Protein expression
An antigen is a molecule recognized by the immune system. At the molecular level, an antigen is characterized by its ability to be "bound" at the antigen-binding site of an antibody. Immunogenicity is the ability to induce a humoral and/or cell-mediated immune response. Antigenicity is the ability to combine specifically with the final products of the immune response(i.e. secreted antibodies and/or surface receptors on T-cells). Although all molecules that have the property of immunogenicity also have the property of antigenicity, the reverse is not true. Antigens are usually proteins that are too large to bind as a whole to any receptor, only specific segments that form the antigen bind with a specific receptor. Such segments are called epitopes.A linear or a sequential epitope is an epitope that is recognized by antibodies by its linear sequence of amino acids, or primary structure. In contrast, antibodies recognize a conformational epitope that has a specific three-dimensional shape and its protein structure. Design and select a suitable antigen is a key to produce antibodies for various applications.
Chinapeptides has the antigen leading technology and advanced equipment, can provide customers with: (1) Protein expression and purification in prokaryotic system; (2) Peptide synthesis and conjugation.
QY-4001, gene synthesis, Oligo DNA sequence synthesis, 2-4 weeks
QY-4002, subclone（expression plasmid construct）, PCR,enzyme restrict, ligate,transform,sequencing, 2-3 weeks
QY-4003, Prokaryotic expression and purification, induction,optimation, expression and purification, 3-4 weeks
The original nucleoprotein express
Prokaryotic protein expression
Our company has Successfully builded a complete variety of protein expression technology platform: including the prokaryotic protein expression and purification, expression and purification of yeastprotein, expression and purification of protein which from insect cells, expression and purification of proteins from mammalian cells. According to customer needs, we can offer one-stop services which comprise gene synthesis, vector, gene expression, protein purification, detection and other . Our company has established a strict quality control system and a set of perfect customer service processes, confidentiality system to ensure that customers would get the quality products and services Our team has very rich experience in optimizing gene codon and expression vector design and construction, integration of objects and tags selection, specific protein purification system selection, protein production process optimization and so on. we canmeet the needs of individual customers.
Expression and purification of prokaryotic protein
prokaryotic protein expression is that interest proteinwhich come from the plants, animals, microorganisms is inserted into an appropriate expression vector ,and then introduced into Escherichia coli to obtain the large amounts of protein. This approach was used for protein purification, localization and function study . Escherichia coli is cultivated easyly,and the experiment has consumed less ; we can choose a variety of strains of Escherichia coliand the matched Plasmids which have a variety of characteristics. prokaryotic protein expression system isthe mature and reliable protein expression systems. our services charges according to the steps,you can choose any of the following steps to start the experiment or only choose one of the single step, please contact us.
For the early molecular biology techniques and services, please refer to the relevant content.
Service features:
1, one stop solution, from the gene synthesis to protein expression and purification, to complete your experimental design.
2,matureand stabletechnique, complete method .
3, efficient production time.
Service of Prokaryotic protein expression (coli E.) content:
1, gene synthesis,
To provide customers: synthetic genes and sequencing.
Time: 1~2
2, vector construction (the design and construction of vector, integration of objects and the selection of tags).
We can amplify and extract the expression plasmid. The expression plasmid was transformed into a highly efficient coli E. expression strain. At least one positive clone was selected to amplify and induce the expression of target protein in LB culture medium. We give the electrophoresis, the expression of target protein isolates, and monoclonal strains suitable for the expression of target protein.
To provide customers:strains for recombinant plasmid expression and expression detection report.
Time: 1~2 weeks
3, cultivation of recombinant bacteria by shake flask, induced expression of target protein.
By AffinityChromatography, Ion Chromatography , Gel chromatography and so on ,the expressed recombinant protein was purified .
We can remove the Tags by the application of Protease, and then target proteins are obtained .We will check the results of each test and control the quality of the final product through SDS-PAGE electrophoresis .
Through the Western-blot , we verify target protein is right or not .
We can provide customers: purified target protein , report ,tag (Tag) or the final protein which has removal of the tag (Tag) . the purity will be more than 95%.
Time: 2~3 weeks
4, large-scale protein preparation.
