CN208949249U

CN208949249U - The pipe cover frame of PCR pipe lid

Info

Publication number: CN208949249U
Application number: CN201820631899.0U
Authority: CN
Inventors: 杨尚鑫; 胡彬; 刘杰; 蔡媛媛; 陶施芳
Original assignee: Shaoxing Xun Xun Kang Biological Technology Co Ltd
Current assignee: Shaoxing Xun Xun Kang Biological Technology Co Ltd
Priority date: 2018-04-28
Filing date: 2018-04-28
Publication date: 2019-06-07
Anticipated expiration: 2028-04-28

Abstract

The utility model discloses the pipe cover frame of PCR pipe lid, pipe cover frame includes pedestal and protrusion, and at least two convex to form a pipe lid positioning unit.Manually PCR reaction tube is covered, not only low efficiency, and easily finished product is polluted, PCR reaction tube is covered automatically by mechanized operation, is necessarily required to pipe cover frame compatible with mechanized operation, the utility model provides a kind of pipe cover frame of PCR pipe lid, when pipe lid is combined with protrusion, pipe lid can not relative positioning part rotation or translation, only after fixture clamping pipe lid, pipe lid is removed from locating piece.

Description

The pipe cover frame of PCR pipe lid

Technical field

The utility model relates to nucleic acid extraction fields, more specifically, it is related to the pipe cover frame of PCR pipe lid.

Background technique

Nucleic acid, such as DNA or RNA, the extraction of nucleic acid mainly use two kinds of extraction sides of centrifugal column method and paramagnetic particle method at present Method.Traditional centrifugation column technology also applies film to carry out nucleic acid purification, and the principle of its film belongs to silicon adsorption method, and this film is only right Nucleic acid has stronger affinity and adsorption capacity.Protease is applied to split under the conditions of 56 DEG C of temperature to sample when operation first Then lysate is added in pillar and is centrifuged by solution, nucleic acid is adsorbed on film, and other impurities are thrown out of, and finally uses eluent Nucleic Acid Elution is got off.The method extraction effect is good, but while cracking needs to heat, and repeatedly centrifugation is needed when nucleic acid purification, whole It is or so a extraction process time-consuming 2 hours, at high cost, complicated for operation, time-consuming, relies on laboratory environment, and be difficult to extract Degradable RNA.

Paramagnetic particle method nucleic acid extraction is a kind of novel nucleic acids extractive technique using nano biological magnetic bead as carrier, and nucleic acid molecules can Specific recognition occurs with the silicone hydroxyl of magnetic bead surfaces and combines, aggregation or dispersion occur under the action of external magnetic field, thus Nucleic acid is carried out to isolate and purify.Paramagnetic particle method nucleic acid extraction kit generally comprises cracking, combination, washing, elution four mainly at present Step, each step can be realized alone or in combination again by a variety of different methods.This method thoroughly gets rid of traditional core extraction process The processes hand-manipulated such as middle centrifugation, extraction supernatant, to realize that the automation of nucleic acid is extracted.

It is sample thermal cracking when heating during nucleic acid extraction, nucleic acid low temperature (or room temperature) Shi Caiyu magnetic bead combines, because This refrigeration is cooling to can be shortened time after nucleic acid cleavage with magnetic bead ining conjunction with, accelerates the speed of nucleic acid extraction, shortening extraction time, but Be the problem is that: manually PCR reaction tube is covered, not only low efficiency, but also easily finished product is polluted, passes through machine Toolization operation automatically covers PCR reaction tube, is necessarily required to pipe cover frame compatible with mechanized operation.

Summary of the invention

The purpose of this utility model is to provide the pipe cover frames for the PCR pipe lid that is suitable for clamping pipe lid and be covered.

The technical scheme adopted by the utility model to solve the technical problem is as follows: the pipe cover frame of PCR pipe lid, pipe cover frame include Pedestal and protrusion, at least two convex to form a pipe lid positioning unit.

Further, there is the base item of protrusion on pedestal, protrusion is set on a base item.

Further, pipe cover frame is set in the operating space of instrument for extracting nucleic acid.

Further, the raised quantity of each pipe lid positioning unit is identical as the prominent lid of pipe lid to be positioned, and each protrusion is corresponding One prominent lid.

Further, there is base item on pedestal, base item protrudes from pedestal, and for bump array on base item, the raised center of circle is located at base On the middle line of item.

Further, the matched of raised height and prominent lid.

Further, pipe cover frame has one or more pipe lid positioning units, has spacing between adjacent pipe lid positioning unit.

Further, pipe cover frame has one or more pipe lid positioning units, and adjacent pipe lid positioning unit is mutually aligned.

Advantage is the utility model compared with prior art: manually PCR reaction tube is covered, not only low efficiency, and And easily finished product is polluted, PCR reaction tube is covered automatically by mechanized operation, is necessarily required to and mechanized operation Compatible pipe cover frame.When pipe lid with protrusion is combined when, pipe lid can not relative positioning part rotation or translate, only when fixture clamping After pipe lid, pipe lid is removed from locating piece.

Detailed description of the invention

Fig. 1 is overall structure figure of the invention.

Fig. 2 is the overall structure figure with door.

Fig. 3 is the schematic diagram of cracking region.

Fig. 4 is the schematic diagram of heating load.

Fig. 5 is bar magnet-bar magnet set of modules and drive transmission device structural schematic diagram.

Fig. 6 is bar magnet-bar magnet set of modules structural schematic diagram.

Fig. 7 is the schematic diagram in pre-treatment region.

Fig. 8 is the structural schematic diagram that the magnetic patch in pre-treatment region is placed.

Fig. 9 is the schematic diagram of capping apparatus.

Figure 10 is the schematic diagram of intermediate plate.

Figure 11 is the schematic diagram of pipe cover frame.

Figure 12 is the schematic diagram of townhouse PCR reaction tube.

Figure 13 is the schematic diagram of townhouse pipe lid.

Figure 14 is the schematic diagram of ultraviolet lamp.

Figure 15 is the sample distribution figure of first group of anti-pollution test of DNA reference material.

Figure 16 is the sample distribution figure of second group of anti-pollution test of DNA reference material.

Figure 17 is the sample distribution figure of the anti-pollution test of RNA reference material.

It is identified in figure: operating space 1, exhaust fan 11, ultraviolet lamp 12, cabinet 14, door 141, top plate 142, bottom plate 143, side Plate 144；Bar magnet-bar magnet set of modules 2, bar magnet 21, bar magnet set 22, bar magnet frame 211, bar magnet stock 221, baffle 23, through-hole 231, Slot 222；Drive transmission device 3；Cracking region 4, heating load 41 heat cooling piece 42, accommodating chamber 411, and sleeve 412 passes Hot plate 413, temperature control plate 421；Pre-treatment region 5, pipe support 51, pedestal 511, jack 512, base item 513；Capping apparatus 6, gland drive Dynamic component 61, clamp assembly 62, gland portion 621, clamping portion 622, intermediate plate 6221, groove 6222, extended segment 6223, clamping driving Portion 623, clamping jaw 6231, upper plate 62211, lower plate 62212；Pipe cover frame 8, pipe cage 81, locating piece 82, base item 83, pipe lid 84, lid Plate 841, lid 842 of dashing forward, folder portion 841 to be installed.

It is described with reference to the accompanying drawings

Structure of the present invention or technical term used in these are described further below.These explanations are only Be only using example way be illustrated it is of the invention by the way of how to realize, any limit can not be constituted to the present invention System.

Sample

It include biofluid (such as casing fluids or clinical sample with the sample that detection device of the invention can detecte This).Liquid sample or fluid sample can from the sample of solid-state or semisolid, including excreta, biological tissue and Food samples.The sample of solid-state or semisolid can be converted to liquid sample using any method appropriate, such as mix, smash It is broken, macerate, be incubated for, dissolution or in a suitable solution (such as water, phosphate solution or other buffer solutions) using enzymatic hydrolysis make With digestion of solid sample." biological sample " includes deriving from animal, plant and food samples, for example including from human or animal Urine, saliva, blood and its ingredient, vaginal fluid, sperm, excrement, sweat, secretion, tissue, organ, tumor, tissue and organ Culture, cell culture and medium.

