CN209652295U - A kind of nucleic acid on-site rapidly extracting pipe - Google Patents
A kind of nucleic acid on-site rapidly extracting pipe Download PDFInfo
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- CN209652295U CN209652295U CN201822164199.8U CN201822164199U CN209652295U CN 209652295 U CN209652295 U CN 209652295U CN 201822164199 U CN201822164199 U CN 201822164199U CN 209652295 U CN209652295 U CN 209652295U
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Abstract
This application provides a kind of nucleic acid on-site rapidly extracting pipes, comprising: push rod, hollow main pipe and nucleic acid absorption device, wherein push rod is nested in main pipe；Independently of at least one fluid reservoir of main pipe's setting, wherein fluid reservoir is connected to main pipe；Control valve is arranged such to control the connection and/or non-interconnected state of fluid reservoir and main pipe.Nucleic acid-extracting apparatus provided by the utility model, it is simple portable, the rapidly extracting from tissue sample processing to nucleic acid can be completed in 20-30 minutes to purify, the quick primary dcreening operation work for being easy to a large amount of samples in one line of the clinic detection of epidemic situation occur especially suitable for farm, port etc., the fluid reservoir being independently arranged simultaneously also can avoid the cross-infection of reagent or sample, be conducive to improve nucleic acid purity.
Description
Technical field
The present invention relates to molecular biology instruments apparatus fields, and in particular to a kind of nucleic acid on-site rapidly extracting pipe.
Background technique
In general, nucleic acid extraction needs the work of multi-step, it is necessary first to the biological samples material such as cell, organization material
Material carries out break process, inactivates nuclease, discharges nucleic acid, so except its hetero-organization such as deproteinized, polysaccharide, lipid or cell at
Point, to obtain high-quality nucleic acid.
From 1869, Switzerland doctor Friedrich Miescher successfully extracted DNA to eighties of last century from cell for the first time
The nineties, nucleic acid extraction be just always one it is cumbersome, time-consuming, need work using toxic reagent.In recent years, with solid
The appearance of phase adsorption technology and biochemical development, greatly alleviate this problem.However, being applicable on site portable
Formula nucleic acid rapid extracting method is still the bottleneck in current nucleic acid extraction field.
Currently, nucleic acid extraction technology can be divided into liquid phase and extracted and two class of solid phase extractions according to the difference of extracting method.Its
In, solid phase extractions can be divided into non magnetic solid phase extractions and Magneto separate again.
1. liquid phase is extracted
Liquid phase extraction refers to through either physically or chemically smudge cells or tissue, then using nucleic acid and other cells or
Different solvents is added in the difference of structural constituent chemical property, by being centrifuged repeatedly, dissolving and precipitate, and then reaches extraction core
The purpose of acid is method more commonly used at present.Liquid phase extraction mainly have CsCl gradient centrifugation, CTAB extraction method, alkaline lysis,
Guanidinium isothiocyanate-phenol chloroform extraction method etc..Liquid phase is extracted needs relatively complicated manual operation step more, to personnel's
Technology and skill requirement are higher, are relatively also easy to produce operation error in operation, various miscellaneous so as to cause being mixed into final product
The loss of matter and nucleic acid substances, it is less reproducible.
2. solid phase extractions
Non magnetic solid phase extractions: solid-phase extraction method mainly utilize solid-phase adsorbent (such as silica, magnetic-particle,
Diatomite, glass fibre, anion-exchange support etc.) with the phases interaction such as electrostatic, affine, ion exchange or hydrogen bond between nucleic acid
With to achieve the purpose that separate nucleic acid.Compared to traditional extracting method, solid phase extraction techniques have rapidly and efficiently excellent
Point, and organic phase and the shortcomings that being not completely separated from of water phase in liquid phase extraction can be overcome.Non magnetic solid phase extractions nucleic acid master
It to be carried out in a manner of spin-column chromatography, by centrifugal action, to achieve the purpose that separation absorption nucleic acid.Solid phase extractions process
Four cracking, combination, cleaning, elution steps are generally divided into, compared to traditional method, this method can greatly shorten nucleic acid
Extraction time.Nowadays numerous nucleic acid extraction kits are developed based on the method.The disadvantage of this method is that must
It must be carried out by centrifuge, when carrying out the operation of a large amount of samples, not can avoid the generation of cross contamination, easily cause false positive knot
Fruit.
Magneto separate: the magnetic-particle for nucleic acid separation need to have superparamagnetism and surface functional groups, and the two will
Property.Firstly, superparamagnetism, guarantees the aggregation and dispersion that can control magnetic-particle by externally-applied magnetic field: secondly, magnetic-particle surface
Functional groups, have an effect under certain condition with nucleic acid molecules, enriched nucleic acid.Applied magnetic particle carries out nucleic acid extraction
Mainly include three processes: one, nucleic acid molecules form magnetic-particle-nucleic acid complexes in conjunction with magnetic-particle；Two, adding outside
Under the action of magnetic field, separating magnetic particles-nucleic acid complexes；Three, Nucleic Acid Elution.Furthermore, it is necessary to be magnetic particle and nucleic acid point
Son combines and isolated solution environmental.For example, Fe3O4Magnetic nanoparticle can be enriched under conditions of PEG-6000 and common salt
DNA (deoxyribonucleic acid, DNA) in cell pyrolysis liquid.Currently, the magnetism of various different modifyings
Particle is ground skilful use in the extracting and developing of DNA DNA and ribonucleic acid (ribonucleic acid, RNA) and richness
Collection.For example, magnetic-particle, the carboxylated magnetic nanoparticle, the magnetic nanoparticle of gelatin coatings, methyl of titanium dioxide Gui package
The magnetic nanoparticle etc. of acrylic acid modification is respectively used to extract DNA, RNA in corn, milk, bacterium or virus.This method
Technical costs is higher, and related kit in the market is expensive, although the nucleic acid extracted is purer, spininess is to micro sample
Product, and the nucleic acid extracted is less, is not particularly suited for clinical detection.
