multiple collected samples to be properly shaken, further ensuring that the cells or tissues in the sample can be effectively lysed, thereby releasing nucleic acids. In addition, the multifunctional sampling tube surface can be stably clamped by the multifunctional clamping pry plate, so that the multifunctional sampling tube can be stably stored in the preheated dispersion cylinder during transportation, thereby reducing the occurrence of problems such as falling and loss of collected samples.

[0025] 3. According to the invention, a copper electrified ring is sleeved on the outer surface of the preheating dispersion cylinder. When the preheating dispersion cylinder slides back and forth on the isolation cover, in order to denature the protein in the sample, thereby destroying the structure of the cell wall and cell membrane, the nucleic acid in the cell is more easily released, and a moderate heating method is adopted for nucleic acid extraction. By passing current into multiple copper electrified rings connected in series, more accurate and efficient heating is achieved. In addition, a temperature sensor is installed near the multifunctional sampling tube, which can monitor and control the internal temperature in real time, thereby ensuring that the heating process is both fast and stable. Compared with traditional heating methods, the present invention greatly shortens the heating time, and the temperature control is more precise, reducing the risk of nucleic acid degradation caused by temperature fluctuations. More importantly, through the design of the present invention, we can not only achieve batch, controllable temperature heating treatment, but also shake the specimen during the heating process. This shaking is conducive to uniform heating inside the sample, so that the nucleic acid can be released more quickly, thereby further improving the efficiency of nucleic acid extraction. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] Figure 1 It is a schematic diagram of the three-dimensional structure of the present invention;

[0027] Figure 2 It is a half-cut auxiliary schematic diagram of the three-dimensional structure of the present invention;

[0028] Figure 3 This is a half-cut schematic diagram of the connection relationship of the partial structure of the uniformly dispersed preheating adjustment mechanism of the present invention;

[0029] Figure 4 This is a schematic diagram of the structural connection relationship of the anti-falling mechanism of the present invention;

[0030] Figure 5 For the present invention Figure 4 Enlarged view of point A in the middle;

[0031] Figure 6 This is a schematic diagram of the structural connection relationship of the auxiliary sampling mechanism of the present invention;

[0032] Figure 7 For the present invention Figure 6 Enlarged view of point B in the middle.

[0033] In the picture:

[0034] 1. Nucleic acid storage table; 11. Isolation cover; 12. Preheating dispersion cylinder; 13. Multifunctional sampling tube; 14. Nucleic acid detection stick;

[0035] 2. Uniformly dispersed preheating adjustment mechanism; 21. Protective cover; 22. Rotating drum; 23. Eccentric hole; 24. Rotating disk; 25. Inclined convex corner block; 26. Buffer spring; 27. Shock absorber block; 28. First return spring; 29. ​​Copper power ring;

[0036] 3. Anti-fall mechanism; 31. Multifunctional clamping and prying plate; 32. Support plate; 33. Second return spring;

[0037] 4. Auxiliary sampling mechanism; 41. Retaining ring; 42. Limiting hole; 43. Sealing cover; 44. Anti-slip limiting plate. DETAILED DESCRIPTION

[0038] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of the present invention.

[0039] The present invention will be described in further detail below with reference to the accompanying drawings and examples.

[0040] First embodiment

[0041] Please refer to Figures 1 to 7 As shown:

[0042] To solve the problems mentioned in the technical solution, the embodiments of the present application provide a nucleic acid extraction device and an extraction method, comprising a nucleic acid storage table 1, an isolation cover 11 fixedly connected to the middle of the upper surface of the nucleic acid storage table 1, a preheating dispersion cylinder 12 provided on the circumference of the upper surface, a multifunctional sampling tube 13 provided in the preheating dispersion cylinder 12, a nucleic acid detection stick 14 provided at the upper end of the multifunctional sampling tube 13, and a uniform dispersion preheating adjustment mechanism 2 and an auxiliary sampling mechanism 4, wherein the uniform dispersion preheating adjustment mechanism 2 is provided in the middle of the isolation cover 11, and the auxiliary sampling mechanism 4 is provided in the multifunctional sampling tube 13;

