CfDNA extraction system and extraction method

Abstract

The invention relates to a cfDNA extraction system and method. The system is provided with a rack, an index plate, a sample deep-hole plate, a magnetic bead deep-hole plate, a rinsing deep-hole plate, an elution deep-hole plate, a magnetic rod sleeve, a reagent sample adding device, a center module, a temperature control module and a control module. The extraction method comprises the steps of incubation, ice bath, magnetic bead adsorption, combination of magnetic beads and cfDNA, rinsing and elution. According to the cfDNA extraction system and the extraction method provided by the invention, related parameters are set in the control module, and the control module controls the index plate of the cfDNA extraction system to cooperate with the center module to execute a magnetic bar sleeve fixing process and controls the index plate to cooperate with the temperature control module to execute incubation and ice bath processes; the index plate is controlled to be matched with the center module to execute the procedures of magnetic bead adsorption, magnetic bead and cfDNA combination, rinsing and elution; the whole cfDNA extraction process does not need manual participation, has the advantages of high automation degree, small personal error and high extraction efficiency, and can adapt to a high-precision and high-efficiency medical examination technology in the future.

Description

Technical Field

[0001] This invention relates to the field of biological detection technology, and in particular to a cfDNA extraction system and extraction method. Background Technology

[0002] cfDNA is a cell-free DNA fragment released into bodily fluids such as blood. Its concentration in the human body changes with tissue damage, cancer, and inflammatory responses. It is currently widely used in areas such as fetal chromosomal abnormality detection and transplant rejection assessment. However, cfDNA is present in extremely low concentrations in bodily fluids such as blood, typically only 10-200 ng / ml, and is highly susceptible to contamination by cellular genomes and other organic components. Furthermore, its small fragment size presents numerous challenges for extraction, including high sample volume requirements, insufficient extraction yield to meet testing requirements, and low extraction stability. Nucleic acid extraction instruments with incubation and ice bath functions are almost nonexistent on the market, preventing true automation of cfDNA extraction. Summary of the Invention

[0003] This invention addresses the technical problem of the difficulty in automatically and efficiently extracting cfDNA in existing technologies by providing a cfDNA extraction system and method.

[0004] To achieve the above objectives, the present invention provides the following technical solution: A cfDNA extraction system includes: a frame, a scale plate, a sample deep well plate, a magnetic bead deep well plate, a rinsing deep well plate, an elution deep well plate, a magnetic rod sleeve, a reagent dispensing device, a central module, a temperature control module, and a control module. The indexing plate has eight workstations evenly distributed around its circumference, which are named sequentially from 1 to 8. The indexing plate is mounted on the machine frame and is rotatable. The sample deep-hole plate is set at station 1; the magnetic bead deep-hole plate is set at station 2; the rinsing deep-hole plate is set at stations 3 to 6 respectively; the elution deep-hole plate is set at station 7; and the magnetic rod sleeve is set at station 8. The reagent dispensing device is mounted on the frame and is used to add reagents to the sample deep well plate; The central module includes a matching magnetic rod and a magnetic rod sleeve, with the magnetic rod positioned above the magnetic rod sleeve. The central module is assembled on the frame and is used to install and remove the magnetic rod sleeve, as well as to drive the magnetic rod and magnetic rod sleeve to move in the vertical direction. The temperature control device is installed on the frame and is located below the indexing plate. The temperature control device is used to incubate or ice bath the sample deep well plate. The control module is electrically connected to the indexing plate, the reagent dispensing device, the central module, and the temperature control module, respectively.

[0005] Furthermore, the central module includes: a magnetic rod, a magnetic rod sleeve, a mounting bracket, a first mounting plate, a second mounting plate, a slide rail, a first slider, a second slider, a first stepper motor, a lead screw, a second stepper motor, and a belt drive assembly; The mounting bracket is fixed on the frame; the slide rail is longitudinally fixed on the mounting bracket, and the first slider and the second slider are disposed on the slide rail, with the first slider disposed above the second slider; The first mounting plate is fixedly connected to the first slider. The first stepper motor is mounted on the upper end of the mounting frame. The first stepper motor is threadedly connected to the first mounting plate through the lead screw and is used to drive the first mounting plate to move up and down. A plurality of evenly distributed magnetic rods are mounted on the lower end of the first mounting plate. The second mounting plate is fixedly connected to the second slider. The second stepper motor is mounted on the side of the mounting frame and is connected to the second mounting plate through the belt drive assembly to drive the second mounting plate to move up and down. The second mounting plate is configured as a square frame structure and is provided with a clamp for holding the magnetic rod sleeve.