Optimization of protein production process
Customer: qualified target protein and service report.
Time: consultation
5, renaturation ofprotein
minimum of target protein was renaturated with different refolding buffer system for customer to detect. According to customer requirements, a small quantity of refolded protein could be preparated according to the renaturation condition of one kind of sample.We can provide the renaturation buffer and renaturation process specific formula. Other: sterilization, freeze-drying and further processing.
With the quality of detection of target proteins.
Customer: the final products after processing and related experiments and test report.
Time: 1~2 weeks
Yeast protein expression
Expression of yeast protein
Expression of yeast protein
Our company has Successfully builded a complete variety of protein expression technology platform: including the prokaryotic protein expression and purification, expression and purification of yeastprotein, expression and purification of protein which from insect cells, expression and purification of proteins from mammalian cells. According to customer needs, we can offer one-stop services which comprise gene synthesis, vector, gene expression, protein purification, detection and other . Our company has established a strict quality control system and a set of perfect customer service processes, confidentiality system to ensure that customers would get the quality products and services Our team has very rich experience in optimizing gene codon and expression vector design and construction, integration of objects and tags selection, specific protein purification system selection, protein production process optimization and so on. we canmeet the needs of individual customers.
Yeast protein expression
Recombinant protein expression and purification technology service (Pichia P. expression system)
The expression system includes: prokaryotic expression system (Escherichia coli expression system and yeast expression system (P. Pichia pastoris expression ) . Our company has established strict product quality control system ,a set of perfect customer service process, security system to ensure to provide our customers with quality products and service. Our services charges according to the steps,you can choose any of the following steps to start the experiment or only choose one of the single step, please contact us.
Yeast expression system -- step 1 subclone of target gene into yeast expression vector:
We can amplify and extract the expression plasmid containing the target gene was.
subclone of target gene into yeast expression vector
The accuracy of plasmid was verified by sequencing.
We can provide expression vector: pPICZ alpha and pPIC9K
To provide customers: the correct construction of the recombinant expression plasmid, the recombinant plasmid containing DH5 strain and sequencing report.
Service cycle: 1 weeks
Yeast expression system - step 2 Pichia P. expression strain electrical transformation:
Enzyme expression plasmid.
The expression plasmid was transformed into a highly efficient Pichia P. expression strain by electroporation.
We can remove the Tags by the application of Protease, and then target proteins are obtained . At least 5 positive clones were identified by PCR.
We can provide expression strains: X33, GS115, KM71 and SMD1168.
To provide customers: PCR positive clones and PCR results report.
Time: 1 weeks
Yeast expression system - step 3 Pichia P. expression strain screening:
The 5 positive clones were amplified in BMGY culture medium, and then were changed to BMMY culture medium, and the expression of target protein was induced by methanol.
The expression of target protein in the supernatant was detected by SDS-PAGE electrophoresis, and the suitable monoclonal strain was selected for the expression of the target protein. If no expression was detected in the culture supernatant, the expression of the target protein was detected by the concentration of the supernatant 20 times, or by the Western-blot method. To provide customers: any customer needs positive clones and expression screening results report.
Time: 2 weeks
Yeast expression system -- step 4 Pichia P. expression optimization:
20 positive clones were selected for routine expression screening, and the optimal expression conditions were optimized. Optimization of culture and induction conditions (culture medium, cell density, methanol concentration, induction time, etc.), and the expression of target protein was improved. To provide customers: any three customers need positive clones strains and optimize the expression of the results of the report.
Time: 2 weeks
Yeast expression system -- step 5 expression and purification of target protein:
1L recombinant yeast was cultured in shake flask , induce the expression of target protein. By AffinityChromatography, Ion Chromatography , Gel chromatography and so on ,the expressed recombinant protein was purified . We will check the results of each test and control the quality of the final product through SDS-PAGE electrophoresis . Through the Western-blot , we verify target protein is right or not . Provide customers: purified target protein 1~10mg and purification report. purified target protein , report ,tag (Tag) or the final protein which has removal of the tag (Tag) . the purity will be more than 90%.
Time: 2~3 weeks
• Yeast expression system -- step 6: remachiningand detection of target protein
The protein product with high biological activity is studied to renature.
