CN104844692A - Dodecapeptide capable of binding zearalenone, and applications thereof - Google Patents
Dodecapeptide capable of binding zearalenone, and applications thereof Download PDFInfo
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Abstract
The invention belongs to the field of biotechnology, and relates to a dodecapeptide capable of realizing specific binding with zearalenone (ZEN), and applications thereof. Amino acid sequence of the dodecapeptide molecules is LLNISRRQIKNL. According to a preparation method, magnetic bead liquid phase affinity elutriation and ZEN antibody competitive elution are combined, and dodecapeptide molecules capable of realizing specific binding with ZEN are obtained via optimization of incubation and elution time, temperature, and the like, and rapid elutriation; the dodecapeptide molecules can be used for replacing conventional ZEN antibodies and used in ZEN immunological analysis systems; linear detection range of the dodecapeptide molecules is wide; immunologic process is unnecessary for obtaining of the dodecapeptide molecules; preparation is simple; cost is low; period is short; large-scale amplification of the dodecapeptide molecules can be realized via biological culturing, and large amount preparation can be realized via chemical synthesis or genetic engineering; a novel method is provided for ZEN antibody preparation; and application value is high.
Description
Technical field
The invention belongs to biological technical field, be specifically related to a kind of dodecapeptide in conjunction with zearalenone and application thereof.
Background technology
Zearalenone (Zearalenone, ZEN) be a kind of oestrogen-like hormone sample mycotoxins produced by sickle-like bacteria, its chemistry 6-(10-hydroxyl-6 oxygen base-undecenyl) β-Lei by name locks acid lactone, is extensively present in the cereal crop such as corn, Chinese sorghum, wheat, oat, barley and milk gone mouldy.Because ZEN pollutes generally in grain, the effect by food chain produces toxic action to poultry and HUMAN HEALTH, therefore just seems particularly important to the monitoring of ZEN.
The method of detection ZEN relatively more conventional at present mainly comprises: high performance liquid chromatography (High performance liquidchromatography, HPLC), vapor-phase chromatography (Gaschromatography, GC), thin layer chromatography (Thin layerchromatography, TLC), tablets by HPLC-MS (High performance liquid chromatography-masssepectrum, HPLC-MS), microbiological method and immunodetection etc., wherein immunological analysis method is highly sensitive with it, simple to operate and can the advantages such as scale selection be realized, be widely used in the field of fast detection of ZEN.
The core of immunological analysis method is specific recognition between Ag-Ab and combination, and the antibody therefore preparing specific binding antigen just becomes prerequisite and the core of development immunological analysis method.The research development in recent years of antibody is swift and violent, from traditional polyclone, monoclonal antibody, and the genetic engineering antibody field that to be developed to single-chain antibody, phage displaying antibody, single domain heavy chain antibody, antiidiotypic antibody etc. be representative.Novel antibody has that molecular weight is little, structure is simple, oneself can evolve, prepare characteristics and evolution trend efficiently.Such as, the molecular weight of monoclonal antibody, single-chain antibody, single domain heavy chain antibody reduces gradually, be respectively 150,30-50,15-20kDa, take Lipocalin as the antibody analog of skelemin be 18-20kDa, aptamer is then only a bit of nucleotide sequence.
Along with developing rapidly of phage-displayed polypeptides storehouse technology, by the elutriation of phage display peptide library, can obtain the peptide molecule be combined with specific target body molecular specificity fast, simply, therefore, peptide molecule can become potential novel antibody molecules.The principal feature of phage display peptide library technology effectively to filter out the phage-displayed polypeptides with target target body specific combination, this technology exploring the interphase interaction binding site of acceptor and part, seek the bioactive ligand molecular of high-affinity, explore agnoprotein matter space structure epi-position, be widely used in the development of new generation vaccine etc., but the application of above-mentioned phage-displayed polypeptides technology is confined to be that target body molecule carries out affine elutriation with macro-molecular protein mostly.Because protein surface has the epi-position of multiple copy, be therefore very easy to obtain the corresponding peptide molecule combined with it.Current peptide molecule is applied in immune analysis as the mimic epitopes (antigen surrogate) of antigen mostly, and the report that peptide molecule is applied to immune analysis as the surrogate of antibody for small molecular substance is less, its reason is, small-molecule substance volume is too small, and surface does not have abundant amino, carboxylic group, be difficult to be combined with peptide molecule, thus cause elutriation difficulty.
