CN110423763A - The preparation method of pathogen protein HIV-GP41 is expressed in Escherichia coli by Optimized Coding Based gene - Google Patents
The preparation method of pathogen protein HIV-GP41 is expressed in Escherichia coli by Optimized Coding Based gene Download PDFInfo
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- CN110423763A CN110423763A CN201910806075.1A CN201910806075A CN110423763A CN 110423763 A CN110423763 A CN 110423763A CN 201910806075 A CN201910806075 A CN 201910806075A CN 110423763 A CN110423763 A CN 110423763A
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- 108090000623 proteins and genes Proteins 0.000 title claims abstract description 85
- 102000004169 proteins and genes Human genes 0.000 title claims abstract description 54
- 241000588724 Escherichia coli Species 0.000 title claims abstract description 26
- 244000052769 pathogen Species 0.000 title claims abstract description 25
- 230000001717 pathogenic effect Effects 0.000 title claims abstract description 24
- 238000002360 preparation method Methods 0.000 title claims abstract description 22
- 230000014509 gene expression Effects 0.000 claims abstract description 17
- 238000004440 column chromatography Methods 0.000 claims abstract description 10
- 239000002808 molecular sieve Substances 0.000 claims abstract description 10
- URGAHOPLAPQHLN-UHFFFAOYSA-N sodium aluminosilicate Chemical compound [Na+].[Al+3].[O-][Si]([O-])=O.[O-][Si]([O-])=O URGAHOPLAPQHLN-UHFFFAOYSA-N 0.000 claims abstract description 10
- 238000000034 method Methods 0.000 claims abstract description 9
- BPHPUYQFMNQIOC-NXRLNHOXSA-N isopropyl beta-D-thiogalactopyranoside Chemical compound CC(C)S[C@@H]1O[C@H](CO)[C@H](O)[C@H](O)[C@H]1O BPHPUYQFMNQIOC-NXRLNHOXSA-N 0.000 claims abstract description 8
- 108091028043 Nucleic acid sequence Proteins 0.000 claims abstract description 7
- 230000001939 inductive effect Effects 0.000 claims abstract description 7
- 230000001580 bacterial effect Effects 0.000 claims abstract description 5
- 230000015572 biosynthetic process Effects 0.000 claims abstract description 5
- 239000001963 growth medium Substances 0.000 claims abstract description 5
- 238000003786 synthesis reaction Methods 0.000 claims abstract description 5
- 230000006698 induction Effects 0.000 claims abstract description 4
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 18
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 claims description 14
- 239000004202 carbamide Substances 0.000 claims description 14
- 210000004027 cell Anatomy 0.000 claims description 12
- 210000003000 inclusion body Anatomy 0.000 claims description 12
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 claims description 9
- 239000011780 sodium chloride Substances 0.000 claims description 9
- 101150048348 GP41 gene Proteins 0.000 claims description 8
- 238000005119 centrifugation Methods 0.000 claims description 8
- 239000000243 solution Substances 0.000 claims description 8
- 239000007853 buffer solution Substances 0.000 claims description 7
- 239000007788 liquid Substances 0.000 claims description 7
- 241000894006 Bacteria Species 0.000 claims description 6
- 238000000502 dialysis Methods 0.000 claims description 6
- 238000000197 pyrolysis Methods 0.000 claims description 6
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 claims description 3
- 102000016943 Muramidase Human genes 0.000 claims description 3
- 108010014251 Muramidase Proteins 0.000 claims description 3
- 108010062010 N-Acetylmuramoyl-L-alanine Amidase Proteins 0.000 claims description 3
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims description 3
- 229930006000 Sucrose Natural products 0.000 claims description 3
- LZCZIHQBSCVGRD-UHFFFAOYSA-N benzenecarboximidamide;hydron;chloride Chemical compound [Cl-].NC(=[NH2+])C1=CC=CC=C1 LZCZIHQBSCVGRD-UHFFFAOYSA-N 0.000 claims description 3
- 239000012535 impurity Substances 0.000 claims description 3
- 229960000274 lysozyme Drugs 0.000 claims description 3
- 235000010335 lysozyme Nutrition 0.000 claims description 3
- 239000004325 lysozyme Substances 0.000 claims description 3
- 230000002101 lytic effect Effects 0.000 claims description 3