3. the automation extraction system of nucleic acid
The original intention of design automation extraction system is the sample of processing high-throughput, can help the extraction for simplifying nucleic acid.
The system is suitable for large and medium-sized laboratory more, can greatly reduce the working time, reduces worker's cost, improves worker's inherently safe,
Increase the reproducibility of result, improve the quality for obtaining nucleic acid, it has become the key method for improving Laboratory efficiencies.It uses suitable
Magnetic-particle processing system handles sample, avoids the cross contamination of sample room, it is only necessary to several simple steps: in Reagent Tube plus
Enter fluid sample；Reagent Tube is put into machine；By start button, elution is finally used.Therefore begin from Jian whole to terminating
A extraction process only about needs minute.
The system needs expensive instrument support, and reagent cost needed for test sample is also higher, and farm, port etc. are existing
Field detecting laboratory general condition is more simple, cannot achieve the universal and daily maintenance of the system.
Currently, laboratory common method for extracting nucleic acid is spin-column chromatography method both at home and abroad, related kit also phase
Work as maturation, multistep requires just to can be carried out by the help of supercentrifuge in operating process, and needs to be repeatedly opened in the process
Pipe lid carries out the addition of each component liquids, and the grinding etc. of tissue sample is also required to additional instrument and equipment and carries out, the preparation of article
And operation is complex.Nucleic acid extraction technology is confined within laboratory more at present, and the epidemic diseases such as farm, port are detected
For one line, detection work is huge with sample size, operating environment is simple, large-scale instrument and equipment lacks, consumptive material processing is inconvenient,
The difficulties such as operator's not enough profession, and the timely discovery of epidemic situation is most important for prevention and control, therefore epidemic disease detection work is again
The high request for facing " examining fastly ", " examining quasi- " has no applicable, mature matched nucleic acid rapidly extracting mode at present, and moves
Object epidemic situation often originates from or from one line of these farms or port, researches and develops a kind of quick, portable, a tubular type nucleic acid extraction
Device and method is simultaneously combined with the nucleic acid amplification technologies being equally adapted to, significant for Disease monitor and prevention and control.
For to sum up, a kind of method for extracting nucleic acid rapidly and efficiently is researched and developed, with nucleic acid rapid amplifying increasingly mature at present
Method (such as loop-mediated isothermal amplification technique, recombinase polymeric enzymatic amplification technology), which matches, is applied to the epidemic disease inspection of a clinical line
It surveys in work, is of great immediate significance for the timely formulation of the rapid screening of epidemic disease, prevention and control measure and implementation, it can
The investment of human and material resources and time cost, social benefit, remarkable in economical benefits are reduced to greatest extent.
The Chinese invention patent application of publication No. CN106318865A discloses a kind of nucleic acid extraction and gene magnification just
Device is taken, including multi-functional heat lid and internal lysate, buffer, the first PCR reaction reagent and the 2nd PCR of prestoring respectively react
Four independent cavitys of reagent, in use, reagent chamber is sequentially ingressed into four separate chambers to inject the intracorporal reagent of chamber.The dress
The interface that reaction chamber is sequentially ingressed into liquid storage chamber is set, still will cause cross contamination, while the liquid being added is extracted there is no being discharged
The purity of nucleic acid is lower.
The Chinese invention patent application of publication No. CN108220125A discloses a kind of nucleic acid rapidly extracting device, including pushes away
Bar, hollow tube body and fluid reservoir, tube body is interior to be equipped with the protrusion structure for being fixed on bottom, and fluid reservoir is placed in tubular body, liquid storage
Room bottom is provided with liquid relieving mechanism, and prominent structure and liquid release structure cooperation are to discharge the liquid inside fluid reservoir.It should
Device extracts nucleic acid using a tubular type, and but its structure is complex, and the sequence of liquid feeding can only fixedly from bottom to top, push and pull push rod
When must be noted that once shift bottom onto, otherwise superposed fluid reservoir is disclosed in the case where residual liquid is not discharged
It is broken, and its sample introduction and outlet are located at the same side, will cause certain loss when collecting nucleic acid from outlet.
Summary of the invention
In order to solve nucleic acid extracting instrument existing in the prior art, structure is complicated, easily causes cross contamination, interval between diagnosis
The problems such as field diagnostic that is long, expensive, not being suitable for one line of animal epidemics, on the one hand, the present invention provides a kind of nucleic acid
Extraction element, comprising: push rod, hollow main pipe and nucleic acid absorption device, wherein push rod is nested in main pipe；It is independent
In at least one fluid reservoir of main pipe's setting, wherein fluid reservoir is connected to main pipe；Control valve, wherein the setting of control valve
It is the connection and/or non-interconnected state in order to control fluid reservoir and main pipe.
Further, fluid reservoir and the junction of main pipe is arranged in control valve.
Further, fluid reservoir is connect with main pipe by communicating pipe, wherein control valve was arranged on communicating pipe, excellent
Choosing is positioned close to the junction of fluid reservoir, or is positioned close to the junction of main pipe.
In one embodiment, fluid reservoir is connect with main pipe by communicating pipe, and control valve can be located at the interior of communicating pipe
Control valve, is preferably located at the junction of communicating pipe and fluid reservoir by portion at this time, and such setting is conducive to as fluid reservoir and main pipe
When in connected state, the liquid in fluid reservoir quickly flows out；Or it is preferably located at communicating pipe and the junction of main pipe, so set
Residual liquid exposure in avoidable communicating pipe is set in air, to be more advantageous to and avoid cross contamination.In another embodiment
In, control valve can also be radially across communicating pipe, to control the connection and/or non-interconnected state of fluid reservoir and main pipe, this time control
Valve processed may be provided in any position among communicating pipe.
