CN102321743A - Solid-phase carrier for surface fixation and preparation and application methods for solid-phase carrier - Google Patents

Solid-phase carrier for surface fixation and preparation and application methods for solid-phase carrier Download PDF

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CN102321743A
CN102321743A CN201110222895A CN201110222895A CN102321743A CN 102321743 A CN102321743 A CN 102321743A CN 201110222895 A CN201110222895 A CN 201110222895A CN 201110222895 A CN201110222895 A CN 201110222895A CN 102321743 A CN102321743 A CN 102321743A
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盛司潼
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Abstract

The invention relates to the field of surface chemical modification and genetic engineering and provides a solid-phase carrier for surface fixation and methods for preparing and applying the solid-phase carrier. The solid-phase carrier for surface fixation is connected with at least one surface group with an activation function, wherein the surface group contains a fixation end and an active end; and the molecular structural formula of the surface group is that: the fixation end is provided with at least one silyloxy group for combining the solid-phase carrier and the active end -R contains an active functional group for combining a surface fixation connection. In the invention, the solid-phase carrier is used for activation, and then resin is used for embedment and the solid-phase carrier is re-activated so as to ensure that the surface group on the surface of the solid-phase carrier is doubled, so that the quantity of the surface fixation connections fixed on the solid-phase carrier is increased.

Description

A kind ofly be used for surperficial fixed solid phase carrier and preparation and application
Technical field
The present invention relates to surface chemistry and genetically engineered field, more particularly, relate to a kind of preparation method who is used for surperficial fixed solid phase carrier and this solid phase carrier, and the application method of this solid phase carrier.
Background technology
The dna sequencing technology has been widely used in the every field of biological study; A lot of biological questions can solve by high-throughput dna sequencing technology, and extensive parallel order-checking platform (massively parallel DNA sequencing platform) has developed into the sequencing technologies of main flow.At present, what comparatively advanced in the world sequenator used all is new order-checking strategy---circulation chip PCR sequencing PCR (cyclic-array sequencing) also can be referred to as " new-generation sequencing technology or s-generation sequencing technologies ".
The new-generation sequencing method obtains parallel sequencing template through microemulsion PCR (emulsion PCR) or bridge-type PCR methods such as (bridge PCR) mostly.Microemulsion PCR forms the independent reaction space through the water-in-oil system, need after the amplification to discharge again to reclaim, and bridge-type PCR is simple to operate relatively, just solid phase carrier is had relatively high expectations.
The solid phase carrier that is applied to bridge-type PCR is that identical primer (nucleotide sequence) is fixed in surface of solid phase carriers through certain function power, directly carries out the PCR reaction at surface of solid phase carriers.Be applied to the solid phase carrier of solid phase bridge-type PCR at present, its preparation method may further comprise the steps: (1) utilizes the vitriol oil and ydrogen peroxide 50 to make the surface of solid phase carriers hydroxylation; (2) utilize aminopropyl triethoxysilane (APTES) to make the surface silicon alkanisation; (3) utilize equal benzene nitrilotriacetic (BTA) to make the surface of solid phase carriers activation; (4) pass through the order-checking detection signal through amino carboxyl effect immobilized oligonucleotide.Because fixed effect is not good, when detecting fluorescent signal, can only uses laser and excite to collect and detect enough signals.This testing conditions is too harsh, is inappropriate for widespread use.There is complex operation in above-mentioned solid phase preparing carriers method, and fixed effect is poor, problems such as production cost height.
Therefore need a kind of new surperficial fixed solid phase carrier and preparation method thereof that is used for; Improve the quantity and the fixed efficiency of solid phase surface fixed group; The fast and convenient signal that detects surface fixed connection thing can simplify procedures, reduce production costs simultaneously.
Summary of the invention
The object of the present invention is to provide a kind of surperficial fixed solid phase carrier that is used for, it is on the low side to be intended to solve the solid phase surface fixed group, and the problem that the detection signal condition is too harsh can simplify the operation simultaneously, reduces production costs.
