CN205223192U

CN205223192U - Nucleic acid extraction device

Info

Publication number: CN205223192U
Application number: CN201520838200.4U
Authority: CN
Inventors: 陈辉
Original assignee: Individual
Current assignee: Individual
Priority date: 2015-10-21
Filing date: 2015-10-27
Publication date: 2016-05-11
Anticipated expiration: 2025-10-27

Abstract

The utility model relates to a nucleic acid extraction device, including the adsorption tube that has first link with adsorb the end, the absorption filter core that sets up nucleic acid composition in the aforesaid is adsorbed the end and can be adsorbed pending sample and the built -in piston pipe that has the piston, this piston pipe is connected with first link and is fed through with the adsorption tube with the opening end and through the second link including the second link, still includes to be used for aspirating the suction components of the liquid of absorption filter core of flowing through. Drawing the in -process among this nucleic acid extraction device and need not the dismouting device, simple structure, convenient operation have realized nucleic acid extraction operation continuity, realize easily that the automation of nucleic acid is drawed, can carry out a lot of rinsing as required, have guaranteed nucleic acid extraction's quality, are swift, efficient nucleic acid extraction method, can extensively popularize and apply.

Description

A kind of nucleic acid-extracting apparatus

Technical field

The utility model relates to biological technical field, particularly relates to a kind of nucleic acid-extracting apparatus.

Background technology

Nucleic acid comprises thymus nucleic acid (DNA) and Yeast Nucleic Acid (RNA) two class, is the genetic material of all living things body, to be mainly present in nucleus and to exist with the state of protein bound in cell.Nucleic acid extraction is a very important technology in life science, biotechnology applications and gene diagnosis, and early stage nucleic acid extraction is based upon in the principle of phenol chloroform extraction decon and alcohol settling nucleic acid, but this method is very burdensome.At present, commercial method for extracting nucleic acid is mainly based upon in solid phase adsorption method, namely under given conditions (as pH, specific salt etc.) nucleic acid molecule is adsorbed on certain solid-phase adsorbent surface, then the purification process of wash-out in addition, mainly comprise following two kinds: (1) centrifugal column method: can the solid of adsorbs nucleic acid be seated in centrifugal column for medium with material or other, and with chaotropic salts such as guanidinesalts as bonding agent, when nucleic acid molecule is by being attracted to during centrifugal column in solid phase, then impurity is washed away with the solution containing ethanol, finally with low salt solutions, Nucleic Acid Elution is got off, the method is simple and convenient, and extraction efficiency is high, and extraction effect is good, is at present to use maximum nucleic acid purification methods, and shortcoming needs repeatedly centrifugal in leaching process, is not easy to high-throughput, automated operation.(2) paramagnetic particle method: utilize the magnetic microsphere of the material of surface parcel silicon materials or other energy bind nucleic acid as solid phase carrier, under chaotropic salt (Guanidinium hydrochloride, guanidinium isothiocyanate etc.) and the effect at externally-applied magnetic field, DNA isolation and RNA from sample; The method is simple to operate, easily be automated, high-throughput operation, shortcoming is that cost is higher, and extraction efficiency is easily influenced, and the magnetic bead extraction effect of different sample, different manufacturers, different batches changes greatly.

The key problem in technology of solid phase adsorption method is: (1) is solid absorbent materials efficiently; (2) auxiliary nucleic acid is in conjunction with solid phase material singular solution.At present, most popular solid absorbent materials is material, as United States Patent (USP) NO6, and 218, silicon-dioxide disclosed in 531; And for example United States Patent (USP) NO5; 503; multiple disclosed in 816 have wetting ability and electropositive material; preferred material comprises borosilicate, pure aluminium silicate, phosphoric acid silicic acid, carbonyl silicon, sulfo group silicon and phosphono silicon, and some of them material can reclaim DNA when not using chaotropic agent, when only using water.In bonding agent, the most important thing is chaotropic agent (chaotrope, chaotropicagent), comprise chaotropic salt (chaotropicsalt), conventional chaotropic agent comprises sodium iodide (NaI), urea, Guanidinium hydrochloride (GuHCl), sodium perchlorate (NaClO ₄) and Potassium Bromide (KBr) etc.In addition, also have use alcohols as bonding agent, as European patent NO0512676A1 and United States Patent (USP) NO5,783, disclosed in 686 100% ethanol.