[0006] Furthermore, it also includes: a third stepper motor, connecting rods, and a dirt baffle; The third stepper motor is mounted on the side of the mounting bracket; The third stepper motor is connected to the baffle plate via the connecting rod, and the baffle plate is used to collect the sample dripping from the magnetic rod sleeve.

[0007] Furthermore, the temperature control module includes: a linear drive mechanism, a height lifting mechanism, a carrier plate, a temperature lifting module, and a contour jig; The linear drive mechanism and the height lifting mechanism are fixed on the frame; The linear drive mechanism is connected to the height lifting mechanism, and the linear drive mechanism is used to drive the height lifting mechanism to perform lifting actions. The carrier plate is connected to the upper end of the height lifting mechanism; The temperature rise and fall module is connected to the upper end of the carrier plate; The contouring fixture is connected to the upper end of the temperature rise and fall module, and the contouring fixture cooperates with the bottom of the deep hole plate.

[0008] Furthermore, the height lifting mechanism includes: a top plate, a first movable arm, a second movable arm, a first limiting frame, a second limiting frame, and a base; The first limiting frame is fixed to the lower end of the top plate, and the second limiting frame is fixed to the upper end of the base. The first limiting frame and the second limiting frame are arranged vertically opposite each other. The first movable arm is connected to the second movable arm via a pin in an "X" shape. The lower end of the first movable arm is connected to the second limiting frame by a pin, and the upper end of the first movable arm is connected to the lower end of the top plate by a pin; the lower end of the second movable arm is connected to the upper end of the base by a pin, and the upper end of the second movable arm is connected to the first limiting frame by a pin; the base is fixed on the frame. The linear drive mechanism is connected to the lower end of the first telescopic arm; the carrier plate is fixed to the upper end of the top plate.

[0009] Furthermore, the temperature control component also includes: a frame connector; The temperature rise and fall module is fixed to the upper end of the carrier plate by the frame connector; The frame connectors include: a first crossbeam, a second crossbeam, a first longitudinal beam, a second longitudinal beam, a third longitudinal beam, and a fourth longitudinal beam; The lower ends of the first and second longitudinal beams are respectively fixed to the first crossbeam, and the lower ends of the third and fourth longitudinal beams are respectively fixed to the second crossbeam; the first and second crossbeams are arranged facing each other. The temperature rise and fall module is fixed to the upper ends of the first longitudinal beam, the second longitudinal beam, the third longitudinal beam and the fourth longitudinal beam; The first and second crossbeams are fixed to the carrier plate.

[0010] This invention also provides a method for extracting cfDNA, which is based on a cfDNA extraction system; the method includes: Step S1. Prepare proteinase K, plasma sample and 20% sodium dodecyl sulfate in a deep-well plate; Step S2. Incubate and ice bath the sample in the deep well plate; Step S3. Add a mixture of binding lysis buffer and isopropanol into the deep well plate of the sample; Step S4. The clamps on the control center module hold and fix the magnetic rod sleeve; Step S5. Control the magnetic rod sleeve to attract the magnetic beads from inside the deep hole plate of the magnetic beads; Step S6. Release the magnetic beads adsorbed by the magnetic rod into the sample deep well plate; Step S7. Shake the magnetic beads in the deep well plate of the sample to mix them well, so that the cfDNA binds to the magnetic beads; Step S8. Release the magnetic beads bound with cfDNA into the rinsing deep well plate for rinsing; Step S9. Release the rinsed magnetic beads into the elution deep well plate for elution. The elution solution left in the elution deep well plate is the purified cfDNA elution solution.

[0011] Furthermore, after rinsing the magnetic beads bound to cfDNA in step 8, the magnetic beads are aspirated by a magnetic rod and separated from the elution deep-well plate, and the alcohol on the surface of the magnetic beads is allowed to evaporate within 5-10 minutes.

[0012] Furthermore, in step S8, the magnetic beads bound to cfDNA need to be rinsed sequentially through four deep-well rinsing plates.

