CN1170460A - Apparatus and method for storage, purification or reaction and processing of biopolymer - Google Patents

Abstract

An apparatus for identifying, tracking and processing a biological sample (104) stored on a solid medium (105) including a sample recognising/recalling identification station (101), a processing station (103), a tracking station (102) that tracks the sample through the other stations (101, 103) and an interactive networking of the stations. Also disclosed is a cannula (1) connectable in fluid communication to an automatic fluid delivery system constructed with a primary fluid aspirating and dispensing port (6) and at least one auxiliary fluid aspirating and dispensing port (7). A method of removing a by-product from a biopolymer reaction mixture using a polymeric gel is also disclosed.

Description

Be used for storage, purification or the reaction of biopolymer and the apparatus and method of processing

Invention field

The present invention is directed to the system that is used for such as storage, purification and the reaction of some such biopolymers of DNA (deoxyribonucleic acid) (DNA).Ingredient of the present invention has: the solid phase storage medium of a kind of DNA of being used for, a kind of intubate of using with some automatic fluid induction systems, these automatic fluid induction systems are used to suck and discharging and a kind of solid material are followed together fluid, ingredient of the present invention also has: a kind of being used for from the device of biopolymer separate low molecular amount molecule, and a kind ofly be used to differentiate the selected biological sample that is stored, handle this selected sample and this sample of spike so that make this sample system relevant with result.Those ingredients of the system of the invention described above can be used as an independent system and use together, perhaps, in some applications, use independently as the independent ingredient of other system.The present invention also is used for the method for purification and the reaction of biopolymer at those ingredients of the system that utilizes the invention described above.

The cross reference of related application

The application is to be the part subsequent application of the U.S. Patent application S N08 159,104 on November 30th, 1993 applying date, here, and in conjunction with the disclosed content of this application as a reference.

Background of invention

In the past, blood DNA is transported with the DNA that purified, liquid blood, freezing blood or the blood of drying on paper.All these methods all have shortcoming.Wish most that the DNA that blood DNA was purified as drying transports, but this requirement there is high-level technology utility appliance can support utilization at the collection scene.When not having the technology utility appliance to support at the collection scene to utilize, usually whole blood and other primary sample are delivered to centralab, DNA there purifies.Transport liquid blood and need aseptic collection.In some cases, for example,, do very inconvenience like this for the situation that baby's heel punctures and takes a sample.Transport liquid blood or freezing blood also require to control temperature and are different from a kind of suitable transportation system of conventional mail system.Even before the consideration hygienic issues, will do like this.In addition, consider the problem that relates to such as some such pathogen of AIDS virus,, generally put behind one any liquid or the freezing sample that potential risk of infection is arranged of transportation except under the situation of suitable and supervision both expensive.

The blood that is dried on the filter paper has shown it is a kind of method of alternative alternative said method, and has proved, can extract from the whole blood spot of drying and DNA isolation with a kind of form, and the DNA that can extract and separate q.s is used for DNA analysis.This method also has a large amount of shortcomings.For example, do not have pathogen careful and that damage the overwhelming majority apace, and do not suppress the process of those degradation of dna except that the process of the degradation of dna that causes by dehydration modestly.Slow dehydration under the situation of high relative humidity or even low rehydration can make the microflora's growth that damages DNA.Or even when the bacteriostatic agent of the sort of type that the protein denaturation of not making is arranged exists, also have such certain situation, that is, and make DNA can enzymatic some non-enzymatic cleavage of self-dissolving fracture and DNA.Enzymatic splits such process that is meant from fusing, and wherein, dry or impaired tissue (people's cell or parasitic animal and plant cell) activates the enzyme of its component of degraded.In addition, if requirement, the suitable difficulty of the DNA of desorb ultra high molecular weight from the paper.The surface adsorption effect can cause losing DNA, and this can cause preferentially losing the minimum DNA that is degraded, that class dna molecular of promptly wanting most.Like this, just need the device of safe, convenient and minimum labour intensity, be used for storing the sample that contains liquid dna.

In the past, after on the filter paper, handle and also have a problem such as the such biopolymer of DNA in stored dry.The sample of Chu Cuning is because the structural reason of the transfer pipet of prior art generally can not be come fast processing by the automatic fluid induction system like this.When handling compound in the back, there is this compound absorption solid-phase media (for example filter paper) in the above can block the suction and the floss hole of transfer pipet, stops sucting reaction fluid effectively.Therefore, before handling stored compound, just must at first from the solid phase storage medium, discharge this compound.Need discharge compound from storage medium and not only cause losing some such compounds such as DNA, and because consuming time, increase the labour and increase the reagent expense and cause increasing breadboard expense.Combine (perhaps wanting this solid-phase media in reactive system) if want to make compound to stay with solid-phase media, because the end that need manually prevent the solid obstruction transfer pipet, so the automatic fluid induction system is low with regard to efficient.

After biopolymer is synthetic, for example, after the PCR method DNA amplification, usually must separate and the assay products compound.At synthetic back overslaugh analysis and the polymeric accessory substance that problem is a separate low molecular amount from needed product of separating bio.For example, be used to analyze and the method for separating the DNA that increases by round pcr, generally require before analyzing DNA, from amplified reaction, to remove byproduct of reaction.Consider the technology Capillary Electrophoresis of newer analyzing DNA, with regard to the easy to understand this point.Successfully adopted this method to be used for the automatic analysis of the DNA behind pcr amplification.But, before Capillary Electrophoresis, key is the salt of removing as the accessory substance of PCR reaction mixture.

Be used for removing the existent method effort, relatively more expensive and of accessory substance for repeatedly sample analysis is unsatisfactory from the biopolymer that is produced by PCR reaction.Moreover these methods are general to require bigger sample volume, and will manually use rather than use automatically.

Handle a large amount of biologies, agricultural or environmental sample simultaneously and become more and more usual.The dna fingerprint identification that is used for court or medical diagnosis purpose is exactly the exemplary of a this respect.Differentiate a kind of specific sample, for example, the sldh gene type usually requires the large quantities of samples of screening.Wish the sample that fast processing is collected, so that reasonably obtaining the result in the time span scope.When screened sample size increased, still just having needed to keep a kind of sample that is verified and by the mutual relationship accurately between the result that this sample produced.

Summary of the invention

According to the present invention, provide a kind of intubate of using with an automatic fluid induction system of being used for.This intubate generally comprises: a member that comprises a shaft determining the fluid passage; One is used for the mechanism that can be operatively connected with above-mentioned intubate, and this intubate and described automatic fluid induction system are used for optionally sucking and exhaust fluid by this intubate by fluid flow communication; A main fluid sucks and floss hole; And at least one auxiliary fluid sucks and floss hole.Described at least one auxiliary fluid sucks and floss hole sucks with respect to above-mentioned main fluid and floss hole is settled like this, promptly make mouth that this is main and mouth that at least one is auxiliary both in use, fluid be discharged into and the zone of therefrom suction in, can not all be blocked the while by solid material.

The various structures that will realize this point have been described.In a kind of structure, main fluid suction and floss hole and fluid suction and floss hole order that at least one is auxiliary connect.For example, by making an end in intubate in those mouthfuls, and another is connected to (or order connects) first mouthful, but extends upward along the side of cannula wall, realizes this point.In an alternative embodiment, described those mouthful are separated from each other, and the geometric position is such, promptly make a solid material can not block two mouths simultaneously.And in another embodiment, provide that (a bit) individual auxiliary mouth and that (a bit) individual main mouth by a kind of porous welded bond that is positioned at intubate one end.If the orientation that the geometry of above-mentioned welded bond is made on-plane surface-exhibition (non-flat), for example, if it is spherical, solid material just can not block the mouth that all pass above-mentioned porous welded bond simultaneously.

Still according to the present invention, provide a kind of and be used for from the automatic fluid induction system of a fluid reaction system suction and exhaust fluid, this fluid reaction system and a kind of solid material are followed together.In general, this automatic fluid induction system comprises an above-mentioned intubate, has flow device, constructs and settle this flow device, so that pass described intubate discharging reacting fluid.For example, described flow device can be a pumping unit or the similar device that is configured in the above-mentioned automatic fluid induction system.

Still according to the present invention, a kind of biopolymer purifying plant or instrument and method are provided.In general, this biopolymer purifying plant comprises a polymer gel that is configured as the fluid that is used for holding a certain volume.In use, biopolymer material to be purified is placed in the gel.Some materials can diffuse through gel with different speed.Under the situation that does not apply electric field, a kind ofly trend towards in container keeping one longer period, and can not diffuse into gel such as the such big biopolymer of DNA.On the other hand, materials with smaller can trend towards diffusing into gel; The result is the big biopolymer in the accommodation hole stay gel of purifying.

The present invention relates to be used for stored DNA, particularly store a kind of solid-phase media of the DNA of blood sample, and relate to the certain methods of using this solid-phase media.In particular, the present invention relates to be used to store and transport a kind of method of the DNA on the above-mentioned solid phase medium, and relate to such certain methods, these methods or relate to (a) and from the above-mentioned solid phase medium, reclaim DNA, or relate to (b) and use DNA on this solid-phase media (for example, utilizing the dna sequence dna amplification of PCR) in position.

DNA in the blood sample (being called " blood DNA " later on) is used for following these purposes: diagnose some genetic diseases, diagnosis and monitoring such as such some of malaria colonize in blood parasitic disease, determine that paternal and monitoring can appear at some other the non-common cell colony in the blood in some neoplasia.For these purposes, term " blood DNA " is used for representing the source of all DNA of being found usually in blood, like this, just comprise the patient's who has been gathered blood sample DNA, and in his/her blood the DNA in any other organic organization of institute's round-robin.

The invention provides a kind of material and method that is used for removing from the reaction that produces some biopolymers accessory substance, this material and method are easy to use, and the automatic sampling and analysing that is used for those biopolymers is through checking.It is particularly useful when this material and method are used for removing the accessory substance that reacts from PCR before an automation process is passed through capillary electrophoresis analysis DNA.If necessary, the feasible product that can handle repeatedly dna amplification reaction simultaneously of this material and method.

