CN101818134A - ppGalNAc-T20 antigen and preparation method of polyclonal antibody thereof - Google Patents

ppGalNAc-T20 antigen and preparation method of polyclonal antibody thereof Download PDF

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CN101818134A
CN101818134A CN201010300097A CN201010300097A CN101818134A CN 101818134 A CN101818134 A CN 101818134A CN 201010300097 A CN201010300097 A CN 201010300097A CN 201010300097 A CN201010300097 A CN 201010300097A CN 101818134 A CN101818134 A CN 101818134A
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ppgalnac
polyclonal antibody
antigen
seq
preparation
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CN101818134B (en
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张延�
彭灿
武功冬
邹霞
谢文娴
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Shanghai Jiaotong University
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Abstract

The invention relates to a ppGalNAc-T20 antigen and a preparation method of a polyclonal antibody thereof. An amino acid sequence of the ppGalNAc-T20 antigen is shown as SEQ ID NO: 2. The invention also relates to separated nucleic acid, a base sequence of which is shown as SEQ ID NO: 1, and also relates to a polyclonal antibody which is prepared by adopting the following steps of: taking protein shown as the amino acid sequence SEQ ID NO: 2 as an antigen, and preparing antiserum by adopting a conventional polyclonal antibody preparation method; and purifying the antiserum to obtain the polyclonal antibody. The invention also relates to a method for preparing the polyclonal antibody, comprising the following steps of: cloning nucleic acid fragments shown as the SEQ ID NO: 1; establishing recombinant plasmids, converting colibacillus, culturing, inducing, pyrolyzing and purifying to obtain protein; taking the protein as the antigen, preparing the antiserum by adopting the conventional polyclonal antibody preparation method, and purifying the antiserum to obtain the polyclonal antibody. The invention has simple method, and the prepared antibody has strong immunogenicity.

Description

PpGalNAc-T20 antigen and Polyclonal Antibody Preparation method thereof
Technical field
The present invention relates to antibody of a kind of biological technical field and preparation method thereof, specifically is a kind of ppGalNAc-T20 antigen and Polyclonal Antibody Preparation method thereof.
Background technology
UDP-semi-lactosi acid amides: N-acetylamino galactosamine transferring enzyme (ppGalNAc-Ts) is the initial glycosyltransferase of the first step of O-type glycoprotein candy chain building-up reactions, and it can be with the group-transfer of N-acetylamino galactosamine on the Serine or threonine residues of protein peptide chain.Oneself has 19 members research that is in the news in the ppGalNAc-Ts family in the human body, they in tissue distribution and to differences in various degree all on the glycosylation modified specificity of external peptide substrate.PpGalNAc-Ts can be used as tumor markers, the ppGalNAc-T13 of specificity overexpression in the strain of neuroblastoma derived cell for example, and it can be used as the early diagnosis sign of spinal cord generation neuroblastoma.In addition, ppGalNAc-T6 can be used as the immunohistochemical methods detection sign of mammary cancer.Current research is the result show, ppGalNAc-T14 regulation and control death receptor Apo2L/TRAIL's is glycosylation modified, crosses and expresses the apoptosis that ppGalNAc-T14 can significantly promote cell.PpGalNAc-Ts also has potential important clinical diagnostic significance except that having the glycosyltransferase function.Therefore, obtain ppGalNAc-Ts each member's of family antibody, it is being distributed with important meaning in tissue and cell to accurate location, thus their biological function of more real reflection.Yet the homology between the ppGalNAc-Ts family member is higher, and the homology between different members can reach 40%～70%, therefore guarantees that the specificity of its antibody is most important.
The ppGalNAc-Ts family member is II type membranin, the N end has 4～22 amino acid whose cytoplasmic domains, continue with 15～25 amino acid whose transmembrane domains, stretch into by a stem district different in size and C end that about 450 amino acid whose catalytic domains are connected in the golgi body.PpGalNAc-T20 belongs to UDP-semi-lactosi acid amides from Primary Structure Analysis: polypeptide N-acetylamino galactosamine transferring enzyme family member, it and ppGalNac-T10 have 70.7% similarity on amino acid levels.Though only in brain and testis tissue, detect the lower ppGalNAc-T20mRNA of expression amount,, can infer that it plays important regulatory role in body differentiation, growth course in view of there is the singularity at position in it.