[0043] The uniform dispersion preheating regulating mechanism 2 is used for uniform mixing and heating of nucleic acid with reagents after extraction;

[0044] The auxiliary sampling mechanism 4 is used to assist in the extraction of nucleic acids;

[0045] The uniformly dispersed preheating adjustment mechanism 2 includes a protective cover 21, the bottom of which is fixedly connected to the upper surface of the isolation cover 11, a motor is installed in the middle of the upper surface of the protective cover 21, and the motor drive shaft passes through the middle of the protective cover 21, and a rotating drum 22 is rotatably connected to the middle of the upper end of the isolation cover 11, and the middle of the upper surface of the rotating drum 22 is fixedly connected to the motor drive shaft;

[0046] An eccentric hole 23 is formed at an eccentric position of the end of the rotating drum 22 away from the motor. A conical sliding post is slidably connected to the eccentric hole 23, and a turntable 24 is fixedly connected to the end of the conical sliding post away from the eccentric hole 23. The turntable 24 is arranged on the inner wall. An inclined convex block 25 is provided on the side of the turntable 24 away from the eccentric hole 23. The inclined convex block 25 is fixedly connected to the bottom of the isolation cover 11 on the side away from the turntable 24.

[0047] The preheating dispersion cylinder 12 is evenly arranged on the upper surface of the turntable 24. A buffer spring 26 is provided on the outer surface of the preheating dispersion cylinder 12 near the inner wall of the isolation cover 11. One end of the buffer spring 26 is fixedly connected to the preheating dispersion cylinder 12, and the other end of the buffer spring 26 is fixedly connected to the inner wall of the isolation cover 11. A copper power ring 29 is provided on the outer surface of the preheating dispersion cylinder 12 near the top of the isolation cover 11. A shock-absorbing block 27 is slidably connected to the inner wall of the preheating dispersion cylinder 12. A first return spring 28 is fixedly connected to the bottom of the shock-absorbing block 27. One end of the first return spring 28 away from the shock-absorbing block 27 is fixedly connected to the bottom of the preheating dispersion cylinder 12.

[0048] Among them, the copper power rings 29 arranged on the circumference of the upper surface of the isolation cover 11 are fixedly connected in series to the non-metallic baffle, and a temperature sensor device is arranged on the circumference of the baffle to detect the internal temperature of the preheating dispersion cylinder 12.

[0049] Compared with the prior art, through the implementation of this embodiment, the present invention greatly shortens the heating time, and the temperature control is more precise, reducing the risk of nucleic acid degradation caused by temperature fluctuations. More importantly, through the design of the present invention, we can not only achieve batch, temperature-controlled heating treatment, but also shake the specimen during the heating process. This shaking is conducive to uniform heating inside the sample, allowing the nucleic acid to be released more quickly, thereby further improving the efficiency of nucleic acid extraction.

[0050] For further examples, please refer to Figures 1 to 7 As shown:

[0051] The upper end of the preheating and dispersing cylinder 12 is also provided with an anti-falling mechanism 3, which is used to prevent the multifunctional sampling tube 13 from falling off. The anti-falling mechanism 3 includes a multifunctional clamping and prying plate 31, and the middle part of the multifunctional clamping and prying plate 31 is rotatably connected to a support plate 32. The support plate 32 is rotatably connected to the outer surface of the upper end of the preheating and dispersing cylinder 12 at one end away from the multifunctional clamping and prying plate 31. A pressing plate is fixedly connected to one end of the multifunctional clamping and prying plate 31, and the other end of the multifunctional clamping and prying plate 31 is fixedly connected to a splint. A card slot is opened at the upper end of the preheating and dispersing cylinder 12, and a second return spring 33 is evenly and fixedly connected in the card slot. The end of the second return spring 33 away from the card slot is fixedly connected to the splint of the multifunctional clamping and prying plate 31;

[0052] The auxiliary sampling mechanism 4 includes a retaining ring 41, which is fixedly connected to the inner wall of the upper end of the multifunctional sampling tube 13. A limiting hole 42 is provided on the retaining ring 41. A sealing cover 43 is rotatably connected to the outer surface of the upper end of the multifunctional sampling tube 13. A non-slip limiting plate 44 is fixedly connected to the side of the sealing cover 43 away from the connection with the multifunctional sampling tube 13.

