CN102824943A - Universal cell rapid breaker using bead vibrating and grinding - Google Patents
Universal cell rapid breaker using bead vibrating and grinding Download PDFInfo
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- CN102824943A CN102824943A CN2012103017880A CN201210301788A CN102824943A CN 102824943 A CN102824943 A CN 102824943A CN 2012103017880 A CN2012103017880 A CN 2012103017880A CN 201210301788 A CN201210301788 A CN 201210301788A CN 102824943 A CN102824943 A CN 102824943A
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Abstract
A universal cell rapid breaker using bead vibrating and grinding mainly comprises a main shell, a driving module, a control module, a power shaft, a rotor, a transmission mechanism, centrifuge tube supports, centrifuge tubes and grinding beads. The driving module fixed on the main shell provides power source for the whole breaker under control of the control module, and transmits motion and torque to the rotor through the power shaft so as to force the rotor to rotate with a high speed. Initiating terminal of the transmission mechanism is connected with the rotating rotor through a higher pair, and transforms the high-speed rotary movement of the rotor to a high-frequency reciprocating linear movement of the terminal of the transmission mechanism. The centrifuge tube supports are fixed on at the terminal of the transmission mechanism and do the high-frequency reciprocating linear movement along with the terminal of the transmission mechanism. The centrifuge tubes with the grinding beads and cell suspension are put in and fixed on the centrifuge tube supports, and do the high-frequency reciprocating linear movement along with the centrifuge tube supports, so that the grinding beads vibrate in a high frequency, and mutually violently shear and impact with cells, thereby resulting in cells disruption to release intracellular active substances.
Description
Technical field
The invention belongs to the biological specimen preparing technical field, be applicable to the clasmatosis in the genetic and cell engineering, specifically is to utilize the quick smudge cells of high frequency flutter grinding bead, to obtain active material in the born of the same parents, like DNA, RNA, enzyme, antibiotic etc.
Background technology
In the genetic and cell engineering technical field, often need obtain intracellular DNA, RNA, enzyme isoreactivity material, i.e. sample preparation.The first step of sample preparation is with extracellular closed structure (cell membrane and cell membrane) fragmentation, and the active material that desire is extracted discharges in cell.This shows that cytoclastic degree and efficient directly affect the quality of sample preparation, even determining the success or failure of sample preparation, its importance is self-evident.
Cytoclastic conventional method mainly contains enzyme and dissolves crush method, chemical reagent crush method, ultrasonic cavitation crush method and bead mill method etc.
It is to utilize the enzyme treated cell of dissolved cell wall earlier that enzyme dissolves crush method, after cell membrane is damaged, utilizes method destruction cell membranes such as osmotic pressure impact again, so that intracellular active material is released.The distinguishing feature that enzyme dissolves crush method is that selectivity is strong, promptly must select specific enzyme according to the structure and the chemical composition of cell, this means that the versatility of the molten crush method of enzyme is very poor.And the price of lyase is higher, has increased the cost of sample preparation to a great extent.
The chemical reagent crush method is to utilize some chemical reagent (like organic solvent, surfactant, metal-chelator etc.), changes the permeability of cell membrane and cell membrane, and some small-molecule active substances infiltrate in the born of the same parents thereby make.The destructiveness of this method pair cell wall and cell membrane is not high, can only obtain some little molecule zymoprotein and polypeptide, and big molecular activity materials such as DNA, RNA still are trapped in the cell.Low through chemical reagent smudge cells length not only consuming time, efficient, and the affiliation that adds of chemical reagent brings difficulty for separation, the purifying of subsequent products, and influence the purity of end product.In addition, some chemical reagent endangers health of operators to toxic elements in human body.
The ultrasonic cavitation crush method is to utilize the ultrasonic Treatment cell suspending liquid, causes the ultrasonic cavitation effect, and local intense impact ripple and the shearing force that produces of suspension makes clasmatosis, and then discharges active material.The ultrasonic cavitation crush method is a kind of strong crumbling method, is applicable to the fragmentation of polytype cell.But the capacity usage ratio of this method is extremely low, can produce great amount of heat in the shattering process, can cause part sensitive materials sex change inactivation, thereby it has quite harsh requirement to cooling.In addition, this method needs supersonic generator, probe, liquid bath and control system, complex structure, and cost is higher, and general is used in the clasmatosis in the laboratory on a small scale.