- 239000005720 sucrose Substances 0.000 claims description 3
- 238000005406 washing Methods 0.000 claims description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 3
- DGVVWUTYPXICAM-UHFFFAOYSA-N β‐Mercaptoethanol Chemical compound OCCS DGVVWUTYPXICAM-UHFFFAOYSA-N 0.000 claims description 3
- 241000725303 Human immunodeficiency virus Species 0.000 description 7
- 208000030507 AIDS Diseases 0.000 description 3
- 241000713772 Human immunodeficiency virus 1 Species 0.000 description 3
- 102000036639 antigens Human genes 0.000 description 3
- 108091007433 antigens Proteins 0.000 description 3
- 238000004519 manufacturing process Methods 0.000 description 3
- 239000000427 antigen Substances 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 238000003745 diagnosis Methods 0.000 description 2
- 208000015181 infectious disease Diseases 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 238000005457 optimization Methods 0.000 description 2
- 108020004705 Codon Proteins 0.000 description 1
- 241000588722 Escherichia Species 0.000 description 1
- 101800001690 Transmembrane protein gp41 Proteins 0.000 description 1
- 241000700605 Viruses Species 0.000 description 1
- 230000006978 adaptation Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000011109 contamination Methods 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000010586 diagram Methods 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 238000013399 early diagnosis Methods 0.000 description 1
- 108700004025 env Genes Proteins 0.000 description 1
- 101150030339 env gene Proteins 0.000 description 1
- 230000002068 genetic effect Effects 0.000 description 1
- 230000003053 immunization Effects 0.000 description 1
- 238000002649 immunization Methods 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 230000000644 propagated effect Effects 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 230000008521 reorganization Effects 0.000 description 1
- 230000019491 signal transduction Effects 0.000 description 1
- 208000011580 syndromic disease Diseases 0.000 description 1
- 230000002194 synthesizing effect Effects 0.000 description 1
- 210000002700 urine Anatomy 0.000 description 1
- 230000003612 virological effect Effects 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/005—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from viruses
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/70—Vectors or expression systems specially adapted for E. coli
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2740/00—Reverse transcribing RNA viruses
- C12N2740/00011—Details
- C12N2740/10011—Retroviridae
- C12N2740/16011—Human Immunodeficiency Virus, HIV
- C12N2740/16211—Human Immunodeficiency Virus, HIV concerning HIV gagpol
- C12N2740/16222—New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
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- Biomedical Technology (AREA)
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- Gastroenterology & Hepatology (AREA)
- Medicinal Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Peptides Or Proteins (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention discloses a kind of preparation methods for expressing pathogen protein HIV-GP41 in Escherichia coli by Optimized Coding Based gene, the step of expressing pathogen protein HIV-GP41 preparation method in Escherichia coli by Optimized Coding Based gene: it first by the DNA sequence dna of synthesis optimizing and is cloned into pET22B carrier, again by utilizing IPTG inducing expression target protein in BL21 bacterial strain, by the microbionation being incubated overnight into 2XYT culture medium, at 37 degree, OD=0.6 is arrived in culture under conditions of shaking table speed is 220rpm；0.5mM IPTG is added, at 20 degree, shaking table speed is inducing expression 4 hours under conditions of 220rpm；Thallus after collecting induction, target protein is therefrom purified into using the method for molecular sieve column chromatography, the preparation method proposed through the invention is allowed to consistent with the encoding gene that the preference of Escherichia coli uses, convenient for Expression product of such pathogen protein in Escherichia coli.