In order to realize the object of the invention, the present invention provides a kind of surperficial fixed solid phase carrier that is used for, and said solid phase carrier is connected with at least one surface group with mobilizing function, and wherein, surface group comprises inboardend and active end, and its molecular structural formula is following:
Figure BSA00000551058300021
Wherein, inboardend has at least one and is used for and solid phase carrier bonded siloxy.
Preferably, active end-R comprises and is used for being fixedly connected thing bonded activity functional groups with the surface, and wherein said activity functional groups preferably combines through amido linkage other part with active end-R; Said surface fixed connection thing is meant and is used to detect or material to be detected after the fixed connection of surface, includes but not limited to oligonucleotide, albumen, antibody, probe.
Preferably, the activity functional groups that comprised of active end-R has at least 2 and is used for and the surperficial thing bonded end group that is fixedly connected.
Preferably, active end-R is the dendritic structure that forms based on resin embedding, connects at least 2 and is used for being fixedly connected thing bonded end group with the surface.
Preferred, saidly be used for that to be fixedly connected thing bonded end group be carboxyl with the surface.
Wherein, said solid phase carrier can be at least a in slide, silicon chip or the plastics (PS or SEPIGEL 305 are processed), and its surface can be the plane or the irregular surface that has radian of rule.
In order to realize the object of the invention, the present invention also provides a kind of preparation method who is used for surperficial fixed solid phase carrier, and the said preparation method who is used for surperficial fixed solid phase carrier may further comprise the steps:
A. utilizing silylating reagent that surface of solid phase carriers is carried out end group modifies;
B. utilize activating reagent to combine end group, make activity functional groups on the surface of solid phase carriers band;
C. activity functional groups is carried out resin embedding, form dendroid embedding body;
D. to the activation once more of dendroid embedding body, obtain being used for surperficial fixed solid phase carrier.
Wherein, said steps A comprises:
A1. solid phase carrier is carried out hydroxylation and handle, make surface of solid phase carriers carry hydroxyl;
A2. utilize silylating reagent to combine, surface of solid phase carriers is carried out end group modify with hydroxyl.
Preferably, surface of solid phase carriers being carried out the amination end group modifies.
Preferably, said step B combines activating reagent through amido linkage with the amination end group of surface of solid phase carriers.
Wherein, said activating reagent comprises activity functional groups.
Preferably, the activity functional groups that comprises of activating reagent is a C-terminal.
Preferably, when activity functional groups was C-terminal, said step C carried out embedding through polyamide resin to the C-terminal of surface of solid phase carriers, formed dendritic structure.
Preferably, based on the activation treatment of said dendritic structure repeating step B, make solid phase carrier connect at least 2 and be used for being fixedly connected thing bonded C-terminal with the surface.
In order to realize goal of the invention, the present invention also provides a kind of above-mentioned method that surperficial fixed solid phase carrier carries out gene sequencing that is used for of utilizing, and said the utilization above-mentionedly is used for the method that surperficial fixed solid phase carrier carries out gene sequencing and may further comprise the steps:
M1. oligonucleotide fragment is connected to solid phase carrier through the activity functional groups effect with the surface;
M2. the said solid phase carrier that is connected with oligonucleotide fragment is carried out signal detection, obtain base sequence information through signal analysis.
Wherein, the signal detection among the said step M2 is meant the signal of measuring base to be measured through the base complementrity principle, thereby obtains the sequence information of base to be measured.
Preferably, signal detecting method comprises but is not limited to connect PCR sequencing PCR or synthetic PCR sequencing PCR.
In order to realize the object of the invention; The present invention also provides a kind of above-mentioned method that surperficial fixed solid phase carrier prepares gene chip that is used for of utilizing; The said utilization above-mentionedly is used for the method that surperficial fixed solid phase carrier prepares chip and comprises: oligonucleotide fragment is connected to solid phase carrier through the activity functional groups effect with the surface, forms gene chip.
Wherein, the said gene chip for preparing can be used for the mensuration of DNA material capture and target nucleic acid sequence.