The bead mill method is a kind of method of cell disruption commonly used, through grinding bead and cell suspending liquid are stirred together fast, makes and shears mutually, collides between grinding bead and the cell, and then with clasmatosis, discharge active material in the born of the same parents.This is the technology of present comparative maturity; The Bullet Blender of U.S. Next Advance company is exactly a based on the theoretical clasmatosis appearance of bead mill, and it adopts the rotary head of rotation at a high speed directly to impact the centrifuge tube that grinding bead is housed and treats smudge cells suspension.In the moment that rotary head impacts, the vibration that the grinding bead in the centrifuge tube will be violent after shearing each other, clash into cell, makes clasmatosis, discharges nucleic acid, protein, subcellular structure isoreactivity material.Though this fragmentation appearance is the cell of broken most of kinds quickly and easily, it still has a lot of defectives and deficiency:
At first, adopt the rotary head of rotation at a high speed directly to impact centrifuge tube, not only can make the centrifuge tube outer wall receive serious wearing and tearing, and centrifuge tube may break or fracture when receiving temporary impact.When particularly some have the microorganism (like gram-positive bacteria, saccharomycete, bacterial spore etc.) of hard outer wall in fragmentation; Rotary head is turned round under higher speed; This moment impact to centrifuge tube is more violent, probably can cause breaking or fractureing of centrifuge tube.
Secondly, the severe of the grinding bead of zones of different vibration is different in the centrifuge tube, causes cytoclastic degree different.Grinding bead vibration near impacting the position is comparatively violent, thereby clasmatosis degree that should the zone is higher; By contrast, because the buffer shock-absorbing effect of liquid, a little less than the grinding bead vibration that impacts the position, because clasmatosis that should the zone maybe be insufficient.
Once more; The rotary head of rotation impacts the moment of centrifuge tube at a high speed; Impact the position and can produce a lot of heats; This part heat is passed to cell suspending liquid through centrifuge tube, can make near the fluid temperature of striking zone to raise, and then possibly cause the sex change inactivations such as subcellular structure, DNA, RNA and enzyme that discharge.
At last, the high speed rotary head directly impacts centrifuge tube, can produce quite ear-piercing noise.Because centrifuge tube is fixed on the broken instrument body, can drives broken instrument body when rotary head impacts centrifuge tube and on workbench, vibrate simultaneously, the body operation is very not steady.In the actual smudge cells process, need operating personnel to push body usually or adopt other frame for movements that body is fixed on the workbench.
In sum; Though at present existing a lot of method of cell disruption; But wherein most methods generally can only broken zooblast, some more breakable cells such as Gram-negative bacteria, usually can not broken gram-positive bacteria, saccharomycete, bacterial spore etc., range of application is quite limited to.Although the bead mill method can adapt to various dissimilar clasmatosises, corresponding apparatus still has a lot of defectives and deficiency, and crushing effect is not really desirable.Therefore, need develop and to adapt to the polytype cell quick breaker of universal cell of (mainly comprising zooblast, Gram-negative bacteria, gram-positive bacteria, saccharomycete, bacterial spore etc.).
Summary of the invention
For solving many defectives of existing method of cell disruption and equipment, theoretical based on bead mill, the present invention provide a kind of simple in structure, easy to operate, operate steadily, lower-cost universal flutter pearl grinds the quick breaker of formula cell.
Technical scheme of the present invention provides the quick breaker of a kind of universal flutter pearl mill formula cell, mainly comprises main casing, driver module, control module, line shaft, rotor, transmission mechanism, centrifuge tube support, centrifuge tube, grinding bead etc.The driver module that is fixed on the main casing provides power source for whole device under the domination of control module, and will move and transmission of torque is given rotor through line shaft, forces the rotor high-speed rotation.The initiating terminal of transmission mechanism links to each other with rotor rotated through higher pair, and the high speed rotary motion of rotor is changed into the terminal high frequency linear reciprocating motion of transmission mechanism.The centrifuge tube support is fixed in the end of transmission mechanism, thereby can do reciprocating linear motion thereupon.The centrifuge tube that grinding bead and cell suspending liquid are housed is placed centrifuge tube support and fixing; Centrifuge tube also can be made the high frequency linear reciprocating motion together, and then causes the flutter of grinding bead high frequency, shears, clashes into cell is fierce each other; Cause clasmatosis, discharge active material in the born of the same parents.
Said main casing is the peripheral structure of device, can not only the insulation blocking effect be provided for interior arrangement, and can also support the man-machine interaction panel.
Said driver module is fixed on the bottom surface of housing, for whole device provides original energy, motion and driving force.In optimized technical scheme, said driver module is the direct current generator of a unidirectional rotation, and the motion and the torque of its output concentrate on the motor output shaft.