Method for extracting nucleic acid

In some embodiments, method for extracting nucleic acid, including cleavage step and transfer step, cleavage step complete nucleic acid The nucleic acid of thermal cracking and magnetic bead absorption thermal cracking release, cleavage step are completed in cracking region；The magnetic bead for being adsorbed with nucleic acid is turned It moves on in pretreatment liquid, complete transfer step；Pretreatment liquid is located at pre-treatment region, and cracking region and pre-treatment region are mutually only It is vertical, as shown in Figure 1.As long as in this way, nucleic acid extraction can be realized in step transfer.Cracking region and pre-treatment region refer to independently of each other Be not intersect between cracking region and pre-treatment region, cracking region is the isolated area cracked, and pre-treatment region is Place the isolated area of pretreatment liquid.Magnetic bead is transferred to after pretreatment liquid, and the solution in the PCR reaction tube in pre-treatment region is made To be the solution that can directly carry out molecular diagnosis processing.Such as quantitative fluorescent PCR, regular-PCR, digital pcr and constant temperature PCR etc..

The volume of each cracking station is 100~200 microlitres；The volume of each pre-treatment station is 100~200 microlitres, The container and the container of pre-treatment station for cracking station are made of independent thin-wall tube respectively, and multiple thin-wall tubes are embarked on journey or in column It is linked to be townhouse pipe.For example, the independent PCR reaction tube or townhouse PCR reaction tube that the use of specification are 100 microlitres or 200 microlitres, such as scheme Shown in 12.

In some embodiments, there is distance between cracking region and pre-treatment region, as shown in figure 12.In this way, cracking zone The temperature change in domain does not interfere with pre-treatment region, and pretreatment liquid is avoided unnecessary evaporation or volatilization occur, place before keeping The liquid measure for managing liquid is accurate.

In some embodiments, in cleavage step, cracking region can accommodate multiple hole locations simultaneously and carry out thermal cracking, each Hole location contains lysate；Pre-treatment area has multiple jacks 512, and each jack 512 can accommodate a container, cleavage step Each hole location is corresponding with a hole location in pre-treatment region 5；The magnetic bead of all hole locations of cracking region 4 is transferred to preceding place simultaneously Manage the corresponding hole site in region.

In some embodiments, each cracking station and pre-treatment station are made of independent thin-wall tube respectively.It can be An individual thin-wall tube is placed in each cracking station (or each pre-treatment station), is also possible to multiple thin-wall tubes and is linked to be Townhouse pipe, each one of thin-wall tube cracked when in station in townhouse pipe.Such as existing 8 townhouse PCR reaction tube or 12 Arranging PCR reaction tube is also multi-hole position container.Each hole location is independent thin-wall tube and refers to: adjacent hole location not share common sidewalls. Therefore, it reduces intersection heat transfer between adjacent hole location to the greatest extent, improves the temperature accuracy of each hole location, make the cracking of all hole locations The temperature consistency of liquid is good.

The magnetic bead for being adsorbed with nucleic acid is transferred in pretreatment liquid by the present invention using bar magnet method from lysate, primary to shift Nucleic acid extraction is completed, it is not necessary that magnetic bead is repeatedly washed or eluted.Pretreatment liquid refers to using for molecular diagnosis anti- Answer liquid.Here nucleic acid can be RNA or DNA.

Cracking step

In some embodiments, lysate and sample are contained in cracking station.Preferably, preset magnetic bead in lysate, Or it is added in pyrolytic process in magnetic bead or the temperature-fall period of thermal cracking and magnetic bead is added.Optimal mode is: preset magnetic in lysate Pearl, thermal cracking is completed, temperature declines, before protein renaturation, and magnetic bead is in conjunction with nucleic acid, when temperature drops to protein renaturation When (room temperature), magnetic bead has been firmly combined with nucleic acid, reduces influence of the protein to nucleic acid, improves magnetic bead to the adsorption rate of nucleic acid, Be conducive to obtain enough, complete nucleic acid.

When thermal cracking, heating temperature needs to reach 80 DEG C or so or above of high temperature, liquid evaporation will occurs and generates Aerosol, to avoid polluting between thin-wall tube, in some preferred schemes, in thermal cracking temperature-rise period, guidance gas is molten Glue discharge.Preferably, guidance aerosol is upwards or upper side is discharged.

In some embodiments, container is inserted into heating load.The abundant wrapping container of heating load is able to maintain entirely The temperature consistency of thin-wall tube avoids occurring aerosol in cooling because of the temperature difference of hole location inner wall and condensing the drop to be formed, from And prevent the possibility for leading to pollution because of aerosol.

In some embodiments, lysate refrigeration is cooled down in cleavage step, after the completion of heating.It is sample hot tearing when heating Solution, nucleic acid low temperature (or room temperature) Shi Caiyu magnetic bead combine, thus freeze it is cooling can be shortened after nucleic acid cleavage in conjunction with magnetic bead when Between, accelerate the speed of nucleic acid extraction, shortens extraction time.Entire nucleic acid extraction Row control is completed within half an hour, is improved normal Temperature is lower to extract the easy in inactivation, the success rate of degradable material such as RNA.

In some embodiments, heating load 41 is heated refrigeration module covering, therefore all hole location quilts of cracking region The temperature rise of Synchronous Heating, all hole locations is consistent.Preferably, heating and cooling module is by the seamless spliced shape of multiple heating refrigeration units At.It heats up or cools down in this manner it is ensured that entire cracking region is synchronous, keep all PCR reaction tube hole location temperature consistent.

It is using the operation that bar magnet and bar magnet cover the magnetic bead in sufficiently absorption lysate: makes the bar magnet set 22 with bar magnet 21 Slowly from up and down motion.The upper limit position of 22 movement of bar magnet set is that bar magnet covers 22 bottoms arrival liquid level, 22 movement of bar magnet set Lower position is the bottom wall and side wall that bar magnet set 22 does not touch container.Lower position can be obtained through test.This way be because are as follows: The suction of bar magnet set 22 concentrates on end, therefore, allows the end of bar magnet set 22 to move up and down in the whole region of lysate, fills The magnetic bead for dividing absorption to be suspended in lysate.It, will not be because of liquid when the slow movement of bar magnet set 22 refers to bar magnet 22 movement of set Tension and cause magnetic bead with respect to bar magnet set 22 displacement.

Bar magnet set 22 and hole location clearance fit, the upper and lower translation in liquid regions of bar magnet set 22.That is, bar magnet set 22 It can move freely hole location is not rebuffed, but the suction of bar magnet set 22 can cover the cross section of entire thin-wall tube substantially, therefore When bar magnet set 22 is mobile, the magnetic bead of each sectional position can be adsorbed onto.Alternatively, bar magnet set 22 and the tube wall of hole location have distance, Bar magnet set 22 had both done upper and lower translation in liquid regions or had done left and right translation or formula movement of drawing a circle.For example it draws circle or draws oval equal It is mobile to belong to formula of drawing a circle.

Magnetic bead transfer process

Magnetic bead is transferred to the correspondence hole location in pre-treatment region using bar magnet method from cracking station, all magnetic beads turn simultaneously It moves.

In some embodiments, in cleavage step, bar magnet 21 and bar magnet set 22 is located at except aerosol discharge path.To Aerosol is avoided to be attached to bar magnet set 22 and pollute.Preferably, lysate is restored to room temperature, the bar magnet set with bar magnet 21 In 22 insertion lysates, magnetic bead is adsorbed.Preferably, the liquid without attaching liq or attachment is only positioned at bar magnet set 22 on bar magnet set 22 Surface is not dripped, then bar magnet set 22 is slowly moved to above the PCR reaction tube for containing pretreatment liquid.Bar magnet set 22 is slow Slowly the speed moved is not fallen with magnetic bead, bar magnet covers the liquid that 22 surfaces are adhered to and do not drip is advisable.

In some embodiments, it when bar magnet set 22 takes out from cracking station, is equipped in the path that bar magnet set 22 takes out Baffle 23, baffle 23 block container, container are avoided to be taken away by bar magnet set 22.

Magnetic bead discharges process

Magnetic bead is transferred to pretreatment liquid from lysate using bar magnet method, before being discharged into from the magnetic bead taken out in lysate In treatment fluid.

