CN111349629A - Cell tissue nucleic acid extraction method for gene sequencing - Google Patents
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Abstract
The invention discloses a cell tissue nucleic acid extraction method for gene sequencing, which comprises the following specific steps: s1, placing animal tissues into a centrifuge tube, adding STE buffer solution, and mixing by using a vortex oscillator to obtain a solution I; s2, adding a sodium dodecyl sulfate solution and proteinase K into the solution I, putting the solution I into an overturning centrifugal device, and uniformly mixing to obtain a solution II; s3, cooling the centrifugal tube to room temperature, adding a mixed solution of Tris-saturated phenol, chloroform and isoamylol into the solution II, turning over, shaking up and centrifuging to obtain a solution III; s4, transferring the supernatant in the solution III to another centrifugal tube, adding ice absolute ethyl alcohol, separating out a cluster, sucking the cluster into a clean centrifugal tube by using a liquid transfer gun, washing twice by using 75% ethyl alcohol, and airing to obtain purified nucleic acid; by using the turnover centrifugal device provided by the invention, the centrifugal tube can be conveniently rotated in the vertical direction and can also be rotated in the horizontal direction, so that the effects of shaking up and centrifuging a sample are achieved.
Description
Technical Field
The invention relates to the technical field of nucleic acid purification, in particular to a cell tissue nucleic acid extraction method for gene sequencing.
Background
DNA and RNA samples used for gene sequencing often depend on obtaining purified nucleic acid samples. Sources of DNA or RNA are numerous, such as tissues, cells, whole blood, plasma, serum, extracts, which may be obtained from an organism (e.g., human, animal, plant, bacteria, virus, etc.), and purification of nucleic acids typically requires separation of the DNA or RNA from other components in the sample source to render the polynucleic acids free of other cellular enzymes, proteins, compounds, and/or debris.
One of the earliest techniques for purifying DNA or RNA was to lyse a cell or tissue sample using, for example, proteinase K, add an equal volume of phenol chloroform solution to the lysate to extract non-nucleic acid material (e.g., proteins, organelles, etc.), and remove the aqueous phase containing the nucleic acids for further use. Nucleic acids are generally recovered from the aqueous phase by precipitation with ethanol or isopropanol. Improvements in this technique include the further addition of small amounts of isoamyl alcohol to the organic phase to aid in the inactivation of RNases. Although phenol chloroform extraction can achieve high purity and high recovery of nucleic acids, the method has disadvantages such as the need for repeated centrifugation and tumble mixing of nucleic acid samples during the operation, and currently there is no device available for both purposes.
Disclosure of Invention
The invention aims to provide a method for extracting cell tissue nucleic acid for gene sequencing, which can effectively purify nucleic acid by a method for extracting nucleic acid by phenol chloroform; by using the turnover centrifugal device provided by the invention, the centrifugal tube can be conveniently rotated in the vertical direction, so that samples in the centrifugal tube can be fully and uniformly mixed, and the centrifugal tube can also be rotated in the horizontal direction, thereby achieving the centrifugal effect.
The purpose of the invention can be realized by the following technical scheme:
a cell tissue nucleic acid extraction method for gene sequencing comprises the following specific steps:
s1, placing animal tissues in a centrifuge tube, adding STE buffer solution, mixing for 0.5-2h by using a vortex oscillator to fully mix tissue cells with the STE buffer solution to obtain a solution I, and adding 8-12mLSTE buffer solution into each gram of animal tissues;
s2, adding 10% sodium dodecyl sulfate solution and 10 mg/mu L proteinase K into the first solution, sealing the opening with a sealing film, placing the solution into a centrifugal tube groove of a turnover centrifugal device, covering a fixed cover, locking the fixed cover with a locking rod, starting an electric heating wire and a fan, heating to 35-50 ℃, starting a second motor, and turning over the centrifugal tube for 5-10 hours to obtain a second solution;
s3, cooling the centrifugal tube to room temperature, adding Tris-saturated phenol into the solution II, placing the solution II in an overturning centrifugal device, starting a second motor, overturning and shaking uniformly for 0.5-1h, adding a mixed solution of chloroform and isoamylol, continuously overturning and shaking uniformly for 0.5-1h, then closing the second motor, starting a first motor, and centrifuging for 15-30 min to obtain a solution III;
s4, transferring the supernatant in the solution III to another centrifuge tube, adding absolute ethyl alcohol at the temperature of-20 to-10 ℃, standing for 15-30 min, separating out a cluster, sucking the cluster into a clean centrifuge tube by using a liquid transfer gun, and washing twice by using 75% ethyl alcohol to obtain the purified nucleic acid.