Description
Technical field
The invention belongs to Optimized Coding Based gene technology fields, and in particular to one kind is by Optimized Coding Based gene in Escherichia coli
In express pathogen protein HIV-GP41 preparation method.
Background technique
AIDS is caused by human immunodeficiency virus (human immunodeficiency virus, HIV) infection
Immune Deficiency Syndrome.Extremely important for the early diagnosis of AIDS, special sensitive diagnosis can allow patient
Treated as soon as possible, while also avoiding being propagated further for HIV.The main pathogens for causing AIDS are HIV-1, env gene institute
The transmembrane protein GP41 of coding is of great significance in the duplication, infection and signal transduction of HIV-1, and GP41 is exposed to HIV
Surface, the epitope cluster comprising a large amount of specificity, is the important antigen of virus surface, can induce body and generates the anti-of specificity
Body.Therefore GP41 albumen is widely used in the diagnostic reagent of vitro detection HIV-1 antibody.It is prepared using technique for gene engineering
Specificity and the preferable Recombinant HIV -1GP41 antigen protein of immunoreactivity mention for the development of HIV quick diagnosis reagent kit and production
Quality raw materials are supplied.
In major expression system, for escherichia expression system because its genetic background understands, destination gene expression is horizontal high,
Cultivation cycle is short, and contamination resistance is strong, and production cost is low and convenient for advantages such as industrialization, becomes production preparation and reorganization viral antigen
The first choice of albumen.General pathogen gene is compiled due to the greatest differences on evolving with the preferred gene used of Escherichia coli
Code difference is very big, is difficult in E. coli, wherein by using the original gene sequence gram of HIV-GP41 gene
It is grand into coli expression carrier, when expressing in BL21 bacterial strain, be difficult to detect the expression of target protein；On the other hand
By analyzing DNA sequence dna, the Codon Adaptation Index (CAI) of HIV-GP41 gene only has 0.6, high far below being suitble to
The CAI value (> 0.8) of expression is imitated, thus it is proposed that a kind of express pathogen egg by Optimized Coding Based gene in Escherichia coli
The preparation method of white HIV-GP41.
Summary of the invention
The purpose of the present invention is to provide one kind to express pathogen protein by Optimized Coding Based gene in Escherichia coli
The preparation method of HIV-GP41, to solve pathogen gene inconvenience mentioned above in the background art efficient table in Escherichia coli
Up to the problem of.
To achieve the above object, the invention provides the following technical scheme: a kind of Optimized Coding Based gene that passes through is in Escherichia coli
In express pathogen protein HIV-GP41 preparation method, pathogen protein is expressed in Escherichia coli by Optimized Coding Based gene
The preparation method step of HIV-GP41:
Step 1: the DNA sequence dna of synthesis optimizing and being cloned into pET22B carrier；
Step 2: utilizing IPTG inducing expression target protein in BL21 bacterial strain, the bacterium being incubated overnight is with the ratio of 1:100
Rate is inoculated into 2XYT culture medium, and OD=0.6 is arrived in culture under conditions of 37 degree of shaking table speed are 220rpm；0.5mM is added
IPTG, under conditions of 20 degree of shaking table speed are 220rpm, inducing expression 4 hours；
Step 3: collecting the thallus after induction, target protein is therefrom purified into using the method for molecular sieve column chromatography.
Preferably, target protein key step is therefrom purified into using the method for molecular sieve column chromatography are as follows:
S1. it cracks bacterium: adding cell pyrolysis liquid, and by after ultrasonic disruption cell, in 4 degree of continuation lytic cells
30min；
S2. it is collected by centrifugation: inclusion body protein is collected by centrifugation, it is each primary with solution and water washing inclusion body protein.
