Reverse suction blending method, nucleic acid magnetic bead purification method and nucleic acid treatment instrument

Abstract

The invention provides a back-suction mixing method, a nucleic acid magnetic bead purification method and a nucleic acid treatment instrument.The back-suction mixing method is applied to a pipette, the pipette comprises a cavity and a piston, and the moving position of the piston in the cavity comprises an initial position, a first position and a second position; the method comprises the following steps: pushing the piston from the initial position to the second position; a pipette head of the pipette is immersed in the liquid in the liquid containing pipe, and the piston is released to suck part of the liquid; the piston is pushed to reciprocate for preset times, the liquid in the liquid containing pipe is evenly mixed, and in the process that the piston is pushed to enable the liquid to be pushed out of the pipette tip, the piston is kept on the side, away from the pipette tip, of the first position; and moving the pipette tip out of the liquid containing tube. Bubbles generated in the uniform mixing process can be reduced, and the uniform mixing effect is improved.

Description

Technical Field

[0001] The invention belongs to the field of biomedicine, and specifically relates to a reverse suction mixing method, a nucleic acid magnetic bead purification method and a nucleic acid processing instrument. Background Art

[0002] High-throughput sequencing technology (Next Generation Sequencing) has been widely used in basic research, prenatal diagnosis, genetic disease diagnosis, tumor diagnosis and precision medicine due to its advantages of high throughput, high accuracy and rich information. Among them, high-throughput sequencing is inseparable from library construction, and fully automatic library construction instrument is a new research direction.

[0003] However, the liquid mixing effect of the existing fully automatic closed cartridge library building instrument needs to be improved. Summary of the invention

[0004] In view of this, the present invention provides a back-absorption mixing method, a nucleic acid magnetic bead purification method and a nucleic acid processing instrument to improve the mixing effect.

[0005] In order to solve the above problems, the present invention provides a reverse suction mixing method, which is applied to a pipette, wherein the pipette includes a cavity and a piston, a pipette head is provided at one end of the cavity, and the other end is used to set the piston, and the moving position of the piston in the cavity includes an initial position, a first position, and a second position, and the distance between the initial position and the second position is greater than the distance between the initial position and the first position. The method includes:

[0006] pushing the piston from the initial position to the second position;

[0007] Dipping the pipette tip of the pipette into the liquid in the liquid holding tube and releasing the piston to aspirate part of the liquid;

[0008] Pushing the piston to reciprocate for a preset number of times to mix the liquid in the liquid holding tube, wherein, in the process of pushing the piston to push the liquid out of the pipette tip, the piston remains at the first position away from the side of the pipette tip;

[0009] Move the pipette tip out of the liquid holding tube.

[0010] Optionally, the step of moving the pipette tip out of the liquid holding tube comprises:

[0011] Release the piston to absorb part of the liquid, and lift the pipette tip out of the liquid in the liquid holding tube;

[0012] Pushing the piston to the first position to obtain a preset pipetting volume;

[0013] Pushing the piston to the second position to blow out the remaining liquid;

[0014] Remove the emptied pipette from the liquid holding tube.

[0015] Optionally, in the step of pushing the piston to reciprocate for a preset number of times to mix the liquid in the liquid holding tube, when the piston pushes out the liquid, the pipette moves to hit the liquid.

[0016] Optionally, the position of the pipette tip in the liquid holding tube is: a first holding position, which is located at the center of the liquid holding tube; or a second holding position, which is located at a side of the liquid holding tube close to the magnet; or a third holding position, which is located at a side of the liquid holding tube away from the magnet.

[0017] To solve the aforementioned problems, the present invention further provides a nucleic acid processing instrument, comprising: a pipette and a main control unit coupled to the pipette, wherein the main control unit is used to issue instructions to control the pipette to execute any of the aforementioned back-suction mixing methods.

[0018] Optionally, the pipette includes at least a first pipette and a second pipette, the capacity of the first pipette is greater than the capacity of the second pipette, wherein the first pipette is used to perform any of the aforementioned back-suction mixing methods.

