CN104928166A - Automatic parameter-adjustable plasmid extracting device - Google Patents

Automatic parameter-adjustable plasmid extracting device Download PDF

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Publication number
CN104928166A
CN104928166A CN201510339213.1A CN201510339213A CN104928166A CN 104928166 A CN104928166 A CN 104928166A CN 201510339213 A CN201510339213 A CN 201510339213A CN 104928166 A CN104928166 A CN 104928166A
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centrifugal
seat
liquid
lifting
liquid adding
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许箴
杨正平
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State Grid Corp of China SGCC
State Grid Jiangsu Electric Power Co Ltd
Changzhou Power Supply Co of Jiangsu Electric Power Co
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State Grid Corp of China SGCC
State Grid Jiangsu Electric Power Co Ltd
Changzhou Power Supply Co of Jiangsu Electric Power Co
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Publication of CN104928166A publication Critical patent/CN104928166A/en
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    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • C12N15/00Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09Recombinant DNA-technology
    • C12N15/10Processes for the isolation, preparation or purification of DNA or RNA
    • C12N15/1003Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09Recombinant DNA-technology
    • C12N15/10Processes for the isolation, preparation or purification of DNA or RNA
    • C12N15/1003Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor
    • C12N15/1006Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor by means of a solid support carrier, e.g. particles, polymers
    • C12N15/101Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor by means of a solid support carrier, e.g. particles, polymers by chromatography, e.g. electrophoresis, ion-exchange, reverse phase
    • CCHEMISTRY; METALLURGY
    • C12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N15/00Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09Recombinant DNA-technology
    • C12N15/10Processes for the isolation, preparation or purification of DNA or RNA
    • C12N15/1003Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor
    • C12N15/1006Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor by means of a solid support carrier, e.g. particles, polymers

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  • Health & Medical Sciences (AREA)
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Abstract

The invention relates to an automatic parameter-adjustable plasmid extracting device. The automatic parameter-adjustable plasmid extracting device comprises a master controller, a centrifugal vibration device, a liquid adding and liquid taking device and a lifting gland rotating lifting mechanism, wherein the liquid adding and liquid taking device and the lifting gland rotating lifting mechanism are arranged on the two sides of the centrifugal vibration device, the centrifugal vibration device is provided with a centrifugal seat, the centrifugal seat is provided with a plurality of centrifugal pipe grooves, a locating inductor is arranged beside one of the centrifugal pipe grooves, the centrifugal seat is provided with a sensor corresponding to the locating inductor, the lifting gland rotating lifting mechanism comprises a rotatable support and a lifting gland capable of lifting, the support is in an L shape, the lifting gland is connected to the support, the lower side of the lifting gland is provided with centrifugal pipe plugs corresponding to the centrifugal pipe grooves, the liquid adding and liquid taking device comprises a rotatable L-shaped support and a liquor relief disk arranged on the L-shaped support, the liquor relief disk is provided with a plurality of liquid adding pipes and a plurality of liquid sucking pipes, and the liquid adding pipes are connected to electromagnetic valves. By means of the automatic parameter-adjustable plasmid extracting device, the extraction of plasmids can be automatically achieved, the production efficiency can be improved, the manual operation can be reduced, and the purity of the extracted plasmids can be improved.