Said control module is the control module of apparatus system.In optimized technical scheme, said driver module is a singlechip controller, can be to startup, the time-out of direct current generator, urgency is stopped and parameters such as the motor speed and the duration of runs are controlled.
Said line shaft is for connecting the power transmission shaft of motor output shaft and rotor, and it gives rotor with the rotation and the transmission of torque of motor output shaft, in order to drive the rotor high-speed rotation.In optimized technical scheme, said line shaft is a vertical circular shaft of placing.Circular shaft one end links with motor output shaft through shaft coupling or clutch etc., and the other end then is fixed on the rotor.Owing to bear certain pressure and bigger moment of torsion on the line shaft, so line shaft need have very high intensity, rigidity and toughness, preferred carbon constructional quality steel modifier treatment of its material or manganese alloy quenched and tempered steel etc.
Said rotor rotates at a high speed around a certain vertical center, the initiating terminal of its direct drive transmission device.In optimized technical scheme, said rotor is the cam or the eccentric wheel of a plate-like.During the rotor driven transmission mechanism; Can bear bigger impact, moment of torsion; Therefore the rotor top layer need have higher hardness and wearability preferably; Rotor heart portion need have higher intensity and toughness preferably, and its material preferably has the alloy carburizing steel of middle and high quenching degree, like 20CrMnTi and 20Cr2Ni4 etc.
Said transmission mechanism is the part of transmitting motion and changing forms of motion, specifically is the high frequency linear reciprocating motion that the high speed rotary motion of rotor is changed into the transmission mechanism end, and this is innovation of the present invention place.In optimized technical scheme, said transmission mechanism is a planar linkage mechanism, further is four-bar linkage, forms the rocking bar slide mechanism by transmission housing, rocking bar, depression bar (also can be described as pull bar) and litter through lower pair.Said lower pair comprises revolute pair and moving sets, and wherein revolute pair is to form through hinge, and moving sets is to cooperate through the plane cooperation or the face of cylinder to form.Housing is that whole transmission mechanism provides fixed constraint and support, and depression bar links up rocking bar and litter through hinge.The high speed rotation of rotating cam, the unidirectional swing of drive rocker under the promotion of depression bar, drives litter and moves along straight line is unidirectional, and the direction that the litter straight line moves is controlled through the guide post that is fixed on the housing.In order to guarantee the continuous operation of transmission mechanism ability; Also one group of back-moving spring need be set on guide post; Litter compresses spring when the guide post straight line moves; Litter moves to after range sentences, and compression spring will drive whole transmission mechanism and reset, and so just realize the high frequency linear reciprocating motion of litter.
Said centrifuge tube support is used to support centrifuge tube, and can receive the forms of motion of transmission mechanism.In optimized technical scheme, the centrifuge tube support is fixed in the end of transmission mechanism, promptly on the litter, the centrifuge tube support can be made the high frequency linear reciprocating motion with litter like this.In the reality, can centrifuge tube support and litter be processed into one, this can strengthen the fastness of the two connection.In order centrifuge tube to be fixed in the centrifuge tube support, need to be equipped with gland on the centrifuge tube support.In addition, if active material is very sensitive to temperature in the born of the same parents that desire is extracted, can be at the peripheral additional cooling jacket of centrifuge tube support, this local type of cooling is not only effective, speed is fast, and very energy-conserving and environment-protective.
Said centrifuge tube is splendid attire grinding bead and the container of treating smudge cells suspension, for smudge cells provides the place.When centrifuge tube was made the high frequency linear reciprocating motion with the centrifuge tube support, grinding bead is violent flutter in centrifuge tube, shears each other, clashes into cell, causes clasmatosis, discharges active material in the born of the same parents.In optimized technical scheme, said centrifuge tube is the standard centrifuge tube of band secure latch, and the cumulative volume of cell suspending liquid and grinding bead is no more than 3/4 of centrifuge tube volume.
Said grinding bead is that violent flutter is with the medium of smudge cells in centrifuge tube, and it should have higher rigidity, hardness and wearability preferably.In optimized technical scheme, said grinding bead is the spherical beads body that quartz glass, pottery, zirconia, stainless steel etc. are processed.In the actual shattering process, need select diameter different polishing pearl for use according to different cell types, generally be that 0.1～1.5mm is advisable, preferred 0.25～0.5mm.