The present invention is by phage-displayed polypeptides storehouse technology, adopt the affine elutriation of magnetic bead liquid phase and in conjunction with the mode of ZEN antibody competition wash-out, by optimize hatch, elution time, the conditions such as temperature, from phage-displayed polypeptides storehouse rapid screening to can with the peptide molecule of ZEN specific binding, this peptide molecule has the immunology detection characteristic similar to traditional ZEN monoclonal antibody molecule, and the surrogate that can be used as traditional Z EN antibody is applied to the immune analysis system of ZEN.This method avoid the required flow process such as immunologic process, cytogamy, mono-clonal screening long through the cycle of the traditional monoclonal antibody molecule of preparation, have without the need to immunologic process, screen the features such as convenient, the cycle is short, preparation cost is low.
Summary of the invention
The present invention with ZEN holoantigen (the conjugate ZEN-BSA of ZEN and bovine serum albumin) for target molecule, target molecule is attached on magnetic bead, drop into phage random and show dodecapeptide storehouse, carry out the affine elutriation of magnetic bead liquid phase, in conjunction with the mode of ZEN antibody competition wash-out, obtain a kind of can the peptide molecule of specific binding ZEN, its aminoacid sequence is:
LLNISRRQIKNL。
In aforementioned polypeptides molecular structure, capitalization English letter represents the one of known natural L-form amino-acid residue or its D-type isomer respectively, namely L represents leucine residue, N represents l-asparagine acid residue, I represents isoleucine residues, and R represents arginine residues, and S represents serine residue, Q represents glutamine residue, and K represents lysine residue.
The invention still further relates to the Nucleotide of encoding such polypeptides molecule aminoacid sequence, its sequence is:
CTT CTT AAT ATT AGT CGT CGT CAG ATT AAG AAT CTT
The present invention mentions that peptide molecule is prepared in a large number by the mode that Phage amplification, chemosynthesis or genetically engineered are recombinant expressed.Phage amplification refers to will show the phage having peptide molecule, the mode increased by biology, and amount reproduction produces the bacteriophage particles shown and have peptide molecule.Chemosynthesis refers to the aminoacid sequence according to the mimic epitopes announced, and carries out Peptide systhesis by the mode of chemically synthesized polypeptide.The recombinant expressed mode of genetically engineered refers to the gene of coded polypeptide molecule, by being cloned into expression vector, carries out a large amount of preparations of peptide molecule with the form of polypeptide-fusion rotein.
The invention still further relates to the application of described peptide molecule in immunology detection is analyzed.The type of immunology detection comprises the immune analysis type of detection based on Ag-Ab specific reaction such as MBP enzyme linked immuno-adsorbent assay.
The invention has the beneficial effects as follows: the alternative traditional ZEN antibody molecule of the peptide molecule mentioned by the present invention, binding molecule as ZEN directly applies to immunology detection analysis, the features such as its linearity range of indirect competitive ELISA typical curve based on peptide molecule is broad, and this peptide molecule has without the need to immunologic process, obtains easily and fast, the cycle is short.
Accompanying drawing explanation
Fig. 1 is based on the ZEN indirect competitive ELISA typical curve of 12 polypeptide.
Embodiment
The affine elutriation of magnetic bead liquid phase of embodiment 1. and ZEN specific binding polypeptide molecule and qualification thereof
1) affine elutriation with the concrete grammar of ZEN specific binding polypeptide molecule is: for efficiently obtaining the peptide molecule that can be combined with ZEN molecular specificity, have employed the affine elutriation of magnetic bead liquid phase and in conjunction with the mode of ZEN antibody competition wash-out, concrete grammar is: get the carboxylated magnetic bead molecule of 1.0mg (diameter 400nm), wash 3 times with PBST.Add 10 μ L phage randoms and show 12 peptide libraries (2 × 10
11pfu, NEB company), after add 900 μ L PBST, mix with magnetic bead, under room temperature, jog shakes anti-60min, after Magneto separate, (object of this step is to remove the peptide molecule be combined with magnetic bead surfaces modification group in phage display peptide library, this kind of peptide molecule is adsorbed on magnetic bead surfaces and is got rid of by Magneto separate), careful Aspirate supernatant (peptide molecule be not combined with magnetic bead surfaces modification group) is to another new centrifuge tube, add magnetic bead (the diameter 400nm of 1.0mg ZEN-BSA coupling, the coupling amount of ZEN-BSA is 1.0mg, the coupling ratio of ZEN:BSA is 15:1, the preparation method of conjugate is see Burkin ea al., 2000, Appliedbiochemistry and microbiology, 36, 282-288)), concussion reaction 45min under room temperature.After Magneto separate, wash 6 times with TBST, wash away unconjugated phage.Add 1mL 150ng/mL ZEN antibody (PBS system), jog concussion 60min under room temperature.After Magneto separate, careful absorption supernatant, obtain the first round affine elutriation product, 10 μ L are stayed to survey titre, residue phage input is in logarithm ER2738 in earlier stage increase, then carry out the 2nd successively to take turns and take turns elutriation with the 3rd, panning step is substantially identical with the first round, and the phagocytosis scale of construction at every turn added is 2 × 10
11pfu, difference is: 2nd, the incubation time of 3 phage-displayed polypeptides and ZEN-BSA coupled bead of taking turns screening is respectively 30min, 15min, and the concentration of competitive elution liquid (ZEN antibody) is respectively 75ng/mL, 50ng/mL.