Description

The automatic plasmid extraction device of Parameter adjustable
The divisional application of the application for a patent for invention of " automatic plasmid extraction device " that the present invention is application number is 201310637644.7, the applying date, to be November 29, invention and created name in 2013 be.
Technical field
The present invention relates to Bioexperiment apparatus field, particularly a kind of plasmid extraction device automatically.
Background technology
Plasmid be attached to acellular karyomit(e) in cell or core district DNA is original can the less DNA molecular of self-replicating, it be a useful target DNA fragment by recombinant DNA technology, send in recipient cell the common carrier going to carry out breeding and expressing to.Thus plasmid extraction is a routine operation in gene cloning procedures, usually uses alkaline lysis to prepare plasmid from intestinal bacteria at present.Principle is: use high ph-values solution to make plasmid DNA and chromosomal DNA sex change, simultaneously precipitating proteins.Again pH value is adjusted to neutrality, plasmid DNA is less, is easy to renaturation and becomes double-strand.And chromosomal DNA is comparatively large, can not renaturation, tangle and reticulate insoluble substance, thus can by centrifugal removing.General step is by centrifugal for inoculum acquisition thalline, adds re-suspension liquid, then adds the bacteria lysis solution of new preparation, finally add ice-cold in and the neutralization solution of PH, then centrifugal, remove precipitation, in supernatant liquor, add DNA adsorption column, centrifugal segregation waste liquid, add washing lotion 1 again and wash one time, centrifugal segregation waste liquid, washing lotion 2 washes 2 times, centrifugal segregation waste liquid, finally obtains pure plasmid DNA with elution.Therefrom visible complex operation step, often extraction 20 plasmids needs are uncapped and are closed lid nearly more than 400 time, and centrifugal more than 10 is secondary.Because plasmid extraction is the laboratory every day of the routine techniques being all engaged in molecular biology research, thus workload is comparatively large, again because its mechanical repeatability is higher, for automatization provides possibility.
Summary of the invention
The object of the invention is to for the existing plasmid extraction process manual operation repeatability described in background technology high, the problem that workload is large, a kind of automatic plasmid extraction device that can solve the Parameter adjustable of foregoing problems is provided.
The technical scheme that can realize goal of the invention of the present invention is as follows:
A kind of automatic plasmid extraction device of Parameter adjustable, it comprises principal controller, Centrifugal vibration device, and be arranged at liquid adding and extracting device and the lifting gland rotary lifting mechanism of Centrifugal vibration device both sides respectively, Centrifugal vibration device is provided with centrifugal seat, centrifugal seat is provided with multiple centrifugal tube seat, one of them centrifugal tube seat is other arranges positioning inductor, centrifugal seat is provided with the sensor corresponding with positioning inductor, lifting gland rotary lifting mechanism comprises rotatable support and liftable lifting gland, support is L shape, lifting gland is connected on support, the centrifugal pipe close corresponding with centrifugal tube seat is provided with below lifting gland, liquid adding and extracting device comprises rotating L bracket and is arranged at L bracket moves liquid dish, move on liquid dish and be provided with multiple liquid adding bottle and pipette, connected electromagnetic valve on liquid adding bottle,
Above-mentioned Centrifugal vibration device comprises base, vibrating motor, vibrating seat and whizzer;
Above-mentioned centrifugal seat being arranged centrifugal tube seat is even number; The corresponding centrifugal pipe close being provided with equal amts below lifting gland; According to the concrete difference extracting the process of DNA plasmid, need the centrifugal tube seat that different quantities is set;
Above-mentioned liquid adding and extracting device is provided with 6 liquid adding bottles and two pipette; According to the concrete difference extracting the process of DNA plasmid, need liquid adding bottle and pipette that different quantities is set.
For the ease of focusing on waste liquid, above-mentioned centrifugal seat being positioned near liquid adding and extracting device and being provided with waste liquid tank.