In some embodiments, as shown in Figure 7 and Figure 8, pre-treatment region 5 has pedestal 511, and the setting of pedestal 511 accommodates The jack of container, each jack 512 are a pre-treatment station, and the quantity of jack 512 has multiple.Preferably, around jack 512 Magnet is set；The corresponding bar magnet 21 of each jack 512 and bar magnet set 22.Therefore, the corresponding cracking of each pre-treatment station All magnetic beads of station, cracking region 4 can be covered by bar magnet-bar magnet while be transferred to pre-treatment region 5.Pedestal 511 has Support stiffness, pedestal 511 are installed on operating space 1.

In some embodiments, pedestal 511 is equipped with base item 513 outstanding, and jack 512 is arranged along base item 513.Pedestal There are multiple base items 513 on 511, there is distance between base item 513, each base item 513 there are the multiple jacks of setting, each base item 513 Jack 512 forms a pipe support unit.Base item 513 all has support stiffness.Pedestal 511 and 513 one of base item or base item 513 with 511 fixing assembling of pedestal.Preferably, base item 513 holds the container being put into it completely, such as PCR reaction tube.

In some embodiments, magnet is set under jack 512, alternatively, magnet is set between adjacent jack 512, Or each jack 512 is surrounded with magnet.Jack 512 is cylinder, alternatively, the cylindrical section of jack 512 and PCR reaction tube Match, alternatively, the shape of jack 512 reacts tube shape matching with the PCR that need to be placed.

Magnetic bead is transferred in pretreatment liquid, and bar magnet 21 22 is withdrawn from bar magnet set, and bar magnet 21 is statically placed in pretreatment liquid, after standing Bar magnet set 22 is axially vibrated.The vibration of bar magnet set 22 is not splashed out outward with pretreatment liquid to be advisable.Bar magnet set 22 vibration, together with The liquid tension of pretreatment liquid is discharged into magnetic bead in pretreatment liquid from 22 surfaces of bar magnet set.Magnet attracts bar magnet to cover 22 surfaces Magnetic bead, accelerate bar magnet set 22 on magnetic bead release rate and efficiency.

In some embodiments, magnetic patch be set to 512 bottom of jack perhaps magnetic patch be set to jack 512 side or insert Magnetic patch is all arranged in the bottom and side in hole 512.When bar magnet is stood, the suction of magnetic patch attracts magnetic bead to be detached from bar magnet set 22.Bar magnet set 22 vibration when, liquid tension, bar magnet set 22 whipping and magnetic patch suction make jointly magnetic bead with respect to bar magnet set 22 generate be displaced, It falls into PCR pretreatment liquid.

Bar magnet 21 is permanent magnet or electromagnet, and bar magnet 21 from bar magnet set 22 when withdrawing, 21 loss of excitation of bar magnet.In this way, magnetic Stick 21 will not attract magnetic bead to be displaced when withdrawing bar magnet set 22, and magnetic bead is concentrated mainly on bottom, be conducive to magnetic patch and attract magnetic bead, magnetic Pearl sufficiently discharges.

The capping in pre-treatment area

After the transfer step, closing step covers all thin-wall tubes of the same pipe support unit to closing step simultaneously. Capping is made of cover board and setting sealing on the cover board, such as the capping of PCR reaction tube, has multiple prominent lids on cover board, lid of dashing forward As sealing, lid of dashing forward is identical as the arrangement of PCR reaction tube to be sealed, and prominent lid reacts seal of tube cooperation (such as mistake with PCR Be full of cooperation etc.).

In some embodiments, the corresponding pipe support unit of each pipe lid of closing step, pipe, which covers, to be had and thin-wall tube One-to-one sealing；When capping, sealing first part is pressed into corresponding thin-wall tube in advance, in advance enter positioning after, again will be all close Envelope portion is pressed into thin-wall tube simultaneously.

In some embodiments, the jack 512 in pre-treatment area is set on pedestal 511, under pedestal 511 supports when covering Pressure.Before capping, pipe lid 84 is taken out, shifted and is aligned corresponding pipe support unit, 8 positioning pipe lid of pipe cover frame from pipe cover frame 84。

Instrument for extracting nucleic acid

In some embodiments, instrument for extracting nucleic acid has operating space 1, has bar magnet-bar magnet cover die in operating space 1 Block 2 and drive transmission device 3, referring to Figures 5 and 6, drive transmission device 3 keeps bar magnet-bar magnet set of modules 2 reciprocal or mobile, operation There are cracking region 4 and pre-treatment region 5 in space, cracking region 4 and pre-treatment region 5 are mutually indepedent, bar magnet-bar magnet set of modules 2 is reciprocal in cracking region 4 and preceding processing region 5.The magnetic bead for being adsorbed with nucleic acid is transferred to by bar magnet and bar magnet set from cracking region Nucleic acid extraction is completed in pre-treatment region.

Cracking region

In some embodiments, cracking region 4 can accommodate multi-hole position container, and lysate is contained in hole location, one or more Multi-hole position carries out thermal cracking simultaneously；Pretreatment liquid is contained in multi-hole position container, each hole location of cracking region with preceding place A hole location for managing region is corresponding；The magnetic bead of all hole locations of cracking region is transferred to the corresponding hole site in pre-treatment region simultaneously.It splits Solution region 4 refers to independently of each other with pre-treatment region 5: cracking station and the hole location in pre-treatment region are not intersected.In this way, The temperature change of cracking region does not interfere with pre-treatment region, and pretreatment liquid is avoided unnecessary evaporation or volatilization occur, protects The liquid measure for holding pretreatment liquid is accurate.

Heating load

Heating load to containing sample lysate heat up, make sample discharge nucleic acid, nucleic acid at low temperature (or room temperature) with Magnetic bead combines.Heating load accommodates thin-wall tube and carries out hot transmitting to thin-wall tube.

In some embodiments, as shown in figure 4, heating load 41 has heat transfer plate 413 and accommodating chamber 411, accommodating chamber is set In heat transfer plate 413.When nucleic acid extraction, first to lysate heat up, after cracking nucleic acid discharge, nucleic acid release after will low temperature (or often Temperature) when, nucleic acid just with magnetic bead combineing with, when cooling down, temperature is consistent everywhere in the thin-wall tube in heating load 41, and steam is avoided to condense In the inner wall of hole location, prevent from polluting.Multiple accommodating chambers 411 form a heating unit；Heating load is with one or more heating Unit, it is independent between adjacent heating unit.It is independently referred between adjacent heating unit not between adjacent heating unit Share common sidewalls reduce to the greatest extent and intersect heat transfer between adjacent heating unit, improve temperature accuracy, improve and crack in all hole locations The temperature consistency of liquid.Each accommodating chamber 411 is surrounded by a sleeve 412, independent between the sleeve 412 of each heating unit, Sleeve is connected with heat transfer plate.As shown in Figure 5.Each hole location is independent thin-wall tube and refers to: adjacent hole location not share common sidewalls. Therefore, it reduces intersection heat transfer between adjacent hole location to the greatest extent, improves the temperature accuracy of each hole location, make the cracking of all hole locations The temperature consistency of liquid is good.

Heat cooling piece

Heating cooling piece heats up or cools down under the control of temperature control plate, and the temperature for heating cooling piece passes to heating load.

In some embodiments, as shown in figure 4, heating load 41 is completely covered in heating cooling piece 42.Heat cooling piece 42 Directly each accommodating chamber 411 of heating load 41 is heated, improves the consistency of temperature.It heats cooling piece 42 (such as Peltier) After the completion of heating, carry out refrigeration cooling.The cooling procedure of thermal cracking just has the heating refrigeration cooling of cooling piece 42, keeps lysate fast Speed is cooled to room temperature, it is time-consuming to shorten cracking, and then entire nucleic acid extraction Row control is completed within half an hour.Heating fin Between it is seamless spliced.Heating cooling piece 42 is seamless spliced to refer to adjacent two panels heating fin by multi-disc heating fin Between without apparent gap or distance.Heating fin is corresponding with radiator fan.

Independent temperature plate

In some embodiments, heating cooling piece 42 is equipped with temperature sensing by independent temperature controller temperature control, heating load 41 Device, temperature sensor are connect with temperature controller.As shown in Fig. 2, temperature control plate 421 and customer-oriented control end communication, temperature control plate 421 Receive the real time temperature that temperature sensor transmits.Independent temperature control plate 421 can be realized the accurate temperature controlling to heating cooling piece 42. Temperature control plate 421 should control heating, also control refrigeration.