In other implementations, fluid reservoir can be directly connected to main pipe, and centre is not provided with communicating pipe, at this point, control
Valve processed may be provided at fluid reservoir and the junction of main pipe.
Further, control valve is check valve or two-way piston.In a preferred embodiment, in order to effectively control liquid storage
The opening and closing of room, so that being closed, control valve preferred one-way valve or two-way when fluid reservoir and main pipe are in non-interconnected state
Piston, particularly preferred check valve.The setting of check valve, can also be intracorporal in supervisor other than the opening and closing that can control fluid reservoir
When liquid level is higher than the height of fluid reservoir, prevent from being responsible for intracorporal liquid reflux.
Further, the quantity of fluid reservoir is 1-10, and more preferable 2-8, particularly preferably, 4-6 is a.Wherein, different storages
Liquid chamber respectively with the independent connection of main pipe.
Further, fluid reservoir is built-in with lysate, in conjunction with any one in liquid, rinsing liquid, eluent.Preferably,
The number of fluid reservoir is 4, distinguishes built-in lysate, in conjunction with liquid, rinsing liquid and eluent.
Wherein, lysate, be the common buffer of nucleic acid extraction in conjunction with liquid, rinsing liquid, eluent, each buffer is to prestore
It is indoor in liquid storage, it is preferable that at least one fluid reservoir is connect with main pipe by communicating pipe, and each fluid reservoir prestores a kind of slow
Fliud flushing.It is highly preferred that fluid reservoir is distributed in the two sides of main pipe and is connect respectively with main pipe by communicating pipe.Due to fluid reservoir
It is independently of main pipe's setting, is not connected between each fluid reservoir, therefore can effective pre- anti-cross-contamination.
In one embodiment, it is successively arranged the first fluid reservoir and the second storage from left to right along the side of main pipe's radial direction
Liquid chamber, the other side are successively arranged third fluid reservoir and the 4th fluid reservoir from left to right, and four fluid reservoirs can be split containing nucleic acid respectively
Buffer, nucleic acid combination buffer, nucleic acid wash buffer and Nucleic Acid Elution buffer are solved, since four fluid reservoirs are independent
It is arranged, therefore each buffer is located at which fluid reservoir does not have rigid requirement, marks at this time in liquid storage chamber outer wall.
In one embodiment, fluid reservoir is detachably connected with communicating pipe.In another embodiment, fluid reservoir and
Be also possible to communicating pipe it is integrally formed, can be with syringe to liquid storage erebro ventricular injection if prestoring buffer into fluid reservoir at this time
It seals afterwards.
Further, fluid reservoir at least partly material is elastic material.It is preferred that the material of fluid reservoir side wall is elastic material
Material, it is further preferred that fluid reservoir is the relatively thin elastic material of locular wall.Wherein, the elastic material is the elastic material that can be punctured, more excellent
Choosing, the elastic material is selected from rubber, plastics.
In one embodiment, fluid reservoir is preferably the relatively thin plastic material of locular wall, is arranged such first is that in control valve
To facilitate extruding fluid reservoir when check valve, second is that in the fluid reservoir sample-adding for inwardly having lysate liquid storage can be punctured with syringe
Locular wall injection sample-adding.Fluid reservoir is not since independently of main pipe, shape is limited by other structures, preferably bottle at this time
Shape is more advantageous to the extruding of bottle body, makes the indoor liquid of liquid storage it is furthermore preferred that the fluid reservoir side wall of ampuliform is annular pleated structure
Body flows into main pipe.
Preferably, fluid reservoir is fixedly connected or is detachably connected with main pipe.Wherein more preferably it is detachably connected, so that
Buffer can be prestored when preparing the device by obtaining.
Further, the bottom that main pipe is set or be removably arranged in main pipe's that nucleic acid absorption device is fixed
Bottom, wherein being fixed at main pipe bottom, more preferably to prevent the pressure that liquid excessively or in push rod pushes in adsorption tube
Under from junction ooze out.
Preferably, nucleic acid-extracting apparatus is syringe construction.Wherein, rubber plug is arranged in push rod lower part, so that push-and-pull is formed
The pressure that the liquid of adsorbent equipment is discharged.In one embodiment, fluid reservoir and the link position of main pipe are located at supervisor
The middle and lower part of body, preferably lower part are arranged such to reduce the liquid for remaining in adsorbent equipment inner wall, reduce the loss of nucleic acid, keep away
Exempt from cross contamination.
In one embodiment, the position that communicating pipe connect with fluid reservoir is higher than the position connecting with main pipe communicating pipe
It sets.The potential energy for being conducive to increase liquid storage indoor liquid is arranged such, when check valve or two-way piston are opened so that fluid reservoir and master
When tube body is in connected state, the liquid in fluid reservoir can quickly flow out.
In one embodiment, connection inside pipe wall is equipped with diversion pipe or diversion trench.Such setting is conducive in fluid reservoir
Liquid flow rapidly into main pipe, reduce connection tube wall residual.Wherein, diversion trench can be linear type, can also be with
It is screw type.
In one embodiment, nucleic acid absorption device is the internal adsorption tube for being equipped with and can capturing the siliceous material of nucleic acid,
Preferably, siliceous material include one of silica gel, silica, silica, glass powder, alkyl silica, alumina silicate or
It is a variety of, it is further preferred that siliceous material is siliceous adsorbed film.Specific adsorption material using siliceous adsorbed film as nucleic acid, and it is right
Other biological material does not adsorb substantially, can ensure the DNA and/or RNA farthest recycled in sample, while removing other
Impurity.
In one embodiment, the bottom of the bottom of fluid reservoir, the bottom of main pipe and adsorption tube is funnel-form,
It can be conducive to the concentration of liquid, reduce nucleic acid loss, improve nucleic acid yield.
On the other hand, the application the invention also provides above-mentioned nucleic acid-extracting apparatus in nucleic acid extraction.Preferably, nucleic acid
For DNA and/or RNA.