The principle of work of the automatic plasmid extraction device of Parameter adjustable of the present invention is: control the rotation of Centrifugal vibration device and vibration, the rotation of liquid adding and extracting device and lifting by primary controller and to injecting in centrifuge tube and taking out liquid, the rotation of lifting gland rotary lifting mechanism and lifting, the switch of magnetic valve, the compression of centrifugal pipe close and unclamping and the switch of centrifugal lid.Centrifugal seat can be provided with the centrifugal tube seat of even number as required, during use, centrifuge tube and DNA adsorption column interval are put into centrifugal tube seat, and be described centrifugal seat to be arranged four centrifugal tube seats below, four centrifugal tube seats are numbered No. one to No. four in turn; There is positioning inductor on a centrifugal tube seat side, can automatically be parked on the position of No. after the sensor sensing corresponding on centrifugal work platform.When extracting plasmid, first, manually cleaning centrifuge tube and No. three centrifuge tubes are put into a centrifugal tube seat and No. three centrifugal tube seats respectively, No. two DNA adsorption columns and No. four DNA adsorption columns are put into No. two centrifugal tube seats and four No. three centrifugal tube seats respectively, and the inoculum (about 1.5mL) of the equal volume fermented is added in No. one and No. three centrifuge tubes, then the support of lifting gland rotary lifting mechanism is rotated, lifting gland is turned to the top of centrifugal seat from the side horizontal of centrifugal seat, move down lifting gland again, be pressed on four corresponding centrifuge tubes after four the centrifugal pipe closes be placed in advance below lifting gland are unclamped, then support moves and lifting gland is horizontally rotated to original position.Then, fasten centrifugal lid, Centrifugal vibration device is to the inoculum rotating centrifugal in No. one and No. three centrifuge tubes, centrifugal complete after, centrifugal lid is opened, lifting gland turns to above centrifugal seat from the side horizontal of centrifugal seat, move down again and four centrifugal pipe closes are unclamped simultaneously and is fixed on below lifting gland, buck covers moves, horizontally rotate support again to original position, at this moment bacterial precipitation is in the bottom of No. one and No. three centrifuge tube, the L bracket of liquid adding and extracting device is rotated subsequently, horizontally rotate above centrifugal seat by moving liquid dish, rotate again and move liquid dish and make the first waste liquid suction pipe turn to directly over a centrifuge tube, move down and move liquid dish, the disposable long plastics tubing loaded onto in advance by first waste liquid pipette tip slowly enters a centrifuge tube, supernatant fluid wherein is slowly blotted, then rotating moves above liquid dish to waste liquid tank, waste liquid is discharged in waste liquid tank, rotate again and move liquid dish and make the first waste liquid suction pipe turn to directly over No. three centrifuge tubes, move down and move liquid dish the long plastics tubing of the first waste liquid suction pipe lower end is slowly entered No. three centrifuge tubes, supernatant fluid wherein is also slowly blotted and is discharged in waste liquid tank.Then move liquid disc spins the first liquid adding bottle is moved to directly over a centrifuge tube and adds quantitative re-suspension liquid 250 μ l by solenoid control, then the re-suspension liquid of equal volume is added in No. three centrifuge tubes, by above-mentioned mode, centrifugal pipe close is pressed on centrifuge tube, start Centrifugal vibration device, centrifuge tube thermal agitation 1 minute, make microorganism even suspension, rotate again and move liquid dish and the second liquid adding bottle is moved to above a centrifuge tube and No. three centrifuge tubes respectively and the bacteria lysis solution 250 μ l being added new preparation by solenoid control, according to the above-mentioned method centrifuge tube that vibrates gently, solution is mixed by Centrifugal vibration device, finally rotate move liquid dish the 3rd liquid adding bottle is moved to above a centrifuge tube and No. three centrifuge tubes respectively and by solenoid control add ice-cold in and the neutralization solution 350 μ l of pH, by Centrifugal vibration device according to above-mentioned method vibrate gently centrifuge tube make solution mix after centrifugal.After centrifugal end, mobile and rotate and move the disposable long plastics tubing that liquid dish makes the second pipette end load onto in advance and extend the supernatant fluid slowly sucking-off wherein containing plasmid DNA in a centrifuge tube, rotate again and move liquid dish and will slowly be added in No. two DNA adsorption columns containing the supernatant fluid of plasmid DNA in the second pipette, same use second pipette is by the liquid sucking-off in No. three centrifuge tubes and transfer in No. four DNA adsorption columns, then centrifugal, at this moment plasmid DNA to be adsorbed onto in DNA adsorption column on DNA binding film, and the waste collection filtered is in