Endotoxin removal, sterilization, freeze-drying and further processing.
The quality of target protein was detected by HPLC, mass spectrometry and other methods. Service content:
1 complex research:
Using up to 18 different types of complex buffer system, a minimum of target protein was renaturated with different refolding buffer system for customer to detect. According to customer requirements, a small quantity of refolded protein could be preparated according to the renaturation condition of one kind of sample. We can provide the renaturation buffer and renaturation process specific formula.
Other: sterilization, freeze-drying and further processing. With the quality of detection of target proteins. 2 endotoxin removal
3 filtration
4 freeze dried
5.HPLC detection
6 mass spectrometry
Provide customer: final products and related experiment and test report.
Time: 1~2 weeks
Yeast expression system - step 7 preparation
Provide customers: qualified target protein and service report.
Time: negotiation
Mammalian cell protein expression
Our company has Successfully builded a complete variety of protein expression technology platform: including the prokaryotic protein expression and purification, expression and purification of yeastprotein,expression and purification of protein which from insect cells, expression and purification of proteins from mammalian cells. According to customer needs, we can offer one-stop services which comprise gene synthesis, vector, gene expression, protein purification, detection and other . Our company has established a strict quality control system and a set of perfect customer service processes, confidentiality system to ensure that customers would get the quality products and services Our team has very rich experience in optimizing gene codon and expression vector design and construction, integration of objects and tags selection, specific protein purification system selection, protein production process optimization and so on. we canmeet the needs of individual customers. Mammalian cell protein expression
Mammalian cell protein expression is suitable for large-scale and fast preparation of protein, we provide the services contained construction of expression vector, transient transfectionand detection of protein expression and purification ofprotein and so on. It can be divided into transient expression, stable expression, stable expression of cell lines.
Transient expression of mammalian cell protein service TGE (HEK293 or CHO cells)
We can provide the amount of Mammalian cell protein from milligrams to grams.Fast transfection can be completed generally in 10 days, without the genetic selection ofDNA from plasmid. Our transient expression using suspended cells, can easily express ml to 100L protein. When construction of the expression vector was completed, we can deliver the product to the customer(the cell culture fluid collection or recombinant protein). If the customer needs, the target gene could be cloned into a highly efficient expression vector designed for the TGE system.
1, the cloning of the coding sequence of target protein to the mammalian transient expression vector;
2, the preparation of plasmid which can be used for transfection of mammalian cells;
3, assessment and optimization of mammalian cell transfection, recombinant protein expression ;
4, expression and purification of small scale protein (500 ml Culture), detection of protein ;
5,expression and purification of large-scale protein.
Stable expression technology service of mammalian cell protein
Throughout the experimental procedure, from transfection, screening and separation, to evaluating the production capacity and adaptability of primary clones, our company provide long-term stable and adjustable scale protein production. According to customer demand. We also have the ability of comprehensive genetic research to ensure the stability of protein production.
1, the cloning of the coding sequence of target protein to the mammalian transient expression vector;
2, the preparation of plasmid which can be used for transfection of mammalian cells;
3, assessment and optimization of mammalian cell transfection, recombinant protein expression ;
4. Monoclonal screening and expression of recombinant protein;
5, expression and purification of small scale protein (500 ml Culture), detection of protein ;
6,expression and purification of large-scale protein.
Construction of stable expression cell lines for scientific research
Our company provide synthetic gene, cloning, vector design construction, protein purification and characterization, and new drug screening a station type service.We can build and screening cell line which would meet your needs , explore faster better experimental solutions, help you save a lot of time in the late of the project .
1, customers can choose the cell by youself orbuy from our company;
2, we can provide the cloning of coding sequence of the target protein to the appropriate vector ;
3, the preparation of plasmid which can be used for transfection of mammalian cells;
4, transfection of mammalian cell , screening stably transfected cells;
5, assessment. of monoclonal screening (optional) andexpression of recombinant protein.