On the other hand, it the invention also provides the method that a kind of pair of nucleic acid is detected and/or expanded, specifically includes: benefit
Nucleic acid is extracted with above-mentioned nucleic acid-extracting apparatus；The nucleic acid obtained to extraction is detected and/or is expanded.Preferably, detection and/or
Amplification includes electrophoresis detection, polymerase chain reaction amplification (polymerase chain reaction, PCR), ring mediated isothermal
Expand (loop-mediated isothermal amplification, LAMP), recombinase polymeric enzymatic amplification
The amplification of (recombinase polymerase amplification, RPA) and/or reverse transcriptase polymerase chain reaction.
On the other hand, the present invention also provides the application methods of the nucleic acid-extracting apparatus.
When extracting nucleic acid using above-mentioned nucleic acid-extracting apparatus, the step including sample to be processed to be applied directly to fluid reservoir
Suddenly；It is furthermore preferred that the locular wall of fluid reservoir can be punctured, sample to be processed is added in fluid reservoir by the place of puncturing.One
In a embodiment, sample to be processed can be injected in fluid reservoir using syringe；Preferably, syringe needle is being extracted
Afterwards, then by pin hole it seals.
Further, the application method of the nucleic acid device further includes squeezing fluid reservoir bottle wall internal liquid is made to enter master
The step of tube body；Preferably, external force squeezes fluid reservoir bottle wall, so that its internal liquid is flowed through check valve and enters main pipe.
Further, the application method of the nucleic acid device further includes push-and-pull push rod, passes liquid through adsorbent equipment and quilt
The step of discharge；Preferably, push rod under the step is specially slowly, makes liquid pass slowly the pellosil in adsorption tube,
It pushes and pulls push rod repeatedly again, makes to be responsible for intracorporal liquid feed and be completely exhausted out；It is highly preferred that when the liquid into nucleic acid device is non-
When eluent, discharge is waste liquid, and when the liquid for entering nucleic acid device is eluent, the liquid of discharge is pure nucleic acid,
It needs to recycle.
On the other hand, the present invention also provides a kind of preparation methods of nucleic acid-extracting apparatus, including prepare push rod, in
The step of empty main pipe, fluid reservoir and nucleic acid absorption device, wherein push rod is nested in main pipe, independently of main pipe
At least one fluid reservoir being arranged, fluid reservoir are connected to main pipe.Nucleic acid-extracting apparatus further includes control valve, to control fluid reservoir
Connection and/or non-interconnected state with main pipe.
Wherein, the sample of nucleic acid to be extracted can also carry out cracking processing only by pretreatment.Work as entrance
The sample of extraction element is when having carried out the sample of cracking processing, directly can be sent into main pipe bottom by the opening of push rod
Adsorbent equipment in, then push and pull push rod discharge waste liquid after, continue that other buffers are added dropwise to adsorbent equipment.In another embodiment party
In formula, lysate is also possible to the environment of with high salt, low pH value, and the pellosil in adsorbent equipment can specifically bind sample at this time
In Plasmid DNA, it is no longer necessary to be added combine liquid, the rinse-added liquid directly into adsorbent equipment.It can be seen that the present invention mentions
The usage mode of the nucleic acid-extracting apparatus of confession is very flexible.
Furthermore it should be noted that carrying out nucleic acid extraction using nucleic acid-extracting apparatus provided by the present invention, it is used for core
Related reagent (such as lysate, rinsing liquid, eluent), concentration, the effect of each ingredient and the process silica gel absorption film that acid extracts
Sequence etc., summarized and refined by nucleic acid extraction method more mature at present, however application scenarios of the invention not office
It is limited to this.Under teachings of the present application, those skilled in the art can also be replaced and/or convert, such as: the present apparatus is combined
Other lysates, rinsing liquid, eluent or maturation nucleic acid extraction kit (such as Trizol method, phenol chloroform method, alkali cracking
Solution etc.) extraction that carries out nucleic acid extraction or other compositions, such as extract protein；Alternatively, since fluid reservoir is independently arranged,
The fluid reservoir quantity being connected with main pipe can also be increased, carry out the filling operation of more complicated, more liquid reagents.
It can bring the following benefits through the invention:
Nucleic acid-extracting apparatus provided by the invention, using identical as the spin-column chromatography method in non magnetic solid phase extraction techniques
Principle, can completely carry out four cracking in nucleic acid extraction, combination, rinsing, elution steps, be quickly obtained the core of high-purity
Acid.The advantage of the present apparatus is:
1, structure is simple, easily operated, easy to carry, does not use the indispensable extraction in the prior art such as supercentrifuge
Instrument, there are no palpus professional technician, large-scale instrument and equipment and stringent laboratory environments, can complete in 15-30 minutes
Rapidly extracting from tissue sample processing to nucleic acid purifies, then cooperates current Rapid nucleic acid amplification technique, such as visual ring
Mediated isothermal amplification detection method can make entire detection of nucleic acids cycle reduction to 1 hour or so, especially suitable for farm,
The quick primary dcreening operation work that port etc. is easy to a large amount of samples in one line of the clinic detection of epidemic situation occur.
2, the fluid reservoir of the present apparatus is independently arranged and is connected to respectively with main pipe, and each reagent used in nucleic acid extraction is close
It closes inside fluid reservoir, effectively prevents the cross-contamination issue between sample or reagent, whole operation process mainly utilizes push rod
Promotion up and down so that nucleic acid by specificity capture on adsorption tube, waste liquid is discharged in time, is finally reached the mesh of purification of nucleic acid
's.