[0019] To solve the above problems, the present invention also provides a method for purifying nucleic acid using magnetic beads, comprising:

[0020] Using any of the above-mentioned back-absorption mixing methods, the magnetic beads, the nucleic acid and the first reagent are mixed;

[0021] Remove the supernatant and retain the magnetic beads adsorbed with nucleic acid;

[0022] Using any of the above-mentioned back-absorption mixing methods, the magnetic beads, the nucleic acid and the second reagent are mixed to separate the magnetic beads from the nucleic acid;

[0023] The supernatant containing the purified nucleic acid is transferred.

[0024] Optionally, in the step of mixing the magnetic beads, nucleic acid and the first reagent, the position of the pipette tip is a first accommodation position, and the first accommodation position is located at the center of the liquid accommodation tube.

[0025] Optionally, in the step of mixing the magnetic beads, nucleic acid and the second reagent, the position of the pipette tip is a second accommodation position, and the second accommodation position is located on a side of the liquid accommodation tube close to the magnet.

[0026] Optionally, in the step of removing the supernatant and the step of transferring the supernatant, the position of the pipette tip is a third accommodation position, and the third accommodation position is located on a side of the liquid holding tube away from the magnet.

[0027] Compared with the prior art, the technical solution of the present invention has the following advantages:

[0028] The embodiment of the present application provides a back-suction mixing method, a nucleic acid magnetic bead purification method and a nucleic acid processing instrument, wherein the back-suction mixing method is applied to a pipette, wherein the pipette includes a cavity and a piston, wherein a pipette tip is provided at one end of the cavity, and the other end is used to provide the piston, wherein the moving position of the piston in the cavity includes an initial position, a first position and a second position, wherein the distance between the initial position and the second position is greater than the distance between the initial position and the first position, and the method includes: pushing the piston from the initial position to the second position; immersing the pipette tip of the pipette into the liquid in a liquid holding tube, and releasing the piston to aspirate part of the liquid; pushing the piston to reciprocate for a preset number of times to mix the liquid in the liquid holding tube, wherein, in the process of pushing the piston to push the liquid out of the pipette tip, the piston remains at the first position away from the side of the pipette tip; and moving the pipette tip out of the liquid holding tube.

[0029] It can be seen that the back-suction mixing method provided in this embodiment uses the second position to suck in the liquid for a liquid with a certain viscosity, and when mixing the liquid, the pipette tip remains in the first position away from the side of the pipette tip while pushing the piston out, and does not cross the first position, so that there is always a part of liquid in the cavity of the pipette, and this part of liquid is located at the liquid outlet of the pipette tip, avoiding mixing of gas into the liquid in the liquid holding tube during the mixing process, thereby reducing the generation of bubbles and further improving the liquid mixing effect. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] In order to more clearly illustrate the embodiments of the present application or the technical solutions in the prior art, the drawings required for use in the embodiments or the description of the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of the present application. For ordinary technicians in this field, other drawings can be obtained based on the provided drawings without paying any creative work.

[0031] Figure 1 It is a schematic diagram of the steps of the forward mixing method used in the prior art;

[0032] Figure 2 is a schematic diagram of the pipette provided in this embodiment;

[0033] Figure 3Schematic diagram of the steps of the back-suction mixing method provided in this embodiment;

[0034] Figure 4 is a schematic diagram of the position of the pipette tip of the pipette provided in this embodiment located in the liquid holding tube;

[0035] Figure 5 is an optional flow chart of step S4 provided in this embodiment;

[0036] Figure 6 is another optional flow chart of the reverse suction and mixing method provided in this embodiment;

[0037] Figure 7 Schematic diagram of the process of the nucleic acid magnetic bead purification method provided in this embodiment;

[0038] Figure 8 This is a schematic diagram of the internal structure of the instrument during nucleic acid magnetic bead purification;

[0039] Fig. 9 This is another optional flow chart of the method for purifying nucleic acid with magnetic beads provided in this embodiment;

[0040] Fig.10 This is another optional flow chart of the method for purifying nucleic acid with magnetic beads provided in this embodiment;

[0041] Fig.11 is another optional flow chart of the method for purifying nucleic acid with magnetic beads provided in this embodiment;

[0042] Fig.12 : is another optional flow chart of the nucleic acid magnetic bead purification method provided in this embodiment;

[0043] Fig.13 It is a schematic diagram of the steps of a pipetting method provided in this embodiment. DETAILED DESCRIPTION

[0044] The following will be combined with the drawings in this embodiment to clearly and completely describe the technical solution in this embodiment. Obviously, the described embodiment is only a part of the embodiments of this application, not all of the embodiments. Based on the embodiments in this application, all other embodiments obtained by ordinary technicians in this field without making creative work are within the scope of protection of the present invention.