It is vented colloidal sol

The volume of thin-wall tube (such as PCR reaction tube) is small, and it is gentle molten that sample and lysate generate steam at a high temperature of cracking Glue guides steam and aerosol discharging operation space to avoid steam and Aerosol Pollution operating space.

In some embodiments, as shown in Fig. 2, operating space 1 sets exhaust fan 11, exhaust fan 11 is directed at cracking region 4；Row Fan 11 is located at top or the upper side of cracking region 4.During cracking, steam that exhaust fan 11 generates high temperature and Or aerosol discharging operation space, it avoids polluting in operating space.Preferably, when exhaust fan 11 works, bar magnet-bar magnet set Module 2 is suspended in outside cracking region 4.Exhaust fan 11 by steam and or aerosol guide the side far from bar magnet-bar magnet set of modules 2 into To avoiding bar magnet set contaminated.

Bar magnet-bar magnet set of modules

Bar magnet is inserted into bar magnet set, and bar magnet set insertion lysate, magnetic bead is adsorbed in bar magnet set under magnetic fields.

In some embodiments, as shown in figure 5, bar magnet-bar magnet set of modules 2 include equipped with bar magnet 21 bar magnet frame 211, can Detachable installs the bar magnet stock 221 and baffle 23 of bar magnet set 22, and baffle 23 is located under bar magnet stock 221, and baffle 23 is higher than Heating load 41, each heating unit have the region stopped by baffle 23, and baffle 23 is equipped with what permission bar magnet set 22 passed through Through-hole 231；When bar magnet set 22 is located at pause region, the first ultraviolet lamp is directed at baffle 23；Top plate 142 or side plate 144 are equipped with second Ultraviolet lamp.First ultraviolet lamp carries out sterilization to the irradiation from bottom to top of baffle 23 and other component, and the second ultraviolet lamp is to behaviour Make the component in space 1 and irradiates progress sterilization from the top down.

Each bar magnet is inserted into a bar magnet set, and each bar magnet covers a corresponding thin-wall tube.In order to guarantee bar magnet and bar magnet Set can obtain magnetic bead in cracking region, and magnetic bead is transferred to pre-treatment region, position arrangement and the cracking region of bar magnet The position of thin-wall tube is arranged and the arrangement of the position of the thin-wall tube in pre-treatment region is identical.

In some embodiments, there are one or more bar magnet units, each bar magnet unit, which has, embarks on journey on bar magnet frame 211 Or the multiple bar magnets 21 arranged in column, it is parallel to each other between bar magnet 21, is mutually aligned between bar magnet unit, in parallel；Bar magnet 21 1 End is on bar magnet frame 211, and the other end of bar magnet 21 is as insertion end；The insertion end of all bar magnets 21 is located on bar magnet frame 211 Approximately the same plane.The corresponding bar magnet set 22 of each bar magnet 21, the corresponding bar magnet of each bar magnet unit cover unit；Bar magnet stock 221 are equipped with the slot 222 of bar magnet set unit, and slot 222 keeps the jack of bar magnet set 22 exposed, as shown in Figure 5.All bar magnet sets 22 bottom end is generally aligned in the same plane.

The pipe support in pre-treatment region

Pipe support is used to place the thin-wall tube (such as PCR reaction tube) in pre-treatment region, and the structure of pipe support will determine thin-wall tube Position arrangement.

In some embodiments, as shown in fig. 7, pipe support includes pedestal 511, pedestal 511 is equipped with jack 512, jack 512 Quantity has multiple；The corresponding bar magnet 21 of each jack 512 and bar magnet set 22.A thin-wall tube can be accommodated in each jack 512. The arrangement of jack 512 is identical as the arrangement of accommodating chamber 411 of heating load 41, all magnetic bead energy of cracking region 4 Enough while being transferred to pre-treatment region 5.Base item 513 and pedestal 511 can bear lower pressure, have support stiffness.

Spacing between adjacent pipe support unit is identical as the spacing between the heating unit in heating load 41, therefore, In multiple heating units thermal cracking simultaneously, all magnetic beads can be disposably transferred in pretreatment liquid with bar magnet method simultaneously, complete At nucleic acid extraction.Heating unit and pipe support unit only need to be extended, container expansion, the scalability of nucleic acid extraction ability can be realized It is good, and hole Bits Expanding has substantially no effect on nucleic acid extraction time-consuming.The entire no matter how many hole locations of extraction apparatus carry out thermal cracking, use Same set of bar magnet-bar magnet set of modules.

In some embodiments, pedestal 511 is equipped with base item 513 outstanding, and jack 512 is arranged along base item 513.Pedestal There are multiple base items 513 on 511, there is distance between base item 513, each base item 513 has the multiple jacks 512 of setting, each base item 513 Jack 512 formed a pipe support unit.Base item 513 all has support stiffness, referring to Fig. 7.513 one of pedestal 511 and base item, Or base item 513 and 511 fixing assembling of pedestal.Preferably, base item 513 holds the PCR reaction tube being put into it completely.

Magnetic frame

It is inserted into bar magnet set with magnetic bead and is discharged into pretreatment liquid.In order to make all magnetic beads rapidly from bar magnet set as far as possible It is discharged into pretreatment liquid, is located at pretreatment liquid in magnetic field, is acted on using magnetic-adsorption, magnetic bead is made to be detached from bar magnet set.

In some embodiments, each jack 512 of pipe support is located in magnetic field.Preferably, magnet is set to 512 bottom of jack Portion or side or magnet surround jack 512, and the number of magnets near each jack is 1 or gives more.Magnetic bead is transferred to When pre-treatment region 5, bar magnet 21 is statically placed in pretreatment liquid from 22 extraction of bar magnet set, bar magnet set 22, and bar magnet covers the absorption of 22 bottoms Magnetic bead is detached from bar magnet set 22 under the suction of magnet, and magnet accelerates the rate of release of magnetic bead and fills magnetic bead from bar magnet set 22 Divide release.

In some embodiments, as shown in figure 8, being equipped with pedestal 514 below jack 512, pedestal 514 is equipped with and container bottom The tube seat 5141 of contact, jack 512 and tube seat 5141 correspond, and are mutually communicated.Preferably, pedestal 514 is provided with receiving magnetic The mounting groove 5142 of body, mounting groove 5142 are arranged along 5141 periphery of tube seat.Preferably, each adjacent two tube seat 5141 is one group of pipe A mounting groove 5142 is arranged in slot unit, every group of tube seat unit side.Preferably, every group of 5141 unit two sides of tube seat are respectively provided with peace Slot 5142 is put, mounting groove 5142 is located at the two sides of tube seat unit midline position, and mounting groove 5142 is symmetrical arranged.Preferably, it places Slot 5142 is diamond shape, and the groove angle of tube seat unit two sides mounting groove 5142 is opposite, and the trough rim of mounting groove 5142 abuts tube seat.It is preferred that , mounting groove 5142 is square.

Automatic capping device

Automatic capping device can be placed directly on follow-up diagnosis to the automatic gland of the thin-wall tube in pre-treatment area, sealing, formation The sample of nucleic acid product handled in equipment (such as PCR equipment).

Automatic capping device, which can be, to be moved alone, is also possible to bar magnet-bar magnet set of modules synchronizing moving.

In some embodiments, automatic capping device 6 includes the clamp assembly 62 of gland driving assembly 61 and clamping pipe lid, Gland driving assembly 61 is with clamp assembly 62 or rises or drops or suspend；Clamp assembly 62 includes gland portion 621, clamping portion 622 With the clamping driving portion 623 for making 622 opening and closing of clamping portion, gland portion 621 is in upper, clamping portion 622 under, and clamping portion 622 is by least Two intermediate plates 6221 form.It holds pipe lid when intermediate plate 6221 closes up tightly, realize clamping, pipe lid is discharged when intermediate plate 6221 opens.Intermediate plate 6221 be in release conditions when, gland portion 621 pushes all prominent lids simultaneously (to closure portion).For example, gland portion 621 is positioned at dress Pressing plate on folder portion 622, pressing plate covering or part cover all prominent lids of pipe lid, and pressing plate can simultaneously apply all prominent lids Power.