Mammalian cell protein expression
Mammalian cell protein expression
Our company has Successfully builded a complete variety of protein expression technology platform: including the prokaryotic protein expression and purification, expression and purification of yeastprotein,expression and purification of protein which from insect cells, expression and purification of proteins from mammalian cells. According to customer needs, we can offer one-stop services which comprise gene synthesis, vector, gene expression, protein purification, detection and other . Our company has established a strict quality control system and a set of perfect customer service processes, confidentiality system to ensure that customers would get the quality products and services Our team has very rich experience in optimizing gene codon and expression vector design and construction, integration of objects and tags selection, specific protein purification system selection, protein production process optimization and so on. we canmeet the needs of individual customers. Mammalian cell protein expression
Mammalian cell protein expression is suitable for large-scale and fast preparation of protein, we provide the services contained construction of expression vector, transient transfectionand detection of protein expression and purification ofprotein and so on. It can be divided into transient expression, stable expression, stable expression of cell lines.
Transient expression of mammalian cell protein service TGE (HEK293 or CHO cells)
We can provide the amount of Mammalian cell protein from milligrams to grams.Fast transfection can be completed generally in 10 days, without the genetic selection ofDNA from plasmid. Our transient expression using suspended cells, can easily express ml to 100L protein. When construction of the expression vector was completed, we can deliver the product to the customer(the cell culture fluid collection or recombinant protein). If the customer needs, the target gene could be cloned into a highly efficient expression vector designed for the TGE system.
1, the cloning of the coding sequence of target protein to the mammalian transient expression vector;
2, the preparation of plasmid which can be used for transfection of mammalian cells;
3, assessment and optimization of mammalian cell transfection, recombinant protein expression ;
4, expression and purification of small scale protein (500 ml Culture), detection of protein ;
5,expression and purification of large-scale protein.
Stable expression technology service of mammalian cell protein
Throughout the experimental procedure, from transfection, screening and separation, to evaluating the production capacity and adaptability of primary clones, our company provide long-term stable and adjustable scale protein production. According to customer demand. We also have the ability of comprehensive genetic research to ensure the stability of protein production.
1, the cloning of the coding sequence of target protein to the mammalian transient expression vector;
2, the preparation of plasmid which can be used for transfection of mammalian cells;
3, assessment and optimization of mammalian cell transfection, recombinant protein expression ;
4. Monoclonal screening and expression of recombinant protein;
5, expression and purification of small scale protein (500 ml Culture), detection of protein ;
6,expression and purification of large-scale protein.
Construction of stable expression cell lines for scientific research
Our company provide synthetic gene, cloning, vector design construction, protein purification and characterization, and new drug screening a station type service.We can build and screening cell line which would meet your needs , explore faster better experimental solutions, help you save a lot of time in the late of the project .
1, customers can choose the cell by youself orbuy from our company;
2, we can provide the cloning of coding sequence of the target protein to the appropriate vector ;
3, the preparation of plasmid which can be used for transfection of mammalian cells;
4, transfection of mammalian cell , screening stably transfected cells;
5, assessment. of monoclonal screening (optional) andexpression of recombinant protein.
Insect cell protein expression
Protein expression in insect cells
Chinapeptides has successfully built a complete variety of protein expression technology platform, including prokaryotic, yeasty, entomic and mammalian protein expression and purification According to customer needs, we offer gene synthesis, vector, gene expression, protein purification, detection and other one-stop service. In order to ensure the quality of protein purification, Chinapeptides establish a strict quality control system and a set of comprehensive customer service processe, to ensure the quality products and excellent service. Our team has a very rich experience in protein gene codon optimization, vector construction design and expression, integration and label, specific protein purification systems optimization, to meet the needs of individual customers.
Insect Cell Protein Expression and Purification
Insect cell expression system is widely-used eukaryotic expression system that can express a variety of exogenous genes including fungi, plants, bacteria, virus genes. Baculovirus is a type of double-stranded DNA that make insect cells as the natural host. Baculovirus is highly species-specific, and do not infect vertebrate, human, animal. More current research are about Autographa californica nuclear polyhedrosis (AcNPV), which host Spodoptera frugiperda. sf9 cells, one of the host cell for baculovirus expression system. sf9 cells, kind of semi-adherent cell line, can be suspension culture or adherent culture with many many advantages。
1、 compared with other eukaryotic expression systems, it has high exogenous gene expression level, especially intracellular proteins. In cases,recombinant protein is soluble, correctly folding have posttranslational modifications (such as glycosylation, etc.), biologically activity, easy separation and purification;
2、insect cells grown in suspension, which is easy for scaled-up cultures and large-scale expression of recombinant protein;
3、it is easy to express heterologous multimeric proteins. Insect cells can be infected through a variety of recombinant virus or can be implemented with a virus containing a plurality of expression cassettes;
4、security. Baculovirus infection do not include vertebrates.