Detailed description of the invention
The drawings described herein are used to provide a further understanding of the present application, constitutes part of this application, this Shen
Illustrative embodiments and their description please are not constituted an undue limitation on the present application for explaining the application.In the accompanying drawings:
Fig. 1 is the perspective view of the preferred embodiment of the present invention；
Fig. 2 is the sectional view of the preferred embodiment of the present invention；
Fig. 3 is the partial enlarged view of Fig. 2；
Fig. 4 is the process for using figure of the preferred embodiment of the present invention；
In figure: 1, push rod；2, rubber plug；3, main pipe；4, adsorption tube；5, liquid outlet；6, control valve；7, communicating pipe；8,
One fluid reservoir；9, the second fluid reservoir；10, third fluid reservoir；11, the 4th fluid reservoir；12, check valve；121, liquid inlet；122,
First sealing ring；123, the second sealing ring；124, valve inner prop；125, valve spring；126, liquid outlet；127, valve body.
Specific embodiment
General idea of the invention is illustrated in order to clearer, is carried out by way of examples with reference to the accompanying drawings of the specification
It is described in detail.In the following description, a large amount of concrete details are given more thoroughly to understand in order to provide to the application.So
And it will be apparent to one skilled in the art that the application may not need one or more of these details and be able to
Implement.In other examples, in order to avoid obscuring with the application, for some technical characteristics well known in the art not into
Row description.
As depicted in figs. 1 and 2, a kind of nucleic acid-extracting apparatus is present embodiments provided, comprising: hollow main pipe 3, supervisor
Body 3 is nested inside with the push rod 1 that bottom is equipped with rubber plug 2, and the adsorbent equipment set on 3 bottom of main pipe adsorbs in the present embodiment
Device is adsorption tube 4.Wherein, one end of adsorption tube 4 is connected to main pipe 3, and the other end is equipped with liquid outlet 5, for waste liquid to be discharged
With the nucleic acid for collecting capture.Preferably, the structure setting of tube body entirety is similar to syringe, and push rod 1 can drive rubber plug 2 being responsible for
It is pushed up and down in body 3, so that push-and-pull forms the pressure that the liquid of adsorbent equipment is discharged.
The present embodiment described device is also independently from the fluid reservoir of the setting of main pipe 3, wherein fluid reservoir is by being connected to
The communicating pipe 7 of 3 side wall of main pipe is connected to main pipe 3, and communicating pipe 7 is equipped with control valve 6, to control fluid reservoir and main pipe 3
Connection and/or non-interconnected state.
In one embodiment, for fluid reservoir with main pipe 3 by connecting communicating pipe 7, control valve 6 can be located at communicating pipe 7
Inside, control valve 6 is preferably located to the junction of communicating pipe 7 Yu fluid reservoir at this time, such setting be conducive to when fluid reservoir with
When main pipe 3 is in connected state, the liquid in fluid reservoir quickly flows out；Or it is preferably located at the connection of communicating pipe 7 Yu main pipe 3
Place, such setting are in order to avoid residual liquid exposure in air, is more advantageous to and avoids cross contamination in communicating pipe 7.In
In another embodiment, control valve 6 can also be radially across communicating pipe, to control the connection of fluid reservoir and main pipe 3 and/or non-
Connected state, control valve 6 may be provided in any position among communicating pipe 7 at this time.
Fluid reservoir is built-in with lysate, in conjunction with any one in liquid, rinsing liquid, eluent.Wherein, lysate, combination
Liquid, rinsing liquid, eluent are the common buffers of nucleic acid extraction, and each buffer is that be pre-stored in liquid storage indoor, preferred real
It applies in mode, at least one fluid reservoir with main pipe 3 by connecting communicating pipe 7, each fluid reservoir prestores a kind of buffer, more excellent
Selection of land, fluid reservoir are distributed in the two sides of main pipe 3 and respectively with main pipe 3 by connecting communicating pipe 7.Since fluid reservoir is independent
It is arranged in main pipe 3, is not connected between each fluid reservoir, it can effective pre- anti-cross-contamination.
In one embodiment, the link position of fluid reservoir and main pipe 3 are located at the middle and lower part of main pipe 3, preferably under
Portion is arranged such to reduce the liquid for remaining in adsorbent equipment inner wall, reduces the loss of nucleic acid, avoid cross contamination.
Preferably, the quantity of fluid reservoir is 1-10, and more preferable 2-8, particularly preferably, 4-6 is a.Wherein, different liquid storages
Room respectively with the independent connection of main pipe.
Be provided with 4 fluid reservoirs in the present embodiment, and be evenly distributed in the left and right sides of main pipe 3, from left to right according to
Secondary is the first fluid reservoir 8, the second fluid reservoir 9, third fluid reservoir 10 and the 4th fluid reservoir 11, and it is slow to prestore nucleic acid cleavage respectively
Fliud flushing, nucleic acid combination buffer, nucleic acid wash buffer and Nucleic Acid Elution buffer, since four fluid reservoirs are to be independently arranged
, therefore each buffer is located at which fluid reservoir does not have rigid requirement, marks at this time in liquid storage chamber outer wall.
In one embodiment, fluid reservoir is with main pipe 3 by connecting communicating pipe 7, and control fluid reservoir is with main pipe's 3
The control valve 6 of connection and/or non-interconnected state may be provided in communicating pipe 7 and fluid reservoir junction, may be provided in communicating pipe 7 and main pipe
3 junction may be alternatively provided at any position among communicating pipe 7.Preferably, fluid reservoir is detachably connected with communicating pipe 7, so that
Buffer must be prestored preparing device Shi Kexiang fluid reservoir.In another embodiment, fluid reservoir and communicating pipe 7 can also be with
It is integrally formed, if prestoring buffer into fluid reservoir at this time, syringe can be used to sealing after liquid storage erebro ventricular injection.
Preferably, control valve 6 may be provided in the junction of communicating pipe 7 Yu fluid reservoir, i.e., fluid reservoir and communicating pipe 7 passes through control
Valve 6 connects, and the liquid be conducive to when fluid reservoir is in connected state with main pipe 3 in fluid reservoir, which is arranged such, quickly to flow
Out.Wherein, in order to effectively control the opening and closing of fluid reservoir, so that being closed when fluid reservoir is in non-interconnected state with main pipe 3
, 6 preferred one-way valve of control valve or two-way piston, as shown in Fig. 2, in the present embodiment, particularly preferred check valve.