For improving existing method for extracting nucleic acid, application number is that the Chinese invention patent " a kind of device for nucleic acid rapid extraction and a kind of nucleic acid rapid extracting method " of 201410333236.7 (application publication number is CN104059849A) discloses a kind of device for nucleic acid rapid extraction and a kind of nucleic acid rapid extracting method, this device comprises syringe, the interconnecting device of hollow, nucleic acid absorbing film pipe, sample needle four parts, through clinical sample cracking process, syringe pump, nucleic acid absorption film adsorbs nucleic acid, rinsing liquid rinsing nucleic acid absorption film, after the steps such as elution nucleic acid, pure nucleic acid solution can be obtained and test for downstream.Although this patent can realize the extraction of nucleic acid in the device of integration, but when utilizing this device to extract nucleic acid, no matter in " two take out one pushes away " method or " three push away " method, finally will all need the syringe more renewed during the nucleic acid component wash-out be adsorbed on filter membrane, in addition in " two take out one pushes away " method because syringe cavity volume is effective, limited to the wash number of sample, therefore to the restriction that the degree of purification of nucleic acid extractive causes, and " push away " each time in " three push away " method all to need syringe to take off and absorb corresponding liquid, cannot automated operation be realized.

Utility model content

Technical problem to be solved in the utility model is the nucleic acid-extracting apparatus providing a kind of structure simple, easy to use for prior art.

The utility model solves the problems of the technologies described above adopted technical scheme: a kind of nucleic acid-extracting apparatus, comprise the adsorption tube with the first coupling end and absorption end, be arranged on absorption filter element and piston tube that above-mentioned absorption end also can adsorb pending nucleic acids in samples composition, the piston that can move up and down movement along piston tube inwall is built-in with in this piston tube, and this piston tube comprises the second coupling end with opening end is connected with the first coupling end by the second coupling end and is communicated with adsorption tube, it is characterized in that, also comprise for the pumping components of suction current through the liquid of absorption filter element.

Or, a kind of nucleic acid-extracting apparatus, comprise the adsorption tube with the first coupling end and absorption end, be located at above-mentioned absorption end and the absorption filter element of pending nucleic acids in samples composition can be adsorbed and be built-in with the piston tube of piston, this piston tube comprises the second coupling end and opening end, and be connected with the first coupling end by the second coupling end and be communicated with adsorption tube, described opening end is provided with a sealing-ring, described piston is embedded and also can moves up and down along piston tube length direction in sealing circle, it is characterized in that, also comprise for the pumping components of suction current through the liquid of absorption filter element.

For making apparatus structure simple and making the suction effect of pumping components better, as preferably, described pumping components comprises suction pipe, check valve and vacuum pump, and this suction pipe one end is used for pumping liquid, and the other end is connected with vacuum pump by check valve; Or described absorbent assembly comprises suction pipe, electrically operated valve and electric air pump, this suction pipe one end is used for pumping liquid, and the other end is connected with electric air pump by electrically operated valve.

Further, form fluid chamber between described absorption filter element and piston, described suction pipe is inserted on the chamber wall of fluid chamber.

Further; one end of described suction pipe vertically to extend in adsorption tube and is positioned at above absorption filter element; thus the speed that the liquid in accelerating liquid chamber is arranged outward through pumping components; and the free end of suction pipe is arranged with disposable protective tube; replaceable disposable protective tube in nucleic acid extraction; avoid straw end to pollute, ensure the DNA purity of nucleic acid.

The mode of connection of the first coupling end and the second coupling end has multiple, for convenience of operation, described first coupling end and the second coupling end removably connect, removably connect in the utility model and can have various ways, as being threaded, namely the first coupling end and the second coupling end are threaded connection, and and for example buckle connects, and the first coupling end snaps in the second coupling end or the second coupling end snaps in the first coupling end.

Further, also make the first coupling end and the second coupling end stable connection for realizing above-mentioned removably connecting simultaneously, as preferably, described first coupling end and the second coupling end are all in round table-like, and the second coupling end snaps in the first coupling end and the inner peripheral surface of its periphery and the first coupling end fits tightly.

As preferably, described absorption filter element is nucleic acid absorption film or nucleic acid absorption filler, can select glass fibre membrane or pellosil when absorption filter element is nucleic acid absorption film.

For being easy to the automated operation realizing this device, as preferably, this device also comprises the drive unit moved up and down along piston tube inwall for driven plunger, such as stepper motor driven endless screw apparatus etc.

Compared with prior art, the utility model has the advantage of: in this nucleic acid-extracting apparatus, pumping components is set, in leaching process, the filtrate of pending sample after absorption filter element filters and the waste liquid after rinsing liquid cleaning all can directly be aspirated by pumping components, without the need to attaching/detaching apparatus in whole leaching process, structure is simple, easy to operate, achieve nucleic acid extraction continuity of operation, the automatization being easy to realize nucleic acid is extracted, in addition, repeatedly rinsing can be carried out as required, ensure that the quality of nucleic acid extraction, a kind of quick, efficient method for extracting nucleic acid, can wide popularization and application.