Find through literature search prior art, employing polypeptide synthetic methods such as N Berois et al. prepare ppGalNAc-T13 antibody (N Berois et al., ppGalNAc-T13:A New Molecular Marker of BoneMarrow Involvement in Neuroblastoma, Clinical Chemistry, 2006,52:1701～1712), this method generally needs synthetic about 15 peptide section, like this than the specificity that is easier to guarantee antibody.But because the molecular weight of polypeptide is less relatively, immunogenicity is relatively poor, obtains relatively difficulty of antibody, needs usually itself and MAP (multipleantigenic peptide) or KLH (keyhole limpet hemocyanin) coupling have still been improved cost like this.Generally be used for preparing the antigen peptide need 10～20mg of antibody, purity is more than 85%, the synthetic price of this other polypeptide of level is 120 a yuan/amino acid, the price of coupling group MAP is also more than 1000 yuan, lower immunogenicity and high cost have limited the application that synthetic polypeptide is used for preparation method for antibody.The research report relevant with the ppGalNAc-T20 Antibody Preparation do not arranged at present as yet.
Summary of the invention
The objective of the invention is to overcome the deficiencies in the prior art, a kind of ppGalNAc-T20 antigen and Polyclonal Antibody Preparation method thereof are provided.Cross reaction takes place in the ppGalNAc-T10 that the antibody discord and the ppGalNAc-T20 of the present invention's preparation has highest homology, more with other member of ppGalNAc-Ts family cross reaction does not take place; The present invention goes out required antigen protein by simple molecular cloning Experiment Preparation, and this antigen protein and GST amalgamation and expression, is easy to purifying; The present invention has overcome that synthetic polypeptide method immunogenicity is poor, shortcoming such as cost an arm and a leg.
The present invention realizes by following technical scheme,
The present invention relates to a kind of ppGalNAc-T20 antigen, this antigen is protein, and its aminoacid sequence is shown in SEQ ID NO:2.
The base sequence of the nucleic acid of code for said proteins is shown in SEQ ID NO:1.
The invention still further relates to the antigenic preparation method of polyclonal antibody of a kind of aforesaid ppGalNAc-T20, comprise the steps:
Step 1 is an antigen with the protein of aminoacid sequence shown in SEQ ID NO:2, adopts conventional preparation method of polyclonal antibody to prepare antiserum(antisera);
Step 2, the purifying antiserum(antisera), promptly.
The invention still further relates to the antigenic preparation method of polyclonal antibody of a kind of aforesaid ppGalNAc-T20, comprise the steps:
Step 1, the nucleic acid fragment of clone shown in SEQ ID NO:1;
Step 2 is utilized step 1 gained nucleic acid fragment construction recombination plasmid, and transformed into escherichia coli is cultivated, induce, and cracking, purifying gets protein;
Step 3 is an antigen with step 2 gained protein, adopts conventional preparation method of polyclonal antibody to prepare antiserum(antisera), the purifying antiserum(antisera), promptly.
In the step 1, described clone, the primer is to being specially upstream primer shown in SEQ ID NO:3 and the downstream primer shown in the SEQ IDNO:4.
In the step 2, described intestinal bacteria are e. coli bl21 (DE3).
In the step 2, the plasmid that described recombinant plasmid uses is pGEX-5X-1.
In the step 2, the process of screening described recombinant plasmid is: in the recombinant plasmid transformed bacillus coli DH 5 alpha that makes up, use contains the LB flat board of 100 μ g/ml penbritins, cultivate 12h down for 37 ℃, shake bacterium behind the picking list bacterium colony, extract plasmid and order-checking, the correct required recombinant plasmid that is checks order.
In the step 2, described cultivation is specially, the consisting of of 1L substratum: peptone 10g, and yeast extract 5g, NaCl10g, surplus is a water; Culture temperature is 37 ℃; The OD value that is cultured to e. coli bl21 (DE3) is 0.8～1.2.
In the step 2, described induce into: add IPTG, making its final concentration is 0.1mM～0.5mM, and temperature is cultivated 3h～6h down for 26 ℃.
Compared with prior art, the present invention has following beneficial effect: the polyclonal antibody that the present invention obtains is that the one section albumen in stem district with ppGalNAc-T20 obtains as antigen; Cross reaction takes place in the ppGalNAc-T10 that the antibody discord and the ppGalNAc-T20 of the present invention's preparation has highest homology, more with other member of ppGalNAc-Ts family cross reaction does not take place; The present invention goes out required antigen protein by simple molecular cloning Experiment Preparation, and this antigen protein and GST amalgamation and expression, is easy to purifying; The present invention has overcome that synthetic polypeptide method immunogenicity is poor, shortcoming such as cost an arm and a leg; The antibody that the present invention obtains not only can be used to detect the ppGalNAc-T20 albumen after the sex change, can also be used to detect the ppGalNAc-T20 albumen with active native conformation.