[0013] Compared with the prior art, the cfDNA extraction system and extraction method provided by the present invention have the following beneficial effects: The cfDNA extraction system provided by this invention includes a graduated disc, a sample deep-well plate, a magnetic bead deep-well plate, a rinsing deep-well plate, an elution deep-well plate, a magnetic rod sleeve, a reagent dispensing device, a central module, a temperature control module, and a control module. During cfDNA extraction, only relevant parameters need to be set in the control module. The control module then controls the graduated disc to work with the central module to fix the magnetic rod sleeve, the temperature control module to perform incubation and ice bath procedures, and the central module to perform magnetic bead adsorption, magnetic bead binding to cfDNA, rinsing, and elution procedures. The entire cfDNA extraction process requires no manual intervention; the program set in the control module controls each module to complete the entire extraction process. It has the advantages of high automation, low human error, and high extraction efficiency, and can adapt to future high-precision and high-efficiency medical testing technologies. Attached Figure Description Figure 1 This is a schematic diagram of the cfDNA extraction system provided in an embodiment of the present invention; Figure 2 This is a schematic diagram of the central module structure provided in an embodiment of the present invention; Figure 3 This is a schematic diagram of the temperature control device provided in an embodiment of the present invention; Figure 4 This is a schematic diagram of the frame connector structure provided in an embodiment of the present invention; Figure 5 This is a schematic diagram of the height lifting mechanism provided in an embodiment of the present invention.

[0014] Among them, 1-frame, 2-indexing plate, 3-sample deep well plate, 4-magnetic bead deep well plate, 5-rinsing deep well plate, 6-elution deep well plate, 7-magnetic rod sleeve, 8-reagent dispensing device, 9-central module, 100-temperature control module, 10-linear drive mechanism, 20-height lifting mechanism, 30-carrier plate, 40-temperature lifting module, 50-contouring fixture, 60-frame connector, 70-base plate, 21-first movable arm, 22-second movable arm, 23-first limiting frame, 24-second limiting frame Position frame, 25-top plate, 26-base, 61-first crossbeam, 62-second crossbeam, 63-first longitudinal beam, 64-second longitudinal beam, 65-third longitudinal beam, 66-fourth longitudinal beam, 91-magnetic rod, 92-mounting bracket, 93-first mounting plate, 94-second mounting plate, 95-slide rail, 96-first slider, 97-second slider, 98-first stepper motor, 99-lead screw, 910-second stepper motor, 911-third stepper motor, 912-connecting rod, 913-pollution baffle. Detailed Implementation

[0015] To facilitate understanding of the present invention, a more complete description will be given below with reference to the accompanying drawings. Embodiments of the invention are shown in the drawings. However, the invention can be implemented in many different forms and is not limited to the embodiments described herein. Rather, these embodiments are provided so that the disclosure of the invention will be thorough and complete.

[0016] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. The terminology used herein in the description of the invention is for the purpose of describing particular embodiments only and is not intended to be limiting of the invention.

[0017] It is understood that spatial relation terms such as "below," "under," "below," "below," "above," "above," etc., can be used here to describe the relationship between one element or feature shown in the figure and other elements or features. It should be understood that, in addition to the orientation shown in the figure, spatial relation terms also include different orientations of the device in use and operation. For example, if the device in the figure is flipped, the element or feature described as "below" or "below" of the other element or feature will be oriented "above" the other element or feature. Therefore, the exemplary terms "below" and "below" can include both upper and lower orientations. Furthermore, the device may also include other orientations (e.g., rotated 90 degrees or other orientations), and the spatial descriptive terms used herein will be interpreted accordingly.

[0018] It should be noted that when one element is considered to be "connected" to another element, it can be directly connected to the other element or connected to the other element through an intermediary element. In the following embodiments, "connection" should be understood as "electrical connection," "communication connection," etc., if the connected circuits, modules, units, etc., have the transmission of electrical signals or data between them.

[0019] When used herein, the singular forms of “a,” “an,” and “the” may also include the plural forms unless the context clearly indicates otherwise. It should also be understood that the terms “comprising,” “including,” or “having,” etc., specify the presence of the stated feature, whole, step, operation, component, part, or combination thereof, but do not preclude the possibility of the presence or addition of one or more other features, wholes, steps, operations, components, parts, or combinations thereof.

[0020] Example 1 See Figure 1 and Figure 2 This invention provides a cfDNA extraction system, which mainly includes: a frame 1, a scale plate 2, a sample deep well plate 3, a magnetic bead deep well plate 4, a rinsing deep well plate 5, an elution deep well plate 6, a magnetic rod sleeve 7, a reagent dispensing device 8, a central module 9, a temperature control module 100, and a control module.