It is more categorical producing above-mentioned medium, and can make the described medium of chunk sizes, so that be fit to needs.

Fig. 1 shows about testing 3.2 (c) above-mentioned analysis (iii).

Fig. 2 is the skeleton view of a kind of preferred transfer pipet of the present invention or intubate.

Fig. 3 be intubate shown in Figure 2 a part amplification part sectioned view.

Fig. 4 is the end-view of Fig. 2 and intubate shown in Figure 3.

Fig. 5 be another embodiment of the present invention intubate amplification part sectioned view.

Fig. 6 is in addition another second embodiment of the present invention, in general is similar to part sectioned view Fig. 3 and Fig. 5, that amplified.

Fig. 7 is the schematic plan top view that can be used for a container of purification of the present invention.

Fig. 8 is in general along the sectional view of the 8-8 line device that got, shown in Figure 7 among Fig. 7.

Fig. 9 is the synoptic diagram of a system of the present invention, as to be used for handling effectively the large number of biological sample.

I, be used for solid-phase media and the method for stored DNA

According to the present invention, provide a kind of and for example comprised that for storing biopolymer blood DNA exists The solid-phase media of interior DNA, this solid-phase media comprises a kind of solid-phase matrix, this solid-phase matrix tool A kind of compound or composition are arranged, and this compound or composition prevent in conjunction with or absorb described base The degraded of biopolymer in the matter.

Solid-phase matrix preferably includes a solid rest, and for example, one is adopted cellulosic absorption Paper (for example filter paper) or a kind of micropore synthetic plastics material, and the compound of protecting biopolymer is arranged Or composition is absorbed on the solid rest. Can be selectively, solid-phase matrix can comprise suitable gluing The mixture material is the form of compressed small pieces or bead thereby make this matrix, and, make protection The compound of biopolymer or composition are attached in described small pieces or the bead or are absorbed in this small pieces Or on the bead.

In a preferred embodiment, the invention provides and a kind ofly comprise blood for storage DNA is in the method for interior DNA, and this method comprises this DNA is applied to a kind of solid-phase media On. Solid-phase media of the present invention comprises the solid-phase matrix with a kind of compound or composition, and is this Compound or composition prevent from being attached in this matrix or are absorbed in dna degradation on this matrix.

In one embodiment of the invention, the DNA that is particularly purified for long term storage, Compound or the composition of protection DNA comprise uric acid, and a kind ofly will be converted into uric acid to this uric acid Salt also provides the weak base of alkaline pH value between 8.0 and 9.5.

In a particularly preferred situation of the present invention, provide a kind of for storing blood DNA Solid-phase media, this solid-phase media comprises a kind of solid-phase matrix, this solid-phase matrix has combination To wherein or absorb a kind of composition on it, this composition comprises a kind of weak base, a kind of chela Mixture and a kind of anion surfactant or detergent and selectively also comprise uric acid or urine Hydrochlorate. Above-mentioned composition gives alkaline pH value, example preferably for example for the blood that is added on the described matrix Such as the pH value between 8.0 and 9.5.

Another kind of situation of the present invention is by soaking into solid-phase media of the present invention, or by solid this Phase medium is embedded in a kind of protective material, and is for example, a kind of such as the such plastics material of polystyrene Material is long-term stored at DNA on the solid-phase media of the present invention.

In practice, when stored DNA, blood sample as blood speckles be applied to of the present invention this On the solid-phase media in the situation of kind, on this solid-phase media, some components (more particularly, are exactly Surfactant) protein and most of any pathogenic organism in the described sample are become The property. But, simultaneously, can protect blood DNA for example to avoid suffering some degradation factors, oxidation And the impact of the processing of ultraviolet (UV) line and the above-mentioned type, thereby so that more stable, become Property dry blood sample can be transported to a diagnostic test chamber. There, can extract DNA, Perhaps can on solid-phase media, use in position DNA.

The composition that is used for this situation of the present invention preferably includes following these materials:

(a) a kind of weak base of unit price (for example, as " Tris " of free alkali or carbonate, i.e. Pehanorm);

(b) a kind of sequestrant (for example EDTA, ethylenediamine tetraacetic acid); With

(c) anionic detergent (for example SDS, sodium dodecylsulphonate); And can select

(d) uric acid or urate.

As an example, a kind of particularly preferred solid-phase media of this situation of the present invention comprises a kind of based on cellulosic absorption paper, for example has the filter paper of minimum every square centimeter of load, and load is as follows:

(a) EDTA:0.5 micromole (free acid of 146.1mg)

(b) Tris:8 micromole (free alkali of 968.8mg)

(c) SDS:1mg; And can select

(d) uric acid: 2 micromoles (acid of 336.24mg).

Have found that, use uric acid of the present invention or urate, although be absolutely necessary for short term stored DNA on solid-phase media,, be ready especially like this for standing storage DNA, because this component has multiple function.At first, it is converted into allantoin, plays the effect of a kind of " free radical " trap, and this free radical trap preferably holds some free radicals, otherwise, will damage the base guanine (for example, 2,3) among the DNA.For example, the spontaneous oxidation by the thio group in the serum proteins of sex change produces some such free radicals, but also can produce some such free radicals by a large amount of iron in the blood.4 uric acid also can play the effect of a kind of component of buffer system, and uric acid is a kind of weak acid in this buffer system.It also plays a kind of effect of erodible surface, and it is soluble in this erodible surface on a small quantity, thereby makes when the urate below them corrodes, the material of the DNA that will be released on its plane of crystal that dry contain.

As previously mentioned, above-mentioned composition can comprise an amount of alkali, can select a kind of weak base of unit price, gives the alkaline pH value of the blood that places on the matrix between 8.0 and 9.5 so that make.This is to guarantee the suitable effect of sequestrant in conjunction with divalent metal the time.It has also prevented from not depend on like this effect of the nuclease of divalent metal.Described alkali can be a kind of weak organic base, for example, and Tris.Can selectively can use a kind of inorganic base, for example, a kind of alkali carbonate or supercarbonate, for example, the carbonate of sodium, lithium or potassium or supercarbonate.

Preferably a kind of strong chelating agent of sequestrant such as EDTA, but, some suitable strong chelating agents of available wide range on the market.The effect of sequestrant is metallic ion magnesium and the calcium in conjunction with divalence, but also in conjunction with transition metal ion, particularly iron.The two all promotes the degraded of DNA by working as the accessory factor that is used for enzyme known calcium and magnesium.Be easy to be subjected to oxidation and reduction such as the such metallic ion of iron, also damage nucleic acid by producing some free radicals 4.

Anionic surfactant or detergent are included in the above-mentioned composition in this case of the present invention as main denaturant.Any and protein bound and to make the reinforcing yin essence ion detergent of protein denaturation all be suitable, and also can use other detergent of such some of above-mentioned SDS, for example, sodium lauroyl sarcosine.This anionic detergent is because the non-special damage of the secondary structure of the coating protein matter of many pathogen, inner protein and any film of being relied on causes these many pathogen to lose activity.Many exceptions are arranged,, some stabilizer pole enterovirus particle is lost activity because anionic detergent does not make most antibacterium spores lose activity.But, these exceptions all are more such reagent, and these reagent may be transferred by common contact, and, also be concerned about that less these reagent constitute the danger that comes autoblood at present.

Cause showing in the test of the present invention at some, can extract blood DNA from the paper of handling through detergent satisfactorily, and urate has prevented that the DNA that was purified from degrading in more than 60 months storage time.In some were further tested, the DNA on filter paper that purifies in position and cross according to special disposal of the present invention then, carried out PCR (PCR) with DNA.In the following embodiments, describe these tests in detail.II, intubate

Provide cannula assembly of the present invention to be used for sample preparation, this sample preparation has utilized a kind of automatic fluid induction system (AFDS) and a kind of and solid material to follow together fluid reaction system.Past, AFDS _sAnd be not suitable for following fluid reaction system together to use with a kind of solid material with one.Problem that overslaugh is used like this is that solid material has blocked intubate or the transfer pipet that fluid flow to and flowed out prior art.Intubate of the present invention or transfer pipet with respect to AFDS _sThe advantage of the transfer pipet of the prior art of Shi Yonging is together, even when fluid reaction system and a kind of solid material are followed together, also can make fluid flow to and flows out intubate.

Here, term " fluid " comprises liquid and gaseous material and flowable suspending liquid." fluid reaction system " is meant any such system, wherein, by adding and/or removing one or more fluid reagents originally material is converted into desired final product.Typically, a fluid reaction system implements in the container of a selected size, and this selected size can change according to the requirement of described fluid reaction system.The size of container and composition can be virtually any size and the materials that is suitable for finishing this process.A kind of suitable containers component is a kind of like this component, this component not with these reagent reactings of fluid reaction system, and these reagent of impermeable this fluid reaction system.Be suitable for adopting the fluid reaction system of intubate of the present invention for example to comprise that pcr amplification, ELISA measure, few ligase (oligoligase) is measured (OLA) and ligase chain reaction (LCR).In general, transfer pipet or intubate are used for guiding fluid to flow to the fluid reaction system or remove fluid from this fluid reaction system.Have therein in the fluid reaction system of solid material, do not wish to want common transfer pipet, this be because when with this transfer pipet from the fluid reaction system during " absorption " fluid, solid material can block transfer pipet.

Term used herein " automatic fluid induction system " comprises manual and automated fluid delivery system.Typically, the automatic fluid induction system is to remove fluid reagent to those each the hole distributing fluids reagent of porous reaction plate with from these each hole.Manually the automatic fluid induction system comprises a plunger handle with a plurality of fluids suctions and discharge end, so that suck and discharge the fluid of a fluid reaction system simultaneously simultaneously from one or more fluid reaction containers.Automated automatic fluid induction system is that computing machine is handled but not manual, and comprises some products like this, for example, picture BIOMEK 2000 (Beckman Instruments, Fullerton is CA) with Zymark Benchmate (Zymark, Hopkinton, MA).