Among the present invention related bacterial strain bacillus coli DH 5 alpha, e. coli bl21 (DE3) " Wang Lingling, poplar is collected, yellow really it, Wang Haihong; Intestinal bacteria holo-ACP crosses the synthetic of expression, separation and purification and long-chain acyl ACP, microorganism journal, 2008,48 (7): 963～969 " open in the document.The bacterial strain that the present invention relates to can be obtained by disclosing commercially available commercial channel, as sky, Shanghai root biotech firm, and CompanyAddress: Room 606, No. 1 building, No. 27 boats star commercial affairs building, Shanghai City Cao Xilu 258 lanes.
Description of drawings
Fig. 1 is a GST-short T20 fusion rotein abduction delivering electrophorogram;
Fig. 2 is a GST-short T20 fusion rotein purifying electrophorogram;
Fig. 3 is a ppGalNAc-T20 antibody purification electrophorogram;
Fig. 4 be ppGalNAc-T20 antibody with other ppGalNAc-T intersect specific detection figure;
Fig. 5 is for detecting ppGalNAc-T20 antibody to the proteic WesternBlotting that tires of the GST-short T20 of different concns figure as a result;
Fig. 6 is for detecting ppGalNAc-T20 antibody to the proteic Western Blotting that tires of FLAG-ppGalNAc-T20 of the different concns of expressing figure as a result in the 293T cell;
Fig. 7 is for detecting ppGalNAc-T20 protein expression distribution results figure in the C57 mouse testis tissue with ppGalNAc-T20 antibody by the immunohistochemical methods method;
Fig. 8 is the Western Blotting that detects MLTC-1 mouse testis Leydig's cell tumor cell pyrolysis liquid by the co-immunoprecipitation method with ppGalNAc-T20 antibody figure as a result.
Embodiment
Following examples will the invention will be further described in conjunction with the accompanying drawings.Present embodiment has provided detailed embodiment and process being to implement under the prerequisite with the technical solution of the present invention, but protection scope of the present invention is not limited to following embodiment.The experimental technique of unreceipted actual conditions in the following example carries out according to normal condition or according to the condition that manufacturer advises usually.
Embodiment
The ppGalNAc-T20 Polyclonal Antibody Preparation
Step 1, the establishment of antigen sequence
Importing the protein sequence of ppGalNAc-T20 (GenBank Accession Number:GU220060) and ppGalNAc-T10 (GenBank Accession Number:AJ505950) in ClustalX software compares, choose one section sequence short T20 as antigen in the proteic stem of ppGalNAc-T20 district, short T20 sequence is (ppGalNAc-T20 holoprotein coding 30AA is to 141AA) shown in SEQ ID NO:2;
Step 2, construction of recombinant plasmid
The aminoacid sequence corresponding DNA sequences of antigen short T20 is shown in SEQ ID NO:1; Design primer according to the multiple clone site of pGEX-5X-1 and the dna sequence dna of short T20:
Upstream primer: 5 '-GTT GCG AGC GGC CGC CTG TAC AAG GAT-3 '; (SEQ ID NO:3);
Downstream primer: 5 '-GATAAT GAG TCG ACC GTT TGG CAG CCTTTC-3 ' (SEQ ID NO:4).
(GenBank Accession Number:GU220060) is template with the ppGalNAc-T20 full-length cDNA, obtains the purpose fragment by pcr amplification, and the PCR reaction conditions is: 94 ℃ of 3min; 94 ℃ of 30s, 56 ℃ of 30s, 68 ℃ of 30s, 25 circulations; 68 ℃ of 10min; Purpose fragment glue behind Sal I (TaKaRa), Not I (TaKaRa) double digestion reclaims, connect into pGEX-5X-1 plasmid (Amersham) with Ligation High ligase enzyme (ToYoBo) through Sal I, Not I double digestion, the ligation condition is: 16 ℃, and 2h;
With constructed pGEX-5X-1-short T20 prokaryotic expression carrier transformed into escherichia coli DH5 α (day root), use the LB flat board that contains 100 μ g/ml penbritins, 37 ℃ of screening and culturing of spending the night; Shake bacterium behind the picking list bacterium colony, extract plasmid and order-checking.