In some embodiments, clamping driving portion 623 is tool there are two the clamping jaw 6231 of movable part, and each clamping jaw 6231 is pacified An intermediate plate 6221 is filled, as shown in Figure 10, intermediate plate 6221 sets groove 6222, and the opening of two grooves 6222 is opposite.When clamping, pipe The folder portion to be installed of lid enters intermediate plate 6221 from the opening of groove 6222, and the bottom surface of groove 6222 holds up the bottom surface of folder portion to be installed.It is excellent Choosing, groove 6222 is opened in 6221 surface of intermediate plate, such as processes groove 6222 with the mode of wire cutting；Alternatively, intermediate plate 6221 Upper installation upper plate 62211 and lower plate 62212, the distance between upper plate 62211 and lower plate 62212 are adapted to folder portion to be installed, lower plate 62212 hold up folder portion to be installed, and upper plate 62211 and lower plate 62212 surround the groove 6222 of a side opening.

When groove 6222 is opened in 6221 surface of intermediate plate, intermediate plate 6221 is equipped with extended segment 6223, extended segment 6223 and groove 6222 top surface flushes, and the extended segment 6223 of two intermediate plates 6221 is opposite, and two extended segments 6223 form gland portion 621；Alternatively, Upper plate 62211 extends compared to lower plate 62212 to the direction far from intermediate plate 6221.When the extended segment of two intermediate plates 6,221 6223 or on When plate closes up, there are the space for accommodating prominent lid between clamping portion 622, intermediate plate 6221 does not squeeze prominent lid 842.When clamping portion 622 is released When putting folder portion 841 (cover board of pipe lid) to be installed, folder portion 841 to be installed is exited in the bottom of groove 6222, at this point, gland portion 621 (prolongs Stretch section) cover board 841 is still covered, therefore following pressures can be applied to all prominent lids 842.

When 622 clamping of clamping portion has pipe lid, pipe cover exposes to intermediate plate 6221.When capping, 62 clamping of clamp assembly Capping alignment container pushes, and the exposed portion of pipe lid is pressed into container in advance.Then, clamping portion 622 discharges pipe lid, and gland portion is by pipe lid It is completely forced into container, sealing container.

Such as container uses townhouse PCR reaction tube (such as 8 townhouse pipes or 12 townhouse pipes), and pipe lid 84 is then townhouse pipe lid, connection Comb lid includes cover board 841 and multiple prominent lids 842, and each prominent lid 842 seals a PCR reaction tube.When capping, townhouse pipe is covered All prominent lids 842 be pressed into corresponding PCR reaction tube simultaneously.

In some embodiments, capping apparatus includes mobile mechanism, and gland driving assembly is installed on mobile mechanism, moving machine Structure keeps clamp assembly reciprocal in pipe cover frame and preceding processing region.

Intermediate plate

Intermediate plate is installed on clamping driving portion (such as clamping jaw), and intermediate plate realizes clamping and release to pipe lid.

Intermediate plate includes ontology, and as shown in figs. 9-10, ontology sets gland portion 621 and clamping portion 622, and gland portion 621 is in upper, dress Folder portion 622 is under.At least two intermediate plates realize the clamping of pipe lid, hold pipe lid when intermediate plate closes up tightly, realize that clamping, intermediate plate 6221 are opened Pipe lid is discharged when opening.When intermediate plate 6221 is in release conditions, gland portion 621 pushes all prominent lids 842 simultaneously (to closure portion).Example Such as, gland portion 621 is the pressing plate on clamping portion 622, and pressing plate covering or part cover all prominent lids 842 of pipe lid 84, Pressing plate can simultaneously exert a force to all prominent lids 842.

In some embodiments, ontology sets groove 6222, and the opening of two grooves 6222 is opposite.When clamping, pipe lid to Clamping portion 841 enters intermediate plate 6221 from the opening of groove 6222, and the bottom surface of groove 6222 holds up the bottom surface of folder portion 841 to be installed.It is excellent Choosing, groove 6222 is opened in body surface, such as processes groove with the mode of wire cutting；Alternatively, installing upper plate on ontology 62211 and lower plate 62212, the distance between upper plate 62211 and lower plate 62212 be adapted to folder portion 841 to be installed, 62212 support of lower plate Folder portion 841 to be installed is played, upper plate 62211 and lower plate 62212 surround the groove 6222 of a side opening.

When groove 6222 is opened in body surface, ontology is equipped with extended segment 6223, the top of extended segment 6223 and groove 6222 Face flushes, and the extended segment 6223 of two ontologies is opposite, and two extended segments 6223 form gland portion 621.Alternatively, 62211 phase of upper plate Extend than lower plate 622 to the direction far from ontology.When the extended segment of two ontologies 6223 or upper plate 62211 close up, clamping portion There are the space for accommodating prominent lid between 622, ontology does not squeeze prominent lid.When clamping portion 622 discharges the 841 (lid of pipe lid of folder portion to be installed Plate) when, folder portion 841 to be installed is exited in the bottom of groove 6222, at this point, gland portion 621 (i.e. extended segment) still covers cover board, therefore Following pressures can be applied to all prominent lids.

When capping, 62 clamping of clamp assembly capping alignment thin-wall tube push, it is all it is prominent lid be pressed into corresponding thin-walled in advance Pipe.Then, clamping portion 622 discharges pipe lid, and prominent lid is completely forced into thin-wall tube by gland portion 621, completes capping.

Pipe cover frame

Before capping, pipe lid is located in stable position with pipe cover frame, mobile mechanism is with clamp assembly in pipe cover frame It is reciprocal between region and pre-treatment region.

In some embodiments, pipe cover frame 8 has pipe cage 81, and pipe cage 81 has locating piece 82, the limitation pipe of locating piece 82 Rotational freedom and translational degree of freedom of the lid 84 on pipe cage 81.When pipe lid 84 is combined with locating piece 82, pipe lid 84 can not The rotation of relative positioning part 82 or translation, only behind intermediate plate clamping pipe lid 84, pipe lid 84 is removed from locating piece 82.Locating piece 82 Quantity it is identical as the quantity of pipe support unit.There is the base item 83 of protrusion on pipe cage 81, each locating piece 82 is set to a base On item 83.The width of base item 83 is matched with 842 size of prominent lid of pipe lid 84, to increase the cover board 841 and pipe cage of pipe lid 84 The distance between 81, cover board 841 is picked up convenient for intermediate plate.

The first positioning method of pipe cover frame

In some embodiments, locating piece 82 is the protrusion with pipe Gai Peihe and relative to pipe cage 81 or base item 83, convex The quantity at least two risen.Pipe cover frame 8 includes pipe cage 81 and protrusion, and at least two convex to form a pipe lid positioning list Member.There is the base item 83 of protrusion on pipe cage 81, protrusion is set on a base item 83.Pipe cover frame 8 is set to instrument for extracting nucleic acid In operating space 1.The raised quantity of each pipe lid positioning unit is identical as the prominent lid 842 of pipe lid to be positioned, and each protrusion is corresponding One prominent lid 842.There is base item 83 on pipe cage 81, base item 83 protrudes from pipe cage 81, and bump array is raised on base item 83 The center of circle is located on the middle line of base item 83.The height of protrusion and the matched of prominent lid 842.Pipe cover frame 8 has one or more pipes Lid positioning unit has spacing between adjacent pipe lid positioning unit, and adjacent pipe lid positioning unit is mutually aligned.

Second of positioning method of pipe cover frame

In some embodiments, locating piece 82 be with pipe Gai Peihe and relative to pipe cage 81 or the pit of base item 83, it is recessed The quantity at least two in hole, base item 83 allow the clamping portion 622 of pipe lid exposed or the clamping portion 622 of base item 83 and pipe lid it Between distance allow intermediate plate be inserted into.At least two pits form a pipe lid positioning unit.Pipe cover frame 8 is set to instrument for extracting nucleic acid Operating space in.The pit quantity of each pipe lid positioning unit is identical as the prominent lid 842 of pipe lid to be positioned, as shown in figure 13, The corresponding prominent lid 842 of each pit；Pit is cylindrical.There is base item 83 on pipe cage 81, base item 83 protrudes from pipe cage 81, Pit is arranged on base item 83, and the center of circle of pit is located on the middle line of base item 83.Pipe cover frame 8 has one or more pipe Gai Dingwei Unit has spacing between adjacent pipe lid positioning unit, and adjacent pipe lid positioning unit is mutually aligned.