According to the present invention, cannula assembly is a kind of like this structure, so that even also can prevent fluid to flow to and flow out the obstruction of intubate when fluid reaction system and a kind of solid material are followed together.Here " follow with a kind of solid material or combine " is meant that a kind of solids are present in the fluid reaction system, thereby makes it can block the suction and the floss hole of intubate, therefore hindered the normal function of AFDS.Follow a kind of solid material together to may reside in this fluid reaction system with a kind of fluid reaction, so that in this fluid reaction system, provide a kind of chemistry or mechanical effect.Follow the example of solid material together to have with the fluid in the fluid reaction system that provides chemical action at: above be adsorbed with the solid material of biologic artifact or biopolymer; A kind of solid active absorbent carbon; C ₁₈The silica of being derived; Ion exchange resin; And/or polystyrene divinylbenzene.Following an example of solid material together with the fluid reaction system that mechanical effect can be provided is magnetic stirring bar.

Reference number 1 an expression transfer pipet of the present invention or the intubate of Fig. 2.A kind of typical intubate 1 is a kind of cylindrical tube.But, adoptable intubate can be other geometric configuration except cylindrical.Shown in the reference number among Fig. 62, intubate comprises the shaft or the wall 2 of a definite internal fluid channels 3 (Fig. 3).Intubate has a far-end 4 and a near-end 5.Far-end 4 is those ends that insert the fluid of fluid reaction system.Near-end 5 is operably connected with AFDS in use.

The far-end 4 of cannula wall 2 comprises two holes at least.These holes or " mouth " but be communicated with 3 streaming flow ground, fluid passage.These mouthful are that main fluid sucks and floss hole 6 (" main mouth ") and fluid suction and floss hole (" mouth of assisting ") 7 that at least one is assisted.Main and auxiliary mouth 6 and 7 can be continuous or discrete." continuous " meaning is meant between main mouth 6 and auxiliary mouth 3 does not have physics and mechanical separation.In addition, auxiliary mouth 7 changes with respect to the position of main mouth 6 specific embodiment with intubate.Main mouth 6 contacts with the fluid of fluid reaction system with auxiliary mouth 7 boths.

The structure of the main and auxiliary mouth 6 and 7 of intubate 1 of the present invention is such, make when applying negative pressure, follow a kind of independent solids together not hinder the far-end that fluid flows to intubate 1 by blocking these two mouths simultaneously with the fluid of fluid reaction system by AFDS.Prevent solid material to block fluid and flow to fluid passage 3, preferably intubate 1 is configured to like this, make main mouth 6 and auxiliary mouth 7 locate toward each other in a kind of preferred mode.The position of mouth 6,7 should be such, makes at least a portion of a mouth can keep not blocked by the solid material of any another mouthful of obstruction.According to the present invention, dispose main relative configuration and/or position and reach this purpose with auxiliary mouth.For Fig. 1 to embodiment shown in Figure 3, by being positioned at an end of intubate 1 to mouthfuls 6 and locating this task of finishing to mouthfuls 7 along a side of shaft 2.

According to the present invention, AFDS is connected to an intubate.The accurate mechanism that this intubate is connected to AFDS changes with employed AFDS.For example, be used for that (DE) coupling arrangement of an intubate of using together comprises 0.5 inch polypropylene tube for ROSYS, Wilmington with RDSYS 3300.The other coupling method comprises and being slidingly matched and " screwing on " coupling arrangement.In Fig. 2,8 show coupling arrangement.When working with coupled intubate, AFDS applies malleation and negative pressure by fluid passage 3 circulations.In general, during the stage, the fluid passage of the opening of the transfer pipet mouth of prior art can get clogged in negative pressure.This blocks generally is that in the transfer pipet of prior art, this negative pressure source is the transfer pipet mouth opening that contacts with the fluid of fluid reaction system because negative pressure is drawn into negative pressure source to solid material.Therefore, the big solids of this transfer pipet mouth opening have covered this transfer pipet mouth opening, and have hindered negative pressure and removed any remaining fluid effectively.

The embodiment of available various intubate reaches described purpose.By several embodiments of explanation intubate, the various relative positions of main and auxiliary mouth can be described.Except the embodiment of being discussed, can make many variations and modification to the present invention and can not deviate from the spirit and scope of the present invention, this point is obvious.

No matter the embodiment of above-mentioned intubate, general guarantee that preferably the edge of the intubate that may contact with solid material is smooth, that is to say do not have burr or some other defective, even do not having under the situation of negative pressure, described burr and some other defective can cause solid material attached on the intubate.

According to the present invention, can make intubate with more such materials, these materials are generally used for making the transfer pipet and the suction nozzle of those exhaust fluid well known in the prior art.But, for any system, the material that constitutes intubate preferably be chemical inertness to the environment that is run into when the reagent with the fluid reaction system contacts.Be suitable for material of the present invention and comprise malleable metal, for example, stainless steel, titanium, aluminium, platinum, gold, nickel and some other inert metal.In addition, can make intubate with plastic resin or glass.For used plastic cannula in containing the fluid reaction system of phenol, the preferred anti-phenol resin of inertia is made intubate.

Those sizes of disclosed intubate are suitable for using with top those listed automatic fir systems of grasping, and for any specific system, those sizes can change on demand.Generally control the preferred length of intubate by the requirement of used AFDS.Determine the preferred sizes of intubate by the diameter of the requirement of AFDS and fluid reaction system container.Certainly, intubate should be a kind of like this size, and when needs, intubate can operationally be inserted in the container of fluid reaction system.

In the embodiment of Fig. 2 to Fig. 4, mouth 6,7 is discontinuous each other, and is spaced on intubate 1, thereby makes them not blocked by an independent particle simultaneously.In another embodiment of intubate 14 (Fig. 6), main mouth 25 is holes at 15 26 places, farthest in the fluid passage.The opening of main mouth 25 is at one in general in the plane perpendicular to the longitudinal axis 27 of fluid passage 15.In this embodiment, the diameter of main mouth is approximately equal to the internal diameter of fluid passage 15, although and do not require like this.To relating to the typical application situation of ROSYS 3300, the internal diameter of main mouth is approximately 0.010mm to 0.1mm, and preferably 0.020mm is to 0.040mm.For the situation of the application that relates to ROSYS 3300, the length of intubate is approximately 6cm to 20cm, preferably 15cm.Such as used herein, term " approximately " is meant described those quantitative sizes, and these sizes are also nonessential quantitatively equal, but can finish a kind of size of similar results.

With further reference to the embodiment of Fig. 6, intubate comprises an auxiliary mouth 24 at least.Auxiliary mouth 24 connects (promptly being connected with main mouthful 25) with main mouth 25 orders, and is the inverted v-shaped shape.Inverted v-shaped has a summit 11 and a base 28.The summit 11 of the inverted v-shaped shape of auxiliary mouth is along 28 cannula wall 19 rises from the base away from the direction of far-end 26.Situation for the preferred embodiment that has adopted ROSYS 3300, the width on the base 28 of inverted v-shaped is about 0.010mm to 0.030mm, preferably be about 0.020mm, and this inverted v-shaped from the base 28 to the limit 11 height be about 1.0mm to 4.0mm, preferably be about 2.0mm.Can place one or more inverted v-shapeds around the periphery of cannula wall.When with a plurality of inverted v-shaped, the section outward appearance of intubate far-end is " sawtooth " shape.Therefore, according to this embodiment, when applying negative pressure, follow solid material together preferably can not block main mouth 25 and auxiliary mouth 24 simultaneously with the fluid reaction system.If under the situation of negative pressure, solid material sticks on the auxiliary mouth 24, and so main mouth 25 and/or any other auxiliary mouth 24 preferably keep not getting clogged.On the other hand, if negative pressure causes solid material to adhere on the main mouth 25, so preferably, one or more auxiliary mouths 24 keep not getting clogged.

Fig. 5 embodiment that another substitutes of the present invention of having drawn.Here, comprise that at far-end 37 places of fluid passage 38 intubate 35 of a main mouth 36 is in the plane perpendicular to the longitudinal axis 39 of fluid passage 38.Then, be contained on the mouth 36 with a porous welded bond 40, the factor of porosity of the welded bond 40 of porous is about 0.20 μ m to 0.60 μ m, preferably is about 0.45 μ m.Because the factor of porosity of welded bond 40 has a plurality of holes to exist.Thereby any one single Kong Douke of welded bond 40 thinks that for one of main mouth auxiliary mouth this main mouth will be some other mouthfuls of same welded bond 40.Can make by such some materials and be suitable for welded bond of the present invention, for example, glass fibre, stainless steel, titanium and resin.According to this embodiment, a non-open and flat part of spherical welded bond 40 remains on outside the farthest of wall 41.By AFDS, will and extract solid facing to welded bond to applying negative pressure with the fluid reaction system fluid in contact passage 38 that contains solid material.Because non-open and flat (sphere) geometric configuration of the welded bond 40 that is inserted, the solid material that is extracted facing to welded bond 40 will tangentially adhere on the welded bond 40, and it is mobile to hinder fluid at the place, point of contact of adhesion place.But, remaining those mouthful of welded bond 40 can keep not getting clogged, and thus, make fluid can flow to not by those holes that hinder by the adhesion solid material.Be appreciated that, can adopt those alternative, non-open and flat geometric configuratioies for welded bond 40.

To in embodiment 5, describe embodiment preferred in detail for cannula structures.III, biopolymer purifying plant

Biopolymer purifying plant assembly of the present invention provides a kind of device, removes salt and those low-molecular-weight compounds in this device those biopolymers from the biopolymer reaction mixture.After making a kind of biopolymer reaction mixture and the biopolymer purifying plant contacts, the polymkeric substance of being purified for example can be used for external and the interior biochemical reaction of body, perhaps, for example can be the object of capillary electrophoresis analysis.

Such as used herein, the meaning of term " biopolymer " is any polymerizable molecular (for example, polypeptide, protein, DNA, RNA, carbohydrates and similar biomolecule) that is used for by identical or different biosystems that monomer derives.Biopolymer can chemosynthesis, obtain or derive out from natural products with gene engineering research.Word " biopolymer reaction mixture " is used for being defined in any biopolymer in the solution that a kind of and salt or some other low molecular weight compound combine here.Such as used herein, term " accessory substance " is meant that with respect to a kind of biopolymer that for example comprises salt and/or residual reactant be low-molecular-weight those compounds.