Step 3 is with recombinant plasmid transformed e. coli bl21 (DE3);
Select the plasmid transformation escherichia coli BL21 (DE3) that order-checking is correct in the step 2 (day root), 1L e. coli bl21 (DE3) nutrient solution is cultured to the OD value at 37 ℃ and reaches 1.0; E. coli bl21 (DE3) nutrient media components is: 1% (w/v) peptone, and 0.5% (w/v) yeast extract, 1% (w/v) NaCl, in the described 1L substratum, peptone 10g, yeast extract 5g, NaCl 10g, surplus is a water; Culture temperature is 37 ℃.
In e. coli bl21 (DE3) nutrient solution, add IPTG to 0.5mM, 26 ℃ of following inducing culture 3h; Cracking e. coli bl21 (DE3): e. coli bl21 (DE3) medium centrifugal after 1L induced is collected thalline, resuspended with the 100ml lysis buffer, the component of this damping fluid is: PBS (pH=7.3), 1% (v/v) Triton X-100, lmM PMSF, in the described 1L damping fluid, Triton X-100 10ml, PMSF 174.2mg, surplus is PBS (pH=7.3); Ultrasonic disruption 20 times, each 5s.Then will be under 4 ℃ of conditions through the thalline behind the ultrasonic disruption, 10, the centrifugal 15min of 000rpm collects supernatant.As shown in Figure 1,1 is marker, and 2 for not using the albumen supernatant behind the IPTG inductive cellular lysate, and 3 are the albumen supernatant behind the cellular lysate after inducing with IPTG.As shown in Figure 1, in e. coli bl21 (DE3), can obtain GST-short T20 fusion rotein by the IPTG abduction delivering.
Step 4, albumen supernatant after the fragmentation of purifying e. coli bl21 (DE3) cellular lysate: the GSTrap HP affinity purification post that uses GE Healthcare company, first this post of PBS balance with 5 times of column volumes, the supernatant that obtains after then last sample ultrasonication is centrifugal, PBS with 5 times of column volumes washes this post again, uses the reductive glutathione eluant solution of 5 times of column volumes then.The component of reductive glutathione solution is: 50mM Tris-HCl, 10mM reductive glutathione, pH are 8.0, detect by UV and collect elution peak.As shown in Figure 2,1 is marker, and 2 are the albumen supernatant behind the cellular lysate of unpurified IPTG after inducing; 3 for penetrating the peak, and promptly not by GSTrap HP post bonded albumen, 4 is elution peak, promptly with reductive glutathione GST-short T20 fusion rotein under the wash-out from the GSTrap HP post.
Step 5 is carried out immunity with the antigen protein of purifying to rabbit, uses antigen protein 200 μ g at every turn, and fortnight once; Get serum after 3 months, 3,000rpm obtains antiserum(antisera) after centrifugal 15 minutes; The purifying antiserum(antisera): use the Hitrap Protein A HP antibody purification post of GE Healthcare company, earlier with 10 times of column volumes in conjunction with this post of liquid balance, be 20mM Na in conjunction with the liquid composition 3PO 4, pH is 7.0, then allows the ppGalNAc-T20 antiserum(antisera) by this post, again with 5 times of column volumes wash this post in conjunction with liquid, use the elutriant wash-out of 5 times of column volumes then, the elutriant composition is 0.1M Na 3C 6H 5O 7, pH is 3.0, detects by UV (254nm) and collects elution peak.As shown in Figure 3,1 is marker, and 2 is unpurified ppGalNAc-T20 antiserum(antisera), and 3 is the ppGalNAc-T20 polyclonal antibody behind the purifying.
Present embodiment gained antibody is that the one section albumen in stem district with ppGalNAc-T20 obtains as antigen, cross reaction with the highest ppGalNAc-T10 of ppGalNAc-T20 homology does not take place in this antibody, with other ppGalNAc-Ts family member cross reaction does not take place yet.Have purposes such as immunoblotting, histocyte dyeing and immunosedimentation.Not only can be used for detecting in the proteic body of ppGalNAc-T20, can also be used for the proteic vitro detection of ppGalNAc-T20.