As shown in figure 3, the check valve 12 in the present embodiment includes hollow valve body 127,127 upper end opening of valve body is liquid
Entrance 121, lower ending opening are liquid outlet 126.The first sealing ring 122 is equipped between valve body 127 and communicating pipe 8, for preventing
Liquid is leaked from fluid reservoir when liquid inlet 121 is closed.It is equipped with outside inside valve body 127 and is coated with valve spring 125
Valve inner prop 124, between valve inner prop 124 and valve body 127 be equipped with the second sealing ring 123, for prevent liquid by fluid reservoir with
It is flowed out in the gap of 12 junction of check valve.Valve spring 125 is used to support valve inner prop 124, makes liquid inlet 121 in normality
It is in closed state down.
Under the supporting role of valve spring 125, valve inner prop 124 is withstood upwards with sealing liquid entrance 121, works as liquid storage
When indoor pressure is greater than the support force of valve spring 125, valve inner prop 124 declines, and liquid can be passed through by valve, from liquid discharge
Mouth 126 flows out.And when the liquid level in main pipe 3 is higher than the height of fluid reservoir, the pressure of liquid outlet 126 makes valve inner prop
124 jack up upwards, sealing liquid entrance 121, make liquid can not adverse current.The opening and closing in addition to can control fluid reservoir is arranged such
Outside, the liquid reflux in main pipe 3 can also be prevented when the liquid level in main pipe 3 is higher than the height of fluid reservoir.
In another embodiment, control valve 6 can also be two-way piston, it is preferred that in order to guarantee that liquid storage is indoor
Air-tightness, two-way piston are preferably tetrafluoro material, or smear vaseline.When needing to be added the indoor reagent of liquid storage, rotation two
Logical piston is connected to fluid reservoir with communicating pipe 7.It is arranged such other than the opening and closing that can control fluid reservoir, can also control
Into the volume of the liquid in main pipe 3, liquid storage chamber outer wall is preferably provided with scale at this time.
Wherein, fluid reservoir at least partly material is elastic material, it is preferable that the material of fluid reservoir side wall is elastic material, more
It is preferred that fluid reservoir is the relatively thin elastic material of locular wall.
In one embodiment, fluid reservoir is preferably the relatively thin plastic material of locular wall, first is that being check valve in control valve
When facilitate extruding fluid reservoir, second is that inwardly have lysate fluid reservoir sample-adding when, can be punctured with syringe liquid storage locular wall injection
Sample-adding.Fluid reservoir is not since independently of main pipe, shape is limited by other structures, preferably ampuliform, more excellent at this time
Choosing, the fluid reservoir side wall of ampuliform is annular pleated structure, is more advantageous to the extruding of bottle body, becomes owner of the indoor liquid flow of liquid storage
Tube body.
In a preferred embodiment, the position that communicating pipe 7 connect with fluid reservoir is higher than to be connect communicating pipe 7 with main pipe 3
Position.The potential energy for being conducive to increase liquid storage indoor liquid is arranged such, when check valve or two-way piston are opened so that fluid reservoir
When being in connected state with main pipe, the liquid in fluid reservoir can quickly flow out.The inner wall of communicating pipe 7 can also be equipped with diversion pipe
Or diversion trench.Such setting is conducive to the indoor liquid of liquid storage and flows rapidly into main pipe, reduces the residual in connection tube wall.
Wherein, diversion trench can be linear type, be also possible to screw type.
As shown in Fig. 2, being equipped with the siliceous material that can capture nucleic acid in adsorption tube 4, it is preferable that the siliceous material includes silicon
One of glue, silica, silica, glass powder, alkyl silica, alumina silicate are a variety of, it is further preferred that siliceous material is
Siliceous adsorbed film.Specific adsorption material using siliceous adsorbed film as nucleic acid, and other biological material is not adsorbed substantially,
It can ensure and farthest recycle in sample
DNA and/or RNA, while removing other impurities.
Preferably, fluid reservoir is fixedly connected or is detachably connected with main pipe 3.Wherein more preferably it is detachably connected, with
So that buffer can be prestored when preparing the device.
Preferably, the bottom that main pipe 3 is set or be removably arranged in main pipe's 3 that nucleic acid absorption device is fixed
Bottom, wherein being fixed at 3 bottom of main pipe, more preferably to prevent the pressure that liquid excessively or in push rod pushes in adsorption tube
Under from junction ooze out.
Preferably, the bottom of the bottom of fluid reservoir, the bottom of main pipe 3 and adsorption tube 4 is funnel-form, can be advantageous
In the concentration of liquid, nucleic acid yield is improved.
It below will be by taking the application nucleic acid-extracting apparatus extracts DNA and RNA as an example, to illustrate nucleic acid extraction provided by the invention
Device bring beneficial effect.If following example will be premised on following setting without special circumstances:
First fluid reservoir: preset lysate is used for lytic cell, discharges nucleic acid.
Second fluid reservoir: preset rinsing liquid, for washing away the nucleic acid fragment and impurity of small molecule.
Third fluid reservoir: preset rinsing liquid, for washing away the impurity such as protein, salt ion.
4th fluid reservoir: preset eluent, the nucleic acid substances for being incorporated on film elute.
The above are according to QIAGEN kit each component and step setting, other cores can also be cooperated in other examples
The component and step of sour extracting method are configured.