To the biopolymer purifying plant be described by means of Fig. 7 and Fig. 8.(Fig. 7 is a vertical view, and Fig. 8 is a sectional view).Fig. 7 and Fig. 8 have shown the biopolymer purifying plant that preferably includes (rectangle) piece 50, and piece 50 contains a kind of polymer gel, preferably polyacrylamide.In piece 50, a plurality of holes or recess 51 are arranged in top surface 52.Hole 51 is formed the shape that can keep the fluid of a certain volume.The method that forms hole 51 comprises any known method that is used for the shaped polymer gel (for example, glass plate casting, injection modulus method and casting mold method).The typical outer appearnce of biopolymer purifying plant is similar to the outward appearance of the flat board of porous.As described below, the biopolymer reaction mixture is put into hole 51.With after the hole contacts a period of time, low-molecular-weight accessory substance will as shown in arrow 60ly diffuse away from this hole.

The biopolymer purifying plant is adaptive, but size is enough to feasible can the use with the automatic fluid induction system of aut.eq. control, so that location and operation on its micropore reliably.

According to the present invention, the biopolymer reaction mixture of a certain volume is put into the hole 51 of biopolymer purifying plant.For typical purposes, it is dark that the size in the typical hole 51 of biopolymer purifying plant is about 0.5mm to 6.0mm, it is dark preferably to be about 4.0cm, and diameter is about 0.25cm to 2.0cm, diameter preferably is about 0.3cm to 1.0cm, with after the biopolymer purifying plant contacts a period of time, the diffusion of the accessory substance of this biopolymer reaction mixture portals 51 at the biopolymer reaction mixture, and makes this biopolymer break away from low-molecular-weight accessory substance relatively.The length of duration of contact between biopolymer reaction mixture and biopolymer purifying plant depends on used situation and changes, but typically is about 5 minutes to 40 minutes, is preferably 10 minutes to 30 minutes.Desired is to have time enough to be used to occur the acceptable extent of purification.Such as used herein, term " is about " length and the number of times that is meant the described time in this context, and the length of this time and number of times are also nonessential equal quantitatively, and still, this time can produce similar result.

According to the present invention, have recognized that the concentration affects of polymkeric substance (polyacrylamide) component that changes polymer gel crosslinked amount between two kinds of polymkeric substance.This crosslinked amount directly influences the factor of porosity of gel.(amount that increases acrylamide causes increasing crosslinked amount, and this has reduced factor of porosity thereby has also reduced can be easy to spread 51 sizes of molecule that enter the remainder of gel piece 50 of portalling.)。Therefore, by controlling crosslinked amount, just can control can spread the size of portalling or staying the molecule in this hole.Although an embodiment preferred of the present invention provides separating bio polymer from low-molecular-weight accessory substance, but the inventor predicts and adopts the biopolymer purifying plant, just such as described here, it is still less discrete but be in the molecule at those relative limit places of given polymer gel pore-size to be used for the isolated molecule amount.

Polyacrylamide (a kind of polymer gel) is generally used in the gel electrophoresis, so that separate little biopolymer molecule.In gel electrophoresis, under the electric field effects that is applied, biopolymer passes Medium Diffusion.Therefore, polyacrylamide gel is used for the little dna molecular of electrophoretic separation by the diffusion of passing this gel.By comparison, polyacrylamide gel is used as the medium of purification biopolymer, and can not cause significantly losing this biopolymer owing to biopolymer diffuses into gel.Need not apply electric field.

Although do not assert any mechanism, a kind of possible reason for the result who is found among the present invention is electric field, when being applied to electric field in the gel electrophoresis, a kind of such as the such biopolymer of DNA by high orientation, and so just can be drawn into gel, move so that pass this gel.But, when such biopolymer was not in the electric field, just as in the present invention, just rotation and the situation of ratio in an electric field that is applied had bigger effective molecular volume (or diameter) at random for it.Like this, just prevented that biopolymer from passing gel and move owing to lacking the orientation electric field.Although gel is obstacles for some biopolymers, it is not an obstacle for less molecule and ion, and molecule that this is less and ion can be easy to diffuse into gel rubber material and biopolymer is waited behind (staying in the container).

Then, available artificial or automatic fluid induction system reclaims the biopolymer solution of being purified from container.This biopolymer can be used in the follow-up biochemical process, and is perhaps, for example analyzed.Illustrated among the embodiment 6 and produced and used the biopolymer purifying plant.IV, puncturing technique (Punch Technology)

Of the present invention this is a system that is used for handling effectively the large number of biological sample on the one hand.With reference to Fig. 9, put up with the ingredient (station) that constitutes this system this system is described.

Provide system 100 to be used for differentiating sample 101, a kind of sample is put into a reaction vessel be used for processing operation 103, handle sample; And, make that sample can be relevant with corresponding result by processing operation 103 spike samples 102.Maximally utilise this system, and can be in each cross pollution between any two of single sample or transfer.

Such as used herein, system 100 will be called puncturing technique system (PTS) or " system ".With regard to its embodiment of wide meaning, system 100 can discriminating, spike and any biological sample that is stored in wherein of processing.Finish those ingredients of the present invention by system ensemble data center (ISDC): differentiate 101, spike 102 and handle 103 interactional network work.Can be by manually finishing the work of finishing by ISDC.Preferably, ISDC is a computer hardware and software systems, and this system provides interactive instruction to PTS.

Such as used herein, term " is differentiated ", " discriminating " and their some similar words are that another ingredient of showing system detects and store or detect and send the ability about the information of sample, is used to make result and sample is relevant accordingly.Term " spike ", " carrying out spike " and their some similar words are meant follows the tracks of and makes the sample ability relevant with its result.Term " is handled " and " processing " is meant any such flow process, flow process thus, and sample stands reaction, purifies, buffering, flushing or some other step, so that be sample changeover originally needed final product.

In a preferred embodiment, the PTS system have the ability fast and differentiate effectively, processing and at least two kinds of samples of spike and be typically a large amount of samples.So-called " in a large number " is meant that described system can preferably differentiate, processing and at least two kinds of spikes, at least 2 to 2000 kinds of samples preferably.The ability that also obtains the result from a large amount of samples rapidly effectively makes system 100 be specially adapted to large-scale screening operation.Such screening have great application prospect in the field of medical diagnosis and treatment, pathology, Biochemical Research, drug screening, design molecular biology, medical jurisprudence, genetic engineering and a large amount of screening samples of some other needs.Some special applications for example comprise antigen/antibody research, histocompatbility, dna fingerprint and gene diagnosis.In a preferred embodiment, said system can compare a unit price DNA sample that is stored on the above-mentioned solid phase medium fast and effectively with the big database that also is stored in the DNA image on a kind of solid-phase media.For example, natural referring to " Britain will set up the DNA database of crime, " 370,588 (on Augusts 25th, 1994).

In a preferred embodiment, system 100 is provided for above-mentioned automatic system, thus, can differentiate, handle the biological sample that also spike is stored, for example, the DNA by the blood source provides the final DNA product that is applicable to pcr amplification or digestion with restriction enzyme.A kind of typical method that is used to store according to the handled biological sample of the present invention is a dry biological sample on a kind of solid-phase media, and this solid-phase media for example is a used medium in the paper of cellulose, nitrocellulose, plastics, glass fibre, anion exchange paper, filter paper, its treated with same, cotton sheet or other prior art.A kind of storage medium that preferably is used for the DNA sample is at the storage medium described in the part I of the present invention.

Fig. 9 synoptic diagram of puncturing technique of the present invention system 100 that drawn.This system is once more by differentiating that station 101, spike station 102 and treating stations 103 constitute.

Will preferably being stored on the sample card 104 by sample by PIS discriminating, spike and processing.Label 106 expressions are stored in " spot " of the single sample on the sample card 104.Sample card 104 preferably comes in addition " sign " with mark 105, is used for by differentiating that station 101 differentiates.

Before using said system, sample card 104 is put into " being subjected to cartridge " 107, be subjected to cartridge 107 to give stored card 108 orientations in such a way, making can be for differentiating 101 acceptance in station.Between the 108 liang of cards of the single sample card that is stored that are being subjected in the cartridge 107, place barrier 109.

Arrow 110 expression is differentiated the route that this sample card is got 101 time of standing when the sample card shift-in.Differentiate the sign 106 of the selected card of station 101 identifications.Give spike station 102 information that is obtained by the discriminating station, relevant after being used for the result who comes out from treating stations 103.

Arrow 111 shows one by the sample card differentiated desirable route when shift-in spike station 102.This sample 105 is followed when a selected sample 105 that is extracted from sample card 104 passes in spike station 102 when treating stations 103 moves.The position of selected sample 105 on sample card 104 at first determined at spike station 102.Then, the effect at spike station 102 is to extract selected sample 105 from sample card 104.Hold selected sample 105 by a container, in this container, handle this sample by treating stations 103.When spike station 102 arrives spike station 102 from selected sample card up to finishing the position of following the trail of selected sample 105 when handling sample.After extracting selected sample card from sample card 104, sample card 104 preferably is subjected to cartridge 114 places along being moved by arrow 112 shown routes, being collected in.A kind of barrier 116 be held in place be subjected in the cartridge 114 in twos between the sample card, so that prevent to be arranged in those the collected sample cards direct contact between any two that is subjected to cartridge 114.Sample card in being subjected to cartridge 114 can turn back to and be subjected to cartridge 107 or some other storage location.

Selected sample 105 can move on to treating stations 103 from spike station 102 along arrow 120 shown routes from sample card.As described below, treating stations 103 is handled this selected sample, so that obtain needed final product result.Then, make for the result for the treatment of stations 103 relevant with selected sample 105, this selected sample 105 from above-mentioned selected sample card just by 102 spikes of spike station.

In some applications, the process of carrying out in treating stations 103 places can directly be carried out on selected box, and need not sample is shifted out from card.A, differentiate station 1, stored sample

Provide that PTS is used to differentiate, processing and spike be stored in the biological sample on a kind of solid-phase media.The medium that is specially adapted to said system comprises solid-phase media, and thus, the processing of sample does not also require the step of at the very start sample being taken out from its formation medium.