[0021] The indexing plate 2 has eight stations evenly distributed around its circumference, named sequentially from station 1 to station 8. The indexing plate 2 is mounted on the frame 1 and is rotatable, rotating one or more stations at a time. The indexing plate 2 is a product of existing technology, and its structure will not be described in detail here. The sample deep-hole plate 3 is located at station 1; the magnetic bead deep-hole plate 4 is located at station 2; rinsing deep-hole plates 5 are located at stations 3 through 6 respectively; the elution deep-hole plate 6 is located at station 7; and the magnetic rod sleeve 7 is located at station 8.

[0022] The reagent dispensing device 8 is mounted on the frame 1 and is used to add reagents to the sample deep well plate 3.

[0023] The central module 9 includes a matching magnetic rod 91 and a magnetic rod sleeve 7, with the magnetic rod 91 positioned above the magnetic rod sleeve 7. The central module 9 is mounted on the frame 1 and is used to install and remove the magnetic rod sleeve 7, as well as to drive the magnetic rod 91 and the magnetic rod sleeve 7 to move in the up and down direction to cooperate in completing different steps of cfDNA extraction.

[0024] The temperature control device is installed on the frame 1, located below the indexing plate 2 and directly below the central module 9. The temperature control device is used to incubate or ice bath the sample deep well plate 3.

[0025] The control module is electrically connected to the indexing plate 2, reagent dispensing device 8, central module 9, and temperature control module 100, respectively. Control commands and control parameters are written into the control module. The control module controls the indexing plate 2, reagent dispensing device 8, central module 9, and temperature control module 100 to execute commands and complete corresponding actions according to the written commands and parameters.

[0026] For details, see Figure 1 and Figure 2 The central module 9 includes: a magnetic rod 91, a magnetic rod sleeve 7, a mounting bracket 92, a first mounting plate 93, a second mounting plate 94, a slide rail 95, a first slider 96, a second slider 97, a first stepper motor 98, a lead screw 99, a second stepper motor 910, and a belt drive assembly. The mounting bracket 92 is fixed on the frame 1; the slide rail 95 is longitudinally fixed on the mounting bracket 92, and the first slider 96 and the second slider 97 are disposed on the slide rail 95, with the first slider 96 positioned above the second slider 97. The first slider 96 and the second slider 97 can move linearly along the slider in the vertical direction.

[0027] The first mounting plate 93 is fixedly connected to the first slider 96. The first stepper motor 98 is installed on the upper end of the mounting frame 92. The first stepper motor 98 is threadedly connected to the first mounting plate 93 through the lead screw 99 and is used to drive the first mounting plate 93 to move up and down. Multiple evenly distributed magnetic rods 91 are installed on the lower end of the first mounting plate 93.

[0028] The second mounting plate 94 is fixedly connected to the second slider 97. The second stepper motor 910 is mounted on the side of the mounting bracket 92. The second stepper motor 910 is connected to the second mounting plate 94 through a belt drive assembly and is used to drive the second mounting plate 94 to move up and down. The second mounting plate 94 is configured as a square frame structure and is provided with a clamp for holding and fixing the magnetic rod sleeve 7.

[0029] In a preferred embodiment of the present invention, to prevent the sample deep-well plate 3 from being contaminated during incubation or washing, a third stepper motor 911, a connecting rod 912, and a baffle plate 913 are also provided. The third stepper motor 911 is mounted on the side of the mounting bracket 92. The third stepper motor 911 is connected to the baffle plate 913 via the connecting rod 912. The third stepper motor 911 drives the baffle plate 913 to cover the sample deep-well plate 3 located below the second mounting plate 94. The baffle plate 913 is used to collect samples dripping from the magnetic rod sleeve 7, preventing samples from accidentally dripping onto other samples and causing contamination during the rotation of the indexing plate 2.

[0030] In this embodiment of the invention, see Figures 3-5The temperature control module includes components such as a linear drive mechanism 10, a height lifting mechanism 20, a carrier plate 30, a temperature lifting module 40, and a contour jig 50. The linear drive mechanism 10 is connected to the height lifting mechanism 20 and drives the height lifting mechanism 20 to perform lifting actions. In this embodiment, the linear drive mechanism 10 uses a stepper motor. The carrier plate 30 is connected to the upper end of the height lifting mechanism 20, and the height lifting mechanism 20 drives the carrier plate 30 to move up and down. The temperature lifting module 40 is connected to the upper end of the carrier plate 30, and the carrier plate 30 supports the temperature lifting module 40. The temperature lifting module 40 is existing technology; for example, a PCR heating and cooling module can be used, which has both heating and cooling functions. Its specific structure will not be described here. The contour jig 50 is connected to the upper end of the temperature lifting module 40, which is used to heat or cool the contour jig 50. The contour jig 50 is made of a metal with good thermal conductivity. Multiple conical grooves that mate with the deep hole plate are distributed on the upper end of the contour jig 50. These conical grooves fit tightly with the bottom of the deep hole plate, thereby achieving the heating or cooling of the deep hole plate. The linear drive mechanism 10 and the base 26 are fixed on the base plate 70.