Automatic sealing method

Automatic sealing method, comprising the following steps:

Pipe lid is transferred to alignment sample transition range by step 1, intermediate plate clamping pipe lid, uplink；

Step 2, clamp assembly downlink, when precompressed of pipe lid, precompressed, wait for that the precompressed space of splenium enters PCR reaction tube hole location, Clamp assembly suspends downlink；

The clamping portion release pipe lid of step 3, intermediate plate；

Step 4, clamp assembly downlink, gland portion are completely forced into corresponding container or cavity, pipe lid to splenium for pipe lid Seal PCR reaction tube；

Step 5, clamp assembly reset.

Preferably, step 1 clamping pipe lid includes:

Step 1.1, clamping jaw and intermediate plate are moved to alignment pipe lid, make the distance between clamping portion of intermediate plate allow to cover into Enter；

Step 1.2, gland driving assembly make clamping jaw downlink, and until gland portion touch pipe lid, gland portion is close to the top of pipe lid Face, simultaneously intermediate plate close up, clamping portion clamping pipe lid.

Operating space

In some embodiments, the inside cabin that band can open the cabinet 14 of door or lid forms aforesaid operations space 1.Preferably, Cabinet 14 is made of door 141, top plate 142, bottom plate 143 and side plate 144, cracking region 4, pre-treatment region 5, drive transmission device 3 and mobile mechanism be respectively arranged on bottom plate 143.

In some embodiments, there is airspace under bottom plate 143, radiator fan and heating fin are mounted on bottom plate 143, heating fin is upper, and radiator fan is under, radiator fan airspace.

In some embodiments, the jack 512 in pre-treatment region 5 is set to pedestal 511, and pedestal 511 is set to bottom plate 143, bottom Pressure when 511 support capping of seat.

In some embodiments, bottom plate 143 is equipped with the first ultraviolet lamp, and the first ultraviolet lamp is located at bar magnet-bar magnet set of modules 2 Suspend region.As shown in figure 14, to bar magnet-bar magnet set of modules 2, radiation sterilization sterilizes ultraviolet lamp 12 from bottom to top.

Embodiment 1

The minimum detection limit of DNA reference material is tested

Be manually operated test using method for extracting nucleic acid of the invention, and using instrument for extracting nucleic acid automatically extract nucleic acid into Row verification experimental verification feasibility:

Experiment is manually operated using method for extracting nucleic acid of the invention, steps are as follows:

Step 1, obtaining the PCR reaction tube equipped with lysate and magnetic bead, (this test uses 8 townhouse PCR pipes, and hole location sequence is A, B, C, D, E, F, G, H), to the addition sample of PCR reaction tube；A4, B4 and C4 hole location of first PCR reaction tube are added 10²CFU/ml, Bordetella pertussis (DNA reference material), D4, E4, F4 hole location are added 10³CFU/ml, Bordetella pertussis (DNA reference Product)；G4, H4 hole location are added 10⁴CFU/ml, Bordetella pertussis (DNA reference material)；The A5 hole location of second PCR reaction tube is added 10⁴CFU/ml, Bordetella pertussis (DNA reference material), B5, C5, D5 hole location are added 10⁵CFU/ml, Bordetella pertussis (DNA reference Product), E5, F5, G5 hole location are added 10⁶CFU/ml, Bordetella pertussis (DNA reference material)；Sample is not added for H5 hole location；

PCR reaction tube after two sample-addings is put into cracking region and carries out thermal cracking by step 2, in thermal cracking processes, guidance Aerosol discharges outward, and after the completion of cracking, in the bar magnet set insertion lysate with bar magnet, bar magnet set and bar magnet stand a period of time Afterwards, it slowly vibrates up and down；

Step 3, bar magnet and bar magnet set take out from lysate, and pre-treatment region is slowly transferred to from cracking region, preceding Processing region has the PCR reaction tube for containing pretreatment liquid, and bar magnet and bar magnet set are inserted into pretreatment liquid, and bar magnet extraction is preceding The PCR reaction tube for the treatment of region is placed around magnet, and bar magnet is covered to be stood a little while in pretreatment liquid, later the vibration up and down of bar magnet set Dynamic, pretreatment liquid will not splash out；

Step 4, bar magnet extract the PCR reaction tube in pre-treatment area out, complete magnetic bead transfer；To the PCR reaction tube in pre-treatment area Capping.

Using instrument for extracting nucleic acid operation experiments of the invention, steps are as follows:

S1, obtain equipped with lysate and magnetic bead PCR reaction tube (this test uses 8 townhouse PCR pipes, and hole location is sequentially A, B, C, D, E, F, G, H), to the addition sample of PCR reaction tube；A2, B2 and C2 hole location of first PCR reaction tube are added 10²CFU/ml, Bordetella pertussis (DNA reference material), D2, E2, F2 hole location are added 10³CFU/ml, Bordetella pertussis (DNA reference Product)；G2, H2 hole location are added 10⁴CFU/ml, Bordetella pertussis (DNA reference material)；The A5 hole location of second PCR reaction tube is added 10⁴CFU/ml, Bordetella pertussis (DNA reference material), B3, C3, D3 hole location are added 10⁵CFU/ml, Bordetella pertussis (DNA reference Product), E3, F3, G3 hole location are added 10⁶CFU/ml, Bordetella pertussis (DNA reference material)；Sample is not added for H3 hole location；It is ready to contain It is filled with the PCR reaction tube of pretreatment liquid, and is placed in the pipe support in pre-treatment area；

S2, two PCR reaction tubes after sample-adding are put into the heating load of cracking region, it is negative that each hole location is put into heating In the sleeve carried；Bar magnet-bar magnet set of modules is located at except cracking region, heats to lysate, and heating, exhaust fan are opened, After heating, freeze cooling lysate；

S3, cracking are completed, and exhaust fan stops, and bar magnet is inserted into bar magnet set, and bar magnet-bar magnet set is moved to cracking region, often In a bar magnet-bar magnet set insertion PCR reaction tube, after standing a period of time, slowly vibrate up and down, to adsorb the every of lysate The magnetic bead in a section；

S4, bar magnet and bar magnet set are extracted out from lysate, and are slowly transferred to pre-treatment area, are directed at pre-treatment area PCR reaction tube；In bar magnet and bar magnet set insertion pretreatment liquid, bar magnet moves up, covers and extract out from bar magnet,

S5, bar magnet are covered to be stood a little while in pretreatment liquid, and bar magnet set vibrates up and down n times later, and bar magnet set moves up, magnetic Stick-bar magnet set of modules leaves pre-treatment area；

S6, capping apparatus clamping pipe lid and the PCR reaction tube for being moved to alignment pre-treatment region, fixture is downward, by pipe lid The sealing for being pressed into PCR reaction tube, completing PCR reaction tube.Instrument for extracting nucleic acid work finishes, and opens the door, the PCR reaction tube that will be sealed Carry out sensitivity technique.

The total time-consuming of instrument for extracting nucleic acid of the invention is 16~18min, and after the completion of instrumentation, pre-treatment region is taken out PCR reaction tube can be directly placed into PCR instrument and carry out subsequent analysis.

Experimental result:

Conclusion: the result of manual operation and instrumentation is without significant difference；The kit sensitivity of instrumentation reaches 100CFU/ml meets kit performance verification.