According to the present invention, being suitable for the suitable medium that stored DNA sample uses for example is the solid-phase matrix described in the part I of the present invention.Past, carry out that PCR amplifies and/or digestion with restriction enzyme before, must at first from storage medium, extract the DNA sample.Although general having at present is used for extracting from solid-phase media the method for DNA in a large number, but many such methods require sample preparation widely, and these processing comprise: boil, repeatedly inhale and move and centrifugation step, desalination chromatography, precipitation, film dialysis and some other method well known in the prior art.Adopt the solid-phase media described in the part I of the present invention, the sample that is stored that is suitable for pcr amplification and/or digestion with restriction enzyme is provided, and need not at first from solid-phase matrix, to extract this sample.Therefore, the matrix of part I of the present invention makes can handle the DNA sample automatically, and the problem of not followed with other those sample stocking systems.

In an embodiment preferred of said system, sample is stored on solid-phase media or " card " 104.Card 104 can have is enough to keep that one of minimum flow is bled or the virtually any size of other biological sample.Typical card is a rectangle.Typical card sizes is about length 1cm to 25cm (preferably being about 5cm to 15cm) and multiply by about 1cm to 20cm (preferably being about 5cm to 10cm).The card of different geometries and size also is suitable for alternative purposes more of the present invention.

Solid-phase media or card 104 are compatible when being preferably used for stored sample, and can not change sample originally.Preferably, coming being placed on the solid-phase media 104 of autoblood, tissue culture, bacterial cultures or other liquid medium such as the such biological sample 105 of DNA.Here, term " liquid medium " meaning is meant a kind of sample, wherein, be stored in that biological sample on the solid-phase media is suspended or dissolved.In general, liquid can stay the DNA that is adsorbed on this solid-phase media from solid-phase media evaporation or other is suspended or dissolved molecule.Single biological sample in liquid medium or a plurality of biological sample are placed on (as mentioned above) on the card with a kind of being suitable for by the setting that the card positioner mechanisms is located to sample.Before or after receiving sample, this card is labeled as 106, in order to discern by discriminator (as described in the above-mentioned part IV) later on.After sample in being contained in liquid medium has been placed on the card, this liquid medium evaporation, and the card that contains the sample after the dehydration stores by suitable device well known in the prior art, for example, by storage external, the polystyrene (all be 25 ℃ time) that applied, and store by some other method that can on solid-phase media, preserve sample.A kind of preferable methods that is used for stored DNA for example is the method described in the part I.2, discriminator

For providing the discriminating station of PTS, system is used for identification and recovery sample card.Each card can comprise at least one or a plurality of same or different biological sample.Preferably, independent card comprises one or more than independent biological sample just.Each sample card preferably has one can be differentiated the sign that the station is differentiated by system.This sign makes may discern and reclaim the biological sample of being made up of card.When differentiating, the work that can reclaim the biological sample of forming by card by discriminator.Can be selectively, discriminator can be given " card sample steady arm " (CSL) mechanism's transmission authentication information (as described below), is used to reclaim the biological sample of being made up of card.

Carry out sample and differentiate, " being subjected to cartridge " and " discriminator " adopted at the discriminating station.Be subjected to cartridge that a kind of device is provided, be used for before card is differentiated by discriminator, holding this card.Be subjected to cartridge can comprise a rack or box, for example, just as the paper duplicating machine or other those hold used rack or box in the system of article of paper or other " similar sheet ".Being subjected to cartridge can be any various shape, as long as its size and dimension can hold card by a certain orientation, makes discriminator can differentiate card.

Card is placed in such a way is subjected in the cartridge, so that prevent in card or sample cross pollution between any two.Can prevent cross pollution by between per two cards, placing a barrier, preferably a kind of barrier that does not cling, for example, cellulose paper, Waxpaper, plastics and some other barrier well known in the prior art.Before or after differentiating, from taking out barrier in twos between the card, this depends on the position of discriminator.

The discriminator of system provides the card that is used to differentiate on it and contained and the device of sample.Card contains a kind of " sign " specially.Such as used herein, the meaning of " sign " is meant other device of a kind of discriminator identification card of encoding or be used for.In a preferred embodiment, card can contain bar code, and discriminator is exactly a bar code reader, this bar code reader is just as used in grocery store's check line, and this discriminator can also be DNA storage container, the bar code scanner on gas chromatograph/mass spectrometer self-actuated sampler and high pressure liquid chromatography (HPLC) self-actuated sampler.Can be selectively, readable digital of available machines used or differentiate card with some other method.

Then, the information of being differentiated by discriminator is delivered to the spike station, so that the sample of spike on the sample card of being differentiated by discriminator is relevant with result after being used for.

In case differentiated sample card by discriminator, just can start the spike station.When sample card when being subjected to cartridge neutralization to move on to the spike station subsequently, can differentiate sample card with discriminator.Can be selectively, sample card can at first move, be carried out discriminating and move on to the spike station then from being subjected to cartridge to pass across discriminator.

With regard to this point, suitable card differentiates that for the effect of bringing into play sample spike part effectively be essential.B, spike station

The spike station of system is provided, when being used for being differentiated that from sample card the station is differentiated when finishing last treatment step by treating stations till the single sample of spike.The ability that said system runs through whole process spike single sample guaranteed when handling a large amount of sample system suitable sample the accuracy when relevant with corresponding results.

There are three major functions at sample spike station.It should at first go up the sample in location at a card (may contain a plurality of samples).Then, the spike station is extracted (selecteed) sample be positioned from sample card and is used for handling (being used for handling if extract sample).At last, when the treating stations that passes system when selected sample moved, the spike station kept reclaiming the position of this selected sample.The technical term of these three functional parts is called " card sample steady arm " respectively, and (CSL) mechanism, " punch block mechanism " be (PM) and " receiving vessel sample steady arm " (RCSL) mechanism.

After differentiating sample card by discriminator, sample card just moves on to the spike station.In case arrive the spike station, card just preferably begins to contact with card sample steady arm (CSL) mechanism.CSL mechanism comprises that an energy localization package contains the device of the spot on the sample card of an independent sample.Preferably, make CSL mechanism can be positioned at which sample on the card be want processed information from ISDC.For example, be applicable to that the CSL mechanism of method of the present invention comprises some devices like this, such as, laser indexing system, mechanical indexing system and optical density (OD) system.For example, used laser and mechanical indexing system on photocopier and facsimile recorder.

In case needed sample is positioned on the card, just start punch block mechanism, put into receiving vessel so that extract sample from sample card.According to the present invention, punch block mechanism comprises a blade, cutting laser instrument, solid perforator and, and some other can cut the device of wearing the solid-phase media that contains sample.Then, the sample that is extracted is put into receiving vessel.The size of the card sample that is extracted from the card obviously is the size that is suitable for putting into receiving vessel.

An embodiment preferred of punch block mechanism comprises an annular blade.The outer appearnce of this blade is similar to Skin biopsy perforator or cork consent.Such blade typically is made of the material that stainless steel, titanium or some other are commonly used to make paper knife.The diameter of perforated solid-phase media is about 2mm to 10mm, and preferred diameter is about 4mm.When using directly (for example), preferably make the surperficial heating disinfection that begins the blade that contacts with card, so that make the cross pollution of sample reduce to minimum with the comparison of laser beam perforation device with blade that the sample card that contains sample contact.Available propane flame or temperature are about 250 ℃ to 300 ℃, and the heater coil that preferably is about 275 ℃ is finished heating disinfection.

As mentioned above, the sample that extracts with punch block mechanism is put into receiving vessel.Such as used herein, term " receiving vessel " comprises that single hole, microcentrifugal tube, PCR pipe and those of ordinary skills of test tube, microtiter plate are through being commonly used to handle other reaction vessel of biological sample.

With " receiving vessel sample steady arm " (RCSL) mechanism detect the sample that is extracted and be placed on wherein receiving vessel.RCSL can send the positional information of selecteed sample to ISDC, and this ISDC can play the effect of spike sample in the entire process process in position during the entire process station and progress by indication.

After having extracted sample, this card is deposited in second again is subjected in cartridge or the box from sample card.Card is placed on described being subjected in the cartridge in such a way, so that prevent in the cross pollution between card or the sample in twos.As mentioned above, can prevent cross pollution by between per two cards, placing a barrier.Then, the receiving vessel that contains biological sample is moved on to the sample preparation station.C, sample preparation station

Provide that the sample preparation station of PTS is used for moving pure, buffering, flushing, PCR amplifies or some other is for being converted into the necessary step of treated final product to sample.After sample being put into receiving vessel, can handle sample in position.Can selectively can at first from solid-phase media, extract and want processed sample.The process that treating stations carried out changes with sample type and the processing target of being wanted.

The treating stations of PTS can comprise an automatic fluid induction system with automatic controller, is used for the mobile response container and is used for sucking and discharging reacting fluid during sample preparation.Preferably utilize among the part II of the present invention disclosed intubation system to finish and in treating stations, use the automatic fluid induction system.

For some sample, the treating stations of PTS can at first make the sample experience extract the step (i.e. Chuan Kong sample card) of sample from solid-phase media.For this situation, the step of processing can relate to use for making and emit the necessary suitable reactant of stored sample handle sample from solid-phase media.Those subsequent steps that sample is emitted from its storage medium of being used for known in the state of the art.

Can be selectively, sample can be adsorbed onto during the entire reaction course on the solid-phase media, and for example, as previously mentioned, the DNA that is adsorbed onto on the solid-phase media finally can increase or being limited property restriction endonuclease digestion in same receiving vessel by PCR.In carrying out such process, preferably make DNA not be contained in protein contained in the sample that is adsorbed and other impurity.After sample being bored a hole and put receiving vessel into, the scheme described in the available embodiment 3 and 4 makes DNA not conform to protein and other impurity.