2) qualification of positive phage clones: measure random picking 80 phage spots the flat board of phage titre after third round elutriation, carry out the amplification of phage, Immunofluorescent antibody detection method is adopted to carry out the qualification of positive phage clones, concrete grammar is: first, ZEN-BSA antigen is diluted with 10mM PBS (pH 7.4), 2 μ g/mL wrap by 96 hole enzyme plates, 4 DEG C of overnight incubation.Within second day, with after PBST (10mM PBS, 0.05%Tween-20 (v/v)) washing 3 times, close with the PBS containing 3% skim-milk, hatch 1 hour for 37 DEG C; Drop into 100 μ l phage spot amplification liquid (1.0 × 10
12pfu), using naive phage peptide storehouse as negative control, hatch 1 hour for 37 DEG C; Add the HRP that 1:5000 doubly dilutes and mark the anti-100 μ l of anti-M13 phage two, hatch 1 hour for 37 DEG C; Add 100 μ l tmb substrate liquid, lucifuge colour developing 5min, microplate reader reads the absorption value at 450nm place.Choose OD
450the phage clone being greater than negative control 2 times is positive colony.
3) qualification of specific binding ZEN peptide molecule: adopt the method for indirect competitive ELISA to carry out the qualification with ZEN specific binding phage-displayed polypeptides, concrete grammar is: with 10mM PBS (pH 7.4) dilution ZEN-BSA, 2 μ g/mL coated elisa plate 100 μ L, 4 DEG C of overnight incubation; Within second day, with after PBST (10mM PBS, 0.05%Tween-20 (v/v)) washing 3 times, close with the PBS containing 3% skim-milk, hatch 1 hour for 37 DEG C; Drop into 50 μ l and be accredited as positive phage clone (1.0 × 10 through indirect ELISA
11pfu) and 50 μ l ZEN standard substance (concentration range is 0-1000ng/mL), hatch 1 hour for 37 DEG C; Add the anti-100 μ l of anti-M13 phage two that 1:5000 dilutes HRP mark, hatch 1 hour for 37 DEG C; Add 100 μ l tmb substrate liquid, lucifuge colour developing 5min, reads OD
450.If phage display peptide molecule can specific binding ZEN, then the OD value adding the hole of ZEN standard substance will lower than control wells (not adding ZEN standard substance).
The order-checking of peptide molecule encoding gene of embodiment 2. and ZEN specific binding and the determination of aminoacid sequence thereof
Show have the phage of specific binding ZEN peptide molecule to increase by through indirect competitive ELISA qualification, extract the DNA sequencing template of phage.Simplified process is as follows: carry out Phage amplification, after the first step is centrifugal, 800 μ l is proceeded to a new centrifuge tube containing phage supernatant.Add 200 μ l PEG/NaCl precipitating phage.After centrifugal, precipitation is resuspended in 100 μ l iodide damping fluid (10mM Tris-HCl (pH 8.0), 1mM EDTA, 4M NaI), add 250 μ l dehydrated alcohol precipitation DNA, again by 70% washing with alcohol precipitation (DNA sequencing template) after centrifugal.Precipitation is finally resuspended in 20 μ l aqua sterilisas, gets 2 μ l and carries out agarose gel electrophoresis analysis; Get 5 μ L phage templates and carry out DNA sequencing, its-96gIII sequencing primer is: 5 '-
hOcCC TCA TAG TTA GCG TAA CG-3 '.DNA sequencing result shows the nucleotide sequence of its amino acid coding: CTT CTT AAT ATT AGT CGT CGT CAG ATT AAG AAT CTT, aminoacid sequence is: LLNISRRQIKNL.