[0053] The nucleic acid detection rod 14 is arranged below the retaining ring 41 near the nucleic acid extraction end. An auxiliary groove is provided on the nucleic acid detection rod 14. The nucleic acid detection rod 14 is arranged at the upper end of the multifunctional sampling tube 13.

[0054] Among them: the rotating connection between the sealing cover 43 and the multifunctional sampling tube 13 is adapted to the size of the card slot opened at the upper end of the preheating dispersion cylinder 12, and the two sides of the bottom of the retaining ring 41 are fixedly connected with limiting blocks to prevent the nucleic acid detection plate 14 from sliding when the nucleic acid is extracted.

[0055] Compared with the prior art, through the implementation of this embodiment, the operator only needs to tilt and align the nucleic acid detection stick 14 with the limiting hole 42, and then press the sealing cover 43 to easily break the handle. This not only avoids the phenomenon of sample falling and contamination caused by the sliding of the nucleic acid detection stick 14 when breaking in the prior art, but also reduces the operator's control over the breaking of the nucleic acid detection stick 14, enhances the operator's stability and anti-falling when breaking the handle of the nucleic acid detection stick 14, and thus reduces the contamination during sample collection.

[0056] Second embodiment

[0057] The nucleic acid extraction device and the extraction method include the following steps:

[0058] Step 1: Sampling of clinical specimens: Use the nucleic acid detection stick 14 to scrape the inside of the pig's mouth and cheeks, and scrape the inside of the nasal cavity. If necessary, such as anal swabs or vaginal swabs, they must also be collected according to the corresponding methods. Then, place the nucleic acid detection stick 14 into the multifunctional sampling tube 13, break off the handle, and mark it clearly;

[0059] Step 2: Extraction of nucleic acid: Extract viral RNA or DNA from the sample. RNA can be reverse transcribed to convert RNA into cDNA to obtain a certain number of DNA or cDNA molecules.

[0060] Step 3: Multiplex quantitative PCR detection and nanopore sequencing library construction. After 1.5 hours of multiplex quantitative PCR detection, a graphical curve display can be obtained on the terminal. After 10 minutes of nanopore sequencing library construction, the ends of these DNA or cDNA molecules are repaired and connected to sequencing adapters to complete the library construction. Then, they are placed in the sequencing chip. The process of molecules entering and passing through the nanopore is monitored in real time by an electrical signal feedback system, and the sequence information of each molecule is recorded.

[0061] Step 4, MinION sequencing: Basic pre-processing of the sequencing data, such as filtering low-quality data, removing sequencing adapter sequences, and splicing and assembling DNA or cDNA sequences, is performed. Finally, the data is compared with a known gene library, and Centrifuge software is used to analyze and identify viral infections and abnormal genes using R language analysis.

[0062] Compared with traditional sequencing technologies, the nanopore sequencing technology in this method has the following characteristics:

[0063] (1) Longer sequencing read length: Nanopore sequencing technology uses the change of electrical signals when bases pass through the nanopore to achieve sequencing. In principle, it can detect all nucleic acid sequences passing through the nanopore. Therefore, there is no limit on the sequencing length. Currently, the longest read length can exceed 2M;

[0064] (2) Real-time sequencing: The sequencing library preparation process is simple, and there is no need to perform PCR amplification on the nucleic acid in the sample. DNA or RNA can be sequenced directly, saving operation time and reducing sequencing costs. The results can be output while sequencing, enabling real-time analysis of sequencing data;

[0065] (3) Direct RNA sequencing: Nanopore sequencing can directly sequence all forms of RNA, avoiding the bias and possible mutations that are caused by reverse transcription of RNA into DNA amplification when sequencing and studying RNA viruses. This feature is extremely attractive in the field of RNA virus detection.