S3. it dissolves: dissolving inclusion body protein with 6M urea buffer solution；
S4. purify: the inclusion body protein that 6M urea is dissolved passes through molecular sieve column chromatography S-300 protein of interest；
S5. dialysis removal of impurities: collecting the component for containing > 95% target protein, removes area's urea by dialysis, finally obtains molten
Solution is in the target protein of 1XPBS.
Preferably, cell pyrolysis liquid includes 100mM Tris-HCl pH7.4,50mM NaCl, 1%TX-100,10mM
EDTA, 8%Sucrose, 5mM benzamidineHCl and 2mg/ml Lysozyme.
Preferably, the solution in being collected by centrifugation includes: 100mM Tris-HCl, pH7.4 and 50mM NaCl.
Preferably, 6M urea buffer solution includes 100mM Tris-HCl pH7.4,50mM NaCl, 6M urine in dissolving step
Element and 10mM beta -mercaptoethanol.
Compared with prior art, the beneficial effects of the present invention are:
The one kind proposed through the invention expresses pathogen protein HIV- by Optimized Coding Based gene in Escherichia coli
The preparation method of GP41 is allowed to consistent with the encoding gene that the preference of Escherichia coli uses, can effectively improve these pathogen
The expression of gene, convenient for producing the pathogen protein in expression in escherichia coli.
Detailed description of the invention
Attached drawing is used to provide further understanding of the present invention, and constitutes part of specification, with reality of the invention
It applies example to be used to explain the present invention together, not be construed as limiting the invention.
Fig. 1 is the contrast schematic diagram that gene code optimization of the invention influences HIV-GP41 protein expression；
Fig. 2 is the active measurement table of Recombinant HIV-GP41 protein immunization in the present invention；
Fig. 3 is the contrast table of HIV-GP41 albumen in the present invention；
Specific embodiment
Following will be combined with the drawings in the embodiments of the present invention, and technical solution in the embodiment of the present invention carries out clear, complete
Site preparation description, it is clear that described embodiments are only a part of the embodiments of the present invention, instead of all the embodiments.It is based on
Embodiment in the present invention, it is obtained by those of ordinary skill in the art without making creative efforts every other
Embodiment shall fall within the protection scope of the present invention.
Fig. 1-Fig. 3 is please referred to, the present invention provides a kind of technical solution: one kind is by Optimized Coding Based gene in Escherichia coli
The preparation method for expressing pathogen protein HIV-GP41, expresses pathogen protein by Optimized Coding Based gene in Escherichia coli
In the step of HIV-GP41 preparation method, first by the DNA sequence dna of synthesis optimizing and it is cloned into pET22B carrier, wherein synthesizing excellent
The DNA sequence dna of change are as follows:
ATGGCAGTTGGAATTGGTGCTATGTTCCTGGGCTTCCTGGGTACCGCAGGTAGCACTATGGGCGCGGC
GTCTATTACGCTGACGGTACAGGCCCGTCAATTATTGTCTGGCATTGTGCAACAGCAAAGCAATCTGCTGCGTGCT
ATTGAGGCTCAACAGCATCTGTTGAAGCTCACTGTCTGGGGTATTAAACAACTGCAGGCACGCGTCCTGGCTGTGG
AAAAGTACCTGAAGGATCAGCAGCTGCTGGGTATTTGGGGCTGCTCTGGAAAACTCATCTGCACCACTACTGTGCC
TTGGAACTCTAGCTGGAGTAATAAATCTTGGAGCGAGATTTGGGACAACATGACCTGGCTGCAATGGGATAAAGAA
ATTTCCAATTACACTGAGACTATTTATGATCTGCTTGAAATTGCACAAATCCAGCAGGAAAAGAATGAGCAAGACC
TGTTGGCACTGGACAAGTGGGCAAGC；
The selection application of Optimized Coding Based and its number are as shown in the table in the HIV-GP41 optimization wherein synthesized
Again by utilizing IPTG inducing expression target protein in BL21 bacterial strain, by the microbionation being incubated overnight to 2XYT
In culture medium, wherein the bacterium being incubated overnight and 2XYT culture medium ratio are 1:100,37 degree at a temperature of, shaking table speed is
OD=0.6 is arrived in 220rpm, culture；0.5mM IPTG is added, at 20 degree, under conditions of shaking table speed is 220rpm, induces table
Up to 4 hours；Thallus after collecting induction, is therefrom purified into target protein using the method for molecular sieve column chromatography.