By on can know; At least 2 activity functional groups of the present invention through being comprised among the activity end-R that connects at surface of solid phase carriers; Add that resin embedding forms dendritic structure reactivate afterwards, reach the purpose that the surface of solid phase carriers activity functional groups is multiplied, thereby can fix the more purpose of multilist face fixed connection thing; Improve strength of signal, reduce requirement condition detection signal.Simultaneously, the above-mentioned method that is used for surperficial fixed solid phase carrier of preparation of the present invention, operating process is simple, and production cost is low.
Description of drawings
Fig. 1 is the structural representation that is used for the surface group of surperficial fixed solid phase carrier in the one embodiment of the invention;
Fig. 2 is the structural representation that is used for the surface group of surperficial fixed solid phase carrier in the one embodiment of the invention;
Fig. 3 is the structural representation that is used for the surface group of surperficial fixed solid phase carrier in the one embodiment of the invention;
Fig. 4 is the structural representation that is used for the surface group of surperficial fixed solid phase carrier in the one embodiment of the invention;
Fig. 5 is that the surface group that is used for surperficial fixed solid phase carrier in the one embodiment of the invention carries out embedding structural representation afterwards;
Fig. 6 is the preparing method's schema that is used for surperficial fixed solid phase carrier in the one embodiment of the invention;
Fig. 7 is the preparing method's schema that is used for surperficial fixed solid phase carrier in a preferred embodiment of the invention;
Fig. 8 is based on being used for the method flow diagram that surperficial fixed solid phase carrier carries out gene sequencing in the one embodiment of the invention;
Fig. 9 carries out the resulting order-checking fluorescent signal of gene sequencing figure based on being used for surperficial fixed solid phase carrier in the one embodiment of the invention;
Figure 10 is based on being used for the method flow diagram that surperficial fixed solid phase carrier carries out the gene chip preparation in the one embodiment of the invention;
Figure 11 is the method flow synoptic diagram that carries out gene sequencing in the one embodiment of the invention based on the preparation that is used for surperficial fixed solid phase carrier.
Embodiment
In order to make the object of the invention, technical scheme and advantage clearer,, the present invention is further elaborated below in conjunction with accompanying drawing and embodiment.
It is following that Fig. 1 shows the universal architecture of the surface group that is used for surperficial fixed solid phase carrier among the present invention:
Figure BSA00000551058300041
Wherein, this surface group comprises inboardend and active end, and inboardend has at least one siloxy that is used for being connected with solid phase carrier use; And active end-R comprises and is used for and the surperficial thing bonded activity functional groups that is fixedly connected; This activity functional groups can be various forms, in following a plurality of embodiment, with setting forth the possible scheme of part in detail.
Fig. 2 shows the specific embodiment that is used for surperficial fixed surface of solid phase carriers group among Fig. 1.This surface group structure is following:
Figure BSA00000551058300042
In the present embodiment, the activity functional groups of the active end-R of surface group links to each other with other parts of activity end-R through amido linkage.In addition, two remaining carboxyls then become the activity functional groups in the subsequent processing steps on the phenyl ring.
Fig. 3 shows the specific embodiment of the surface of solid phase carriers group that is used for surperficial immobilized oligonucleotide among Fig. 1.This surface group structure is following:
Figure BSA00000551058300051
In the present embodiment; Activity functional groups in the active end-R group of surface group partly is connected through different sulphur cyanogen key other parts with active end-R; After the different sulphur cyanogen key is the straight chain of three carbon atoms; Be connected with ring texture with ester bond, remaining carboxyl is as the activity functional groups of follow-up use on the ring texture.
Fig. 4 shows the specific embodiment of the surface of solid phase carriers group that is used for surperficial immobilized oligonucleotide among Fig. 1.This surface group structure is following:
Figure BSA00000551058300052
In the present embodiment; The surface group inboardend is connected through two siloxies with active end-R; Activity functional groups among the active end-R partly is connected through amido linkage other parts with active end-R, encircles the activity functional groups of remaining two carboxyls as follow-up use.