Phenol is a strong protein denaturant, can effectively denature and remove proteins in animal tissues, chloroform has strong fat solubility and can remove lipid impurities in the animal tissues, phenol and water have high intersolubility, if phenol is used alone to extract the proteins, a large amount of phenol is dissolved into a water phase after extraction, phenol can inhibit a large number of enzyme reactions in subsequent operations, and the residual amount of phenol can be greatly reduced when phenol and chloroform are mixed for use; the addition of isoamyl alcohol can reduce the surface tension between interfaces, thereby reducing bubbles generated in the process of protein denaturation operation, and in addition, the isoamyl alcohol is helpful for phase separation of the mixed solution after centrifugation; the sodium dodecyl sulfate solution can dissolve lipids and proteins on cell membranes, so that the cell membranes are damaged, nucleoprotein in cells is dissociated, and the sodium dodecyl sulfate can be combined with the proteins to precipitate, so that the effect of separating the proteins is achieved; proteinase K hydrolyzes digestive proteins, particularly histones bound to DNA, and thus achieves protein removal.
Preferably, in step S2, the turning centrifugal device includes a housing, two first rotating shafts are fixedly connected to a bottom end surface of the housing, the first rotating shafts are rotatably connected to belt pulleys, the belt pulleys are connected to each other by a belt transmission, a first motor is disposed above one of the belt pulleys, a driving shaft of the first motor is fixedly connected to a top end surface of the belt pulley, a connecting block is fixedly connected to a top end surface of the other belt pulley, a supporting plate is fixedly connected to a top end surface of the connecting block, two electric telescopic rods are respectively disposed at two ends of the supporting plate, a rotating block is fixedly connected to one of telescopic ends of the two electric telescopic rods, a second motor is fixedly connected to the other of the telescopic ends of the two electric telescopic rods, a centrifuge tube frame is rotatably connected between the rotating block and the second motor, a driving shaft of the second motor, the centrifugal tube rack is provided with the electric telescopic rod, the centrifugal tube rack can be extended when needing to be turned, the end part of the centrifugal tube rack cannot touch the supporting plate and the sampling cover, and the electric telescopic rod is shortened when a sample needs to be centrifuged, so that the resistance is reduced, and the centrifugation process is more stable;
the top end face of the shell is provided with a notch, a sampling cover is arranged on the notch and is rotationally connected to the top end face of the shell through a second rotating shaft, the inner side wall of the top end face of the shell is fixedly connected with a vertical clapboard, the lower end of the vertical clapboard extends to the lower end in the shell and is fixedly connected with a first horizontal clapboard, one end of the vertical clapboard is fixedly connected to the lower end of the inner side wall of the shell, a connecting block penetrates through the first horizontal clapboard, the upper end of the vertical clapboard is provided with an air outlet, one side of the air outlet, away from the centrifuge tube rack, is fixedly connected with a heating wire, one side of the heating wire is provided with a fan, the fan is fixedly connected to the inner side wall of the shell, the side wall of the shell, which is positioned at one side of the fan, is provided with a ventilation grid, through setting up heating wire and fan, can rise the device internal temperature, satisfy the best active temperature who uses proteinase K in purification process, through setting up the ventilation grid, can dispel the heat in the device, avoid the heating wire to cause the condition of local overheat.
Preferably, the pulley groove has been seted up at backup pad bottom end face both ends, and pulley groove inside wall fixedly connected with metal crate installs the pulley in the metal crate, and the pivot of pulley is rotated and is connected on the metal crate inside wall, and the groove that rolls has been seted up to first horizontal baffle top face, and the groove that rolls and the pulley phase-match, through setting up the pulley, can make the backup pad remain stable at the rotation in-process, through setting up the groove that rolls, makes the pulley at the inslot motion that rolls, makes the backup pad rotate more stably.
Preferably, the centrifuge tube rack includes chassis and fixed lid, and fixed lid block is on chassis top face, and chassis top face is opened has a plurality of centrifuge tube groove, and chassis top face rotates and is connected with the check lock lever, through setting up fixed lid and check lock lever, makes the centrifuging tube fix at the centrifuge tube inslot, and the centrifuging tube falls out from the centrifuge tube rack when preventing upset or centrifugal rotation.