In the present embodiment, it is preferred that be therefrom purified into target protein key step using the method for molecular sieve column chromatography are as follows:
S1. it cracks bacterium: adding cell pyrolysis liquid, and by after ultrasonic disruption cell, in 4 degree of continuation lytic cells
30min, wherein cell pyrolysis liquid includes 100mM Tris-HCl, pH7.4,50mM NaCl, 1%TX-100,10mM EDTA,
8%Sucrose, 5mM benzamidineHCl and 2mg/ml Lysozyme；
S2. it is collected by centrifugation: inclusion body protein is collected by centrifugation, it is each primary with solution and water washing inclusion body protein, wherein molten
Liquid includes: 100mM Tris-HCl, pH7.4 and 50mM NaCl；
S3. it dissolves: dissolving inclusion body protein with 6M urea buffer solution, wherein 6M urea buffer solution includes 100mM Tris-
HCl, pH7.4,50mM NaCl, 6M urea and 10mM beta -mercaptoethanol；
S4. purify: the inclusion body protein that 6M urea buffer solution is dissolved passes through molecular sieve column chromatography S-300 purification of target egg
It is white；
S5. dialysis removal of impurities: collecting the component for containing > 95% target protein, removes urea by dialysis, finally obtains dissolution
In the target protein of 1XPBS.
It although an embodiment of the present invention has been shown and described, for the ordinary skill in the art, can be with
A variety of variations, modification, replacement can be carried out to these embodiments without departing from the principles and spirit of the present invention by understanding
And modification, the scope of the present invention is defined by the appended.
Claims (6)
1. a kind of preparation method for expressing pathogen protein HIV-GP41 in Escherichia coli by Optimized Coding Based gene, feature
Be: described the step of expressing pathogen protein HIV-GP41 preparation method in Escherichia coli by Optimized Coding Based gene, wraps
It includes:
Step 1: the DNA sequence dna of synthesis optimizing and being cloned into pET22B carrier；
Step 2: utilizing IPTG inducing expression target protein in BL21 bacterial strain, the bacterium being incubated overnight is connect with the ratio of 1:100
Kind is into 2XYT culture medium, and at 37 degree, OD=0.6 is arrived in culture under conditions of shaking table speed is 220rpm；0.5mM IPTG is added
Afterwards, at 20 degree, under conditions of shaking table speed is 220rpm, inducing expression 4 hours；
Step 3: collecting the thallus after induction, target protein is therefrom purified into using the method for molecular sieve column chromatography.