Should be noted that Fig. 2, Fig. 3 and Fig. 4 have only provided several specific embodiments of surface group, but the present invention is not so limited.In principle, have two or above can getting final product with the surface group that the surface is fixedly connected the activity functional groups that thing is connected.
Fig. 5 shows the structure after the universal architecture process subsequent disposal that is used for surperficial fixed solid phase carrier among the embodiment.Preferably carry out embedding, make surface of solid phase carriers bonded surface group form dendritic structure through the activity end-R of polyamide resin after to the surface of solid phase carriers activation.Further after the activation, on the dendritic structure again activation become the activity functional groups of original active end-R, thereby make the increase of its quantity generation multiple, can fix more surface fixed connection thing.
Fig. 6 has provided the method flow that is used for surperficial fixed solid phase carrier preparation in the one embodiment of the invention.Present method may further comprise the steps:
S1. utilizing silylating reagent that surface of solid phase carriers is carried out end group modifies;
S2. utilize activating reagent to combine, surface of solid phase carriers is connected be used for being fixedly connected thing bonded activity functional groups with the surface with end group;
S3. activity functional groups is carried out resin embedding, form dendroid embedding body;
S4. to the activation once more of dendroid embedding body, obtain being used for surperficial fixed solid phase carrier.
Need to prove, before institute begins in steps, used solid phase carrier is carried out the cleaning and removing removal of impurity, then surface of solid phase carriers is carried out hydroxylation and handle, make hydroxyl on the surface of solid phase carriers band.Described solid phase carrier comprises but is not limited to glass, silicon chip or plastic material, and simultaneously surface of solid phase carriers can be the plane of rule or irregular as have a surface of radian.In one embodiment of the invention, alternation rule planar glass is as solid phase carrier; In another embodiment, select for use and be with arcual plastic sheet as solid phase carrier.
Wherein, hydroxylation is handled the alcohols alkaline process and is handled, also can select for use simultaneously other like sour oxygen method methods such as (vitriol oil and dioxygen water laws).About hydroxylated preferred version, will describe in detail in another preferred embodiment.
Among the step S1, surface of solid phase carriers is carried out end group modify.Utilize the hydroxyl on alkylating reagent and the solid phase carrier to react, form amidized siloxy inboardend.
Wherein, Silylating reagent among the step S1; Therefore siloxanes or siloxanes acid can be selected in principle, DIOXANE (dioxane) solution of carbonyl dimidazoles (CDI), 3-Racemic glycidol propyl trimethoxy silicane (GOPS) the toluene liquid that contains triethylamine (TEA), aminopropyl triethoxysilane (APTES) reagent can be selected to use as silylanization.For the preferred version of step S1, can in follow-up preferred embodiment, provide detailed description.
Among the step S2, utilize activating reagent, make activity functional groups on the surface of solid phase carriers band.Surface of solid phase carriers siloxy inboardend is amination when end modified, preferably through amido linkage activating reagent is combined with the amination end group of surface of solid phase carriers, thereby makes surface of solid phase carriers band upper surface group-R.
Wherein, active end-R has two or more can be fixedly connected the activity functional groups that thing is connected with the surface.Therefore; Can select different activated reagent; For example can select saturated Pyroglutaric acid (DMF)/NHS/N for use, N '-dicyclohexyl carbodiimide (DCC), 3-Racemic glycidol propyl trimethoxy silicane/silane coupling agent (GOPTS), inferior phenylene diisothiocyanic acid salts solution (PDITC) are as activating reagent.About the preferred version of step S2, will in follow-up preferred embodiment, provide.
Among the step S3, the activity end-R of surface group is carried out resin embedding, form dendroid embedding body.Through resin embedding, the activity functional groups that activity end-R is comprised carries out the numerical value amplification, in the dendritic structure that guarantees simultaneously to form, connects at least 2 and is used for and the surperficial thing bonded activity functional groups that is fixedly connected.