Preferably, the volume ratio of the sodium dodecyl sulfate solution to the proteinase K to the first solution in the step S2 is 0.1-0.3: 0.02-0.05: 1.
preferably, the volume ratio of Tris-saturated phenol, chloroform and isoamylol in step S3 is 25-30: 20-24: 1.
the invention has the beneficial effects that:
the nucleic acid can be effectively purified by a method for extracting the nucleic acid by phenol chloroform, phenol is a strong protein denaturant and can effectively denature and remove proteins in animal tissues, chloroform has strong fat solubility and can remove lipid impurities in the animal tissues, phenol and water have large mutual solubility, if phenol is used alone to extract the proteins, a large amount of phenol is dissolved in a water phase after extraction, phenol can inhibit a plurality of enzyme reactions in subsequent operations, and phenol and chloroform are mixed for use, so that the residual amount of phenol can be greatly reduced; the addition of isoamyl alcohol can reduce the surface tension between interfaces, thereby reducing bubbles generated in the process of protein denaturation operation, and in addition, the isoamyl alcohol is helpful for phase separation of the mixed solution after centrifugation; the sodium dodecyl sulfate solution can dissolve lipids and proteins on cell membranes, so that the cell membranes are damaged, nucleoprotein in cells is dissociated, and the sodium dodecyl sulfate can be combined with the proteins to precipitate, so that the effect of separating the proteins is achieved; proteinase K hydrolyzes and digests proteins, especially histones combined with DNA, thereby achieving the effect of removing proteins;
by using the overturning centrifugal device provided by the invention, the centrifugal tube can be conveniently rotated in the vertical direction, so that samples in the centrifugal tube are fully and uniformly mixed, and the centrifugal tube can also be rotated in the horizontal direction, so that a centrifugal effect is achieved; the sampling cover is arranged and is rotatably connected to the shell through the second rotating shaft, so that sampling and sample loading operations can be facilitated, the temperature in the device can be raised through the arrangement of the heating wire and the fan, the optimal active temperature of protease K used in the purification process is met, the device can be internally cooled through the arrangement of the ventilation grating, and the situation that the heating wire causes local overheating is avoided; the pulley is arranged, so that the support plate can be kept stable in the rotating process, and the pulley moves in the rolling groove by arranging the rolling groove, so that the support plate is more stable to rotate; through setting up fixed lid and check lock lever, make the centrifuging tube fix at the centrifuging tube inslot, the centrifuging tube falls out from the centrifuge tube rack when preventing upset or centrifugation rotation.
Drawings
In order to facilitate understanding for those skilled in the art, the present invention will be further described with reference to the accompanying drawings.
FIG. 1 is a schematic side view of the present invention;
FIG. 2 is a side view of the front structure of the present invention;
FIG. 3 is an enlarged view of the detail of area A in FIG. 1;
FIG. 4 is an enlarged view of the detail of the area B in FIG. 1;
in the figure: 1. a housing; 2. a first rotating shaft; 3. a belt pulley; 4. a belt; 5. a first motor; 6. connecting blocks; 7. a support plate; 8. an electric telescopic rod; 9. rotating the block; 10. a centrifuge tube rack; 11. a second motor; 12. a sampling cover; 13. a second rotating shaft; 14. a vertical partition; 15. a first horizontal partition; 16. an air outlet; 17. an electric heating wire; 18. a fan; 19. a ventilation grille; 20. a second horizontal partition; 21. a pulley groove; 22. a metal frame; 23. a pulley; 24. a rolling groove; 25. a chassis; 26. a fixed cover; 27. a centrifugal pipe groove; 28. a locking lever.