[0045] As described in the background art, the liquid mixing effect of the existing fully automatic closed cartridge library building instrument needs to be improved.

[0046] The inventor believes that the mixing method using the pipette tip to blow and beat in the prior art is a forward aspiration mixing method. Figure 1 , Figure 1This is a schematic diagram of the steps of the forward mixing method used in the prior art, as shown in the figure, S10: push the piston to the first position of the cavity; S20: immerse the pipette tip of the pipette gun into the liquid in the liquid holding tube for 2-3mm; S30: release the piston to absorb part of the liquid; S40: push the piston up and down; S50: slide along the container wall of the holding tube to lift the pipette tip; S60: push the piston through the first position to the second position to blow out the residual liquid. This method is not friendly to viscous or easily foaming liquids. Viscous liquids are easy to remain and cannot achieve the pipetting accuracy, or the residual viscous liquid when the pipette tip is removed is easy to cause pollution to the surrounding area. If the speed is too fast, it is easy to generate bubbles.

[0047] In view of this, the present invention provides a back-suction mixing method, a nucleic acid magnetic bead purification method and a nucleic acid processing instrument, wherein the back-suction mixing method is applied to a pipette, the pipette includes a cavity and a piston, one end of the cavity is provided with a pipette head, and the other end is used to set the piston, the moving position of the piston in the cavity includes an initial position, a first position and a second position, the distance between the initial position and the second position is greater than the distance between the initial position and the first position, the method includes: pushing the piston from the initial position to the second position; immersing the pipette head of the pipette into the liquid in the liquid holding tube, and releasing the piston to absorb part of the liquid; pushing the piston to reciprocate for a preset number of times to mix the liquid in the liquid holding tube, wherein, in the process of pushing the piston to push the liquid out of the pipette head, the piston remains at the first position away from the side of the pipette head; moving the pipette head out of the liquid holding tube.

[0048] For liquids with a certain viscosity, this method uses the second position to suck in the liquid, and when mixing the liquid, the piston is kept in the first position away from the side of the pipette tip during the process of the pipette tip pushing out the piston, and does not cross the first position, so that there is always a part of liquid in the cavity of the pipette, and this part of liquid is located at the liquid outlet of the pipette tip, avoiding mixing of gas into the liquid in the liquid holding tube during the mixing process, thereby reducing the generation of bubbles and improving the liquid mixing effect.

[0049] refer to Figure 2 , Figure 201 is a schematic diagram of the pipette provided in this embodiment. As shown in the figure, the pipette 1 includes a cavity 05 and a piston 03. A pipette head 04 is provided at one end of the cavity 05, and the other end is used to set the piston 03. When the piston 03 is not pushed, the piston 03 is located at the initial position 06. 01 is the first position, and 02 is the second position, wherein, in the direction toward the pipette head 04, the initial position 06, the first position 01 and the second position 02 are arranged in sequence. It should be noted that in a specific example, the first position and the second position can be set based on the specific liquid situation and the specific structure of the pipette, and are not necessarily completely fixed. The piston 03 is driven by the instrument, and the instrument motor determines the distance to push the piston 03 to move according to the pipetting volume requirement. It can be understood that the cavity 05 can also include a hose with an extended length, and the piston 03 can be located at one end of the extended hose of the cavity 05.

[0050] refer to Figure 3 , Figure 3 is a schematic diagram of the steps of the back-sucking and mixing method provided in this embodiment, wherein the back-sucking and mixing method can be based on Figure 2 The pipette implementation provided in Figure 2-Figure 3 As shown, the method includes:

[0051] Step S1: pushing the piston from the initial position to the second position;

[0052] Push the piston to the second position of the cavity. For liquids with a certain viscosity, use the second position to suck in more liquid, so that less liquid can be pushed out in the subsequent mixing process, reducing the possibility of air entering the liquid in the liquid holding tube during the liquid pushing process, thereby reducing the generation of bubbles.