Embodiment 2

The anti-pollution test of instrument for extracting nucleic acid

First group of anti-pollution test of 2.1DNA reference material

S1, obtain equipped with lysate and magnetic bead PCR reaction tube (this test uses 8 townhouse PCR pipes, and hole location is sequentially A, B, C, D, E, F, G, H), to the addition sample of PCR reaction tube, the arrangement of sample is as shown in figure 15,

The A3 hole location of first PCR reaction tube is DNA strong positive reference material 10⁶CFU/m (Bordetella pertussis), B3 hole location are Negative sample, C3 hole location are DNA strong positive reference material 10⁶CFU/m (Bordetella pertussis), D3 hole location are negative sample, E3 hole location For DNA strong positive reference material 10⁶CFU/m (Bordetella pertussis), F3 hole location are negative sample, and G3 hole location is the reference of DNA strong positive Product 10⁶CFU/m (Bordetella pertussis), H3 hole location are negative sample；

The A4 hole location of second PCR reaction tube is negative sample, and B4 hole location is DNA strong positive reference material 10⁶CFU/m (hundred Day cough bacillus), C4 hole location is negative sample, and D4 hole location is DNA strong positive reference material 10⁶CFU/m (Bordetella pertussis), E4 hole location For negative sample, F4 hole location is DNA strong positive reference material 10⁶CFU/m (Bordetella pertussis), G4 hole location are negative sample, the hole H4 Position is DNA strong positive reference material 10⁶CFU/m (Bordetella pertussis)；

The A5 hole location of third PCR reaction tube is DNA strong positive reference material 10⁶CFU/m (Bordetella pertussis), B5 hole location are Negative sample, C5 hole location are DNA strong positive reference material 10⁶CFU/m (Bordetella pertussis), D5 hole location are negative sample, E5 hole location For DNA strong positive reference material 10⁶CFU/m (Bordetella pertussis), F5 hole location are negative sample, and G5 hole location is the reference of DNA strong positive Product 10⁶CFU/m (Bordetella pertussis), H5 hole location are negative sample；

The A6 hole location of 4th PCR reaction tube is negative sample, and B6 hole location is DNA strong positive reference material 10⁶CFU/m (hundred Day cough bacillus), C6 hole location is negative sample, and D6 hole location is DNA strong positive reference material 10⁶CFU/m (Bordetella pertussis), E6 hole location For negative sample, F6 hole location is DNA strong positive reference material 10⁶CFU/m (Bordetella pertussis), G6 hole location are negative sample, the hole H6 Position is DNA strong positive reference material 10⁶CFU/m (Bordetella pertussis)；

PCR reaction tube after four sample-addings is anti-by first PCR reaction tube, second PCR reaction tube, third PCR It should manage and successively be put in heating load with the 4th PCR reaction tube；

It gets out 4 and contains 8 townhouse PCR reaction tubes of pretreatment liquid, and be placed in the pipe support in pre-treatment area；

S2, four PCR reaction tubes after sample-adding are put into the heating load of cracking region, it is negative that each hole location is put into heating In the sleeve carried；Bar magnet-bar magnet set of modules is located at except cracking region, heats to lysate, and heating, exhaust fan are opened, After heating, freeze cooling lysate；

S6, capping apparatus clamping pipe lid and the PCR reaction tube for being moved to alignment pre-treatment region, fixture is downward, by pipe lid The sealing for being pressed into PCR reaction tube, completing PCR reaction tube.Instrument for extracting nucleic acid work finishes, and opens the door, the PCR reaction tube that will be sealed Carry out pollution detection.

Experimental result A3-H3 is first row, and A4-H4 is that second row A5-H5 is third row, and A6-H6 is the 4th row

Conclusion: no cross contamination between the Kong Yukong of the PCR reaction tube of instrumentation.

2.2, second group of anti-pollution test of DNA reference material

S1, obtain equipped with lysate and magnetic bead PCR reaction tube (this test uses 8 townhouse PCR pipes, and hole location is sequentially A, B, C, D, E, F, G, H), to the addition sample of PCR reaction tube, the arrangement of sample is as shown in figure 15, the A3 of first PCR reaction tube Hole location, B3 hole location, C3 hole location, D3 hole location, E3 hole location, F3 hole location, G3 hole location and H3 hole location are DNA strong positive reference material (hundred Day cough bacillus)；

A4 hole location, B4 hole location, C4 hole location, D4 hole location, E4 hole location, F4 hole location, G4 hole location and the H4 of second PCR reaction tube Hole location is negative sample；

A5 hole location, B5 hole location, C5 hole location, D5 hole location, E5 hole location, F5 hole location, the G5 hole location, H5 of third PCR reaction tube Hole location is DNA strong positive reference material (Bordetella pertussis)；

A6 hole location, B6 hole location, C6 hole location, D6 hole location, E6 hole location, F6 hole location, the G6 hole location, H6 of 4th PCR reaction tube Hole location is negative sample；

PCR reaction tube after four sample-addings is successively put in heating load；

Experimental result:

Conclusion: sample and lysate, pretreatment liquid use townhouse PCR reaction tube as container (such as 8 townhouses, 12 townhouses), Cross contamination is not deposited between each group of townhouse PCR reaction tube and other group of townhouse PCR reaction tube.2.3, the pollution of RNA reference material Test

The A3 hole location of first PCR reaction tube is RNA reference material (Respiratory Syncytial Virus(RSV) reference material), and B3 hole location is feminine gender Sample, C3 hole location are RNA reference material (Respiratory Syncytial Virus(RSV) reference material), and D3 hole location is negative sample, and E3 hole location is RNA reference Product (Respiratory Syncytial Virus(RSV) reference material), F3 hole location are negative sample, and G3 hole location is RNA reference material (Respiratory Syncytial Virus(RSV) ginseng Examine product), H3 hole location is negative sample；

The A4 hole location of second PCR reaction tube is negative sample, and B4 hole location is RNA reference material (Respiratory Syncytial Virus(RSV) ginseng Examine product), C4 hole location is negative sample, and D4 hole location is RNA reference material (Respiratory Syncytial Virus(RSV) reference material), and E4 hole location is negative sample This, F4 hole location is RNA reference material (Respiratory Syncytial Virus(RSV) reference material), and G4 hole location is negative sample, and H4 hole location is RNA reference material (Respiratory Syncytial Virus(RSV) reference material)；

The A5 hole location of third PCR reaction tube is RNA reference material (Respiratory Syncytial Virus(RSV) reference material), and B5 hole location is feminine gender Sample, C5 hole location are RNA reference material (Respiratory Syncytial Virus(RSV) reference material), and D5 hole location is negative sample, and E5 hole location is RNA reference Product (Respiratory Syncytial Virus(RSV) reference material), F5 hole location are negative sample, and G5 hole location is RNA reference material (Respiratory Syncytial Virus(RSV) ginseng Examine product), H5 hole location is negative sample；

The A6 hole location of 4th PCR reaction tube is negative sample, and B6 hole location is RNA reference material (Respiratory Syncytial Virus(RSV) ginseng Examine product), C6 hole location is negative sample, and D6 hole location is RNA reference material (Respiratory Syncytial Virus(RSV) reference material), and E6 hole location is negative sample This, F6 hole location is RNA reference material (Respiratory Syncytial Virus(RSV) reference material), and G6 hole location is negative sample, and H6 hole location is RNA reference material (Respiratory Syncytial Virus(RSV) reference material)；

Embodiment 3

3.1, the precision test of DNA reference material

S1, obtain equipped with lysate and magnetic bead PCR reaction tube (this test uses 8 townhouse PCR pipes, and hole location is sequentially A, B, C, D, E, F, G, H), to the addition sample of PCR reaction tube, the arrangement of sample is as shown in figure 15, gets out 4 and is loaded with cracking DNA reference material (10 is added in 48 townhouse PCR reaction tubes in 8 townhouse PCR reaction tubes of liquid and magnetic bead⁶CFU/ml, pertussis Bacillus)；

S6, capping apparatus clamping pipe lid and the PCR reaction tube for being moved to alignment pre-treatment region, fixture is downward, by pipe lid The sealing for being pressed into PCR reaction tube, completing PCR reaction tube.Instrument for extracting nucleic acid work finishes, and opens the door, the PCR reaction tube that will be sealed Carry out precision detection.

Experimental result:

Conclusion: being 2.63% < 5% using the CV value of instrument for extracting nucleic acid, instrument essence precision complies with standard.

3.2, the precision test of RNA reference material

S1, obtain equipped with lysate and magnetic bead PCR reaction tube (this test uses 8 townhouse PCR pipes, and hole location is sequentially A, B, C, D, E, F, G, H), to the addition sample of PCR reaction tube, the arrangement of sample is as shown in figure 15, gets out 4 and is loaded with cracking RNA reference material (Respiratory Syncytial Virus(RSV)) is added in 48 townhouse PCR reaction tubes in 8 townhouse PCR reaction tubes of liquid and magnetic bead；

Conclusion: being 1.60% < 5% using the CV value of instrument for extracting nucleic acid, instrument essence precision complies with standard.

Embodiment 4

The silent winged KingFisher of the sample and match that instrument for extracting nucleic acid of the invention obtains^TMDuo Prime magnetic beads for purifying instrument obtains The sample obtained；And two instruments are simultaneously to DNA reference material (10⁴CFU/ml, Bordetella pertussis) carry out nucleic acid extraction after carry out QPCR detection.