Among the step S4, dendroid embedding body is carried out activatory operation repeat according to the method for step S2.
Above-mentionedly among the present invention be used for surperficial fixed solid phase carrier, its surface is fixedly connected thing and is meant by surface of solid phase carriers and is used to detect or material to be detected after fixing, includes but not limited to oligonucleotide, albumen, antibody and probe.
Fig. 7 has provided the method flow that one of the present invention is used for the preferred embodiment of surperficial fixed solid phase carrier preparation.Concrete step comprises:
S101. adopt NaOH and ethanol to handle, make the surface of glass slide hydroxylation;
S102. utilize silylating reagent APTMS, make the hydroxyl aminoization of surface of glass slide;
S103. utilize acvator TMA/NHS/DCC to carry out activation, make activity functional groups on the surface of glass slide band;
S104. with polymeric amide PAMAM to the activity functional groups embedding, form dendroid embedding body;
S105. to the activation once more of dendroid embedding body, obtain being used for surperficial fixed solid phase carrier.
Need to prove, before institute begins in steps, need clean, remove the impurity of surface of glass slide testing used slide.
Among the step S101, preferably make hydroxyl on the surface of glass slide band through pure alkaline process.Specific operation process is: preparation 18% NaOH solution, join then mix in the ethanol solution stir limpid until liquid; Slide is put into wherein, secluding air, 2h is soaked in vibration, takes out the slide washed several times with water then and removes remaining alcohol, alkali, dries up.
In this step, the concrete reaction formula that relates to is following:
Figure BSA00000551058300071
Among the step S102, utilize silylating reagent to handle slide and make its surface hydroxyl amination.The acetone solution of preferred 1% aminopropyl trimethoxysilane (APTMS) carries out Silanization reaction.Concrete operations are: slide is put into 1% aminopropyl trimethoxysilane (APTMS) and 95% acetone solution immersion 2min, its surface is with-NH 2, N is cleaned in the acetone vibration then 2Dry.Concrete chemical reaction is as follows:
Figure BSA00000551058300081
Among the step S103, utilize activating reagent to handle surface of glass slide and make its surface active and be with activity functional groups.Preferred trimesic acid (TMA), N-hydroxy-succinamide (NHS), N, N '-NSC 57182 (DCC) solution and diisopropylethylamine (DIEA) make the surface of glass slide activation as surfactant, are with activity functional groups.
Wherein, Concrete operations comprise: with trimesic acid (TMA), N-hydroxy-succinamide (NHS), N; N '-NSC 57182 (DCC) solution mixes back air-isolation oscillatory reaction 2h, and the spinning product is mixed with into Acibenzolar with diisopropylethylamine (DIEA) solution equal-volume; Slide single face at silylanization drips an amount of Acibenzolar then, and reaction 3h makes the surface of solid phase carriers radical functino, N, and N-dimethylformamide (DMF) and ethanol vibration are cleaned.Use 5% then, the sodium hydrogen carbonate solution of pH8.8 is handled, and makes carboxyl on the surface of glass slide band.Concrete reaction formula is as follows:
Figure BSA00000551058300082
Among the step S104, the carboxyl of surface of glass slide is carried out embedding, form dendroid embedding body through polymeric amide PAMAM resin.Drip methanol solution that 100 μ l are dissolved with 10% polymeric amide PAMAM in surface of glass slide, lump together the formation sandwich structure with second slide again, room temperature is put 12h.Use washed with methanol then 2 times, N2 is dry.
The related reaction of step S104 is following:
Figure BSA00000551058300091
Among the step S105, dendroid embedding body is carried out activatory operation repeat according to the method for step S103.
Fig. 8 has provided in the one embodiment of the invention method flow that is applied to gene sequencing based on a kind of solid phase carrier of surperficial immobilized oligonucleotide.Concrete step comprises:
Among the step S201, carry out the point sample of testing sample.Testing sample mixes with order-checking buffer, on the solid phase carrier for preparing and be positioned in the container, carries out point sample.