2. a kind of Optimized Coding Based gene that passes through according to claim 1 expresses pathogen protein HIV- in Escherichia coli
The preparation method of GP41, it is characterised in that: the DNA sequence dna of the synthesis optimizing are as follows:
ATGGCAGTTGGAATTGGTGCTATGTTCCTGGGCTTCCTGGGTACCGCAGGTAGCACTATGGGCGCGGCGTCT
ATTACGCTGACGGTACAGGCCCGTCAATTATTGTCTGGCATTGTGCAACAGCAAAGCAATCTGCTGCGTGCTATTG
AGGCTCAACAGCATCTGTTGAAGCTCACTGTCTGGGGTATTAAACAACTGCAGGCACGCGTCCTGGCTGTGGAAAA
GTACCTGAAGGATCAGCAGCTGCTGGGTATTTGGGGCTGCTCTGGAAAACTCATCTGCACCACTACTGTGCCTTGG
AACTCTAGCTGGAGTAATAAATCTTGGAGCGAGATTTGGGACAACATGACCTGGCTGCAATGGGATAAAGAAATTT
CCAATTACACTGAGACTATTTATGATCTGCTTGAAATTGCACAAATCCAGCAGGAAAAGAATGAGCAAGACCTGTT
GGCACTGGACAAGTGGGCAAGC。
3. a kind of Optimized Coding Based gene that passes through according to claim 1 expresses pathogen protein HIV- in Escherichia coli
The preparation method of GP41, it is characterised in that: the method using molecular sieve column chromatography is therefrom purified into target protein and mainly walks
Suddenly are as follows:
S1. it cracks bacterium: adding cell pyrolysis liquid, and by after ultrasonic disruption cell, in 4 degree of continuation lytic cell 30min；
S2. it is collected by centrifugation: inclusion body protein is collected by centrifugation, it is each primary with solution and water washing inclusion body protein；
S3. it dissolves: dissolving inclusion body protein with 6M urea buffer solution；
S4. purify: the inclusion body protein that 6M urea is dissolved passes through molecular sieve column chromatography S-300 protein of interest；
S5. dialysis removal of impurities: collecting the component for containing > 95% target protein, removes urea by dialysis, finally obtains and be dissolved in
The target protein of 1XPBS.
4. a kind of Optimized Coding Based gene that passes through according to claim 3 expresses pathogen protein HIV- in Escherichia coli
The preparation method of GP41, it is characterised in that: the cell pyrolysis liquid includes 100mM Tris-HCl, pH7.4,50mM NaCl,
1% TX-100,10mM EDTA, 8% Sucrose, 5mM benzamidineHCl and 2mg/ml Lysozyme.
5. a kind of Optimized Coding Based gene that passes through according to claim 3 expresses pathogen protein HIV- in Escherichia coli
The preparation method of GP41, it is characterised in that: it is described be collected by centrifugation in solution include: 100mM Tris-HCl, pH7.4 and 50mM
NaCl。
6. a kind of Optimized Coding Based gene that passes through according to claim 3 expresses pathogen protein HIV- in Escherichia coli
The preparation method of GP41, it is characterised in that: 6M urea buffer solution includes 100mM Tris-HCl in the dissolving step, pH7.4,
50mM NaCl, 6M urea and 10mM beta -mercaptoethanol.
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Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20060286631A1 (en) * | 2003-09-01 | 2006-12-21 | Kang Choong K | Method for preparating t-20 peptide by high cell density cultivation of recombinant e. coli containing t-20 peptide coding gene |
| CN102370979A (en) * | 2011-10-10 | 2012-03-14 | 中国人民解放军第四军医大学 | Building method for autovaccine by aiming at human TNF(Tumor Necrosis Factor)-alpha molecule |
| CN105039379A (en) * | 2015-08-31 | 2015-11-11 | 浙江大学 | Method for producing HIV-1 gp41 recombinant antigen by means of Escherichia coli cell-free system |
-
2019
- 2019-08-29 CN CN201910806075.1A patent/CN110423763A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20060286631A1 (en) * | 2003-09-01 | 2006-12-21 | Kang Choong K | Method for preparating t-20 peptide by high cell density cultivation of recombinant e. coli containing t-20 peptide coding gene |
| CN102370979A (en) * | 2011-10-10 | 2012-03-14 | 中国人民解放军第四军医大学 | Building method for autovaccine by aiming at human TNF(Tumor Necrosis Factor)-alpha molecule |
| CN105039379A (en) * | 2015-08-31 | 2015-11-11 | 浙江大学 | Method for producing HIV-1 gp41 recombinant antigen by means of Escherichia coli cell-free system |
Non-Patent Citations (1)
| Title |
|---|
| 李文利等: "HIV-1 GP41第二优势表位簇在大肠杆菌中的高效表达", 《青岛大学医学院学报》 * |
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Application publication date: 20191108 |