Accompanying drawing explanation

Fig. 1 is the utility model embodiment 1 amplifying nucleic acid extraction element structural representation;

Fig. 2 is the utility model embodiment 1 amplifying nucleic acid extraction element local structure schematic diagram;

Fig. 3 is the utility model embodiment 3 amplifying nucleic acid extracting method operating process schematic diagram;

Fig. 4 is the utility model embodiment 2 amplifying nucleic acid extraction element structural representation.

Embodiment

Below in conjunction with accompanying drawing embodiment, the utility model is described in further detail.

Embodiment 1:

As depicted in figs. 1 and 2, a kind of nucleic acid-extracting apparatus, comprising adsorption tube 1, the absorption filter element 3 with the first coupling end 12 and absorption end 11 and be built-in with can along the piston tube 2 of the piston 4 of inside pipe wall upper and lower displacement.In the utility model, adsorption tube 1 can be disposable adsorption tube, and its absorption end 11 is provided with the absorption filter element 3 for adsorbing pending nucleic acids in samples composition.Piston tube 2 comprises the second coupling end 22 and opening end 21, this second coupling end 22 and above-mentioned first coupling end 12 are all in round table-like and be connected by removably, particularly, second coupling end 22 snaps in the first coupling end 12 and the inner peripheral surface of its periphery and the first coupling end 12 fits tightly, and piston tube 2 is communicated with adsorption tube 1.In addition, this device outside is also provided with the drive unit 7 that energy driven plunger 4 moves up and down along piston tube 2 inwall, and handled easily, the automatization for nucleic acid provides basis.

Fluid chamber 5 is formed between above-mentioned absorption filter element 3 and piston 4, this fluid chamber 5 is provided with the pumping components 6 for liquid in aspirated liquid body cavity 5, this pumping components 6 comprises suction pipe 61, check valve 62 and vacuum pump 63, suction pipe 61 is inserted in fluid chamber 5 chamber wall, one end vertically to extend in adsorption tube 1 and is positioned at above absorption filter element 3, the other end is connected with vacuum pump 63 by check valve 62, certain absorbent assembly also can be other possible array configurations, as: absorbent assembly comprises suction pipe, electrically operated valve and electric air pump, this suction pipe one end is used for pumping liquid, the other end is connected with electric air pump by electrically operated valve, do not repeat them here.In leaching process, the filtrate of pending sample after absorption filter element 3 filters and the waste liquid after rinsing liquid cleaning all can directly be aspirated by pumping components 6.Polluted in nucleic acid extraction process for avoiding the end of suction pipe 61; thus affect the purity of nucleic acid extraction; the free end (being namely positioned at the end above absorption filter element 3) of suction pipe 61 is arranged with disposable protective tube 8; by changing disposable protective tube 8 in nucleic acid extraction, ensure the purity of nucleic acid extraction further.

Embodiment 2:

As shown in Figure 4, with above-described embodiment 1 unlike, in embodiment 2, piston tube 2 is built-in with piston 4, and the opening end 21 of piston tube 2 is provided with a sealing-ring 9, and piston 4 to be embedded in sealing circle 9 and can to move up and down along piston tube 2 length direction.

Nucleic acid-extracting apparatus in the utility model can be used as independently device, for the extraction of nucleic acid, also as a part for nucleic acid automatic extracting device, thus can realize the automatization extraction of nucleic acid.

Utilize the process of carrying out nucleic acid extraction of above-described embodiment 1 amplifying nucleic acid extraction element as follows:

(1) adsorb: pending sample adds in test tube, the absorption end 11 of adsorption tube 1 is inserted pending sample, open vacuum pump 63, the suction utilizing suction pipe 61 to produce makes the nucleic acid component in pending sample flow through absorption filter element 3 to be attracted in absorption filter element 3 because of the specific adsorption of absorption filter element 3, filtrate is the filtered liquid containing the impurity such as cell debris and protein, and filtrate is aspirated through suction pipe 61 and discharged;

(2) rinsings: rinsing liquid A joins in another clean test tube, absorption end 11 is inserted in rinsing liquid A, open vacuum pump 63, protein or other impurity components that the suction utilizing suction pipe 61 to produce makes rinsing liquid A be cleaned by absorption filter element 3 to remain on absorption filter element 3, the rinsing liquid A after cleaning enters to aspirate through suction pipe 61 and discharges;