[0053] Step S2: immersing the pipette tip of the pipette into the liquid in the liquid holding tube and releasing the piston;

[0054] In a specific example, the pipette tip of the pipette gun is immersed in the liquid in the liquid holding tube, and the position of the pipette tip in the liquid holding tube can be: a first holding position, the first holding position is located at the center of the liquid holding tube; or a second holding position, the second holding position is located at a side of the liquid holding tube close to the magnet; or a third holding position, the third holding position is located at a side of the liquid holding tube away from the magnet. Among them, the first holding position can be called None, the second holding position can be called Toward, and the third holding position can be called Away.

[0055] refer to Figure 4 , Figure 4Schematic diagram of the position of the pipette tip of the pipette provided in this embodiment in the liquid containing tube, (a) is a schematic diagram of the first containing position, (b) is a schematic diagram of the second containing position, and (c) is a schematic diagram of the third containing position. It can be seen from Figure (a) that the pipette tip 04 is located in the center of the liquid containing tube 10, which is called None; from Figure (b) it can be seen that the pipette tip 04 is located on the side of the liquid containing tube 10 close to the magnet 20, which is called Toward; from Figure (c) it can be seen that the pipette tip 04 is located on the side of the liquid containing tube 10 away from the magnet 20, which is called Away.

[0056] Continue to refer Figure 2-Figure 3 , executing step S3: pushing the piston to reciprocate for a preset number of times;

[0057] Pushing the piston 03 up and down to mix the liquid in the liquid holding tube; wherein, in the process of pushing the piston 03 downward, the pipette tip 04 pushes out the liquid, and the piston 03 remains at the first position away from the side of the pipette tip, and does not go beyond the first position 01;

[0058] In an optional implementation, in the step of pushing the piston 03 to reciprocate for a preset number of times to mix the liquid in the liquid holding tube, the pipette gun 1 can also be moved to hit the liquid when the piston 03 pushes out the liquid to improve the mixing efficiency.

[0059] It should be noted that when the piston 03 is pushed to move back and forth, the pipette tip 04 also moves up and down synchronously relative to the liquid, that is, when the piston is pushed to push out the liquid, the pipette tip also moves downward to hit the liquid; when the piston is pushed to inhale the liquid, the pipette tip also moves upward synchronously. In the process of the pipette tip moving up and down, the pipette tip is always lower than the liquid level in the liquid holding tube, so that the tip hits the liquid up and down, improving the mixing efficiency while avoiding mixing air into the liquid to generate bubbles.

[0060] Step S4: moving the pipette tip out of the liquid holding tube.

[0061] refer to Figure 5 , Figure 5 is an optional flow chart of step S4 provided in this embodiment. As shown in the figure, the step of moving the pipette tip out of the liquid holding tube includes:

[0062] Step S41, releasing the piston to absorb part of the liquid, and lifting the pipette tip from the liquid in the liquid holding tube;

[0063] When the pipette tip is moved out of the liquid holding tube, it can be moved out along the tube wall of the liquid holding tube. By sliding along the tube wall of the liquid holding tube to lift the pipette tip, it is possible to prevent the residual droplets on the pipette tip from falling from the air into the liquid in the liquid holding tube, thereby preventing bubbles from being generated. By sliding along the tube wall to lift the pipette tip, the risk of residual droplets falling and generating bubbles can be effectively reduced.

[0064] Step S42, pushing the piston to the first position to obtain a preset pipetting volume;

[0065] Step S43, pushing the piston to the second position to blow out the residual liquid;

[0066] The preset pipetting volume is obtained by first pushing the piston to the first position, pausing for a while to allow the liquid remaining in the pipette chamber to slowly flow down, and then pushing the piston to the second position to blow out the remaining liquid to avoid liquid residue. During the process of pushing the piston to the first position, the liquid flow rate and pressure can be reduced to make the liquid discharged more smoothly, thereby improving the precision and accuracy of the pipetting.

[0067] Step S44, removing the emptied pipette from the liquid holding tube.

[0068] It should be noted that after the pipette is removed from the liquid holding tube, the tip of the pipette may be removed and replaced with a new tip, or the tip of the pipette may not be removed.

[0069] Wherein, in this step, the liquid volume pushed out by the pipette tip is 85%-98% of the liquid volume sucked in, preferably 95%, so as to avoid pushing out air and generating bubbles.