Wherein, when biased sample, can add a spot of microballon, be used for focusing and make sample can concentrate on the certain position place.
Among the step S202, the sealing lucifuge is carried out the room temperature of testing sample and is fixed.Utilize masking foil will be placed with the container of putting excellent solid phase carrier and encase lucifuge, fixing 16h under the room temperature.
Among the step S203, last appearance is carried out gene sequencing, detection signal.Solid phase carrier after fixing the end washs with TE, and uses ddH 2O washed.Washed solid phase carrier is packed in the order-checking cell, carry out gene sequencing, collect fluorescent signal.
Wherein, gene order surveying method described in the step S103 can be to connect PCR sequencing PCR, also can be synthetic PCR sequencing PCR.The two concrete principle and operation about this can be given unnecessary details this no longer doing with reference to existing detailed description in the prior art more.
Wherein, figure is as shown in Figure 9 as a result according to connecting the resulting fluorescent signal of PCR sequencing PCR.
Figure 10 has provided that the solid phase carrier based on a kind of surperficial immobilized oligonucleotide carries out the method flow that the gene chip preparation is used in the one embodiment of the invention.Concrete step comprises:
Among the step S301, will be fixed on the solid phase carrier through the active function group of surface of solid phase carriers, be prepared into gene chip with target DNA complementary oligonucleotide fragment.
Among the step S302, testing sample hatched with gene chip combine, catch the target dna fragmentation.
Among the step S303, catch after the end, reaction system is washed, remove and be not attached to the testing sample on the gene chip, thereby obtain the target dna fragmentation with damping fluid.
Figure 11 is based on a kind of solid phase carrier preparation of oligonucleotide and synoptic diagram that is applied to the gene order surveying method flow process of being used for fixing in the one embodiment of the invention.This figure has represented the preparation process that is used for fixing the solid phase carrier of oligonucleotide based on a kind of intuitively, and then carries out the fixing also process of detection signal of oligonucleotide:
Step 1, handle surface of solid phase carriers, make hydroxyl on the surface of solid phase carriers hydroxylation band through the mixed clear liquid of sodium hydroxide and absolute ethyl alcohol.
The solid phase carrier of step 2, the acetone solution that utilizes silylating reagent APTMS and surface hydroxylation reacts, and the shrink reaction through siloxanes and hydroxyl makes the surface of solid phase carriers amination.
Step 3, adopt the mixed solution of activating reagent TMA/NHS/DIEA, the amino through its carboxyl and surface of solid phase carriers carries out condensation reaction, makes surface of solid phase carriers formation activity functional groups; And then pass through NaHCO 3Processing, make its activity functional groups become pendant carboxylic group.
Step 4, carboxyl (in the present embodiment-the R group is a carboxyl) is carried out embedding with polymeric amide PAMAM resin, and then to resin surface with amino according to the method in the step 3 activation again, make the pendant carboxylic group quantity of surface of solid phase carriers become multiple to increase.
Step 5, utilize condensation amino and carboxyl, connect the immobilized oligonucleotide fragment, carry out the detection of signal through the method that connects order-checking or synthetic order-checking then at aforesaid method activatory surface of solid phase carriers.
Should be noted that typical application of the present invention but be not limited to the fixedly amplification of oligonucleotide, contain in the amino compound at other similar protein molecules and other and also can use the method that the present invention sets forth.
The above is merely preferred embodiment of the present invention, not in order to restriction the present invention, all any modifications of within spirit of the present invention and principle, being done, is equal to and replaces and improvement etc., all should be included within protection scope of the present invention.

Claims (12)

1. one kind is used for surperficial fixed solid phase carrier, it is characterized in that, said solid phase carrier is connected with at least one surface group with mobilizing function, and said surface group comprises inboardend and active end, and its molecular structural formula is following:
Figure FSA00000551058200011
Said inboardend has at least one and is used for and solid phase carrier bonded siloxy;
Said active end-R comprises and is used for being fixedly connected thing bonded activity functional groups with the surface.