Finish the pcr amplification of the DNA that is purified, sample must experience the PCR process described in embodiment 3 and the embodiment 4.Finish this step, treating stations preferably is equipped with the hot/cold zone of four independent controls.In these zones, a zone preferably is about 4 ℃, and second area preferably is about 54 ℃, and the 3rd zone preferably is about 96 ℃, and the 4th zone preferably is about 37 ℃.

Adopt the DNA cloning of round pcr to be vulnerable to cross pollution.Here, the meaning of " cross pollution " is meant that a sample is by another sample contamination.Can be selectively, the meaning of " cross pollution " is meant that a kind of reagent is by another kind of reagent contamination., make the situation that this pollution occurs reduce to minimum herein, be used for during pcr amplification, sucking and the transfer pipet suction nozzle of exhaust fluid preferably only is used for a kind of independent reagent and also only is used for a kind of independent sample.Available automatic fluid induction system prevents such cross pollution.Preferably, finish this result, can produce for transfer pipet suction nozzle special purpose, that be similar to the aforesaid automatic fluid induction system of the present invention intubate.According to this situation of the present invention, behind transfer pipet suction nozzle of independent use, be connected this most advanced and sophisticated disengaging with the automatic fluid induction system.Then, before AFDS moves on to next sample or reagent, the suction nozzle of a new transfer pipet is placed on the automatic fluid induction system.In case install, just discharge a kind of independent fluid, and repeat to replace the process of the suction nozzle of transfer pipet with new transfer pipet suction nozzle.

After finishing processing, the result of selected sample and the sample of before being discerned by the discriminating station are interrelated.

Although the present invention can have various improvement and alternative form, will describe in detail and show specific embodiments of the invention by example later on.But, should understand, do not plan to make those examples that the present invention is confined to disclosed those specific forms, but opposite, the present invention has covered all forms of improving and can supplying to substitute that are in the spirit and scope of the present invention of being explained by appending claims.

EXAMPLE Example 1, collection and extraction DNA

Form with solution is applied to sodium dodecylsulphonate on the Whatman 3mm paper, thereby make 2% the sodium dodecyl sulfate solution of every square centimeter of nearly 50 μ l, the EDTA of 10mM, and the tris (free alkali) of 60mM, that is to say the sodium dodecylsulphonate of about every square centimeter of 1mg.Then, make described paper drying.

Use drop of blood to soak the above-mentioned paper of handling from various primates.Making to dye has the paper of blood drying, sends by common postal service, makes the time of posting be at least three days, then, comes to extract DNA from described paper with some standard methods, and these standard methods relate to the auxiliary proteolysis and the phenol extraction of detergent of described paper.Then, by digesting resulting DNA with restriction endonuclease and analyzing fragment, check the quality of this resulting DNA by agarose gel electrophoresis.

The quality of finding dna fragmentation is with the same high by the quality of the DNA that new blood produced.This shows, can extract DNA from the paper of being crossed by detergent-treatment, and the quality of this DNA is enough to be used in the vast majority of conventional purpose.The standing storage of embodiment 2, DNA half purification or that purify

Utilized some recording cards to store such as plasmid or the such DNA of some other virus replication type, these recording cards are made by the absorption paper that soaks in advance in a kind of solution of uric acid and tris (free alkali).Then make the further plasticising of these cards be used for further protection.In the time of in the time of later for a long time far away certain, may needing original material, and when in order, volume is little, preserve like this some one after another extremely during the clone of many gatherings, just established this flow process, the clone who comes those Dot blots of self aggregation for standing storage.Successfully stored sample by this way about 4 years.2.1 preparation DNA recording card

But preparation in quantity and store recording card are used when need waiting.The Whatman l paper that size is about 10cm * 15cm and has some correct positions that usefulness " prepared Chinese ink " (be colloid carbon ink note) marks is suitable, and available common " lead " pen (being the graphite pen) is made any special record on those cards.Preferably, mark card, so that help system ground stores and reclaim the DNA sample with the form of regular pattern.

The card that mark is crossed wraps in the clean paper, seals with paper tinsel then, and use autoclaving in the circulation of a drying.Afterwards, with the solution-treated of the tris (free alkali) of the uric acid of 40mM and 100mm they.Uratic effect is to make DNA avoid wearing out, and, if necessary, help desorb from the paper.Then, can preserve the recording card that these were handled, when needs.2.2DNA sample and their application

In the alkaline buffer of the dilution that contains EDTA, for example, in TE damping fluid (tris-EDTA, pH value 8.0), the DNA that taking-up will be stored.As an example, use the alkaline lysis method, and, handle the bacterial cultures of the about 1ml that contains plasmid, so that obtain plasmid and other DNA of the about 50 μ l in the TE damping fluid with a phenol extraction and a precipitation with alcohol.Aliquot with 5 μ l of every kind of DNA sample is made spot on the recording card of being handled by urate.2.3 soak into the card that is loaded with DNA with plastics

In case fill a card with the DNA sample,, then, this card immersed in the polystyrene of 12%w/v of chloroform that 5ml do not contain acid with regard to thorough air-dry DNA spot.Preferably by card being placed in the suitable tygon pouch, then polystyrene solution is poured in the described pouch, the polystyrene solution that scatters is so that thoroughly apply described card, and peel the tygon of having been made dirty then off, so just reached above-mentioned purpose.At room temperature make described card drying then.2.4 condition of storage

Card is stored in an airtight container of the refrigerating chamber that places a refrigerator (-15 ℃ approximately) easily, has in this airtight container such as the such drying agent of silica gel and the sodium carbonate particle of some a spot of dryings, so that remove the acid vapor of any trace.2.5 use the DNA on the card of plasticising

Make reservoir vessel can rise to room temperature, so that when the standing storage card, make unnecessary soaking reduce to minimum with the drying process process.Take out suitable card, differentiate relevant DNA spot, and downcut its small amount of sample.

Because the spot of 5 μ l is spread out regularly, in fact just can use scalpel blade they cross sections fully, so that extract a part of sample, for example, 1/4th spots.Then, this disposable plastic tube of putting into anacidity methenyl choloride, close the lid, and at room temperature on the blood spinner, rotated at least 30 minutes with about 5ml.This has just removed the polystyrene of protectiveness, but also the sample of having sterilized effectively.Then, eluted dna from sample, perhaps, for example (face as follows) handled sample and used this sample in position in dna sequence dna amplification reaction (PCR).

In an example, by utilizing plasmid pUC19 as the source of standard double-stranded DNA and utilize the source of M13 as single stranded DNA, check the two desorb from No. 1 paper of Whatman of strand and double-stranded DNA, this paper was soaked by the solution of the tris of the uric acid of 40mM and 100mM (free alkali).

For above-mentioned two kinds of situations, the sample of DNA is dried on the paper from those solution the TE damping fluid, then this wrapper in the polystyrene of protectiveness, just as above-mentioned.After, extract described paper with methenyl choloride, so that remove protective seam, with fresh TE buffer extraction DNA, and, assess the efficient of extracting by observing being converted of coli strain JM101 of causing by the DNA that is extracted.With undressed paper with soak with above-mentioned uric acid solution in advance and the dry paper of crossing carries out this flow process.Successfully reclaimed the active DNA that is converted into from the undressed paper, and the paper of above-mentioned processing provides about 100% the recovery far fewer than 10%.

In the environment of drying ,-15 ℃ store 36 months after, further check has been applied on the paper of handling also the DNA sample of packaged mistake as mentioned above.Make the plastics peel sample with methenyl choloride once more, by reclaiming sample DNA (just pUC19) with the simple extraction of TE damping fluid, and, the activity of check sample DNA as previously mentioned.The activity of DNA is again high, and the loss activity is not consistent with in fact having.Embodiment 3, original position is used stored blood DNA in PCR

Can come original position amplification to be stored in blood DNA on the filter paper of handling according to the present invention by PCR (PCR) technology.The paper that is used for the processing of this example is Whatman 3mm paper, and this solution comprises: the SDS of every square centimeter 2 micromolar uric acid, 8 micromolar tris (free alkali), 0.5 micromolar EDTA and 1 milligram.Handle stored blood DNA,, wash this stored blood DNA then, so that before DNA cloning, remove phenol and add suitable ion so that remove protein.3.1 solution solution A:

Single-phase phenol solution.Suitable mixture is the phenol of the oxine that contains 120mg of 50gm, and this potpourri has been that the 2 mercapto ethanol of 8.0 tris-acetic acid and 1.0ml is saturated with 10ml, 1.0M, pH value.After saturated, at room temperature shake, thoroughly remove and aqueous phase discarded.Solution B:

The isopropyl alcohol of 75%v/v, 25%v/v, 0.1M, pH value are 7.8 potassium acetate.Solution C:

The isopropyl alcohol of 75%v/v, the magnesium acetate of 25%v/v, 0.01M.

(note, available other some alcohol or similarly can mix water-soluble solvent, for example, n-propanol replaces the isopropyl alcohol in these solution.) 3.2 methods

Preferably in a test tube of making by a kind of material (for example tygon) of suitable anti-phenol, carry out all steps.(a) remove protein: for example, handle square shape paper piece that soaked by blood DNA, 0.5cm * 0.5cm, in the time of 45 ℃, handle about 1.5 hours (this temperature and time is not crucial) with the solution A of 1ml.Then dirty solution A is drawn in the refuse and goes, and wash described square shape paper piece three times rapidly with the solution A of 0.25ml again.Each flushing only needs several seconds, and washing fluid is drawn in the refuse goes immediately.(b) remove phenol and add suitable ion: the paper in the test tube in the flushing above-mentioned steps (a) rapidly in the solution B of three crowdes of 1ml.Flushing is to carry out in room temperature, and just adds subsequently washing fluid to be drawn in the refuse and go.Then, (this is the DNA that will use on the saturated described paper of magnesium ion, and removes last phenol to wash described paper in room temperature, with solution C 20 minutes.)。Solution C is drawn in the refuse goes, and once make the solvent seasoning of paper, make described paper vacuum drying then with pure isopropyl alcohol flushing.