S1, obtain equipped with lysate and magnetic bead PCR reaction tube (this test uses 8 townhouse PCR pipes, and hole location is sequentially A, B, C, D, E, F, G, H), to the addition sample of PCR reaction tube, the arrangement of sample is as shown in figure 15, gets out 1 and is loaded with cracking 8 townhouse PCR reaction tubes of liquid and magnetic bead；DNA reference material (10 is added in A, B, C, D, E, F, G, this 7 hole locations⁴CFU/ml, hundred Day cough bacillus), H hole location is negative sample, and sample size is 20 microlitres；It is anti-to get out the 8 townhouse PCR that 1 contains pretreatment liquid Ying Guan, and it is placed in the pipe support in pre-treatment area；

The instrument time-consuming of instrument for extracting nucleic acid of the invention is 16min；Sample-adding time-consuming about 5min manually, after the completion of instrumentation Obtain the sample that can be directly placed into QPCR processing.Although this test has only used 7 sample inventions can handle 32 simultaneously Kind or more sample.

Use the silent winged KingFisher of match^TMThe operating operation test of Duo Prime magnetic beads for purifying instrument, steps are as follows:

Step 1 is loaded this: obtaining 96 orifice plates, sequentially adds 96 by lysate, in conjunction with liquid, cleaning solution and eluent manually In the hole location of orifice plate, the hole location one of a usual row 8 region, if 8 holes of first row accommodate lysate, 8 holes of second row are accommodated In conjunction with liquid, third arranges 8 holes and accommodates the first cleaning solution, and the 4th hole of row 8 accommodates second of cleaning solution, the 5th Kong Rong of row 8 Receive eluent；The position for accommodating lysate is corresponding with heating module；Sample is added in lysate again, sample size requirements are in 5 millis It rises；

Step 2, cracking/combination: lysate lysed sample (reaction a period of time, during which bottom can heat acceleration cracking), Period bar magnet set (only bar magnet set, without bar magnet), which is inserted under liquid level, to carry out oscillation up and down and mixes, and nucleic acid is made to be suspended in lysate In；After the completion of cracking, the bar magnet set with bar magnet protrudes into magnetic bead hole and picks up magnetic bead, is transferred in cracking fluid apertures, bar magnet takes out, magnetic Oscillation mixes so that magnetic bead, which is evenly spread to, adsorbs nucleic acid in lysate stick set up and down, and after standing a period of time, bar magnet insertion is right It stands afterwards or upper and lower oscillation is all adsorbed on bar magnet to magnetic bead and puts on；

Step 3, salt are washed: will be transferred in the hole that salt is washed from absorption magnetic bead in lysate, bar magnet takes out, the vibration up and down of bar magnet set Swing mixing so that magnetic bead is evenly spread in salt washing lotion, after standing a period of time, bar magnet insertion be then allowed to stand or vibrate up and down to Magnetic bead is all adsorbed on bar magnet and puts on；

Step 4, washing: the magnetic bead after salt is washed takes out, and is transferred in cleaning solution, and oscillation mixes so that magnetic bar magnet set up and down Pearl evenly spreads in cleaning solution, and after standing a period of time, bar magnet insertion, which is then allowed to stand or is vibrated up and down to magnetic bead, to be all adsorbed on Bar magnet is put on；

Step 5, elution: the magnetic bead after washing is taken out, and bar magnet takes out, and oscillation mixes so that magnetic bead is uniform bar magnet set up and down It is distributed in eluent, is generally heated to that 65 DEG C/5min is stood, heating makes nucleic acid and magnetic bead disengaging, at a higher temperature, magnetic Stick insertion, which is then allowed to stand or is vibrated up and down to magnetic bead, to be all adsorbed on bar magnet and puts on, and magnetic bead is recycled；After magnetic bead recycles, instrument behaviour Work terminates.

Instrument is opened, reaction plate is taken out, the eluent containing nucleic acid is transferred to PCR reaction tube with pipettor by hand In, it is handled being put into QPCR analysis instrument after PCR reaction tube by hand capping.Sai Mo flies KingFisher^TMDuo Prime magnetic The instrumentation time-consuming that pearl purifies instrument is about 45min, and it is about 15min that sample-adding is time-consuming by hand, and transfer is for QPCR processing by hand Sample time-consuming is about 25min.In the instrument, a reaction plate most multipotency is loaded 12 kinds of samples.

Testing result is as follows；

As a result:

The QPCR analysis sample and Sai Mo that instrument for extracting nucleic acid of the invention obtains fly KingFisher^TMDuo Prime magnetic bead It purifies the QPCR that instrument obtains and analyzes sample, the analysis sample that nucleic acid instrument for extracting nucleic acid of the invention obtains is silent better than match to fly KingFisher^TMThe analysis sample that Duo Prime magnetic beads for purifying instrument obtains.

Meanwhile to equipment and control experiment of the invention, for the nucleic acid extraction amount and integrality under identical sample size Investigation, it is found that the amount for the DNA that equipment of the invention is extracted is to compare the 2 times or more for the DNA that instrument extracts.Meanwhile for phase The extraction of RNA in the sample of same volume, find RNA that equipment of the invention is extracted be 10 times of the DNA extracted than instrument with On, this of the invention may not use many washing process, RNA exposure be reduced, to reduce the probability of RNA degradation.

The integrality of DNA and RNA is investigated, find nucleic acid that this equipment is extracted almost without fragment and fracture, 85% It all keeps complete above, and uses conventional methods and the method for comparative apparatus, nucleic acid only only has that 30-45%'s is complete Property.In this way, in order to effectively carry out following amplification, it is necessary to which more samples extract, and guarantee more nucleic acid Integrality can be only achieved the purpose of amplification.In addition, if nucleic acid (such as DNA or RNA) is imperfect, target target nucleic acid The efficiency of amplification just need the longer time.It was found that the nucleic acid extracted using the present invention, in the case where identical sample size, Such as 20 microlitres, the nucleic acid that the present invention extracts can be expanded effectively, and using comparative apparatus to extract nucleic acid cannot be effective Amplification or the result for feminine gender occur.

In the case where lacking any element specifically disclosed herein, limitation, may be implemented illustrated and described herein Invention.Used terms and expressions method is used as the term of explanation rather than limits, and is not intended in these terms and table Up to any equivalent for excluding shown and described feature or part thereof in the use of method, and it should be realized that various remodeling exist It is all feasible in the scope of the present invention.It is therefore to be understood that although specifically being disclosed by various embodiments and optional feature The present invention, but the modifications and variations of concept as described herein can be used by those of ordinary skill in the art, and recognize It is fallen into for these modifications and variations within the scope of the present invention of the appended claims restriction.

It is described herein or record article, patent, patent application and every other document and can electronically obtain The content of information to a certain extent in full include herein by reference, just as each individual publication by specific and single Solely point out by reference.Applicant retains from any of any this article, patent, patent application or other documents And all material and information are incorporated into the right in the application.

Claims

The pipe cover frame of 1.PCR pipe lid, it is characterised in that: pipe cover frame includes pedestal and protrusion, and at least two convex to form a pipe Lid positioning unit.
2. the pipe cover frame of PCR pipe lid as described in claim 1, it is characterised in that: have the base item of protrusion, protrusion setting on pedestal In on a base item.
3. the pipe cover frame of PCR pipe lid as described in claim 1, it is characterised in that: pipe cover frame is set to the behaviour of instrument for extracting nucleic acid Make in space.
4. the pipe cover frame of PCR pipe lid as described in claim 1, it is characterised in that: the raised quantity of each pipe lid positioning unit It is identical as the prominent lid of pipe lid to be positioned, corresponding one prominent lid of each protrusion.
5. the pipe cover frame of PCR pipe lid as described in claim 1, it is characterised in that: have base item on pedestal, base item protrudes from bottom Seat, on base item, the raised center of circle is located on the middle line of base item bump array.
6. the pipe cover frame of PCR pipe lid as described in claim 1, it is characterised in that: the matched of raised height and prominent lid.
7. the pipe cover frame of PCR pipe lid as described in claim 1, it is characterised in that: pipe cover frame has one or more pipe Gai Ding Bit location has spacing between adjacent pipe lid positioning unit.
8. the pipe cover frame of PCR pipe lid as described in claim 1, it is characterised in that: pipe cover frame has one or more pipe Gai Ding Bit location, adjacent pipe lid positioning unit are mutually aligned.