Detailed Description
The technical solutions of the present invention will be described clearly and completely with reference to the following embodiments, and it should be understood that the described embodiments are only a part of the embodiments of the present invention, and not all of the embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Please refer to FIGS. 1-4
A cell tissue nucleic acid extraction method for gene sequencing comprises the following specific steps:
s1, taking 0.05g of animal tissue to be placed in a centrifuge tube, adding 0.5mL of STE buffer solution, mixing for 0.5h by using a vortex oscillator, and fully mixing the histiocytes and the STE buffer solution;
s2, adding 0.1mL of 10% sodium dodecyl sulfate solution and 20 μ L of 2mg/mL proteinase K into the first solution, sealing the first solution by using a sealing film, placing the first solution into a centrifugal tube groove 27 of an overturning centrifugal device, covering a fixing cover 26, locking the fixing cover 26 by using a locking rod 26, starting an electric heating wire 17 and a fan 18, heating the first solution to 37 ℃, starting a second motor 11, overturning the centrifugal tube for 5 hours to obtain a second solution, wherein the sodium dodecyl sulfate solution can dissolve lipids and proteins on cell membranes, so that the cell membranes are damaged, nucleoproteins in the cells are dissociated, and the sodium dodecyl sulfate can be combined with the proteins to precipitate, so that the effect of separating the proteins is achieved; proteinase K hydrolyzes and digests proteins, especially histones combined with DNA, thereby achieving the effect of removing proteins;
s3, cooling the centrifugal tube to room temperature, adding 250 mu L of Tris-saturated phenol into the solution II, placing the solution II in an overturning centrifugal device, starting the second motor 11, overturning and shaking uniformly for 0.5h, adding a mixed solution of 240 mu L of chloroform and 10 mu L of isoamyl alcohol, continuously overturning and shaking uniformly for 0.5h, then closing the second motor 11, starting the first motor 5, and centrifuging for 15min to obtain a solution III, wherein phenol is a strong protein denaturant and can effectively denature and remove proteins in animal tissues, chloroform has strong lipid solubility and can remove lipid impurities in animal tissues, phenol and water have high intersolubility, if phenol is used alone to extract proteins, a large amount of phenol is dissolved in a water phase after extraction, phenol can inhibit a large number of enzyme reactions in subsequent operations, phenol and chloroform are mixed for use, the residual amount of phenol can be greatly reduced, isoamyl alcohol is added, and the surface tension between interfaces can be reduced, thereby reducing bubbles generated in the protein denaturation operation process, and in addition, isoamylol is helpful for phase separation of the mixed solution after centrifugation;
s4, transferring the supernatant in the solution III to another centrifuge tube, adding absolute ethyl alcohol at the temperature of-20 ℃, standing for 30min, sucking the separated cluster-shaped objects into a clean centrifuge tube by using a pipette gun, and washing twice by using 75% ethyl alcohol to obtain the purified nucleic acid.
The overturning centrifugal device comprises a shell 1, two first rotating shafts 2 are fixedly connected to the bottom end face of the shell 1, belt pulleys 3 are rotatably connected to the first rotating shafts 2, the belt pulleys 3 are in transmission connection through a belt 4, a first motor 5 is arranged above one belt pulley 3, a driving shaft of the first motor 5 is fixedly connected to the top end face of the belt pulley 3, a connecting block 6 is fixedly connected to the top end face of the other belt pulley 3, a supporting plate 7 is fixedly connected to the top end face of the connecting block 6, two electric telescopic rods 8 are fixedly connected to the top end face of the supporting plate 7, the two electric telescopic rods 8 are respectively positioned at two ends of the supporting plate 4, a rotating block 9 is fixedly connected to the telescopic ends of the two electric telescopic rods 8, a second motor 11 is fixedly connected to the other end face of the two electric telescopic rods, a centrifugal pipe frame 10 is rotatably connected between the, by arranging the first motor 5, the first motor 5 is started to drive the belt pulley 3 and the belt 4 to rotate, so that the support plate 4 is driven to rotate, the centrifuge tube rack 10 is driven to rotate, a centrifugal effect is achieved, by arranging the rotating block 9 and the second motor 11, the centrifuge tube rack 10 can rotate in the vertical direction, so that the effect of uniformly shaking the centrifuge tube in a turnover manner can be realized, by arranging the electric telescopic rod 8, the centrifuge tube rack 10 can be extended when needing to be turned over, the end part of the centrifuge tube rack 11 cannot touch the support plate 7 and the sampling cover 12, and when a sample needs to be centrifuged, the centrifuge tube rack is shortened, so that the resistance is reduced, and the centrifugation process is more stable;
a gap is arranged on the top end face of the shell 1, a sampling cover 12 is arranged on the gap, the sampling cover 12 is rotatably connected on the top end face of the shell 1 through a second rotating shaft 13, a vertical clapboard 14 is fixedly connected on the inner side wall of the top end face of the shell 1, the lower end of the vertical clapboard 14 extends to the lower end in the shell 1 and is fixedly connected with a first horizontal clapboard 15, one end of the vertical clapboard 14 is fixedly connected at the lower end of the inner side wall of the shell 1, a connecting block 6 is arranged by penetrating through the first horizontal clapboard 15, an air outlet 16 is arranged at the upper end of the vertical clapboard 14, one side of the air outlet 16, which is far away from the centrifuge tube rack 10, is fixedly connected with an electric heating wire 17, a fan 18 is arranged at one side of the electric heating wire 17, the fan 18 is fixedly connected on the inner side, through setting up sample lid 12 to rotating sample lid 12 through second pivot 13 and connecting on shell 1, can conveniently take a sample and adorn a kind operation, through setting up heating wire 17 and fan 18, can rise the temperature in the device, satisfy the best active temperature who uses proteinase K in the purification process, through setting up ventilation grid 19, can dispel the heat in the device, avoid heating wire 17 to cause the local overheated condition.