the bottom of DNA adsorption column.Then rotate move liquid dish make the 4th liquid adding bottle move to successively No. two DNA adsorption columns and No. four DNA adsorption columns top and by Controlling solenoid valve first washing lotion added containing Virahol in DNA adsorption column clean one time remove foreign protein, then centrifugal, rotate again and move liquid dish and to DNA adsorption column, add the second washing lotion containing ethanol again from the 5th liquid adding bottle, wash more than twice and remove foreign protein, then centrifugal, artificial taking-up DNA adsorption column top, have the DNA adsorption column bottom of waste liquid to take out collection to abandon, change and put a new DNA collection tube, DNA collection tube is put on DNA adsorption column top containing plasmid DNA, elutriant is added respectively to No. two and No. four DNA collection tubes from the 6th liquid adding bottle, then after centrifugal wash-out, pure plasmid DNA is obtained in DNA collection tube, the collection tube that artificial taking-up contains plasmid DNA saves backup.Such single job can extract the plasmid DNA of two volumes, adds output, has saved the time.Above-mentioned for four centrifuge tubes, but according to actual needs, more centrifugal tube seat can be set in centrifugal seat, to increase higher output, also as required, several different plasmid DNA can be extracted simultaneously.
Beneficial effect of the present invention: compact construction of the present invention, the process of whole extraction plasmid completes automatically mainly through machine, do not need artificial participation, in plasmid extraction process, the add-on of each solution and washing lotion can be regulated by the parameter of change control device, centrifugation time and vibration intensity can also be regulated, the wash number of scavenging solution can be regulated, to improve the quality of the plasmid of extraction.
Accompanying drawing explanation
Fig. 1 is structural representation of the present invention;
Fig. 2 is the structural representation of the Centrifugal vibration device in invention.
Reference numeral in above-mentioned accompanying drawing is as follows: 1 is principal controller, and 2 is centrifugal seat, and 3 is centrifugal tube seat, 4 is positioning inductor, and 5 is sensor, and 6 is lifting gland, 7 is centrifugal pipe close, and 8 is L bracket, and 9 for moving liquid dish, 10 is liquid adding bottle, and 11 is pipette, and 12 is magnetic valve, 13 is waste liquid tank, and 14 is base, and 15 is vibrating motor, 16 is vibrating seat, and 17 is whizzer.
Embodiment
Be further elaborated below in conjunction with accompanying drawing 1, accompanying drawing 2 pairs of the specific embodiment of the present invention:
(embodiment 1)
The automatic plasmid extraction device of a kind of Parameter adjustable as shown in Figure 1, it comprises principal controller 1, Centrifugal vibration device, and be arranged at liquid adding and extracting device and the lifting gland rotary lifting mechanism of Centrifugal vibration device both sides respectively, Centrifugal vibration device is provided with centrifugal seat 2, centrifugal seat 2 is provided with multiple centrifugal tube seat 3, one of them centrifugal tube seat 3 is other arranges positioning inductor 4, centrifugal seat 2 is provided with the sensor 5 corresponding with positioning inductor 4, described lifting gland rotary lifting mechanism comprises rotatable support and liftable lifting gland 6, support is L shape, lifting gland 6 is connected on support, lifting gland 6 is provided with the centrifugal pipe close 7 corresponding with centrifugal tube seat 3 below, liquid adding and extracting device comprises rotating L bracket 8 and is arranged at L bracket 8 moves liquid dish 9, move on liquid dish 9 and be provided with multiple liquid adding bottle 10 and pipette 11, connected electromagnetic valve 12 on liquid adding bottle 10.
(embodiment 2)
As shown in Figure 2, in the scheme that previous embodiment 1 is recorded, Centrifugal vibration device comprises base 14, vibrating motor 15, vibrating seat 16 and whizzer 17.Drive vibrating seat 16 to vibrate by vibrating motor 15, whizzer 17 is arranged on vibrating seat 16, can realize vibrating and centrifugal operation, simplify the structure of automatic plasmid extraction device, make whole apparatus structure compacter by this set.
(embodiment 3)
In the scheme that previous embodiment 1 is recorded, liquid adding and extracting device is provided with 6 liquid adding bottles 10 and two pipette 11, centrifugal seat 2 is arranged altogether 4 centrifugal tube seats 3, below lifting gland 6, correspondence arranges 4 centrifugal pipe closes 7.
(embodiment 4)
In the scheme that previous embodiment 1 is recorded, for the ease of focusing on waste liquid, described centrifugal seat 2 being positioned near liquid adding and extracting device and being provided with waste liquid tank 13.