[0066] Everything in the above example works as follows:

[0067] In the initial state: the motor is not started, the multifunctional sampling tube 13 is not placed in the preheating dispersion cylinder 12, and the copper power ring 29 is not energized.

[0068] The following is the working process of the auxiliary sampling mechanism 4 for assisting in the extraction of nucleic acids:

[0069] When collecting nucleic acid samples from imported pigs, a special nucleic acid detection stick 14 and a multifunctional sampling tube 13 are used. The sampling personnel need to wear protective clothing and gloves to ensure aseptic operation. At this time, the operator uses the collection end of the nucleic acid detection stick 14 to scrape the cheeks inside the pig's mouth and scrape inside the nasal cavity. According to needs, such as anal swabs or vaginal swabs, they also need to be collected according to the corresponding methods. Then, the collection end of the nucleic acid detection stick 14 is placed in the inner wall of the multifunctional sampling tube 13 below the retaining ring 41 and the handle is broken off. Figure 7As shown, by placing the nucleic acid detection end of the nucleic acid detection stick 14 at the bottom of the retaining ring 41 and in the limiting hole 42, the limiting hole 42 and the anti-slip limiting plate 44 limit the detection rod of the nucleic acid detection stick 14, so that the detection rod can be stably tilted at the upper end of the multi-functional sampling tube 13. At this time, when the sealing cover 43 is closed, the anti-slip limiting plate 44 presses and engages the auxiliary groove of the nucleic acid detection stick 14. At this time, the operator can easily break the handle of the nucleic acid detection stick 14. At this time, the broken handles are collected uniformly, and the specimen collection end located in the multi-functional sampling tube 13 is removed by the nucleic acid detection stick 14. After breaking, the specimen begins to fall into the bottom of the multifunctional sampling tube 13 to be mixed with the reagent. Compared with the prior art, the present invention only requires the operator to tilt and align the nucleic acid detection stick 14 with the limiting hole 42, and then press the sealing cover 43 to easily break the handle. This not only avoids the phenomenon of specimen falling and contamination caused by the sliding of the nucleic acid detection stick 14 when breaking in the prior art, but also reduces the operator's control over the breaking of the nucleic acid detection stick 14, enhances the operator's stability and anti-falling when breaking the handle of the nucleic acid detection stick 14, and thus reduces the contamination during specimen collection.

[0070] Please refer to the above working process Figures 1 to 7 .

[0071] The following is the working process of the uniform dispersion preheating adjustment mechanism 2 for uniform mixing and heating of nucleic acid with reagents after extraction:

[0072] As described above, after the specimens are uniformly collected, the operator uniformly puts the multifunctional sampling tube 13 into the preheated dispersion cylinder 12. At this time, by gently pressing the multifunctional sampling tube 13, as shown in FIG. Figure 4 Figure 5As shown, at this time, by pressing the nucleic acid detection stick 14, the operator places the multifunctional sampling tube 13 and the card slot opened in the preheating dispersion cylinder 12 on the same motion trajectory. When the multifunctional sampling tube 13 slides into the card slot opened in the preheating dispersion cylinder 12, since one end of the multifunctional clamping pry plate 31 is fixedly connected to the pressing plate, the other end of the multifunctional clamping pry plate 31 is fixedly connected to the splint, and the second return spring 33 is evenly fixedly connected in the card slot, and the second return spring 33 is fixedly connected to the splint of the multifunctional clamping pry plate 31 away from one end of the card slot. When the connection of the sealing cover 43 at the upper end of the multifunctional sampling tube 13 is gradually squeezed to the splint, the squeezing of the splint will drive the second return spring 33 to be compressed, and at this time the bottom of the multifunctional sampling tube 13 In contact with the upper surface of the shock-absorbing block 27, the multifunctional sampling tube 13 can be stably stored in the preheating dispersion cylinder 12 under the elastic action of the second return spring 33 in the clamping plate; by placing the multifunctional sampling tube 13 in the preheating dispersion cylinder 12 for storage, not only can the contamination from external pollution be reduced, but the rotation of the turntable 24 can also drive multiple collected samples to be properly shaken, further ensuring that the cells or tissues in the sample can be effectively lysed, thereby releasing nucleic acids, and the multifunctional clamping plate 31 can stably clamp the surface of the multifunctional sampling tube 13, so that the multifunctional sampling tube 13 can be stably stored in the preheating dispersion cylinder 12 during the transportation process, thereby reducing the problem of the collected samples falling or being lost.