The use method of the turnover centrifugal device comprises the following steps: firstly, opening a sampling cover 12, extending an electric telescopic rod 8 to enable a centrifuge tube rack 10 to reach a position convenient for placing a centrifuge tube, placing the centrifuge tube into a centrifuge tube groove 27, covering a fixed cover 26, locking the fixed cover 26 by a locking rod 28, shortening the electric telescopic rod 8 to a proper position, starting a second motor 11, enabling the centrifuge tube rack 10 to rotate on a vertical plane, thereby achieving the purpose of uniformly mixing liquid in the centrifuge tube, when centrifugal operation is needed, controlling the second motor 11 to enable the centrifuge tube rack 10 to turn over until the fixed cover 26 faces upwards, then shortening the electric telescopic rod 8 to the shortest, starting a first motor 5, enabling the first motor 5 to drive a belt pulley 3 to rotate, enabling another belt pulley 3 to rotate through transmission of a belt 4, enabling a supporting plate 7 to rotate through a connecting block 6, thereby enabling the centrifuge tube rack 10 to rotate to achieve the centrifugal purpose, and enabling a pulley 23 in a pulley groove 21 to roll in a rolling groove 24 in the process, the supporting plate 7 is supported and stabilized; when the overturning or centrifugal function is used, the heating wire 17 and the fan 18 can be started, the temperature in the device is raised through hot air, so that the temperature reaches the highest temperature of the activity of the reagent enzyme, and the fan 18 can be independently started to cool the interior of the equipment after the use is finished.
The preferred embodiments of the invention disclosed above are intended to be illustrative only. The preferred embodiments are not intended to be exhaustive or to limit the invention to the precise forms disclosed. Obviously, many modifications and variations are possible in light of the above teaching. The embodiments were chosen and described in order to best explain the principles of the invention and the practical application, to thereby enable others skilled in the art to best utilize the invention. The invention is limited only by the claims and their full scope and equivalents.
Claims (6)
1. A cell tissue nucleic acid extraction method for gene sequencing is characterized by comprising the following specific steps:
s1, placing animal tissues into a centrifuge tube, adding STE buffer solution, mixing for 0.5-2h by using a vortex oscillator to obtain a solution I, and adding 1mLSTE buffer solution into each mg of animal tissues;
s2, adding 10% sodium dodecyl sulfate solution and 10 mg/mu L proteinase K into the first solution, sealing the first solution by using a sealing film, placing the first solution into a centrifugal tube groove (27) of an overturning centrifugal device, covering a fixed cover (26), locking the fixed cover (26) by using a locking rod (26), starting an electric heating wire (17) and a fan (18), heating to 35-50 ℃, starting a second motor (11), and overturning the centrifugal tube for 5-10 hours to obtain a second solution;
s3, cooling the centrifugal tube to room temperature, adding Tris-saturated phenol into the solution II, placing the solution II in an overturning centrifugal device, starting a second motor (11), overturning and shaking uniformly for 0.5-1h, adding a mixed solution of chloroform and isoamylol, continuously overturning and shaking uniformly for 0.5-1h, then closing the second motor (11), starting a first motor (5), and centrifuging for 15-30 min to obtain a solution III;
s4, transferring the supernatant in the solution III to another centrifuge tube, adding absolute ethyl alcohol at the temperature of minus 20 to minus 10 ℃, starting a first motor (5) for centrifugation for 15 to 30min, separating out a cluster, sucking the cluster into a clean centrifuge tube by using a pipette gun, washing the cluster twice by using 75% of ethyl alcohol, and airing to obtain the purified nucleic acid.