Claims (1)

1. the automatic plasmid extraction device of a Parameter adjustable, it is characterized in that: it comprises principal controller, Centrifugal vibration device, and be arranged at liquid adding and extracting device and the lifting gland rotary lifting mechanism of Centrifugal vibration device both sides respectively, Centrifugal vibration device is provided with centrifugal seat, centrifugal seat is provided with multiple centrifugal tube seat, one of them centrifugal tube seat is other arranges positioning inductor, centrifugal seat is provided with the sensor corresponding with positioning inductor, described lifting gland rotary lifting mechanism comprises rotatable support and liftable lifting gland, support is L shape, lifting gland is connected on support, the centrifugal pipe close corresponding with centrifugal tube seat is provided with below lifting gland, liquid adding and extracting device comprises rotating L bracket and is arranged at L bracket moves liquid dish, move on liquid dish and be provided with 6 liquid adding bottles and 2 pipette, connected electromagnetic valve on liquid adding bottle,
Described Centrifugal vibration device comprises base, vibrating motor, vibrating seat and whizzer;
The centrifugal tube seat that described centrifugal seat is arranged is even number; The corresponding centrifugal pipe close being provided with equal amts below lifting gland;
Described centrifugal seat is positioned near liquid adding and extracting device and is provided with waste liquid tank.
CN201510339213.1A 2013-11-29 2013-11-29 Automatic parameter-adjustable plasmid extracting device Pending CN104928166A (en)

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CN201510339214.6A Pending CN104893969A (en) 2013-11-29 2013-11-29 Automatic plasmid extracting device with compact structure
CN201510338181.3A Pending CN105018340A (en) 2013-11-29 2013-11-29 Automatic plasmid extracting apparatus with high extracting quality
CN201510339040.3A Active CN104893967B (en) 2013-11-29 2013-11-29 Labor-saving automatic plasmid extraction element
CN201510338858.3A Pending CN104893966A (en) 2013-11-29 2013-11-29 Automatic plasmid extraction device with compact structure
CN201510339212.7A Pending CN104893968A (en) 2013-11-29 2013-11-29 Automatic plasmid extraction device with high extraction quality
CN201510339213.1A Pending CN104928166A (en) 2013-11-29 2013-11-29 Automatic parameter-adjustable plasmid extracting device
CN201310637644.7A Expired - Fee Related CN103602582B (en) 2013-11-29 2013-11-29 Automatic plasmid-extracting device

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CN201510339040.3A Active CN104893967B (en) 2013-11-29 2013-11-29 Labor-saving automatic plasmid extraction element
CN201510338858.3A Pending CN104893966A (en) 2013-11-29 2013-11-29 Automatic plasmid extraction device with compact structure
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CN114326847A (en) * 2021-12-29 2022-04-12 浙江旅游职业学院 Intelligent pH value regulating and controlling device and method for protein modification research

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CN107115977A (en) * 2017-05-08 2017-09-01 浙江兰博生物科技股份有限公司 A kind of centrifuge
CN107523494B (en) * 2017-09-26 2020-12-04 北京龙基高科生物科技有限公司 Fluid system and nucleic acid sequencing system with same
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CN110003305A (en) * 2019-04-01 2019-07-12 中国科学技术大学 The equipment that polypeptide after synthesis in solid state is post-processed
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US20030091473A1 (en) * 2001-02-08 2003-05-15 Downs Robert Charles Automated centrifuge and method of using same
JP3952019B2 (en) * 2001-12-28 2007-08-01 株式会社日立ハイテクノロジーズ Extraction apparatus, chemical analysis apparatus, and chemical analysis method
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CN104893967B (en) 2019-07-23
CN105018340A (en) 2015-11-04
CN103602582A (en) 2014-02-26
CN104893968A (en) 2015-09-09
CN104893969A (en) 2015-09-09
CN104893967A (en) 2015-09-09
CN103602582B (en) 2015-07-15

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Application publication date: 20150923