[0073] At this time, in order to speed up the decomposition of nucleic acid, the operator can start the switch button and temperature control switch for controlling the start of the motor on the nucleic acid storage table 1, and set the optimal nucleic acid extraction temperature according to different specimens. At this time, the motor installed on the upper surface of the protective cover 21 starts to rotate under the electrical control of the switch. Because the motor drive shaft passes through the protective cover 21 and is fixedly connected to the middle part of the upper end of the drum 22, the drum 22 will start to rotate in the same direction when the motor rotates. An eccentric hole 23 is provided at the eccentric part of the bottom of the drum 22, and a conical sliding column fixedly connected to the middle part of the turntable 24 is slidably connected in the eccentric hole 23. Moreover, the inclined convex corner block 25 is fixedly connected to the bottom of the isolation cover 11, and the middle part of the turntable 24 is set on the inclined convex corner block 25. Since the inclined convex corner block 25 adopts an inclined conical setting, At this time, the rotation of the rotating cylinder 22 will drive the conical sliding column to drive the turntable 24 to rotate with different slopes on the inclined convex corner block 25. Furthermore, because the preheating dispersion cylinder 12 slides circumferentially in the isolation cover 11, and the bottom of the preheating dispersion cylinder 12 is arranged on the upper surface of the turntable 24, the preheating dispersion cylinder 12 will be driven to slide in the isolation cover 11 by the rotation of the turntable 24 with different amplitudes. At this time, the multifunctional sampling tube 13 engaged in the preheating dispersion cylinder 12 begins to shake the specimen inside the multifunctional sampling tube 13 under the movement of the preheating dispersion cylinder 12. Shaking the inside of the multifunctional sampling tube 13 by the preheating dispersion cylinder 12 can not only reduce the damage to the sample caused by excessive physical force generated during the shaking process, especially for those fragile cells or tissue structures, but also increase the integrity and efficiency of nucleic acid extraction.

[0074] And at this time, with the start of the stable control switch and the threshold control of the temperature sensor device, the copper electrified ring 29 can quickly heat the preheating dispersion tube 12, and the temperature sensor control can control the internal temperature of the preheating dispersion tube 12, and then the heat transfer inside the preheating dispersion tube 12 can gradually heat the inside of the multi-functional sampling tube 13. At this time, the preheating dispersion tube 12 slides in the isolation cover 11, and the copper electrified ring 29 heats it. At the same time, the preheating dispersion tube 12 slides back and forth in the copper electrified ring 29, so that the inside of the preheating dispersion tube 12 can be evenly heated. The copper electrified ring 29 is sleeved on the outer surface of the preheating dispersion tube 12. When the preheating dispersion tube 12 slides back and forth on the isolation cover 11, in order to denature the protein in the sample, the structure of the cell wall and cell membrane is destroyed, making the nucleic acid in the cell more easily released. Released, thereby adopting a moderate heating method for nucleic acid extraction, by passing current into multiple copper power rings 29 connected in series, more accurate and efficient heating is achieved, and a temperature sensor is installed near the multifunctional sampling tube 13, which can monitor and control the internal temperature in real time, thereby ensuring that the heating process is both fast and stable. Compared with traditional heating methods, the design of the present invention not only greatly shortens the heating time, but also makes the temperature control more precise, reducing the risk of nucleic acid degradation caused by temperature fluctuations, and more importantly, through the design of the present invention, we can not only achieve batch, controllable temperature heating treatment, but also make the specimen shake during the heating process, and this shaking is conducive to uniform heating inside the sample, so that the nucleic acid can be released more quickly, thereby further improving the efficiency of nucleic acid extraction.