The application of embodiment 3. specific binding ZEN peptide molecule in ELISA
(1) sample extraction
Take 5g testing sample, add the methyl alcohol-PBS solution of 25ml 60%, 200rpm vibrates 5 minutes; After extracting solution whatman No. 1 filter paper is filtered, get 1ml filtrate and add 4ml PBS (phosphate buffered saline buffer, pH=7.2)
After mixing, be sample extracting solution, stand-by.
(2) wrap by and close
The ZEN-BSA (2 μ g/ml) diluted with 10mM PBS (pH 7.4), coated elisa plate, 4 DEG C of overnight incubation.Within second day, with after PBST (10mM PBS, 0.05%Tween-20 (v/v)) washing 3 times, close with containing the PBS of 3% skim-milk, 37 DEG C hatch 1 hour after, wash plate 6 times with PBST stand-by.
(3) foundation of typical curve
Take out the lath handled well through step (2), every hole is dropped into 50 μ l respectively and is shown the phage (1.0 × 10 having specific binding ZEN peptide molecule
12pfu) and 50 μ l ZEN standard substance (0-1000ng/ml) of a series of different concns, 20min is hatched for 37 DEG C.The anti-M13 phage two adding 1:5000 dilution HRP mark resists, and hatches 1 hour for 37 DEG C.Then with tmb substrate colour developing, OD is read
450.With ZEN log concentration for X-coordinate, combination rate (adds the OD in the hole of ZEN
450/ do not add the OD in the hole of ZEN
450× 100%) be ordinate zou, set up indirect competition typical curve.
(4) detection of sample
Take out the lath handled well through step (2), every hole is dropped into 50 μ l respectively and is shown the phage (1.0 × 10 having specific binding ZEN peptide molecule
12pfu) and testing sample extracting solution, 1 hour is hatched for 37 DEG C.The anti-M13 phage two adding 1:5000 dilution HRP mark resists, and hatches 1 hour for 37 DEG C.Then with tmb substrate colour developing, OD is read
450, calculations incorporated rate, and according to typical curve, the content of ZEN in sample of retrodicting out.
Embodiment 4 and ZEN specific binding polypeptide molecule a large amount of preparations
1) in the mode of Phage amplification
Phage with ZEN specific binding being added to 20ml inoculates in the culture of ER 2738,37 degree of 220rpm shaking culture 4.5h.Proceeded to by culture in another centrifuge tube, 4 DEG C of centrifugal 10min of 10000rpm, proceed to the top of supernatant 80% in a fresh tube, add the PEG/NaCl of 1/6 volume, leave standstill 120min at 4 DEG C.4 DEG C of centrifugal PEG/NaCL of 10000rpm leave standstill solution 15min.Abandon supernatant, of short duration centrifugal after suck residual supernatant liquor.Adding 1mL TBS carries out resuspended, is Phage amplification liquid.
2) be prepared in the mode of polypeptide-fusion rotein
The external source encoding gene of A.PCR amplification peptide molecule
PCR reaction system: (50 μ L)
PCR reaction conditions:
Adopt PCR primer to reclaim kits PCR primer, trace dna quantitative instrument is quantitative.The coding gene sequence of peptide molecule is: CTT CTT AAT ATT AGT CGT CGT CAG ATT AAG AAT CTT.
B. the double digestion of external source encoding gene and expression vector
ACC65I and the Eag external source code gene of I enzyme and expression vector (MBP fusion rotein can be expressed by pMAl-pIII, NEB company) is adopted to carry out double digestion respectively.
C. enzyme cuts connection and the conversion of after product
Plasmid pMal-PIII and object fragment are mixed with 1: 20 (mol ratio), connects 12h in 20 DEG C of water-baths, get 15 μ L connection products and add in 100 μ L competent cell DH5 α, fully mix.After ice bath 30min, 42 DEG C of water-bath heat shock 40s, 800 μ L LB liquid mediums are added immediately after ice bath 3min, 37 DEG C, 200rpm cultivates the centrifugal 10min of 1h, 8000rpm, suck supernatant and leave and take about 300 μ L, coat in LB-A solid (Amp) substratum, 37 DEG C of incubated overnight, obtain subclone positive colony.