(3) repeatedly rinsing: according to pending sample preparation needs, multiple different rinsing liquid is injected in new test tube accordingly by substep, inserted by absorption end 11 successively in above-mentioned each drift liquid, substep carries out repeatedly rinsing by the operation in step (2);

(4) wash-out: after rinsing terminates, close vacuum pump 63, adsorption tube 1 is inserted and to be equipped with in the new test tube of elutriant and under making absorption end 11 stretch into liquid level, open drive unit driven plunger 4 to move up and down, elutriant flows through absorption filter element 3 under the suction of piston tube 2, and will be adsorbed on the nucleic acid component wash-out on absorption filter element 3, the nucleic acid component of repeatedly absorption on wash-out absorption filter element 3, be collected in nucleic acid collection container or proofing unit, complete the extraction of nucleic acids in samples.

Below by way of specific embodiment, the utility model is further elaborated:

Embodiment 3:

1, sample pre-treatments

Get 60 μ l anticoagulated whole bloods, add isopyknic lysate GL and mix, then adding appropriate proteolytic enzyme, mixing 15s, reacts 10min at 37 DEG C, adds the dehydrated alcohol of GL volume 1.5 times, mix to obtain pending sample.Above-mentioned lysate GL, proteolytic enzyme are all from test kit ZD-XY-Micro-50 (purchased from Ningbo Chong Ding Bioisystech Co., Ltd).

2, nucleic acid extraction

Utilize the nucleic acid-extracting apparatus in the utility model, treat processing sample with reference to said extracted method to process, the reagent used in leaching process is all from test kit ZD-XY-Micro-50 (purchased from Ningbo Chong Ding Bioisystech Co., Ltd), and as shown in Figure 3, detailed process is as follows:

(1) adsorb: above-mentioned obtained pending sample is added in test tube I, the absorption end 11 of adsorption tube 1 is inserted pending sample, open vacuum pump 63, the suction utilizing suction pipe 61 to produce makes the nucleic acid component in pending sample flow through absorption filter element 3 (purchased from Ningbo Chong Ding Bioisystech Co., Ltd) to be attracted in absorption filter element 3 because of the specific adsorption of absorption filter element 3, filtrate is the filtered liquid containing the impurity such as cell debris and protein, and filtrate is aspirated through suction pipe 61 and discharged;

(2) rinsings: rinsing liquid A (in the present embodiment, rinsing liquid A is the washing lotion W1 in test kit ZD-XY-Micro-50) joins in test tube II, absorption end 11 is inserted in rinsing liquid A, open vacuum pump 63, protein or other impurity components that the suction utilizing suction pipe 61 to produce makes rinsing liquid A be cleaned by absorption filter element 3 to remain on absorption filter element 3, the rinsing liquid A after cleaning enters to aspirate through suction pipe 61 and discharges;

(3) repeatedly rinsing: according to pending sample preparation needs, substep is by rinsing liquid B (in the present embodiment, rinsing liquid C is the washing lotion W2 in test kit ZD-XY-Micro-50) and rinsing liquid C (in the present embodiment, rinsing liquid A is the dehydrated alcohol in test kit ZD-XY-Micro-50) injecting tube III and test tube IV, inserted by absorption end 11 successively in above-mentioned each drift liquid, substep carries out repeatedly rinsing by the operation in step (2);

(4) wash-out: after rinsing terminates, suction makes the absolute ethanol volatilizes on filter core for two minutes, then vacuum pump 63 is closed, adsorption tube 1 is inserted and to be equipped with in elutriant (elutriant in the present embodiment is the TE solution in the test kit ZD-XY-Micro-50 of 50 μ l) test tube V and under making absorption end 11 stretch into liquid level, open drive unit 7 driven plunger 4 to move up and down, elutriant flows through absorption filter element 3 under the suction of piston tube 2, and will the nucleic acid component wash-out on absorption filter element 3 be adsorbed on, the repeatedly nucleic acid component of absorption on wash-out absorption filter element 3, be collected in nucleic acid collection container, complete the extraction of nucleic acids in samples.Utilize spectrophotometry to detect the DNA extracted, detected result is as shown in table 1.

Comparative example:

Utilize MICRO centrifugal column method to extract the nucleic acid in whole blood, detailed process is as follows:

(1) get 60 μ l anticoagulated whole bloods, add isopyknic lysate GL and mix, then adding appropriate proteolytic enzyme, mixing 15s, reacts 10min at 37 DEG C, adds the dehydrated alcohol of GL volume 1.5 times, mix to obtain pending sample.

(2) pending sample all being moved into cover has in the microcentrifugation post of collection tube, and centrifugal 2min under rotating speed 12000g takes out centrifugal column (not allowing centrifugal column encounter waste liquid below), discards in collection tube and put back in collection tube by centrifugal column after waste liquid.