[0031] For details, see Figure 3 and Figure 5 The height lifting mechanism 20 includes: a top plate 25, a first movable arm 21, a second movable arm 22, a first limiting frame 23, a second limiting frame 24, and a base 26. The first limiting frame 23 is fixed to the lower end of the top plate 25, and the second limiting frame 24 is fixed to the upper end of the base 26. The first limiting frame 23 and the second limiting frame 24 are arranged vertically opposite each other. The first movable arm 21 is connected to the second movable arm 22 by a pin in an "X" shape. The lower end of the first movable arm 21 is connected to the second limiting frame 24 by a pin, and the pin at the lower end of the first movable arm 21 can move laterally within the second limiting frame 24. The upper end of the first movable arm 21 is connected to the lower end of the top plate 25 by a pin. The lower end of the second movable arm 22 is connected to the upper end of the base 26 by a pin, and the upper end of the second movable arm 22 is connected to the first limiting frame 23 by a pin, and the pin at the upper end of the second movable arm 22 can move laterally within the first limiting frame 23. The linear drive mechanism 1 is connected to the lower end of the first telescopic arm; the carrier plate 3 is fixed to the upper end of the top plate 25.

[0032] See Figure 3 and Figure 5The working process of the height lifting mechanism 20 is as follows: During the upward movement, the movable end of the linear drive mechanism 10 extends outward, pushing the lower end of the first movable arm 21 towards the lower end of the second movable arm 22. The upper ends of the first movable arm 21 and the second movable arm 22 move closer together and move upward synchronously. During the upward movement of the upper ends of the first movable arm 21 and the second movable arm 22, the carrier plate 30 moves upward synchronously. During the downward movement, the movable end of the linear drive mechanism 10 retracts inward, driving the lower end of the first movable arm 21 away from the lower end of the second movable arm 22. The upper ends of the first movable arm 21 and the second movable arm 22 separate and move downward synchronously. During the downward movement of the upper ends of the first movable arm 21 and the second movable arm 22, the carrier plate 30 moves downward synchronously.

[0033] In a preferred embodiment provided by the present invention, see [reference needed]. Figure 3 and Figure 4 To ensure reliable installation of the temperature lifting module 4, a frame connector 60 is also provided; the temperature lifting module 40 is fixed to the upper end of the carrier plate 30 via the frame connector 60. Specifically, the frame connector 60 includes: a first crossbeam 61, a second crossbeam 62, a first longitudinal beam 63, a second longitudinal beam 64, a third longitudinal beam 65, and a fourth longitudinal beam 66. The lower ends of the first longitudinal beam 63 and the second longitudinal beam 64 are respectively fixed to the first crossbeam 61, and the lower ends of the third longitudinal beam 65 and the fourth longitudinal beam 66 are respectively fixed to the second crossbeam 62; the first crossbeam 61 and the second crossbeam 62 are arranged facing each other. The temperature lifting module 40 is fixed to the upper ends of the first longitudinal beam 63, the second longitudinal beam 64, the third longitudinal beam 65, and the fourth longitudinal beam 66. The first crossbeam 61 and the second longitudinal beam 62 are fixed to the carrier plate 30.

[0034] See Figure 3 The working process of the temperature control module is as follows: the height lifting mechanism 20 moves upward, causing the contour jig 50 and the temperature control module to move upward; the lifting mechanism stops moving when the contour jig 50 contacts the deep hole plate; the temperature lifting module 40 starts working to heat or cool the deep hole plate; when the temperature reaches the expected value, the temperature lifting module 40 stops working; the height lifting mechanism 20 moves downward, causing the contour jig 50 and the temperature lifting module 40 to move downward; the contour jig 50 separates from the deep hole plate.

[0035] When the cfDNA extraction system provided in this embodiment of the invention is working, relevant parameters are set in the control module. The control module controls the indexing plate 2 of the cfDNA extraction system to work with the central module 9 to perform the process of fixing the magnetic rod sleeve 7, controls the indexing plate 2 to work with the temperature control module to perform the incubation and ice bath processes, and controls the indexing plate 2 to work with the central module to perform the processes of adsorbing magnetic beads, binding magnetic beads with cfDNA, rinsing and elution.