[0070] An embodiment of the present invention also provides a nucleic acid processing instrument, including: a pipette, and a main control unit coupled to the pipette, wherein the main control unit is used to issue instructions to control the pipette to perform the back-suction mixing method provided in the embodiment of the present invention. The pipette includes at least a first pipette and a second pipette, the capacity of the first pipette is greater than the capacity of the second pipette, wherein the first pipette is used to perform the back-suction mixing method provided in the embodiment of the present invention. The liquid mixing effect can be further improved by selecting a first pipette with a larger capacity. Specifically, refer to Figure 6 , Figure 6 is another optional flow chart of the back-absorption mixing method provided in this embodiment. Before step S1, this embodiment of the present invention further includes:

[0071] Step S11: Select a first pipette with a larger capacity.

[0072] Furthermore, the present invention also provides a method for purifying nucleic acid using magnetic beads. Figure 7 and Figure 8 , Figure 7 : is a schematic diagram of the process of the nucleic acid magnetic bead purification method provided in this embodiment, Figure 8 This is a schematic diagram of the internal structure of the instrument during nucleic acid magnetic bead purification. Table 1 below is Figure 8 The reagent wells and their corresponding reagent names are shown in the table. As shown in the figure, the method includes:

[0073] Step 100: using the back-absorption mixing method provided in an embodiment of the present invention to mix the magnetic beads, the nucleic acid and the first reagent;

[0074] Specifically, the first reaction well P1 is mixed by pipetting and beating using the reverse suction mixing method, wherein the position of the pipette tip is None, and incubated for a certain time.

[0075] Step 200: removing the supernatant and retaining the magnetic beads adsorbed with nucleic acid;

[0076] Remove the supernatant with the pipette tip in the Away position to avoid removing the magnetic beads when removing impurities.

[0077] Step 300: using the reverse aspiration and mixing method provided in an embodiment of the present invention, the magnetic beads, the nucleic acid and the second reagent are mixed to separate the magnetic beads from the nucleic acid;

[0078] Specifically, the second capture reagent is transferred from well 13 to the fifth capture reagent B5, and the mixture is mixed by blowing and beating using the reverse suction mixing method, wherein the position of the pipette tip is None; the second reagent here is the capture reagent mix1 prepared by the second capture reagent and the fifth capture reagent. Transfer the capture reagent mix1 from B5 to the first reaction well P1; mix by blowing and beating using the reverse suction mixing method, wherein the position of the pipette tip is Toward. At this time, the mixing volume is small and the liquid viscosity is high. Using the reverse suction mixing method and setting the position of the pipette tip to Toward can better mix the magnetic beads and separate the nucleic acid from the magnetic beads.

[0079] Step 400: Transferring the supernatant, which contains the purified nucleic acid.

[0080] Specifically, the supernatant is transferred to the second reaction well P2, wherein the position of the pipette tip is Away, that is, the purified nucleic acid is transferred to the second reaction well P2.

[0081] Table 1

[0082] Reagent wells Reagent name Reagent wells Reagent name 1 First end fill-in reagent 35 Second capture wash reagent 2 Second end fill-in reagent 36 Third capture wash reagent 3 First ligation reagent 37 Fourth capture wash reagent 4 Secondary ligation reagent 38 Fifth magnetic bead washing reagent 5 Third Linking Reagent 39 Second library elution reagent 6 Fourth Linking Reagent 40 Fourth blocking reagent 7 First amplification reagent 45 Waste hole 8 First library elution reagent 46 Waste hole 10 First blocking reagent 47 Waste hole 11 Secondary blocking reagent 48 Waste hole 12 First capture reagent 49 Fourth washing reagent 13 Second capture reagent 50 Fifth washing reagent 14 The third capture agent 51 Fifth washing reagent 15 Fourth capture reagent 52 Seventh washing reagent 16 Second amplification primer P1 First reaction well 17 Secondary amplification reagent P2 Second reaction well 18 The third blocking reagent P3 Second capture wash reagent 26 First purification reagent P4 Fifth capture wash reagent 27 First magnetic bead washing reagent P5 Fifth capture wash reagent 28 Second Magnetic Bead Washing Reagent B1 Library storage well 29 Third magnetic bead washing reagent B2 First magnetic beads 30 Fourth magnetic bead washing reagent B3 First amplification primer 31 First wash reagent B4 Second magnetic bead 32 Second washing reagent B5 Fifth capture agent 33 Third washing reagent B6 Capture beads 34 First capture wash reagent B8 The third magnetic bead