One, DNA is extracted using nucleic acid-extracting apparatus
With above-mentioned nucleic acid-extracting apparatus, DNA rapid extraction, wherein the preset liquid in each fluid reservoir is as shown in table 1:
Each fluid reservoir of table 1 is for extracting preset liquid component when DNA
| First fluid reservoir | + 200 μ L Buffer AL of 20 μ L Proteinase K |
| Second fluid reservoir | 200 μ L ethyl alcohol |
| Third fluid reservoir | 70% ethyl alcohol of 1.5mL |
| 4th fluid reservoir | 200 μ L deionized-distilled waters |
Extracting DNA using nucleic acid-extracting apparatus, specific step is as follows:
(1) it takes 50-100 μ L not include the blood sample with nucleated red blood cell or 5-10 μ L and includes the blood with nucleated red blood cell
Sample, or it is no more than 1 × 107A artificial culture cell sample can must extract sample with PBS adjustment volume to 220 μ L, or
By 25mg animal tissue, homogenization processing is shredded, also can be used as extraction sample.
(2) said extracted sample is injected in the first fluid reservoir with the syringe of 2.5ml, after extracting syringe needle, is used
Glass cement, sealing paste etc. seal the pin hole left, then by the indoor liquid blending of the first liquid storage.If cracking is insufficient, can stand
It is carried out again after 10min in next step.
(3) the first fluid reservoir bottle wall is squeezed, its internal liquid is made to enter main pipe followed by check valve and communicating pipe, then
The second fluid reservoir bottle wall is squeezed, its internal liquid is made to enter main pipe followed by check valve and communicating pipe.
(4) intracorporal liquid blending will be responsible for, slowly under push rod, so that liquid is passed slowly the pellosil in adsorption tube,
It pushes and pulls push rod repeatedly again, makes to be responsible for intracorporal liquid feed and be completely exhausted out.
(5) third fluid reservoir bottle wall is squeezed, its internal liquid is made to enter main pipe followed by check valve and communicating pipe, then
Push rod under slowly, makes liquid pass slowly the pellosil in adsorption tube, then push and pull push rod repeatedly, and it is most to make to be responsible for intracorporal liquid
Amount is completely exhausted out.
(7) the 4th fluid reservoir bottle wall is squeezed, its internal liquid is made to enter main pipe followed by check valve and communicating pipe, it is quiet
After setting 1 minute, then push rod is slowly pushed, pass liquid through pellosil, collected filtrate, obtain pure DNA.
Through experiment can obtain, using nucleic acid-extracting apparatus of the present invention carry out aforesaid operations when be about 20-30 minutes, then
In conjunction with loop-mediated isothermal amplification technique, so that the detection cycle duration of DNA is about 1 hour.
It can be obtained through experiment, the DNA of aforesaid operations acquisition is carried out using nucleic acid-extracting apparatus of the present invention, measures it
OD260/OD280Ratio illustrates no protein residues in 1.8-2.0, and the DNA purity of acquisition is higher.Two, using nucleic acid-extracting apparatus
Extract RNA
With above-mentioned nucleic acid-extracting apparatus, rapidly extracting RNA, wherein the preset liquid in each fluid reservoir is as shown in table 2:
Each fluid reservoir of table 2 is for extracting preset liquid component when RNA
Extracting RNA using nucleic acid-extracting apparatus, specific step is as follows:
(1) it takes 50-100 μ L not include the blood sample with nucleated red blood cell or 5-10 μ L and includes the blood with nucleated red blood cell
Sample, or it is no more than 1 × 107A artificial culture cell sample can must extract sample with PBS adjustment volume to 220 μ L, or
By 25mg animal tissue, homogenization processing is shredded, also can be used as extraction sample.
(2) said extracted sample is injected in the first fluid reservoir with the syringe of 2.5ml, after extracting syringe needle, is used
Glass cement, sealing paste etc. seal the pin hole left, then by the indoor liquid blending of the first liquid storage.If cracking is insufficient, can stand
It is carried out again after 10min in next step.
(3) the first fluid reservoir bottle wall is squeezed, its internal liquid is made to enter main pipe followed by check valve and communicating pipe, then
The second fluid reservoir bottle wall is squeezed, its internal liquid is made to enter main pipe followed by check valve and communicating pipe.
(4) intracorporal liquid blending will be responsible for, slowly under push rod, so that liquid is passed slowly the pellosil in adsorption tube,
It pushes and pulls push rod repeatedly again, makes to be responsible for intracorporal liquid feed and be completely exhausted out.
(5) third fluid reservoir bottle wall is squeezed, its internal liquid is made to enter main pipe followed by check valve and communicating pipe, then
Push rod under slowly, makes liquid pass slowly the pellosil in adsorption tube, then push and pull push rod repeatedly, and it is most to make to be responsible for intracorporal liquid
Amount is completely exhausted out.
(7) the 4th fluid reservoir bottle wall is squeezed, its internal liquid is made to enter main pipe followed by check valve and communicating pipe, it is quiet
After setting 1 minute, then push rod is slowly pushed, pass liquid through pellosil, collected filtrate, obtain pure RNA.
It should be noted that the Buffer RLT that 2 mercapto ethanol is added can be stored at room temperature one month, therefore preferably existing
With now matching, the first fluid reservoir is injected together.
Through experiment can obtain, using nucleic acid-extracting apparatus of the present invention carry out aforesaid operations when be about 20-30 minutes, then
In conjunction with loop-mediated isothermal amplification technique, so that the detection cycle duration of RNA is about 1 hour.
It can be obtained through experiment, the RNA of aforesaid operations acquisition is carried out using nucleic acid-extracting apparatus of the present invention, measures it
OD260/OD280Ratio is greater than 2.0, illustrates no protein residues, the RNA purity of acquisition is higher.