D. Cloning Transformation
Subclone sky root test kit above-mentioned steps obtained extracts object plasmid, gets in 10 μ L to 100 μ L competent cell TB1, fully mixes.After ice bath 30min, 45 DEG C of water-bath heat shock 50s, 800 μ L LB liquid mediums are added immediately after ice bath 5min, 37 DEG C, 200rpm cultivates the centrifugal 10min of 45min, 7000rpm, suck supernatant and leave and take about 300 μ L, coat in LB-A solid (Amp) substratum, 37 DEG C of incubated overnight, obtain positive colony.
E. the expression of polypeptide-MBP fusion rotein
By the positive colony of above-mentioned acquisition, a single colony inoculation is chosen in 5mL LB-A from flat board, in 0.5% sucrose, 37 DEG C, 220r/min, shaking culture is spent the night, overnight culture is inoculated in the LB-A of 50mL by 1% inoculum size (v/v), in 0.2% sucrose medium, inoculate 3 bottles respectively, 37 DEG C, 220r/min shaking culture, when culture bacterial concentration OD600 reaches 0.6, in three bottles of cultures, add IPTG to final concentration is 0.5mmol/L, 120r/min shaking culture, by inductor (IPEG solution) in 4 DEG C, 5000g, centrifugal 15min collects bacterial sediment, abandon supernatant.Re-suspended cell in 400mL 30mM Tris-HCl, 25% sucrose, pH 8.0 (80mL/g wet cell weight), add EDTA to 1mM, shake 5-10min under room temperature, 8000g, 4 DEG C of centrifugal 15min, abandon supernatant, and precipitation is resuspended in the 5mM MgSO4 of 400ml precooling, shake 15min on ice, 8000g, 4 DEG C, centrifugal 20min, retains supernatant, adds 8mL 1M Tris-HCl in supernatant liquor, pH 7.4, obtains polypeptide-MBP fusion rotein.
The immunology detection stability experiment of embodiment 5 and ZEN specific binding polypeptide molecule
1) resistance to acids and bases experiment
Chemosynthesis can be used pH=4.0 respectively with the dodecapeptide of ZEN specific binding and ZEN monoclonal antibody, 5.0,6.0,7.4, the damping fluid of 8.0 is diluted to working fluid, carries out indirect competitive ELISA mensuration respectively, and concrete grammar is: the ZEN-BSA (2 μ g/ml) diluted with 10mM PBS (pH 7.4), coated elisa plate, 4 DEG C of overnight incubation.Within second day, with after PBST (10mM PBS, 0.05%Tween-20 (v/v)) washing 3 times, close with containing the PBS of 3% skim-milk, 37 DEG C hatch 1 hour after, wash plate 6 times with PBST stand-by.Take out the lath handled well, 50 μ l are dropped in every hole respectively can the peptide molecule/ZEN monoclonal antibody (20 μ g/ml) of specific binding ZEN and 50 μ l ZEN standard substance (0-1000ng/ml) of a series of different concns, hatch 20min for 37 DEG C.Add 1:5000 dilute the anti-M13 phage two of HRP mark anti-/sheep anti-mouse igg two of HRP mark resists, and hatches 1 hour for 37 DEG C.Then with tmb substrate colour developing, OD is read
450.With ZEN log concentration for X-coordinate, combination rate (adds the OD in the hole of ZEN
450/ do not add the OD in the hole of ZEN
450× 100%) be ordinate zou, set up indirect competition typical curve.
Experimental result shows: the indirect competitive ELISA using peptide molecule as ZEN recognition component, at pH=4.0, and 5.0,6.0,7.4, when 8.0, its IC
50be respectively 60,55,53,63,58ng/ml; With the indirect competitive ELISA that ZEN monoclonal antibody is recognition component, at pH=4.0,5.0, do not have good blocking effect when 6.0, present smooth curve, the immune analysis performance of antibody is subject to remarkably influenced, at pH=7.4, when 8.0, its IC
50be respectively 46,39ng/ml, surface is compared to traditional ZEN monoclonal antibody, and the peptide molecule of ZEN antibody surrogate thing has better soda acid patience.