Utilize the quick breaker of above-mentioned universal flutter pearl mill formula cell to carry out fragmentation to extract active material in the born of the same parents to most cells (like zooblast, Gram-negative bacteria, gram-positive bacteria, bacterial spore, saccharomycete etc.), the concrete operations step is following:
1) early-stage preparations: if original sample is dry state powder or blocks of solid; Should with physiological saline (mass fraction is 0.4% the NaCl aqueous solution) it be prepared into suspension earlier; Change in the centrifuge tube of sterile-processed mistake after evenly mixing, insert in the centrifuge tube according to the grinding bead of the corresponding specification of type selecting of cell then and compress the secure latch of centrifuge tube.
2) setting-out is broken: the centrifuge tube that cell suspending liquid and grinding bead will be housed places among the centrifuge tube support of breaker, and with the fixing centrifuge tube of gland.Then, according to the type of treating smudge cells, set motor speed and broken time.Then, start START button and carry out fragmentation, after the broken time reached preset value, motor stopped automatically, accomplishes shattering process.
3) post-processed: after motor thoroughly stops operating, waited 1～2 minute so that the grinding bead natural sedimentation.Take out centrifuge tube then, draw supernatant (active material that DNA, RNA, enzyme etc. discharge all is included in wherein), obtain end product after purified with pipettor.The grinding bead that is deposited in the centrifuge tube bottom can reuse after repeatedly cleaning, and also can directly abandon.
Compare with existing same device, the quick breaker of universal flutter pearl mill formula cell that the present invention proposes has the following advantages and good effect:
1) versatility is good: existing cell breaking plant generally can only be handled a spot of several kinds of cells; No matter the device that the present invention proposes is to more breakable cells such as zooblast, Gram-negative bacterias; Still to difficult broken cell such as gram-positive bacteria, bacterial spore, saccharomycete; Can both be well broken, have good versatility.
2) broken fast: the quick breaker of cell that the present invention proposes, the high frequency linear reciprocating motion of centrifuge tube force grinding bead violent flutter in centrifuge tube, thereby can accomplish the fragmentation of pair cell within a short period of time.
3) be easy to expansion: the quick breaker of cell that the present invention proposes; Its modular construction is simple, compact in design; Be easy to be extended to the quick breaker of multichannel cell,, can greatly increase the processing flux of this device like this like four-way, six passages, eight passages and 12 passages etc.
4) operate steadily: the quick breaker of cell that the present invention proposes; After expansion; Each passage becomes the even distribution of circumference along motor output shaft; And each passage all is radially to do reciprocating linear motion, so the active force that each passage is applied on the housing cancels out each other, and housing can the held stationary state.
5) need not clean: the quick breaker of cell that the present invention proposes, directly sample (sample is stored in the centrifuge tube) is not handled in contact, and therefore in whole clasmatosis process, device can be maintained in the clean condition by sample contamination, need not clean.
Description of drawings
Fig. 1 is the perspective view of a kind of specific embodiment of the present invention;
Fig. 2 is the kinematic sketch of transmission mechanism among Fig. 1;
Fig. 3 is the distribution schematic diagram of each broken passage among Fig. 1;
Fig. 4 is after utilizing the preferred embodiment of the invention that bacillus subtilis black variety gemma is carried out fragmentation, extracts the DNA that purifying discharges, and carries out the reacted agarose gel electrophoresis figure of PCR again.
Among the figure: 1---direct current generator, 2---motor output shaft, 3---line shaft, 4---rotating cam, 5---rocking bar; 6---depression bar, 7---litter, 8---guide post, 9---the centrifuge tube support; 10---centrifuge tube, 11---pivoting support, 12---back-moving spring, 13---housing.
The specific embodiment
For further setting forth the characteristics and the effect of the quick breaker of the universal flutter pearl mill formula cell of the present invention, do more detailed explanation below in conjunction with embodiment.
The overall structure of the quick breaker embodiment of the universal flutter pearl mill formula cell of a kind of four-way such as Fig. 1 are to shown in Figure 3, and this device mainly is made up of direct current generator 1, motor output shaft 2, line shaft 3, rotating cam 4, rocking bar 5, depression bar 6, litter 7, guide post 8, centrifuge tube support 9, centrifuge tube 10, pivoting support 11, back-moving spring 12, housing 13 and control module and man-machine interaction panel (not shown).Under the domination of control module, direct current generator 1 is passed to line shaft 3 through motor output shaft 2 with forms of motion and torque, driving cam 4 rotation at a high speed whereby.Rocking bar 5, depression bar 6, litter 7 and guide post 8 are formed four-bar linkage, the high speed rotary motion of cam 4 are changed into the high frequency linear reciprocating motion of litter 7.Centrifuge tube support 9 is fixed in above the litter 7, be that the two is processed as one in the present embodiment, thereby centrifuge tube support 9 also will be made the high frequency linear reciprocating motion.With grinding bead being housed and treating that the centrifuge tube 10 of smudge cells suspension places centrifuge tube support 9; And fix with the gland (not shown); The high frequency linear reciprocating motion of centrifuge tube support 9 will make grinding bead violent flutter in centrifuge tube 10; Shear each other, clash into cell, cause clasmatosis, discharge active material in the born of the same parents.