Final DNA paper should be complete white, without any the tangible residue of the rufous of blood.Just all set it is used in the PCR reaction mixture now.(C) amplification of the DNA on the paper of handling:

Showed already that the DNA paper of handling as mentioned above was the suitable substrate of archaeal dna polymerase chain reaction (PCR) amplification of a kind of DNA of being used for.(i) DNA that sample extracted: use the DNA of the scheme extraction of standard on one's body from the resulting 10ml blood of male volunteers.The DNA filter paper of handling: the blood sample from same volunteer is applied directly on the filter paper of handling, and carries out follow-up processing as mentioned above.Described paper is cut into about 1mm ²Small pieces be used for PCR reaction.(ii) be used to No. 1 target of target of increasing: the zone of exon 2 on No. 1 chromosome, the n-Ras proto-oncogene.Used primer is: R1:5 ' TGA CTG AGT ACA AAC TGG TGG TG3 ' and R2:5 ' CTC TAT GGT GGG ATC ATA TTC A 3 '.

Size with the dna fragmentation of the resulting amplification of these primers is 110bp.No. 2 targets: the Y chromosome repetitive sequence that a kind of male sex is special.Primer is: 007:5 ' TGG GCT GGA ATG GAA AGG AAT GCA AAC 3 ' and 008:5 ' TCC ATT CGA TTC CAT TTT TTT CGA GAA 3 '.

Size with the dna fragmentation of the resulting amplification of these primers is 124bp.No. 3 targets: the Y chromosome repetitive sequence that a kind of male sex is proprietary.Used primer is: 004:5 ' GAA TGT ATT AGA ATG TAA TGA ACT TTA 3 ' and 006:5 ' TTC CAT TCC ATT CCA TTC CTT TCC TTT 3 '.

Size with the dna fragmentation of the resulting amplification of these primers is 250bp.(iii) PCR flow process

The DNA that is extracted (1 μ g) or about 1mm ²The DNA filter paper of processing put into the Eppendorf pipe of 0.5ml, and in the PCR reaction mixture, make 25 μ l, this PCR reaction mixture composed as follows: the Tris-HCl of 67mM (8.8@25 ℃ of pH value) 16.6mM ammonium sulfate 2mM MgCl ₂The deoxynucleotide of 0.01% (w/v) gelatin 0.1mM (dATP, dTTP, dCTP, dGTP) the Taq archaeal dna polymerase of every kind of primer of 0.25 μ g (for target separately) 0.25U.

Light mineral oil with 25 μ l covers said mixture, and carries out DNA cloning by 30 amplification cycles on " thermal cycler " of Pekin Elmer-Cetus company.

First circulation comprises:

DNA sex change 6 minutes.@94℃

Primer-DNA annealed 1 minute.@55℃

Taq archaeal dna polymerase extension 1 minute.@75℃

Be above-mentioned 29 circulations subsequently, before being cooled to 4 ℃ DNA sex change 1 Fen Zhong @94 ℃ and Fen Zhong @72 ℃ of circulation extension time 10 in the end at reaction mixture.

After amplification, analyze the aliquot of the PCR potpourri of 10 μ l by the electrophoresis on the polyacrylamide gel of 15% (w/v).By with ultraviolet ray irradiation ethidium bromide institute stained gel, can visually see the target DNA that increased.For being analyzed as follows as shown in Figure 1: swimming lane: 1-3:1 target, swimming lane 1:1 μ gDNA, swimming lane 2:1mm from the product of the PCR of the DNA that is extracted and the DNA filter paper handled ²Filter paper, swimming lane 3: do not have the contrast of DNA; Swimming lane 4-5:2 target, swimming lane 4:1 μ g DNA, swimming lane 5:1mm ²Filter paper; Swimming lane 6: do not have the contrast of DNA; Swimming lane 7-9:3 target, swimming lane 7:1 μ g DNA, swimming lane 8:1mm ²Filter paper, swimming lane 9: do not have the contrast of DNA.The big tick marks of swimming lane S:DNA (pUC19/HpaII digestion).

Shown in the result clearly illustrated that DNA and do not changed by any way owing to be stored on the filter paper of the present invention, and, as described here, can use this DNA in position.Embodiment 4: method and material

Collect and store the other blood sample.Extract hematoglobin protein by the method for using solution A as in Example 3 from the paper of handling.Also utilize two flow processs that relate to a kind of improved solution A (solution A 4 and solution A 7) from sample, to extract hematoglobin protein.(B.Y.T.Co.Baycity MI) extracts on those samples in the Eppendorf pipe prepared and that be placed on 1.7 μ l at the small opening with 0.5 * 0.5mm.

Then, the DNA that does not have adsorbed proteins with the PCR method amplification.4.1 the collection of sample and storage

All blood samples are stored on No. 1 paper of Whatman that the tris (free alkali) with the EDTA of sodium dodecylsulphonate, 10mM and 60mM handled.Then, make described paper drying, and handle with the trihydroxy methyl aminomethane buffer solution of the uric acid of 40mM and 100mM that (uric acid prevents the aging and desorb (Flinders technology, Australia) from the described paper of DNA.Then, use the blood that provides by lab assistant to soak the filter paper of handling, and seal up for safekeeping at 4 ℃ with paper tinsel.4.2 extracting method

Method 1: add the solution A (oxine of the ultrarapture liquid phenol of 50ml, the TE damping fluid of 10ml 1M, 120mg, the 2-β mercaptoethanol of 1ml) of 500 μ l for each test tube.Shelve 15 minutes, so that the water-bearing zone is separated with organic matter layer.Remove the water-bearing zone.55 ℃ of incubations 1.5 hours.With solution A flushing 1 time.With solution B (the pH value of 75%v/v isopropyl alcohol, 25%v/v is 7.8 potassium acetate) flushing 3 times.With solution C (isopropyl alcohol of 75%v/v, the magnesium acetate of 25%v/v) room temperature incubation 20 minutes.Then, with the paper vacuum drying of pure isopropanol solvent drying 10 minutes.Paper should be white, and is easy to carry out PCR.

Method 2: give the solution A 4 that each test tube adds 500 μ l (the TE damping fluid of the 1M of phenol/methenyl choloride of 50ml/isopentyl, 10ml, the oxine of 120mg, do not add the 2-beta-mercaptoethanol).Shelved 15 minutes, and removed the upper strata.55 ℃ of incubations 1.5 hours.With solution A 4 flushings once.With solution B (the pH value of the isopropyl alcohol of 75%v/v, 25%v/v is 7.8 potassium acetate) flushing three times.With solution C (isopropyl alcohol of 75%v/v, the magnesium acetate of 25%v/v) room temperature incubation 20 minutes.Then, make the paper vacuum drying 10 minutes of solvent seasoning with pure isopropyl alcohol.Paper should be white, and is easy to carry out PCR.

Method 3: add 500 μ l solution A 7 (a kind of suitable digestion damping fluid, for example, the tris of 10mM, the EDTA of 5mM, 0.5% pH value are the Proteinase K that 7.8 SDS adds 3 μ l) for each test tube.55 ℃ of incubations 1 hour.Made enzyme-deactivating 10 minutes at 95 ℃.55 ℃ of incubations 1.5 hours.With solution A 7 flushings once.With solution B (the pH value of the isopropyl alcohol of 75%v/v, 25%v/v is 7.8 potassium acetate) flushing three times.With solution C (isopropyl alcohol of 75%v/v, the magnesium acetate of 25%v/v) room temperature incubation 20 minutes.Make the paper vacuum drying 10 minutes of solvent seasoning with pure isopropyl alcohol.Paper should be white, and is easy to carry out PCR.4.3PCR amplification

In 9600 type thermal cyclers of Perkin Elmer company, carry out all reactions.In order to ensure the quality of products, guarantee to carry out all reactions with positive and negative experiment contrast.

Amplitype ^HLA DQ _α(CA) magnesium chloride+paper of the 8mM of 25 μ lMaster Mix+25 μ l mineral oil with 40 μ l in the gene magnification thin-walled test tube of 0.5 μ l covers for Pekin Elmer company, Foster city.The parameter of thermal cycler is: 94 ℃/30 seconds, 94 ℃/10 seconds, 60 ℃/10 seconds, 72 ℃/10 seconds (32X) 72 ℃/10 minutes, soaked at 4 ℃ subsequently.

Amplitype ^TM(CA) magnesium chloride+paper of the 60mM of 40 μ l MasterMix+40 μ l is in little amplification test tube (Micro-Amp) for Perkin Elmer company, Foster city for PM.The parameter of thermal cycler is: 95 ℃/1 minute, 95 ℃/30 seconds, 63 ℃/30 seconds, 72 ℃/30 seconds (32X) 72 ℃/10 seconds, soaked at 4 ℃ subsequently.Use suggested scheme amplification by Perkin Elmer company.Make DQ _αAnd Amplitype ^TMTypical case representative.

Gene Print ^TM(Promega, Madison WI) increase according to the Promega scheme of revising in September, 93 in STR system (TH01).The preparation of embodiment 5 intubate

This example is a preferred embodiment about preparation intubate of the present invention as shown in Figures 2 and 3.(Wilmington Delaware) obtains that (ROSYS, Wilmingron Delaware) work the standard cannula of using with ROSYS 3300 by ROSYS.The long 15cm of this intubate.The internal diameter of main mouth 6 is 0.040mm.

Use the titanium drill bit, at contiguous main mouth 62mm place, by the wall of intubate upwards, getting out two, each other approximately to become that 180 ° of angles separate, diameter be auxiliary mouthful 7 of 0.020mm.Sliding with file main and auxiliary mouth 6,7 file finishings.

Ready-made intubate and ROSYS 3300 (ROSYS, Wilmington, Delaware) automatic fluid induction system connection.In case on connecting, just use automatic fluid induction system to come to suck and exhaust fluid from fluid reaction system with solid with intubate disclosed in this invention.Described solid is the little sample that contains as the solid phase storage medium of embodiment 1 described DNA.The preparation of embodiment 6 biopolymer purification devices

This example is the standard polypropylene acrylamide gel that will be used for constituting the biopolymer purifying plant about preparation.Can use other to have the hydrophilic polymer of similarity.