2. The method for extracting nucleic acid from cell tissues for gene sequencing according to claim 1, wherein the turning centrifugal device in step S2 comprises a housing (1), two first rotating shafts (2) are fixedly connected to the bottom end face of the housing (1), belt pulleys (3) are rotatably connected to the first rotating shafts (2), the belt pulleys (3) are in transmission connection through belts (4), a first motor (5) is arranged above one belt pulley (3), a driving shaft of the first motor (5) is fixedly connected to the top end face of the belt pulley (3), a connecting block (6) is fixedly connected to the top end face of the other belt pulley (3), a supporting plate (7) is fixedly connected to the top end face of the connecting block (6), two electric telescopic rods (8) are fixedly connected to the top end face of the supporting plate (7), and the two electric telescopic rods (8) are respectively located at two ends of the supporting plate (4), one telescopic end of each of the two electric telescopic rods (8) is fixedly connected with a rotating block (9), the other telescopic end of each electric telescopic rod is fixedly connected with a second motor (11), a centrifuge tube rack (10) is rotatably connected between the rotating block (9) and the second motor (11), and a driving shaft of the second motor (11) is fixedly connected with the side wall of the centrifuge tube rack (10);
a gap is formed in the top end face of the shell (1), a sampling cover (12) is arranged on the gap, the sampling cover (12) is rotatably connected to the top end face of the shell (1) through a second rotating shaft (13), a vertical partition plate (14) is fixedly connected to the inner side wall of the top end face of the shell (1), the lower end of the vertical partition plate (14) extends to the inner lower end of the shell (1), a first horizontal partition plate (15) is fixedly connected to the lower end of the inner side wall of the shell (1), a connecting block (6) penetrates through the first horizontal partition plate (15), an air outlet (16) is formed in the upper end of the vertical partition plate (14), an electric heating wire (17) is fixedly connected to one side, away from the centrifuge tube rack (10), a fan (18) is arranged on one side of the electric heating wire (17), the fan (18) is fixedly connected to the inner side wall of the shell (1), a ventilation grating, the electric heating wire (17) and the fan (18) are positioned above the first motor (5), and a second horizontal clapboard (20) is arranged in the middle.
3. The method for extracting nucleic acid from cell tissues for gene sequencing according to claim 2, wherein pulley grooves (21) are formed at two ends of the bottom end face of the support plate (7), the inner side walls of the pulley grooves (21) are fixedly connected with metal frames (22), pulleys (23) are installed in the metal frames (22), rotating shafts of the pulleys (23) are rotatably connected to the inner side walls of the metal frames (22), rolling grooves (24) are formed in the top end face of the first horizontal partition plate (15), and the rolling grooves (24) are matched with the pulleys (23).
4. The method for extracting nucleic acid from cell tissues for gene sequencing according to claim 2, wherein the centrifuge tube rack (10) comprises a bottom frame (25) and a fixing cover (26), the fixing cover (26) is clamped on the top end surface of the bottom frame (25), a plurality of centrifuge tube slots (27) are formed on the top end surface of the bottom frame (25), and a locking rod (28) is rotatably connected to the top end surface of the bottom frame (25).
5. The method for extracting nucleic acid from cell tissues for gene sequencing according to claim 2, wherein the volume ratio of the sodium dodecyl sulfate solution to the proteinase K to the first solution in step S2 is 0.1-0.3: 0.02-0.05: 1.
6. the method for extracting nucleic acid from cell tissues for gene sequencing according to claim 2, wherein the volume ratio of Tris-saturated phenol, chloroform and isoamyl alcohol in the step S3 is 25-30: 20-24: 1.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN112175812A (en) * | 2020-10-13 | 2021-01-05 | 山西农业大学 | Fluid control device for researching genetic diversity of pigs based on high-throughput gene sequencing |
| CN115463588A (en) * | 2022-08-24 | 2022-12-13 | 中启源(宁夏)生物科技有限公司 | 3D rotation type thermostated container suitable for multiple tissue digestion preparation mesenchymal stem cell |
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