In the present embodiment; Control module is the SCM system that adopts the STM32 chip to form; Be used for velocity of rotation, the rotation time of direct current generator 1 are controlled; Concrete control parameter is through the input of man-machine interaction panel, and the man-machine interaction panel is provided with broken time knob, motor speed knob and " beginning ", " time-out " and " suddenly stopping " button.
In the present embodiment, line shaft 3 is a vertical circular shaft of placing, and diameter is 15mm; One end links through shaft coupling and motor output shaft; The other end is fixed in the pivot of cam 4, gives cam 4 with the motion and the transmission of power of motor output shaft 2, makes cam 4 rotation at a high speed.
In the present embodiment, cam 4 be one around the axle center rotational symmetric disk-like structure, maximum radial dimension is 80mm, smallest radial is of a size of 60mm, thickness is 20mm, material therefor is alloy carburizing steel 20CrMnTi.
In the present embodiment, the four-bar linkage that transmission mechanism is made up of rocking bar 5, depression bar 6, litter 7 and guide post 8 is used for the high speed rotary motion of cam 4 is changed into the high frequency linear reciprocating motion of litter 7.Rocking bar 5 one ends are fixed on the housing 13 through pivoting support 11, and the other end is close on the outer surface of cam 4, tangent with it composition higher pair, and the high speed rotation of cam 4 will promote the unidirectional swing of rocking bar 5 high speeds.Depression bar 6 links to each other with litter 7 with rocking bar 5 respectively through hinged, forms revolute pair.Litter 7 is formed moving sets through internal openings and guide post 8; And it is the two is continuous through back-moving spring 12; Withstand shocks load and cyclic loading of back-moving spring 12 here; Need higher mechanical property, so back-moving spring 12 material therefors are selected silicomangan spring steel (60Si2Mn) in the present embodiment.Guide post 8 is fixed on the housing 13, in the present embodiment is through welding the two to be fixed the pivot of guide post 8 place straight-line pass cams 4.The unidirectional swing of the high speed of rocking bar 5 promotes litter 7 through depression bar 6 and moves along guide post 8 straight lines, compresses spring 12 storage power simultaneously.After litter 7 moves to the range place, compression spring 12 will release energy, and promote whole transmission mechanism and reset, and get into the circulation of next cycle then again.Through the periodic alternately storage power of spring 12 with release energy, realized that litter 7 makes the high frequency linear reciprocating motion along guide post 8.
In the present embodiment, centrifuge tube support 9 and litter 7 are processed into an integral body, the cylindrical hole that it is 11mm that centrifuge tube support 9 has a diameter is used to hold centrifuge tube 10, and passes through fixedly centrifuge tube of gland (not shown).Be furnished with the cooling jacket (not shown) on centrifuge tube support 9 outer walls, the loss of active material can eliminated or reduce to the cell suspending liquid in the cooling centrifuge tube 10.
In the present embodiment, centrifuge tube 10 is the standard centrifuge tubes that have secure latch, is used for splendid attire grinding bead and cell suspending liquid, for smudge cells provides the place.Litter 7 will force grinding bead violent flutter in centrifuge tube along the high frequency linear reciprocating motion of guide post 8, shear each other, will clash into cell, make clasmatosis, discharge active material in the born of the same parents.
In the present embodiment, the spherical beaded that grinding bead adopts zirconia to process, its diameter has different sizes such as 0.20mm, 0.25mm, 0.40mm, 0.50mm, 0.75mm, 1.00mm, to adapt to the fragmentation of dissimilar cells.
In the present embodiment, the quick breaker of universal flutter pearl mill formula cell is expanded to four-way, can handle the fragmentation of 4 road cells simultaneously, each passage evenly distributes in same plane, and its distribution is as shown in Figure 3.