[0075] Please refer to the above working process Figures 1 to 7 .

[0076] It should be noted that, in this document, relational terms such as first and second, etc., are used only to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply the existence of any such actual relationship or order between these entities or operations. Moreover, the terms "comprises," "comprising," or any other variants thereof are intended to cover non-exclusive inclusion, so that a process, method, article, or device comprising a series of elements includes not only those elements, but also other elements not explicitly listed, or elements inherent to such process, method, article, or device. In the absence of further limitations, an element defined by the phrase "comprising a ..." does not exclude the presence of other identical elements in the process, method, article, or device comprising the element.

[0077] While embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that various changes, modifications, substitutions, and variations may be made to these embodiments without departing from the principles and spirit of the invention, and that the scope of the invention is defined by the appended claims and their equivalents.

Claims

1. A nucleic acid extraction device for collecting and accessing nucleic acid, comprising a nucleic acid storage table (1), wherein an isolation cover (11) is fixedly connected to the middle of the upper surface of the nucleic acid storage table (1), a preheating dispersion cylinder (12) is arranged on the circumference of the upper surface, a multifunctional sampling tube (13) is arranged in the preheating dispersion cylinder (12), and a nucleic acid detection stick (14) is arranged on the upper end of the multifunctional sampling tube (13), characterized in that: It also includes a uniform dispersion preheating adjustment mechanism (2) and an auxiliary sampling mechanism (4), wherein the uniform dispersion preheating adjustment mechanism (2) is arranged in the middle of the isolation cover (11), and the auxiliary sampling mechanism (4) is arranged in the multifunctional sampling tube (13); The uniform dispersion preheating regulating mechanism (2) is used for uniform mixing and heating of nucleic acid with reagents after extraction; the uniform dispersion preheating regulating mechanism (2) comprises a protective cover (21), the bottom of the protective cover (21) is fixedly connected to the upper surface of the isolation cover (11), a motor is installed in the middle of the upper surface of the protective cover (21), and the motor drive shaft passes through the middle of the protective cover (21), the middle of the upper end of the isolation cover (11) is rotatably connected to a rotating drum (22), and the middle of the upper surface of the rotating drum (22) is fixedly connected to the upper surface of the isolation cover (11). Connected to the motor drive shaft; an eccentric hole (23) is provided at an eccentric position of one end of the rotating drum (22) away from the motor, a conical sliding column is slidably connected in the eccentric hole (23), and a turntable (24) is fixedly connected to one end of the conical sliding column away from the eccentric hole (23), the turntable (24) is arranged on the inner wall, and an inclined convex block (25) is provided on one side of the turntable (24) away from the eccentric hole (23), and the inclined convex block (25) is fixedly connected to the bottom of the isolation cover (11) on one side away from the turntable (24); The auxiliary sampling mechanism (4) is used to assist in the extraction of nucleic acids.

2. A nucleic acid extraction device according to claim 1, characterized in that: The preheating dispersion cylinder (12) is evenly arranged on the upper surface of the turntable (24); a buffer spring (26) is sleeved on the outer surface of the preheating dispersion cylinder (12) near the inner wall of the isolation cover (11); one end of the buffer spring (26) is fixedly connected to the preheating dispersion cylinder (12); the other end of the buffer spring (26) is fixedly connected to the inner wall of the isolation cover (11); a copper power ring (29) is sleeved on the outer surface of the preheating dispersion cylinder (12) near the top of the isolation cover (11); a shock-absorbing block (27) is slidably connected to the inner wall of the preheating dispersion cylinder (12); a first return spring (28) is fixedly connected to the bottom of the preheating dispersion cylinder (12); and one end of the first return spring (28) away from the shock-absorbing block (27) is fixedly connected to the bottom of the preheating dispersion cylinder (12).