(3) in centrifugal column, add 500 μ l solution W 1, centrifugal 2min under rotating speed 12000g, discard waste liquid in collection tube after taking out centrifugal column, centrifugal column is put back in collection tube.

(4) in centrifugal column, add 500 μ l solution W 2, centrifugal 10sec under rotating speed 12000g, discard waste liquid in collection tube after taking out centrifugal column, centrifugal column is put back in collection tube.

(5) centrifugal 2min under rotating speed 12000g.

(6) taken out by microcentrifugation post and put into new 1.5ml centrifuge tube, add 50 μ l solution TE, the static 2min of room temperature, under rotating speed 12000g, centrifugal 1min, DNA are namely by wash-out.Utilize spectrophotometry to detect the DNA extracted, detected result is as shown in table 1.

The reagent lysate GL used in comparative example, proteolytic enzyme, solution W 1, solution TE are also all from test kit ZD-XY-Micro-50.

The detected result of DNA of table 1 for extracting in embodiment and comparative example

From table 1, method for extracting nucleic acid in the utility model is not only easy to operate, extraction rate is fast, OD260/OD280 is about 1.8, show that the DNA purity extracted is high, without impurity such as protein, extraction effect and centrifugal column method basically identical, the extraction of various biological specimen amplifying nucleic acid can be widely used in.

Claims

1. a nucleic acid-extracting apparatus, comprise the adsorption tube (1) with the first coupling end (12) and absorption end (11), be located at above-mentioned absorption end (11) and absorption filter element (3) and the piston tube (2) of pending nucleic acids in samples composition can be adsorbed, this piston tube (2) is built-in with the piston (4) that can move up and down along its inwall, and this piston tube (2) comprises the second coupling end (22) and opening end (21), and be connected by the second coupling end (22) with the first coupling end (12) and be communicated with adsorption tube (1), it is characterized in that, also comprise for the pumping components (6) of suction current through the liquid of absorption filter element (3).

2. a nucleic acid-extracting apparatus, comprise the adsorption tube (1) with the first coupling end (12) and absorption end (11), be located at above-mentioned absorption end (11) and the absorption filter element (3) of pending nucleic acids in samples composition can be adsorbed and be built-in with the piston tube (2) of piston (4), this piston tube (2) comprises the second coupling end (22) and opening end (21), and be connected by the second coupling end (22) with the first coupling end (12) and be communicated with adsorption tube (1), described opening end (21) is provided with a sealing-ring (9), described piston (4) is embedded and also can moves up and down along piston tube (2) length direction in sealing circle (9), it is characterized in that, also comprise for the pumping components (6) of suction current through the liquid of absorption filter element (3).

3. nucleic acid-extracting apparatus as claimed in claim 1 or 2, it is characterized in that, described pumping components (6) comprises suction pipe (61), check valve (62) and vacuum pump (63), this suction pipe (61) one end is used for pumping liquid, and the other end is connected with vacuum pump (63) by check valve (62); Or described pumping components (6) comprises suction pipe (61), electrically operated valve and electric air pump, this suction pipe (61) one end is used for pumping liquid, and the other end is connected with electric air pump by electrically operated valve.

4. nucleic acid-extracting apparatus as claimed in claim 3, it is characterized in that, described suction pipe (61) is inserted in by the chamber wall of the fluid chamber (5) formed between absorption filter element (3) and piston (4).

5. nucleic acid-extracting apparatus as claimed in claim 4, is characterized in that, one end of described suction pipe (61) vertically to extend in adsorption tube (1) and is positioned at absorption filter element (3) top.

6. nucleic acid-extracting apparatus as claimed in claim 5, it is characterized in that, the free end of described suction pipe (61) is arranged with disposable protective tube (8).

7. nucleic acid-extracting apparatus as claimed in claim 1 or 2, is characterized in that, described first coupling end (12) and the second coupling end (22) removably connect.

8. nucleic acid-extracting apparatus as claimed in claim 7, it is characterized in that, described first coupling end (12) and the second coupling end (22) are all in round table-like, and the second coupling end (22) snaps in the first coupling end (12) and the inner peripheral surface of its periphery and the first coupling end (12) fits tightly.

9. nucleic acid-extracting apparatus as claimed in claim 1 or 2, it is characterized in that, described absorption filter element (3) is nucleic acid absorption film or nucleic acid absorption filler.

10. nucleic acid-extracting apparatus as claimed in claim 1 or 2, is characterized in that, also comprising the drive unit (7) for driving described piston (4) to move up and down along piston tube (2) inwall.