In order to verify the effect of smudge cells of the present invention; Now utilize the quick breaker of the universal flutter pearl mill formula cell of present embodiment four-way that bacillus subtilis var (ATCC9372) gemma is carried out fragmentation; Extract the DNA of hay bacillus, carry out pcr amplification then, at last with pcr amplification product electrophoresis (100V on 1.5% Ago-Gel; 30min), the pcr amplification product behind the electrophoresis uses ethidium bromide staining under uviol lamp, to observe.Whole implementing procedure is specific as follows:
One, smudge cells extracts DNA.
1) early-stage preparations: from concentration is 10
3Cfu/mL and 10
4Respectively get the standard centrifuge tube 10 that 800 μ L inject 2 1.5mL respectively in the bacillus subtilis black variety gemma suspension of cfu/mL, add the zirconium oxide abrasive pearl that the 200mg diameter is 0.2mm more respectively in every centrifuge tube.
2) setting-out is broken: 2 centrifuge tubes that will configure are put into the centrifuge tube support 9 of passage 1 and passage 3 respectively, and fix centrifuge tube with gland.It is 2000r/min that motor speed is set then, and the broken time is 10min, and starting drive carries out fragmentation to gemma.
3) DNA extracts purifying: after broken the completion, and pearl deposition to be ground, careful sucking-off supernatant from centrifuge tube utilizes phenol-chloroform method to extract and purify DNA.
Two, PCR reaction, DNA amplification.
1) design of primers: according to bacillus subtilis var rrnE gene order design primer P
1And P
2, extension increasing sequence length is 494bp.
P
1：5′-GGATCCTAATACATGCAAGTCGAGCGG-3′
P
2：3′-GGATCCACGTATTACCGCGGCTGCTGGC-5′
2) PCR reaction system: the bacillus subtilis var dna profiling liquid 10 μ L behind the purifying, the mixing dNTP4 μ L of 1pmol/ μ L, the primer P of 200pmol
1And P
2Each 1 μ L, 10 * Buffer3 μ L, Taq polymerase 1 μ L adds aseptic deionized water and is supplemented to 25 μ L.
3) PCR controlled condition: 94 ℃ of preparatory sex change 5min; 94 ℃ of sex change 30s, 56 ℃ of annealing 30s, 72 ℃ are extended 30s, 35 circulations; 72 ℃ are extended 8min, stop at 4 ℃.
Three, electrophoresis detection PCR product
With pcr amplification product electrophoresis on 1.5% Ago-Gel, electrophoretic voltage is 100V, and electrophoresis time is 30min.Pcr amplification product behind the electrophoresis uses the ethidium bromide (EB) of 0.5 μ g/mL to dye, observation under uviol lamp then.
The agarose gel electrophoresis result of pcr amplification product is as shown in Figure 4, and M is a dna molecular amount mark (Marker) among the figure, C
1(concentration is 10 for the bacillus subtilis black variety gemma suspension without break process
4Cfu/mL) directly go up appearance negative control, C
2For not adding the PCR reaction system negative control of dna profiling, C
3, C
4Then corresponding concentration is 10 respectively
3Cfu/mL and 10
4The gemma suspension of cfu/mL is through going up an appearance electrophoresis behind clasmatosis, DNA extraction purifying and the pcr amplification.
Can know C from Fig. 4
3, C
4All run out of the purpose band about 500bp, two negative control C
1And C
2Bands visible does not then appear.This shows that the quick breaker of the universal flutter pearl mill of present embodiment four-way formula cell can be accomplished the fragmentation to bacillus subtilis black variety gemma within a short period of time (10min), makes it discharge DNA.
Although more than combine accompanying drawing that the preferred a kind of embodiment of the present invention is described in detail, the present invention is not limited to the above-mentioned specific embodiment, and the specific embodiment only is schematically, is not restrictive.The engineers and technicians of this area under the situation that does not break away from aim of the present invention and claim institute protection domain, can also make a variety of improved forms under enlightenment of the present invention, these all belong to protection scope of the present invention.
Claims (10)
1. the quick breaker of universal flutter pearl mill formula cell is characterized in that: mainly comprise main casing, driver module, control module, line shaft, rotor, transmission mechanism, centrifuge tube support, centrifuge tube and grinding bead; The driver module that is fixed on the main casing provides power source for whole breaker under the domination of control module, and will move and transmission of torque is given rotor through line shaft, forces the rotor high-speed rotation; The initiating terminal of transmission mechanism links to each other with rotor rotated through higher pair, and the high speed rotary motion of rotor is changed into the terminal high frequency linear reciprocating motion of transmission mechanism; The centrifuge tube support is fixed in the terminal of transmission mechanism and makes the high frequency linear reciprocating motion with the end of transmission mechanism; The centrifuge tube that grinding bead and cell suspending liquid are housed places centrifuge tube support and fixing; Centrifuge tube is made the high frequency linear reciprocating motion with the centrifuge tube support, and then causes the flutter of grinding bead high frequency, shears, clashes into cell is fierce each other; Cause clasmatosis, discharge active material in the born of the same parents.