[0036] Example 2 See Figures 1-5 This invention provides a method for extracting cfDNA, which is based on the cfDNA extraction system provided in Example 1; the method includes the following steps: Step S1. Prepare proteinase K, plasma sample and 20% sodium dodecyl sulfate in sample deep well plate 3; Step S2. Incubate and ice bath the sample in the deep well plate 3.

[0037] Specifically, the sample deep well plate 3 is rotated above the temperature control module 100, and the sample in the sample deep well plate 3 is incubated and subjected to an ice bath according to the set time and temperature.

[0038] Step S3. Add a mixture of binding lysis buffer and isopropanol into the sample deep well plate 3.

[0039] Specifically, the sample deep well plate 3 is rotated to a position below the reagent loading device 8, and a mixture of binding lysis buffer and isopropanol is added into the sample deep well plate 3.

[0040] Step S4. The clamps on the control center module 9 hold and fix the magnetic rod sleeve 7.

[0041] Specifically, the magnetic rod sleeve 7 is rotated to the bottom of the central module 9, the clamp on the central module 9 is controlled to move downward, and the magnetic rod sleeve 7 is fixed by the clamp, and then the clamp moves upward to reset.

[0042] Step S5. Control the magnetic rod sleeve 7 to attract the magnetic beads from inside the magnetic bead deep hole plate 4.

[0043] Specifically, the magnetic bead deep hole plate 4 is rotated to the bottom of the central module 9, and the magnetic rod 91 of the central module 9 is controlled to be inserted into the inside of the magnetic rod sleeve 7 from above. The magnetic rod sleeve 7 is controlled to move downward so that it is inserted into the magnetic bead deep hole plate 4 to attract the magnetic beads. Then the magnetic rod sleeve 7 is controlled to move upward to reset, and the magnetic rod sleeve 7 is separated from the magnetic bead deep hole plate 4.

[0044] Step S6. Release the magnetic beads adsorbed by the magnetic rod sleeve 7 into the sample deep well plate 3.

[0045] Specifically, the sample deep well plate 3 is rotated to the bottom of the central module 9, and the magnetic rod 91 and the magnetic rod sleeve 7 are controlled to move downward synchronously, so that the magnetic rod sleeve 7 is inserted into the sample deep well plate 3; the magnetic rod 91 is controlled to move upward and separate from the magnetic rod sleeve 7, releasing the magnetic bead into the sample deep well plate 3; then the magnetic rod sleeve 7 is controlled to move upward and separate from the sample deep well plate 3.

[0046] Step S7. Shake the magnetic beads in the sample deep well plate 3 to mix them well, so that the cfDNA binds to the magnetic beads.

[0047] Specifically, the magnetic rod sleeve 7 is controlled to move up and down reciprocally, continuously engaging and disengaging with the sample deep well plate 3, shaking and agitating the magnetic beads inside the sample deep well plate 3 to allow cfDNA to bind to the magnetic beads; then the magnetic rod 91 and the magnetic rod sleeve 7 are controlled to move downwards synchronously, allowing the magnetic rod sleeve 7 to insert into the magnetic bead deep well plate 4 to adsorb the magnetic beads; the magnetic rod 91 and the magnetic rod sleeve 7 are controlled to move upwards synchronously, allowing the magnetic rod sleeve 7 to separate from the magnetic bead deep well plate 4.

[0048] Step S8. Release the magnetic beads bound with cfDNA into the rinsing deep well plate 5 for rinsing.

[0049] Specifically, rinsing stations 3-6 are rotated sequentially to below the central module 9. Magnetic rod 91 and its sleeve 7 are simultaneously moved downwards, allowing the sleeve 7 to insert into the rinsing deep-well plate 5. Magnetic rod 91 is then moved upwards to separate from the sleeve 7, releasing the magnetic beads bound to cfDNA into the rinsing deep-well plate 5 for rinsing. After rinsing, magnetic rod 91 is moved downwards again, inserting into the sleeve 7, which adsorbs the magnetic beads within the magnetic bead deep-well plate 4. Magnetic rod 91 and its sleeve 7 are then simultaneously moved upwards, separating the sleeve 7 from the magnetic bead deep-well plate 4. In this step, the magnetic beads bound to cfDNA need to pass through four rinsing deep-well plates 5 sequentially during rinsing. After rinsing the magnetic beads bound to cfDNA in step 8, the magnetic beads are aspirated by the sleeve 7 and separated from the elution deep-well plate 6. 5-10 minutes are allowed for the alcohol on the surface of the magnetic beads to evaporate.