Furthermore it should be noted that carrying out nucleic acid extraction using nucleic acid-extracting apparatus provided by the present invention, it is used for core
Related reagent (such as lysate, rinsing liquid, eluent), concentration, the effect of each ingredient and the process silica gel absorption film that acid extracts
Sequence etc., summarized and refined by nucleic acid extraction method more mature at present, however application scenarios of the invention not office
It is limited to this.Under teachings of the present application, those skilled in the art can also be replaced and/or convert, such as: the present apparatus is combined
Other lysates, rinsing liquid, eluent or maturation nucleic acid extraction kit (such as Trizol method, phenol chloroform method, alkali cracking
Solution etc.) extraction that carries out nucleic acid extraction or other compositions, such as extract protein；Alternatively, since fluid reservoir is independently arranged,
The fluid reservoir quantity being connected with main pipe can also be increased, carry out the filling operation of more complicated, more liquid reagents.
The above description is only an example of the present application, is not intended to limit this application.For those skilled in the art
For, various changes and changes are possible in this application.All any modifications made within the spirit and principles of the present application are equal
Replacement, improvement etc., should be included within the scope of the claims of this application.
Claims (16)
1. a kind of nucleic acid on-site rapidly extracting pipe characterized by comprising
Push rod, hollow main pipe and nucleic acid absorption device, the push rod are nested in main pipe；
Independently of at least one fluid reservoir of main pipe's setting, the fluid reservoir is connected to main pipe；
The nucleic acid-extracting apparatus further includes control valve, and the control valve is arranged such to control the connection of fluid reservoir and main pipe
And/or non-interconnected state.
2. nucleic acid on-site rapidly extracting pipe according to claim 1, which is characterized in that the control valve is arranged in fluid reservoir
With the junction of main pipe.
3. nucleic acid on-site rapidly extracting pipe according to claim 2, which is characterized in that the fluid reservoir passes through with main pipe
Communicating pipe connection, the control valve were arranged on communicating pipe.
4. nucleic acid on-site rapidly extracting pipe according to claim 3, which is characterized in that the control valve was arranged in communicating pipe
Close to the junction of fluid reservoir, or junction of the communicating pipe close to main pipe is set.
5. nucleic acid on-site rapidly extracting pipe according to claim 1 to 4, which is characterized in that the control valve is unidirectional
Valve or two-way piston.
6. nucleic acid on-site rapidly extracting pipe according to claim 1, which is characterized in that the quantity of the fluid reservoir is 1-10
It is a.
7. nucleic acid on-site rapidly extracting pipe according to claim 6, which is characterized in that the quantity of the fluid reservoir is 2-8
It is a.
8. nucleic acid on-site rapidly extracting pipe according to claim 6, which is characterized in that the quantity of the fluid reservoir is 4-6
It is a.
9. nucleic acid on-site rapidly extracting pipe according to claim 6, which is characterized in that the fluid reservoir respectively with main pipe
Independent connection.
10. nucleic acid on-site rapidly extracting pipe according to claim 1, which is characterized in that the fluid reservoir is built-in with cracking
Liquid, in conjunction with any one in liquid, rinsing liquid, eluent.
11. nucleic acid on-site rapidly extracting pipe according to claim 1, which is characterized in that the fluid reservoir at least partly material
Matter is elastic material.
12. nucleic acid on-site rapidly extracting pipe according to claim 11, which is characterized in that the elastic material is that can puncture
Elastic material.
13. nucleic acid on-site rapidly extracting pipe according to claim 11, which is characterized in that the elastic material is selected from rubber
Glue, plastics.
14. nucleic acid on-site rapidly extracting pipe according to claim 11, which is characterized in that the fluid reservoir and main pipe are solid
It connects or is detachably connected calmly.
15. nucleic acid on-site rapidly extracting pipe according to claim 1, which is characterized in that the nucleic acid absorption device is fixed
The bottom that main pipe is set or the bottom of main pipe is removably set.
16. nucleic acid on-site rapidly extracting pipe according to claim 15, which is characterized in that the nucleic acid-extracting apparatus is note
Emitter structure.
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| CN201822164199.8U CN209652295U (en) | 2018-12-20 | 2018-12-20 | A kind of nucleic acid on-site rapidly extracting pipe |
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| CN201822164199.8U CN209652295U (en) | 2018-12-20 | 2018-12-20 | A kind of nucleic acid on-site rapidly extracting pipe |
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Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109456880A (en) * | 2018-12-20 | 2019-03-12 | 中国检验检疫科学研究院 | Nucleic acid on-site rapidly extracting pipe and its application method |
| CN112322473A (en) * | 2020-11-11 | 2021-02-05 | 苏州雅睿生物技术有限公司 | One-step POCT (point of care testing) real-time nucleic acid detection device |
| CN112342128A (en) * | 2020-11-11 | 2021-02-09 | 苏州雅睿生物技术有限公司 | Nucleic acid extraction amplification test tube and working method thereof |
| CN112501007A (en) * | 2020-12-01 | 2021-03-16 | 杭州康金来技术有限公司 | Rotation type structure of nucleic acid extraction element |
-
2018
- 2018-12-20 CN CN201822164199.8U patent/CN209652295U/en not_active Expired - Fee Related
Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109456880A (en) * | 2018-12-20 | 2019-03-12 | 中国检验检疫科学研究院 | Nucleic acid on-site rapidly extracting pipe and its application method |
| CN109456880B (en) * | 2018-12-20 | 2022-07-19 | 中国检验检疫科学研究院 | On-site rapid nucleic acid extraction tube and use method thereof |
| CN112322473A (en) * | 2020-11-11 | 2021-02-05 | 苏州雅睿生物技术有限公司 | One-step POCT (point of care testing) real-time nucleic acid detection device |
| CN112342128A (en) * | 2020-11-11 | 2021-02-09 | 苏州雅睿生物技术有限公司 | Nucleic acid extraction amplification test tube and working method thereof |
| CN112342128B (en) * | 2020-11-11 | 2021-07-20 | 苏州雅睿生物技术有限公司 | Nucleic acid extraction amplification test tube and working method thereof |
| CN112501007A (en) * | 2020-12-01 | 2021-03-16 | 杭州康金来技术有限公司 | Rotation type structure of nucleic acid extraction element |
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