2. the quick breaker of universal flutter pearl mill formula cell as claimed in claim 1, it is characterized in that: said transmission mechanism is a planar linkage mechanism, comprises the rocking bar slide mechanism of being made up of through lower pair transmission housing, rocking bar, depression bar and litter; Said lower pair comprises revolute pair and moving sets, and wherein revolute pair is to form through hinge, and moving sets is to cooperate through the plane cooperation or the face of cylinder to form; The transmission housing is that whole transmission mechanism provides fixed constraint and support, and depression bar links up rocking bar and litter through hinge; The unidirectional swing of rotating cam drive rocker of rotation at a high speed under the promotion of depression bar, drives litter and moves along straight line is unidirectional, and the direction that the litter straight line moves is controlled through the guide post that is fixed on the transmission housing; One group of back-moving spring is set on guide post, and litter is along the guide post straight line moves time compression spring, and litter moves to after range sentences, and the spring of pressurized will drive whole transmission mechanism and reset.
3. the quick breaker of universal flutter pearl mill formula cell as claimed in claim 1, it is characterized in that: said driver module is the direct current generator of a unidirectional rotation, and the motion of its output and torque concentrate on the motor output shaft; Said control module is the control module of breaker, and it is a singlechip controller, can to the startup of direct current generator, time-out, anxious stop and motor speed and the duration of runs each parameter control.
4. the quick breaker of universal flutter pearl mill formula cell as claimed in claim 3; It is characterized in that: said line shaft is for connecting the power transmission shaft of motor output shaft and rotor; Said line shaft is given rotor with the rotation and the transmission of torque of motor output shaft, in order to drive the rotor high-speed rotation; Said line shaft is the circular shaft vertically placed, and circular shaft one end links through shaft coupling or clutch and motor output shaft, and the other end then is fixed on the rotor.
5. the quick breaker of universal flutter pearl mill formula cell as claimed in claim 1, it is characterized in that: the material of said line shaft is the structural carbon steel or the manganese alloy quenched and tempered steel of modifier treatment.
6. the quick breaker of universal flutter pearl mill formula cell as claimed in claim 1, it is characterized in that: said rotor is the cam or the eccentric wheel of a plate-like, and its material is the alloy carburizing steel of 20CrMnTi or the middle and high quenching degree of 20Cr2Ni4.
7. the quick breaker of universal flutter pearl mill formula cell as claimed in claim 2 is characterized in that: said centrifuge tube support be fixed in transmission mechanism end, be on the litter, make the high frequency linear reciprocating motion with litter; Said centrifuge tube is the standard centrifuge tube of band secure latch, and the cumulative volume of cell suspending liquid and grinding bead is no more than 3/4 of centrifuge tube volume.
8. the quick breaker of universal flutter pearl mill formula cell as claimed in claim 1, it is characterized in that: said grinding bead is the spherical beads body that quartz glass, pottery, zirconia or stainless steel are processed.
9. the quick breaker of universal flutter pearl mill formula cell as claimed in claim 8, it is characterized in that: the diameter of said grinding bead is 0.1～1.5mm, is preferably 0.25～0.5mm.