[0083] refer to Figure 8-Figure 9 , Fig. 9 1 is another optional flow chart of the method for purifying nucleic acid with magnetic beads provided in this embodiment. Before step 100, the embodiment of the present invention further comprises:

[0084] Step S110: injecting nucleic acid into the first reaction well P1;

[0085] A sample is transferred from the outside to the first reaction well P1, wherein the sample can be understood to contain nucleic acid.

[0086] Step S120: transferring the first capture reagent (first reagent) into the first reaction well P1;

[0087] Specifically, the first capture reagent can be transferred from well 12 to the first reaction well P1.

[0088] Step S130: mixing the magnetic bead reagent by pipetting and mixing using a reverse suction mixing method;

[0089] Specifically, the Beads turntable was rotated, and the B4 magnetic beads were mixed by blowing using the reverse suction mixing method, wherein the position of the pipette tip was None.

[0090] Step S140: Transfer the magnetic beads to the first reaction well P1.

[0091] Specifically, beads are transferred from well B4 to the first reaction well P1.

[0092] refer to Figure 8 and Fig.10 , Fig.10 This is another optional flow chart of the method for purifying nucleic acid with magnetic beads provided in this embodiment. After step 100 and before step 200, this embodiment of the present invention further includes:

[0093] Step S101: The magnet is brought close to the reaction well;

[0094] Specifically, the turntable is rotated to the position of the magnetic module, and the magnetic module is placed under the first reaction well P1, wherein the magnetic module is a module with a magnet. The magnet outside the first reaction well P1 is brought close to the first reaction well P1, so that the magnet attracts the magnetic beads adsorbed with nucleic acid in the reagent well, and is left to stand for a certain period of time.

[0095] Step S102: removing the supernatant containing other impurities;

[0096] When removing supernatant containing other impurities, the position of the pipette tip is Away;

[0097] Step S103: adding magnetic bead washing reagent;

[0098] Specifically, the fourth magnetic bead washing reagent is transferred from 30 to the first reaction well P1, wherein the position of the pipette tip is Away, and the mixture is left to stand for a certain period of time.

[0099] refer to Figure 8 and Fig.11 , Fig.11This is another optional flow chart of the method for purifying nucleic acid with magnetic beads provided in this embodiment. After step 200 and before step 300, this embodiment of the present invention further includes:

[0100] Step S201: The magnet leaves the reaction hole.

[0101] Specifically, the magnetic module leaves the first reaction well P1 and stands for a certain period of time;

[0102] refer to Figure 8 and Fig.12 , Fig.12 This is another optional flow chart of the method for purifying nucleic acid with magnetic beads provided in this embodiment. After step 300 and before step 400, this embodiment of the present invention further includes:

[0103] Step S301: The magnet is close to the reaction hole.

[0104] Specifically, the turntable is rotated to the magnetic module position, and the magnetic module is placed below the first reaction well P1. The magnet outside the first reaction well P1 is brought close to the first reaction well P1, so that the magnet attracts the magnetic beads. It should be noted that no nucleic acid is adsorbed on the magnetic beads at this time, and it is left to stand for a certain period of time.

[0105] When the pipette tip is used to transfer various liquids in a reagent tube, the reagents can be mixed in the reagent tube. The reagents can be mixed first and then transferred, or directly transferred, or only mixed.

[0106] To solve the aforementioned problem, the present invention also provides a pipetting method, comprising: mixing the liquid in the liquid holding tube using the back-suction mixing method provided in an embodiment of the present invention; wherein, after executing the step of lifting the pipette tip from the liquid in the liquid holding tube, moving the pipette to the target container, pushing the piston to the first position, and obtaining a second preset pipetting volume; removing the pipette, pushing the piston to the second position, and blowing out the residual liquid.