Be prepared as follows polymkeric substance:

The water of 12ml

The acrylamide of 2g

The N of 40mg, the N-methylene-bisacrylamide

0.2ml tris EDTA (500mm, the pH value is 8.0)

The TEMED of 12ul

The 7mg ammonium persulfate

Polymer reaction potpourri cast drilling depth cun suitable (for example, 8.0mm it is dark, 50mm is long and 30mm is wide) rectangular block in, this rectangular block has some rules dimples isolated, little, that rise (these dimples generation micropores) in its basal surface.Each about 4.0mm in single hole is dark, and diameter is 4.0mm.Make polyacrylamide can place mould about 30 minutes, and, be placed in the distilled water then and spend the night.Final product is about 14% water-soluble plastics, but after dehydration, said apparatus is about 12.2% dry weight and polymkeric substance.

For the purpose that stores and distribute, for example, said apparatus can be sealed in the polybag of being made by tygon.The material of described device has extremely long shelf-life MIN looking after when storing, but preferably at 25 ℃ or be lower than 25 ℃, and in the dark.

Adopt the method for biopolymer purifying plant

Typically, putting in each independent micropore of the said apparatus that is in room temperature, with artificial or use by the intubate of automatic manipulation control and accomplish this point from the DNA reaction mixture of the DNA by pcr amplification.

For after will reducing pollution level and being the sufficiently long time (for example 2 to 30 minutes), can come the sampling of the dna solution in the micropore by artificial or automated intubate, and DNA isolation, perhaps, for example analyze this DNA with Capillary Electrophoresis.

But, in order to reuse, can clean the material of said apparatus by the flushing of spending the night with distilled water, described manufacture of materials is more cheap, and abandons after use usually, and any of those samples may subsequently to avoid polluting.

To it will be evident to one of ordinary skill in the art that above-mentioned amplification and the Overall Steps that is used for the purification of blood DNA is particularly suitable for finishing automatically.

List of references:

1.McCabe, E.R.B., Huang, S.E., Seltzer, W.K. and Law, M.L (1987).Human genetics.75：213-216。

2.Ames, wait people (1981).Institute of NAS periodical.78：6858-6862。

3.Kwok Lai Lam, Fong, D., Lee, A and Ken Ming D.Lui. (1984), inorganic biochemistry magazine

22：241-248

4.Singer, B. and Fraenkel-Conrat, H. (1965). and biological chemistry 4:226-223.

Sequence table (1) general information

(i) applicant:

(A) title: Flinders Technologies Pty incorporated company

(B) block: the Flinders university of South Australia

(C) city: Bedford Park

(D) state: South Australia

(E) country: Australia

(F) postcode (postal region):

(ii) denomination of invention: the device and the method that are used for storage, purification or the reaction of biopolymer

(iii) sequence number: 6

(iv) computer-readable form:

(A) media type: floppy disk

(B) computing machine: IBM PC compatible

(C) operating system: PC-DOS/MS-DOS

(D) software: PatentIn Release#1.0, version: #1.25 (EPO)

(v) current application materials:

Application number:

(vi) application materials formerly:

(A) application number: US 08/320,041

(B) applying date: on October 7th, 1994

(C) classification number:

(vi) application materials formerly

(A) application number: US 08/159,104

(B) submission date: on November 30th, 1993 (2) SEQ ID NO:1 information:

(i) sequence signature:

(A) length: 23 base-pairs

(B) type: nucleic acid

(C) chain: strand

(D) topological structure: linearity

(ii) molecule type: DNA (genomic)

(xi) sequence explanation: the information of SEQ ID NO:1:TGACTGAGTA CAAACTGGTG GTG 23 (2) SEQ ID NO:2:

(i) sequence signature:

(A) length: 22 base-pairs

(B) type: nucleic acid

(C) chain: strand

(D) topological structure: linearity

(ii) molecule type: DNA (genomic)

(xi) sequence explanation: the information of SEQ ID NO:2:CTCTATGGTG GGATCATATT CA 22 (2) SEQ ID NO:3:

(i) sequence signature:

(A) length: 27 base-pairs

(B) type: nucleic acid

(C) chain: strand

(D) topological structure: linearity

(ii) molecule type: DNA (genomic)

(xi) sequence explanation: the information of SEQ ID NO:3:TGGGCTGGAA TGGAAAGGAA TGCAAAC 27 (2) SEQ ID NO:4:

(i) sequence signature:

(A) length: 27 base-pairs

(B) type: nucleic acid

(C) chain: strand

(D) topological structure: linearity

(ii) molecule type: DNA (genomic)

(xi) sequence explanation: the information of SEQ ID NO:4:TCCATTCGAT TCCATTTTTT TCGAGAA 27 (2) SEQ ID NO:5:

(i) sequence signature:

(A) length: 27 base-pairs

(B) type: nucleic acid

(C) chain: strand

(D) topological structure: linearity

(ii) molecule type: DNA (genomic)

(xi) sequence explanation: the information of SEQ ID NO:5:GAATGTATTA GAATGTAATG AACTTTA 27 (2) SEQ ID NO:6:

(i) sequence signature:

(A) length: 27 base-pairs

(B) type: nucleic acid

(C) chain: strand

(D) topological structure: linearity

(ii) molecule type: DNA (genomic)

(xi) sequence explanation: SEQ ID NO:6:TTCCATTCCA TTCCATTCCT TTCCTTT 27

Claims (18)

1. One kind be used to differentiate, system that spike and processing are stored in the biological sample on a kind of solid-phase media, described system comprises:

   (a) store the solid-phase media of biological sample above a kind of;

   (b) discern and recall the discriminating station that is stored in the above-mentioned biological sample on the described solid-phase media for one;

   (c) processing is stored in the treating stations of the described biological sample on the described solid-phase media;

   (d) one from described discriminator until described treating stations all spike be stored in the spike station of the biological sample on the described solid-phase media; And

   (e) system ensemble data center, this system ensemble data center provides the interactive network of described discriminating, spike and treating stations.
2. 2. the system as claimed in claim 1 is characterized in that described system ensemble data center is by manually carrying out work.
3. 3. the system as claimed in claim 1 is characterized in that described system ensemble data center is a computer hardware and software systems.
4. 4. the system as claimed in claim 1 is characterized in that described system ensemble data center carries out work by artificial and computer hardware and software systems.
5. 5. the system as claimed in claim 1 is characterized in that the described biological sample that is stored on the described solid-phase media is the DNA sample.
6. 6. the system as claimed in claim 1 is characterized in that the described solid-phase media that is used to store described biological sample comprises a kind of solid-phase matrix, and this solid-phase matrix has absorption or is attached to the component of a kind of effective dose on it, and this component prevents dna degradation.
7. 7. system as claimed in claim 6 is characterized in that the described component of dna degradation that prevents comprises a kind of protein denaturant.
8. 8. component as claimed in claim 6 also comprises a kind of free radical trap.
9. 9. system as claimed in claim 5, the DNA in the described sample that is stored in the DNA on the described solid-phase media that it is characterized in that in described treating stations purifying.
10. 10. the system as claimed in claim 1 is characterized in that by the people by manually finishing the work at described discriminating station and described spike station.
11. 11. the system as claimed in claim 1 is characterized in that described treating stations is an automated automatic fluid induction system.
12. 12., comprising as automated automatic fluid induction system as described in the claim 11:

    (a) a kind of flow device of constructing and being positioned to suction and discharging reacting fluid;

    (b) intubate that be communicated with the flowable fluid of described flow device, that operationally locate, described intubate comprises:

    (i) wall of determining a fluid passage;

    The (ii) mechanism of the described intubate that is operably connected, described intubate is communicated with the flowable fluid of described automated automatic fluid exhaust system;

    (iii) a main fluid sucks and floss hole;

    (iv) at least one auxiliary fluid sucks and floss hole, it is characterized in that making the main and auxiliary fluid suction and the floss hole of described intubate to construct so each other and settle, that is, make and when a fluid reaction system sucks also exhaust fluid, to prevent that fluid from flowing to or flowing out the possible obstruction in described fluid passage of above-mentioned intubate when above-mentioned automated automatic fluid induction system.
13. 13. one kind that use with a kind of automatic fluid induction system, be used for optionally sucking and the intubate of exhaust fluid; Described intubate comprises:

    (a) wall of determining a fluid passage;

    (b) mechanism of the described intubate that is operably connected, described intubate is communicated with by streaming flow with above-mentioned automatic fluid induction system;

    (c) a main fluid sucks and floss hole;

    (d) at least one auxiliary fluid sucks and floss hole, it is characterized in that making the suction and the floss hole main and that assist of described intubate to construct so each other and settle, that is, make and to flow to or to flow out may the blocking of described fluid passage of above-mentioned intubate when described automatic fluid induction system prevents fluid during from a fluid reaction system suction and exhaust fluid.
14. 14. intubate as claimed in claim 13 is characterized in that described main fluid sucks and described at least one the auxiliary fluid of floss hole distance sucks and the interval of floss hole is at least about 2mm.
15. 15. intubate as claimed in claim 13 is characterized in that the described main fluid suction and the fluid suction and the floss hole in floss hole and described at least one auxiliary distally are connected together continuously.
16. 16. intubate as claimed in claim 13 is characterized in that described main fluid suction and floss hole and described at least one auxiliary fluid sucks and floss hole is the assembly of the porous welded bond of some not open and flat (non-flat).
17. 17. intubate as claimed in claim 13 comprises:

    (a) elongated shaft with a far-end;

    (i) described main fluid sucks and floss hole is included in a hole in the described far-end of above-mentioned shaft; And

    (ii) described at least one auxiliary mouth is spaced apart with described far-end and described main mouth along described shaft.
18. 18. one kind is used for adopting polymer gel to remove the method for accessory substance from the biopolymer of biopolymer reaction mixture, described method comprises:

    (a) provide shape can be used for holding the polymer gel of the fluid of a certain volume;

    (b) in the polymer gel of described shape, put into the biopolymer reaction mixture of a certain volume;

    (c) make above-mentioned biopolymer reaction mixture contact a period of time, be enough to make material some diffusions when not applying electric field, to occur during this period of time from described biopolymer reaction mixture with the polymer gel of described shape, and

    (d) from the polymer gel of above-mentioned shape, remove described biopolymer.

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