2) high temperature resistance experiment
By chemosynthesis can be placed in temperature respectively with the peptide molecule of ZEN specific binding and ZEN monoclonal antibody be 37 DEG C, 45 DEG C, 50 DEG C, 60 DEG C, hatch 30min in the water-bath of 70 DEG C, carry out indirect ELISA experiment respectively after taking-up, concrete grammar is: the ZEN-BSA (2 μ g/ml) diluted with 10mM PBS (pH 7.4), coated elisa plate, 4 DEG C of overnight incubation.Within second day, with after PBST (10mM PBS, 0.05%Tween-20 (v/v)) washing 3 times, close with containing the PBS of 3% skim-milk, 37 DEG C hatch 1 hour after, wash plate 6 times with PBST stand-by.Take out the lath handled well, the 50 μ l of hatching through differing temps are dropped in every hole respectively can the peptide molecule/ZEN monoclonal antibody (20 μ g/ml) of specific binding ZEN, hatches 20min for 37 DEG C.Add 1:5000 dilute the anti-M13 phage two of HRP mark anti-/sheep anti-mouse igg two of HRP mark resists, and hatches 1 hour for 37 DEG C.Then with tmb substrate colour developing, OD is read
450, with OD value when 37 DEG C for 100%, OD under calculating differing temps
450the stability % of value.
Experimental result shows: the indirect ELISA using peptide molecule as ZEN recognition component is 37 DEG C in temperature, 45 DEG C, 50 DEG C, 60 DEG C, and when 70 DEG C, its temperature degree is respectively 100%, 98%, 95%, 88%, 86%; Indirect ELISA using ZEN monoclonal antibody as ZEN recognition component is 37 DEG C in temperature, 45 DEG C, 50 DEG C, 60 DEG C, when 70 DEG C, its temperature degree is respectively 100%, 75%, 30%, 25%, 16%, show that, compared to traditional ZEN monoclonal antibody, the peptide molecule of ZEN antibody surrogate thing has better high temperature resistance.
SEQUENCE LISTING
<110> University Of Nanchang
<120> mono-kind can in conjunction with the dodecapeptide of zearalenone and application thereof
<130> 1
<160> 1
<170> PatentIn version 3.3
<210> 1
<211> 12
<212> PRT
<213> artificial sequence
<400> 1
Leu Leu Asn Ile Ser Arg Arg Gln Ile Lys Asn Leu
1 5 10
Claims (6)
1. in conjunction with the dodecapeptide of zearalenone, can it is characterized in that its aminoacid sequence is:
LLNISRRQIKNL。
2. the Nucleotide of peptide molecule aminoacid sequence described in coding claim 1.
3. nucleotide sequence as claimed in claim 2 corresponds to: CTT CTT AAT ATT AGT CGT CGT CAG ATT AAG AAT CTT.
4. peptide molecule preparation method as claimed in claim 1, is characterized in that the mode by Phage amplification, chemosynthesis or genetically engineered are recombinant expressed is prepared in a large number; Described Phage amplification refers to the phage of displayed polypeptides molecule, the mode increased by biology, and amount reproduction is produced and shown that have can the bacteriophage particles of specific binding ZEN peptide molecule; Described chemosynthesis refers to the aminoacid sequence according to peptide molecule, carries out Peptide systhesis by the mode of chemically synthesized polypeptide; The recombinant expressed mode of described genetically engineered refers to the gene of coded polypeptide molecule, by being cloned into expression vector, prepares in a large number with the form of polypeptide-fusion rotein.
5. the application of peptide molecule described in claim 1 in immunology detection is analyzed.
6. apply as claimed in claim 5, it is characterized in that the application of peptide molecule replacement of corn zeranol antibody in immunology detection is analyzed.
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102443585A (en) * | 2011-11-25 | 2012-05-09 | 国家纳米技术与工程研究院 | Zearalenone nucleic acid aptamer and application thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN102443585A (en) * | 2011-11-25 | 2012-05-09 | 国家纳米技术与工程研究院 | Zearalenone nucleic acid aptamer and application thereof |
Non-Patent Citations (2)
| Title |
|---|
| 何庆华等: "利用噬菌体肽库淘选玉米赤霉烯酮的模拟表位", 《食品科学》 * |
| 徐富勇等: "噬菌体展示系统表达玉米赤霉烯酮模拟表位肽", 《生物技术通报》 * |
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