[0050] Step S9. Release the rinsed magnetic beads into the elution deep well plate 6 for elution. The elution solution left in the elution deep well plate 6 is the purified cfDNA elution solution.

[0051] Specifically, the elution deep-well plate 6 is rotated below the central module 9. The magnetic rod 91 and its sleeve 7 are simultaneously moved downwards, allowing the sleeve 7 to insert into the elution deep-well plate 6. The magnetic rod 91 is then moved upwards to separate from the sleeve 7, releasing the magnetic beads into the elution deep-well plate 6 for elution. The magnetic rod 91 is then moved downwards, allowing the sleeve 7 to insert into the plate. The sleeve 7 adsorbs the magnetic beads within the elution deep-well plate 6. The magnetic rod 91 and its sleeve 7 are then moved upwards simultaneously, separating the sleeve from the magnetic beads in the deep-well plate 4. The eluent remaining in the elution deep-well plate 6 is the purified cfDNA eluent.

[0052] See Figure 1 The cfDNA extraction system and method provided in this invention have the following beneficial effects: The present invention provides a cfDNA extraction system and method. The system comprises a graduated disc 2, a sample deep-well plate 3, a magnetic bead deep-well plate 4, a rinsing deep-well plate 5, an elution deep-well plate 6, a magnetic rod sleeve 7, a reagent dispensing device 8, a central module 9, a temperature control module 100, and a control module. During cfDNA extraction, only relevant parameters need to be set within the control module. The control module controls the graduated disc 2 of the cfDNA extraction system to work with the central module 9 to perform the fixing of the magnetic rod sleeve 7, the incubation and ice bath processes with the temperature control module, and the adsorption of magnetic beads, binding of magnetic beads to cfDNA, rinsing, and elution processes with the central module. The entire cfDNA extraction process requires no manual intervention; the program set within the control module controls each module to complete the entire extraction process. It has the advantages of high automation, low human error, and high extraction efficiency, and can adapt to future high-precision and high-efficiency medical testing technologies. The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

Claims

1. A cfDNA extraction system, characterized in that: include: The equipment includes a frame, indexing plate, sample deep well plate, magnetic bead deep well plate, rinsing deep well plate, elution deep well plate, magnetic rod sleeve, reagent dispensing device, central module, temperature control module, and control module. The indexing plate has eight workstations evenly distributed around its circumference, which are named sequentially from 1 to 8. The indexing plate is mounted on the machine frame and is rotatable. The sample deep-hole plate is set at station 1; the magnetic bead deep-hole plate is set at station 2; the rinsing deep-hole plate is set at stations 3 to 6 respectively; the elution deep-hole plate is set at station 7; and the magnetic rod sleeve is set at station 8. The reagent dispensing device is mounted on the frame and is used to add reagents to the sample deep well plate; The central module includes a matching magnetic rod and a magnetic rod sleeve, with the magnetic rod positioned above the magnetic rod sleeve. The central module is assembled on the frame and is used to install and remove the magnetic rod sleeve, as well as to drive the magnetic rod and magnetic rod sleeve to move in the vertical direction. The temperature control device is installed on the frame and is located below the indexing plate. The temperature control device is used to incubate or ice bath the sample deep well plate. The control module is electrically connected to the indexing plate, the reagent dispensing device, the central module, and the temperature control module, respectively.

2. The cfDNA extraction system according to claim 1, characterized in that: The central module includes: a magnetic rod, a magnetic rod sleeve, a mounting bracket, a first mounting plate, a second mounting plate, a slide rail, a first slider, a second slider, a first stepper motor, a lead screw, a second stepper motor, and a belt drive assembly; The mounting bracket is fixed on the frame; the slide rail is longitudinally fixed on the mounting bracket, and the first slider and the second slider are disposed on the slide rail, with the first slider disposed above the second slider; The first mounting plate is fixedly connected to the first slider. The first stepper motor is mounted on the upper end of the mounting frame. The first stepper motor is threadedly connected to the first mounting plate through the lead screw and is used to drive the first mounting plate to move up and down. A plurality of evenly distributed magnetic rods are mounted on the lower end of the first mounting plate. The second mounting plate is fixedly connected to the second slider. The second stepper motor is mounted on the side of the mounting frame and is connected to the second mounting plate through the belt drive assembly to drive the second mounting plate to move up and down. The second mounting plate is configured as a square frame structure and is provided with a clamp for holding the magnetic rod sleeve.