3. A nucleic acid extraction device according to claim 1, characterized in that: The upper end of the preheating dispersion tube (12) is also provided with an anti-falling mechanism (3), and the anti-falling mechanism (3) is used to prevent the multifunctional sampling tube (13) from falling off. The anti-falling mechanism (3) includes a multifunctional clamping and prying plate (31), and the middle part of the multifunctional clamping and prying plate (31) is rotatably connected to a support plate (32). The support plate (32) is rotatably connected to the outer surface of the upper end of the preheating dispersion tube (12) at one end away from the multifunctional clamping and prying plate (31). The multifunctional clamping and prying plate (31) is fixedly connected to a pressing plate at one end, and the multifunctional clamping and prying plate (31) is fixedly connected to a splint at the other end. A card slot is provided at the upper end of the preheating dispersion tube (12), and a second return spring (33) is evenly and fixedly connected in the card slot. The second return spring (33) is fixedly connected to the splint of the multifunctional clamping and prying plate (31) at one end away from the card slot.

4. A nucleic acid extraction device according to claim 1, characterized in that: The auxiliary sampling mechanism (4) includes a retaining ring (41), the retaining ring (41) is fixedly connected to the inner wall of the upper end of the multifunctional sampling tube (13), a limiting hole (42) is provided on the retaining ring (41), a sealing cover (43) is rotatably connected to the outer surface of the upper end of the multifunctional sampling tube (13), and a non-slip limiting plate (44) is fixedly connected to the side of the sealing cover (43) away from the connection with the multifunctional sampling tube (13); The nucleic acid detection stick (14) is arranged below the retaining ring (41) near the nucleic acid extraction end, an auxiliary groove is provided on the nucleic acid detection stick (14), and the nucleic acid detection stick (14) is arranged at the upper end of the multifunctional sampling tube (13); The nucleic acid detection end of the nucleic acid detection stick (14) is placed at the bottom of the retaining ring (41) and in the limiting hole (42). The detection rod of the nucleic acid detection stick (14) is limited by the limiting hole (42) and the anti-slip limiting plate (44), so that the detection rod is stably tilted at the upper end of the multi-functional sampling tube (13). When the sealing cover (43) is closed, the auxiliary groove of the nucleic acid detection stick (14) is pressed by the anti-slip limiting plate (44), thereby breaking the handle of the nucleic acid detection stick (14).

5. A nucleic acid extraction method, applicable to a nucleic acid extraction device according to any one of claims 1 to 4, characterized in that: The following steps are involved: Step 1: Sampling of clinical specimens: Use the nucleic acid detection stick (14) to scrape the inside cheeks of the pig's mouth and the inside of the nasal cavity, then put the nucleic acid detection stick (14) into the multifunctional sampling tube (13), break off the handle, and mark it clearly; Step 2: Extraction of nucleic acid: Extract viral RNA or DNA from the sample and reverse transcribe the RNA to obtain a certain number of DNA or cDNA molecules; Step 3: Multiplex quantitative PCR detection and nanopore sequencing library construction: After 1.5 hours of multiplex quantitative PCR detection, a graphical curve is obtained on the terminal. After 10 minutes of nanopore sequencing library construction, the ends of these DNA or cDNA molecules are repaired and ligated with sequencing adapters to complete the library construction. The library is then placed on the sequencing chip. The process of molecules entering and passing through the nanopore is monitored in real time by an electrical signal feedback system, and the sequence information of each molecule is recorded. Step 4, MinION sequencing: Basic preprocessing of the data obtained through sequencing. In step 3, low-quality data is filtered, the sequencing adapter sequences are removed, and the DNA or cDNA sequences are spliced ​​and assembled. Finally, the data results are compared with the known gene library, and the viral infection status and abnormal genes are analyzed and identified through Centrifuge software.

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