Implementation result
(1)Western?Blotting
Fig. 4, Fig. 5 and Fig. 6 are specificity and the sensitivity that detects the polyclonal antibody of present embodiment preparation with Western Blotting.As shown in Figure 4, from left to right be followed successively by the 293T cell pyrolysis liquid of ppGalNAc-T1, the ppGalNAc-T2, ppGalNAc-T3, ppGalNAc-T4, ppGalNAc-T6, ppGalNAc-T8, ppGalNAc-T9, ppGalNAc-T10, ppGalNAc-T12, ppGalNAc-T13, ppGalNAc-T14, ppGalNAc-T15, ppGalNAc-T16, ppGalNAc-T17, ppGalNAc-T18 and the ppGalNAc-T20 that contain band Flag label.Figure A, figure B and the 293T cell pyrolysis liquid of scheming to be respectively among the C transfection Flag-ppGalNAc-Ts plasmid, figure A is anti-ppGalNAc-T20 antibody test figure, figure B is anti-FLAG antibody test figure.
Wherein: figure A is for the proteic Western Blotting of ppGalNAc-T1, ppGalNAc-T2, ppGalNAc-T3, ppGalNAc-T4, ppGalNAc-T6, ppGalNAc-T8, ppGalNAc-T9, ppGalNAc-T10, ppGalNAc-T12, ppGalNAc-T13, ppGalNAc-T14, ppGalNAc-T15, ppGalNAc-T16, ppGalNAc-T17, ppGalNAc-T18 and ppGalNAc-T20 of ppGalNAc-T20 antibody test band Flag label figure as a result.One anti-ratio is 1: 2000, and anti-rabbit two anti-ratios are 1: 10000, development time 10min;
Figure B is for the proteic Western Blotting of ppGalNAc-T1, ppGalNAc-T2, ppGalNAc-T3, ppGalNAc-T4, ppGalNAc-T6, ppGalNAc-T8, ppGalNAc-T9, ppGalNAc-T10, ppGalNAc-T12, ppGalNAc-T13, ppGalNAc-T14, ppGalNAc-T15, ppGalNAc-T16, ppGalNAc-T17, ppGalNAc-T18 and ppGalNAc-T20 of anti-Flag antibody test band Flag label figure as a result; One anti-ratio is 1: 2000 (HRPconjugated), need not two and resists development time 10min;
Figure C is for the proteic Western Blotting of ppGalNAc-T1, ppGalNAc-T2, ppGalNAc-T3, ppGalNAc-T4, ppGalNAc-T6, ppGalNAc-T8, ppGalNAc-T9, ppGalNAc-T10, ppGalNAc-T12, ppGalNAc-T13, ppGalNAc-T14, ppGalNAc-T15, ppGalNAc-T16, ppGalNAc-T17, ppGalNAc-T18 and ppGalNAc-T20 of anti-actin antibody test band Flag label figure as a result; One anti-ratio is 1: 2000, and anti-mouse two anti-ratios are 1: 2000, development time 10min.
Fig. 4 illustrates that the ppGalNAc-T20 antibodies specific is good, with other ppGalNAc-Ts family member cross reaction does not take place; Fig. 5 illustrates that ppGalNAc-T20 antibody has good sensitivity, can detect antigen protein GST-short T20 behind the purifying that is low to moderate 0.625ng; Fig. 6 illustrates that ppGalNAc-T20 antibody has good sensitivity, can detect the FLAG-ppGalNAc-T20 albumen in the transfectional cell cracking supernatant that is low to moderate 1.0 μ g;
(2) application of ppGalNAc-T20 polyclonal antibody in immunohistochemical methods
Fig. 7 is for detecting ppGalNAc-T20 protein expression distribution results figure in the C57 mouse testis tissue with ppGalNAc-T20 antibody by the immunohistochemical methods method, locate shown in the arrow to be use the ppGalNAc-T20 antibody test to the expression and distribution of ppGalNAc-T20 albumen in C57 mouse testis tissue.This figure illustrates that ppGalNAc-T20 antibody can be applied to immunohistochemical experiment and detect the proteic expression and distribution situation of ppGalNAc-T20 in the animal tissues.
(3) application of ppGalNAc-T20 polyclonal antibody in immunoprecipitation
Fig. 8 is the Western Blotting that detects MLTC-1 mouse testis Leydig's cell tumor cell pyrolysis liquid by immuno-precipitation with ppGalNAc-T20 antibody figure as a result, and wherein MLTC-1 mouse testis mesenchymal cell oncocyte 1 and 2 is respectively transfection pcDNA3.1-Full-length-T20 and contrast pcDNA3.1 carrier.This figure illustrates that ppGalNAc-T20 antibody can be applied to the ppGalNAc-T20 albumen in the immunoprecipitation experiment detection cell pyrolysis liquid.