Chinapeptides provides baculovirus expression vector, expression detecting and mass expression and purification of protein and other one-stop technical services.
1、coding sequence of target protein was cloned into the baculovirus transfer vector;
2、Preparation of a recombinant plasmid
3、transfected into insect cells, the virus modification;
4、viral titer test;
5、the recombinant protein expression verification and Optimization;
6、small-scale protein expression and purification (500 ml culture);
7、large-scale protein expression and purification (optional, suitable for biomedical research enterprise).
Technical services:
Baculovirus technical services
Protein expression optimization services
1、5-10ml recombinant baculovirus is provided by the customer;
2、insect cells are infected to obtain high titer virus;
3、optimization of expression conditions.
Optimization factors include:
Different insect cell lines;
Different expression time;
Different multiplicity of infection (MOI);
Virus titer test services;
Plaque assay titer test.
Protein purification services
Prokaryotic proteins purification services
prokaryotic expression is the method of inserting cloned gene into suitable vector and introducing into E. coli for expression of large amounts of protein.prokaryotic expression system is by far the mature and reliable protein expression system which rapidly express and purify a variety of protein sources. Prepared recombinant protein can be used for drug screening, structural biology, cell biology, proteomics and other fields of biomedical.Based on different expression requirements, such as volume of expression, viability of targeted protein and different purification method, we choose proper expression system and corresponding expression strategy. We make use of our mature and steady protein purification system to optimize expression terms and purify targeted protein.
Service Number: QY-4003
Service Items:
Select of plasmid transformation and high expression strain
Cultivate Transformed bacteria flask and Inducible expression
Purify of Targeted proteins
Detect expression and purification through SDS-PAGE
Final material delivery:
Initial culture conditions and reasonable purification process; purified protein (1mg, more than 85% purity)
SDS-PAGE protein electrophoresis results and detailed experimental records
Product Report (including relevant data, SDS-PAGE and plasmid sequencing results, etc.)
Sample requirement:
Provide cloned prokaryotic expression vector or plasmid or strain in cloning vector
Provide relevant information and Nucleic acid/protein sequence,use bioinformatics methods to analyze and make concrete and feasible program of the experiment
Attention:
1.Expression vector provided by client shall be transformed and expressed, proving with the expression of the target protein, plasmid ≥1ug
2.Expression protein should be equipped with His and GST label.
Protein markers service
Protein markers service Radioisotope-labelled Protein
Radioactive Isotope Labeling Method(RILM) is used to measure lipoprotein lipase activity(LPLA) in adipose and other tissues. Titrimetry and colorimetry, which are usually used to measure LPLA, are not sensitive or reliable enough. Therefore, RILM is first created in order to measure the LPLA accurately in China. What is more, it is also used to measure gene expression of LPLA in vitro and LPLA of mice’s heparin plasma, myocardial, adipose and liver tissue in 57℃. It uses glycerin emulsion, which contains 3H labeling olein, as substrate. After the Incubation Reaction among sample and substrate in 37℃, glyceride and free fatty acid are separated by extraction. The radioactive activity of free fatty acid which produced from steatolysis has positive relationship with activity of lipase in sample. 293A cell, which infected by recombinant adenovirus for the expression of lipoprotein lipase shows high level of LPLA in heparin nutrient solution.The data of LPLA in 5 mice’s heparin plasma, myocardial, adipose tissue shows 314. 3U/L, 349 and 61mU/g respectively, which indicates that RILM is sensitive, accurate and stable in lipoprotein lipase gene expression. RILM in myocardial tissue sample of mice provides a series of information for the further study in lipoprotein lipase.