10. the quick breaker of universal flutter pearl mill formula cell is characterized in that: comprise housing, driver module, centrifuge tube and grinding bead; The centrifuge tube that grinding bead and cell suspending liquid are housed places centrifuge tube support and fixing; Centrifuge tube is made the high frequency linear reciprocating motion with the centrifuge tube support, and then causes the flutter of grinding bead high frequency, shears, clashes into cell is fierce each other; Cause clasmatosis, discharge active material in the born of the same parents.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201210301788.0A CN102824943B (en) | 2012-08-23 | 2012-08-23 | Universal cell rapid breaker using bead vibrating and grinding |
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|---|---|---|---|
| CN201210301788.0A CN102824943B (en) | 2012-08-23 | 2012-08-23 | Universal cell rapid breaker using bead vibrating and grinding |
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| CN102824943A true CN102824943A (en) | 2012-12-19 |
| CN102824943B CN102824943B (en) | 2014-12-10 |
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| CN201210301788.0A Expired - Fee Related CN102824943B (en) | 2012-08-23 | 2012-08-23 | Universal cell rapid breaker using bead vibrating and grinding |
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107724144A (en) * | 2017-10-18 | 2018-02-23 | 济南大学 | A kind of pulping equipment based on reciprocal reverse agitating function and upset disturbed flow type pulping |
| CN114874897A (en) * | 2022-07-11 | 2022-08-09 | 南京邦康生物技术有限公司 | Selenium-enriched yeast wall breaking control system and method |
| CN115069430A (en) * | 2022-06-25 | 2022-09-20 | 烟台罗博特智能设备有限公司 | Yeast centrifuge control method, system and device and storage medium |
Citations (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2000202314A (en) * | 1999-01-18 | 2000-07-25 | Yasui Kikai Kk | Cell-crushing apparatus |
| US6235501B1 (en) * | 1995-02-14 | 2001-05-22 | Bio101, Inc. | Method for isolation DNA |
| CN1473659A (en) * | 2003-08-12 | 2004-02-11 | 西安交通大学 | Multifunctional scraper stirring ball mill |
| JP2004202497A (en) * | 2004-04-12 | 2004-07-22 | Yasui Kikai Kk | Apparatus for crushing cell and method therefor |
| JP2004313966A (en) * | 2003-04-17 | 2004-11-11 | Yasui Kikai Kk | Cell disintegrator |
| JP2006043554A (en) * | 2004-08-03 | 2006-02-16 | Yasui Kikai Kk | Apparatus for disintegrating cell |
| CN100344357C (en) * | 2002-08-01 | 2007-10-24 | 贝尔坦技术有限公司 | Device for fast vibration of tubes containing samples |
| CN101650274A (en) * | 2009-08-31 | 2010-02-17 | 宁波新芝生物科技股份有限公司 | High pass tissue grinder and grinding method |
| CN102159931A (en) * | 2008-09-18 | 2011-08-17 | 恰根有限公司 | Method and apparatus for the simultaneous, automated decomposition of a plurality of biological samples |
-
2012
- 2012-08-23 CN CN201210301788.0A patent/CN102824943B/en not_active Expired - Fee Related
Patent Citations (10)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US6235501B1 (en) * | 1995-02-14 | 2001-05-22 | Bio101, Inc. | Method for isolation DNA |
| US20020172949A1 (en) * | 1995-02-14 | 2002-11-21 | Bio 101, Inc. | Method for isolating DNA |
| JP2000202314A (en) * | 1999-01-18 | 2000-07-25 | Yasui Kikai Kk | Cell-crushing apparatus |
| CN100344357C (en) * | 2002-08-01 | 2007-10-24 | 贝尔坦技术有限公司 | Device for fast vibration of tubes containing samples |
| JP2004313966A (en) * | 2003-04-17 | 2004-11-11 | Yasui Kikai Kk | Cell disintegrator |
| CN1473659A (en) * | 2003-08-12 | 2004-02-11 | 西安交通大学 | Multifunctional scraper stirring ball mill |
| JP2004202497A (en) * | 2004-04-12 | 2004-07-22 | Yasui Kikai Kk | Apparatus for crushing cell and method therefor |
| JP2006043554A (en) * | 2004-08-03 | 2006-02-16 | Yasui Kikai Kk | Apparatus for disintegrating cell |
| CN102159931A (en) * | 2008-09-18 | 2011-08-17 | 恰根有限公司 | Method and apparatus for the simultaneous, automated decomposition of a plurality of biological samples |
| CN101650274A (en) * | 2009-08-31 | 2010-02-17 | 宁波新芝生物科技股份有限公司 | High pass tissue grinder and grinding method |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107724144A (en) * | 2017-10-18 | 2018-02-23 | 济南大学 | A kind of pulping equipment based on reciprocal reverse agitating function and upset disturbed flow type pulping |
| CN115069430A (en) * | 2022-06-25 | 2022-09-20 | 烟台罗博特智能设备有限公司 | Yeast centrifuge control method, system and device and storage medium |
| CN115069430B (en) * | 2022-06-25 | 2023-06-16 | 烟台罗博特智能设备有限公司 | Control method, system, device and storage medium of centrifugal machine for yeast |
| CN114874897A (en) * | 2022-07-11 | 2022-08-09 | 南京邦康生物技术有限公司 | Selenium-enriched yeast wall breaking control system and method |
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