Sequence table
＜110〉Shanghai Communications University
＜120〉ppGalNAc-T20 antigen and Polyclonal Antibody Preparation method thereof
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Claims (10)

1. a ppGalNAc-T20 antigen is characterized in that, this antigen is protein, and its aminoacid sequence is shown in SEQ ID NO:2.
2. the antigenic nucleic acid of the coding described ppGalNAc-T20 of claim 1 is characterized in that the base sequence of this nucleic acid is shown in SEQ ID NO:1.
3. the antigenic Polyclonal Antibody Preparation method of ppGalNAc-T20 as claimed in claim 1 is characterized in that, comprises the steps:
Step 1 is an antigen with the protein of aminoacid sequence shown in SEQ ID NO:2, adopts conventional preparation method of polyclonal antibody to prepare antiserum(antisera);
Step 2, the purifying antiserum(antisera), promptly.
4. the antigenic Polyclonal Antibody Preparation method of ppGalNAc-T20 as claimed in claim 1 is characterized in that, comprises the steps:
Step 1, the nucleic acid fragment of clone shown in SEQ ID NO:1;
Step 2 is utilized step 1 gained nucleic acid fragment construction recombination plasmid, and transformed into escherichia coli is cultivated, induce, and cracking, purifying gets protein;
Step 3 is an antigen with step 2 gained protein, adopts conventional preparation method of polyclonal antibody to prepare antiserum(antisera), the purifying antiserum(antisera), promptly.
5. the antigenic Polyclonal Antibody Preparation method of ppGalNAc-T20 according to claim 4, it is characterized in that, in the step 1, described clone, the primer is to being specially upstream primer shown in SEQ ID NO:3 and the downstream primer shown in the SEQID NO:4.
6. the antigenic Polyclonal Antibody Preparation method of ppGalNAc-T20 according to claim 4 is characterized in that, in the step 2, described intestinal bacteria are e. coli bl21 (DE3).
7. the antigenic Polyclonal Antibody Preparation method of described ppGalNAc-T20 according to claim 4 is characterized in that in the step 2, the plasmid that described recombinant plasmid uses is pGEX-5X-1.
8. the antigenic Polyclonal Antibody Preparation method of ppGalNAc-T20 according to claim 4, it is characterized in that, in the step 2, the process of screening described recombinant plasmid is: in the recombinant plasmid transformed bacillus coli DH 5 alpha that makes up, use contains the LB flat board of 100 μ g/ml penbritins, cultivates 12h down, shakes bacterium behind the picking list bacterium colony for 37 ℃, extract plasmid and order-checking, the correct required recombinant plasmid that is checks order.
9. the antigenic Polyclonal Antibody Preparation method of ppGalNAc-T20 according to claim 4 is characterized in that in the step 2, described cultivation is specially, the consisting of of 1L substratum: peptone 10g, and yeast extract 5g, NaCl 10g, surplus is a water; Culture temperature is 37 ℃; The OD value that is cultured to e. coli bl21 (DE3) is 0.8～1.2.
10. the antigenic Polyclonal Antibody Preparation method of ppGalNAc-T20 according to claim 4 is characterized in that, in the step 2, described induce into: add IPTG, making its final concentration is 0.1mM～0.5mM, and temperature is cultivated 3h～6h down for 26 ℃.
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CN102296052A (en) * 2011-08-17 2011-12-28 上海交通大学 Specific antigenic peptide of polyclonal antibody aiming at ppGalNAc-T13, and its preparation method and use
CN107988180A (en) * 2017-12-19 2018-05-04 湖北工业大学 Acyl group ACP synzyme and its polyclonal antibody
CN108179154A (en) * 2011-12-23 2018-06-19 Kws种子欧洲股份公司 Develop pathogen-response chimeric promoters novel plant source it is cis--regulating element

Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN102296052A (en) * 2011-08-17 2011-12-28 上海交通大学 Specific antigenic peptide of polyclonal antibody aiming at ppGalNAc-T13, and its preparation method and use
CN102296052B (en) * 2011-08-17 2013-04-10 上海交通大学 Specific antigenic peptide of polyclonal antibody aiming at ppGalNAc-T13, and its preparation method and use
CN108179154A (en) * 2011-12-23 2018-06-19 Kws种子欧洲股份公司 Develop pathogen-response chimeric promoters novel plant source it is cis--regulating element
CN107988180A (en) * 2017-12-19 2018-05-04 湖北工业大学 Acyl group ACP synzyme and its polyclonal antibody

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