[0107] refer to Fig.13 , Fig.13 Schematic diagram of the steps of a liquid transfer method provided in this embodiment. As shown in the figure, the method includes:

[0108] Step S51: moving the pipette gun to the target container and pushing the piston to the first position;

[0109] Push the piston to the first position to obtain a preset pipetting volume.

[0110] Step S52: remove the pipette gun and push the piston to the second position.

[0111] Push the piston to the second position to blow out the remaining liquid.

[0112] The above describes multiple implementation schemes provided by the embodiments of the present invention. The various optional methods introduced in the implementation schemes can be combined and cross-referenced with each other without conflict, thereby extending a variety of possible implementation schemes, which can all be considered as implementation schemes disclosed and open in the embodiments of the present invention.

[0113] Although the embodiments of the present invention are disclosed above, the present invention is not limited thereto. Any person skilled in the art can make various changes and modifications without departing from the spirit and scope of the present invention. Therefore, the protection scope of the present invention shall be subject to the scope defined by the claims.

Claims

1. A back suction mixing method, characterized in that: Applied to a pipette, the pipette comprises a cavity and a piston, a pipette head is arranged at one end of the cavity, and the piston is arranged at the other end, the moving position of the piston in the cavity comprises an initial position, a first position and a second position, the distance between the initial position and the second position is greater than the distance between the initial position and the first position, and the method comprises: pushing the piston from the initial position to the second position; Dipping the pipette tip of the pipette into the liquid in the liquid holding tube and releasing the piston to aspirate part of the liquid; Pushing the piston to reciprocate for a preset number of times to mix the liquid in the liquid holding tube, wherein, in the process of pushing the piston to push the liquid out of the pipette tip, the piston remains at the first position away from the side of the pipette tip; Move the pipette tip out of the liquid holding tube.

2. The method according to claim 1, characterized in that The step of moving the pipette tip out of the liquid holding tube comprises: Release the piston to absorb part of the liquid, and lift the pipette tip out of the liquid in the liquid holding tube; Pushing the piston to the first position to obtain a preset pipetting volume; Pushing the piston to the second position to blow out the remaining liquid; Remove the emptied pipette from the liquid holding tube.

3. The method according to claim 1, characterized in that In the step of pushing the piston to reciprocate for a preset number of times to mix the liquid in the liquid holding tube, when the piston pushes out the liquid, the pipette moves to hit the liquid.

4. The method according to claim 1, characterized in that The position of the pipette tip in the liquid holding tube is: a first holding position, which is located at the center of the liquid holding tube; or a second holding position, which is located at a side of the liquid holding tube close to the magnet; or a third holding position, which is located at a side of the liquid holding tube away from the magnet.

5. A nucleic acid processing instrument, characterized in that: include: A pipette and a main control unit coupled to the pipette, wherein the main control unit is used to issue instructions to control the pipette to execute the back-suction mixing method according to any one of claims 1 to 4.

6. The nucleic acid processing apparatus according to claim 5, characterized in that: The pipette includes at least a first pipette and a second pipette, the capacity of the first pipette is greater than the capacity of the second pipette, wherein the first pipette is used to perform the back-suction mixing method described in any one of claims 1-4.

7. A method for purifying nucleic acid using magnetic beads, characterized in that: include: Using the back-absorption mixing method described in any one of claims 1 to 4, the magnetic beads, the nucleic acid and the first reagent are mixed; Remove the supernatant and retain the magnetic beads adsorbed with nucleic acid; Adopting the back-absorption mixing method described in any one of claims 1 to 4, the magnetic beads, the nucleic acid and the second reagent are mixed to separate the magnetic beads from the nucleic acid; The supernatant containing the purified nucleic acid is transferred.

8. The method according to claim 7, characterized in that In the step of mixing the magnetic beads, the nucleic acid and the first reagent, the position of the pipette tip is a first accommodation position, and the first accommodation position is located at the center of the liquid accommodation tube.

9. The method according to claim 7, characterized in that In the step of mixing the magnetic beads, nucleic acid and the second reagent, the position of the pipette tip is the second accommodation position, and the second accommodation position is located on a side of the liquid accommodation tube close to the magnet.

10. The method according to claim 7, characterized in that In the step of removing the supernatant and the step of transferring the supernatant, the position of the pipette tip is a third accommodating position, and the third accommodating position is located on a side of the liquid containing tube away from the magnet.

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