3. The cfDNA extraction system according to claim 2, characterized in that: Also includes: The third stepper motor, connecting rod, and dirt baffle; The third stepper motor is mounted on the side of the mounting bracket; The third stepper motor is connected to the baffle plate via the connecting rod, and the baffle plate is used to collect the sample dripping from the magnetic rod sleeve.

4. The cfDNA extraction system according to claim 1, characterized in that: The temperature control module includes: a linear drive mechanism, a height lifting mechanism, a carrier plate, a temperature lifting module, and a contour jig. The linear drive mechanism and the height lifting mechanism are fixed on the frame; The linear drive mechanism is connected to the height lifting mechanism, and the linear drive mechanism is used to drive the height lifting mechanism to perform lifting actions. The carrier plate is connected to the upper end of the height lifting mechanism; The temperature rise and fall module is connected to the upper end of the carrier plate; The contouring fixture is connected to the upper end of the temperature rise and fall module, and the contouring fixture cooperates with the bottom of the deep hole plate.

5. The cfDNA extraction system according to claim 4, characterized in that: The height lifting mechanism includes: a top plate, a first movable arm, a second movable arm, a first limiting frame, a second limiting frame, and a base; The first limiting frame is fixed to the lower end of the top plate, and the second limiting frame is fixed to the upper end of the base. The first limiting frame and the second limiting frame are arranged vertically opposite each other. The first movable arm is connected to the second movable arm via a pin in an "X" shaped structure; The lower end of the first movable arm is connected to the second limiting frame by a pin, and the upper end of the first movable arm is connected to the lower end of the top plate by a pin; the lower end of the second movable arm is connected to the upper end of the base by a pin, and the upper end of the second movable arm is connected to the first limiting frame by a pin; the base is fixed on the frame. The linear drive mechanism is connected to the lower end of the first telescopic arm; the carrier plate is fixed to the upper end of the top plate.

6. The cfDNA extraction system according to claim 5, characterized in that: The temperature control component also includes: a frame connector; The temperature rise and fall module is fixed to the upper end of the carrier plate by the frame connector; The frame connectors include: a first crossbeam, a second crossbeam, a first longitudinal beam, a second longitudinal beam, a third longitudinal beam, and a fourth longitudinal beam; The lower ends of the first and second longitudinal beams are respectively fixed to the first crossbeam, and the lower ends of the third and fourth longitudinal beams are respectively fixed to the second crossbeam; the first and second crossbeams are arranged facing each other. The temperature rise and fall module is fixed to the upper ends of the first longitudinal beam, the second longitudinal beam, the third longitudinal beam and the fourth longitudinal beam; The first and second crossbeams are fixed to the carrier plate.

7. A method for extracting cfDNA, the method being implemented based on the cfDNA extraction system of claim 1; characterized in that: The method includes: Step S1. Prepare proteinase K, plasma sample and 20% sodium dodecyl sulfate in a deep-well plate; Step S2. Incubate and ice bath the sample in the deep well plate; Step S3. Add a mixture of binding lysis buffer and isopropanol into the deep well plate of the sample; Step S4. The clamps on the control center module hold and fix the magnetic rod sleeve; Step S5. Control the magnetic rod sleeve to attract the magnetic beads from inside the deep hole plate of the magnetic beads; Step S6. Release the magnetic beads adsorbed by the magnetic rod into the sample deep well plate; Step S7. Shake the magnetic beads in the deep well plate of the sample to mix them well, so that the cfDNA binds to the magnetic beads; Step S8. Release the magnetic beads bound with cfDNA into the rinsing deep well plate for rinsing; Step S9. Release the rinsed magnetic beads into the elution deep well plate for elution. The elution solution left in the elution deep well plate is the purified cfDNA elution solution.

8. The method for extracting cfDNA according to claim 7, characterized in that: After rinsing the magnetic beads bound to cfDNA in step 8, the magnetic beads are picked up by the magnetic rod and separated from the elution deep well plate. Wait 5-10 minutes for the alcohol on the surface of the magnetic beads to evaporate.

9. The method for extracting cfDNA according to claim 7, characterized in that: In step S8, the magnetic beads bound to cfDNA need to be rinsed sequentially through four deep-well rinsing plates.

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