2. according to claim 1ly be used for surperficial fixed solid phase carrier, it is characterized in that said activity functional groups combines through amido linkage other part with activity end-R.
3. according to claim 1 and 2ly be used for surperficial fixed solid phase carrier, it is characterized in that, said activity functional groups has at least 2 and is used for and the surface is fixedly connected thing bonded end group.
4. according to claim 3ly be used for surperficial fixed solid phase carrier, it is characterized in that said active end-R be the dendritic structure based on resin embedding formation, connect at least 2 and be used for and the surperficial thing bonded end group that is fixedly connected.
5. according to claim 3ly be used for surperficial fixed solid phase carrier, it is characterized in that, said solid phase carrier adopts at least a in slide, silicon chip or the plastics, and its surface is the plane or the irregular surface that has radian of rule.
6. the preparation method of solid phase carrier according to claim 1 is characterized in that, said method comprising the steps of:
A. utilizing silylating reagent that surface of solid phase carriers is carried out end group modifies;
B. utilize activating reagent to combine end group, make activity functional groups on the surface of solid phase carriers band;
C. activity functional groups is carried out resin embedding, form dendroid embedding body;
D. to the activation once more of dendroid embedding body, obtain being used for surperficial fixed solid phase carrier.
7. the preparation method of solid phase carrier according to claim 6 is characterized in that, said steps A comprises:
A1. solid phase carrier is carried out hydroxylation and handle, make surface of solid phase carriers carry hydroxyl;
A2. utilize silylating reagent to combine, surface of solid phase carriers is carried out end group modify with hydroxyl.
8. the preparation method of solid phase carrier according to claim 7 is characterized in that, modifies if in the steps A 2 surface of solid phase carriers is carried out amidized end group, and then said step B comprises:
Through amido linkage activating reagent is combined with the amination end group of surface of solid phase carriers;
Wherein, said activating reagent comprises activity functional groups.
9. the preparation method of solid phase carrier according to claim 8 is characterized in that, if the activity functional groups that activating reagent comprises is a C-terminal, then step C comprises:
C1. through polyamide resin the C-terminal of surface of solid phase carriers is carried out embedding, form dendritic structure;
C2. based on the activation treatment of said dendritic structure repeating step B, make solid phase carrier connect at least 2 and be used for being fixedly connected thing bonded C-terminal with the surface.
10. method of utilizing the described solid phase carrier of claim 1 to carry out gene sequencing is characterized in that said method comprises:
M1. oligonucleotide fragment is connected to solid phase carrier through the activity functional groups effect with the surface;
M2. the said solid phase carrier that is connected with oligonucleotide fragment is carried out signal detection, and through the signal in the testing process is analyzed, thereby base sequence information obtained.
11. the method for gene sequencing according to claim 10 is characterized in that, the signal detection among the said step M2 is meant the signal of measuring base to be measured through the base complementrity principle, thereby obtains the sequence information of base to be measured.
12. a method of utilizing the described solid phase carrier of claim 1 to prepare gene chip is characterized in that said method is: oligonucleotide fragment is connected to solid phase carrier through the activity functional groups effect with the surface, forms gene chip.
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CN110511973A (en) * 2019-07-16 2019-11-29 艾吉泰康生物科技(北京)有限公司 A kind of solid phase carrier and preparation method thereof for nucleic acid fabricated in situ
CN113109574A (en) * 2021-04-15 2021-07-13 北京森美希克玛生物科技有限公司 Kit for detecting anti-mutant citrullinated vimentin antibody and detection method

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CN102787068A (en) * 2012-07-23 2012-11-21 盛司潼 Sample applicator
CN110511973A (en) * 2019-07-16 2019-11-29 艾吉泰康生物科技(北京)有限公司 A kind of solid phase carrier and preparation method thereof for nucleic acid fabricated in situ
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CN113109574A (en) * 2021-04-15 2021-07-13 北京森美希克玛生物科技有限公司 Kit for detecting anti-mutant citrullinated vimentin antibody and detection method

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