Producing recombinant proteins is not always simple. Optimization is typically time-consuming and labor-intensive because various factors can affect protein expression and purification. GenScript's protein expression specialists provide specialized bacterial expression services with state-of-the-art solutions to help with some unknowns and difficulties.
Users can now freely select a solution that meets their needs, including protein purification and characterization, gene synthesis, codon optimization, small-scale testing, and scale-up.
Researchers have created a unique protein refolding technique that can be produced on a large scale to recover biologically active proteins from inclusion bodies. A high-throughput E. coli expression platform provides an affordable choice for small-scale expression to speed up target screening projects.
E. coli is a well-known expression system that is simple to genetically modify and facilitates the low-cost and high-yield synthesis of various recombinant proteins, including enzymes, cytokines, immunogens, and antibody fragments.
The tailored workflow can meet the downstream applications. GenScript has successfully delivered more than 50,000 batches of recombinant proteins using the E. coli expression system.
Highlights
- Quick delivery in just two weeks
- Development of enzymes
- High yield of protein >15 g/L
- Scale up to kg levels with flexibility
General workflow
Image Credit: GenScript
Service details
Source: GenScript
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Milestones |
Specifications |
Deliverables |
| 1 |
Gene synthesis, Cloning, Plasmid Prep |
- Gene synthesis and Codon optimization
- Subcloning into an appropriate expression vector
- Plasmid Prep and DNA QC
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- Optimized gene sequence report
- Protein in buffered solution with your specified amount & purity
- QC data
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| 2 |
Protein Expression
Evaluation |
- Transform plasmids into appropriate bacterial expression strain
- Protein expression evaluation and optimization
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| 3 |
Protein Purification
Tag Removal (Optional)
Refolding (Optional) |
- Scale up protein expression using optimized conditions
- Purification to reach desired protein amounts and purity
- Tag removal and separation of tag-free protein (if requested)
- Refolding in case of insoluble protein (performed only if required)
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| 4 |
QC & Delivery |
- Purity analysis and Detection using SDS-PAGE and Western Blot (for tagged protein)
- Bradford assay for quantitation
- Protein delivery
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| |
Turnaround time* |
- 2 weeks for protein sequence < 1500 bp with one-step purification
- 3 weeks for protein sequence > 1500 bp with one-step purification
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*Turnaround times are approximate. In most cases, GenScript delivers within 2-3 weeks. However, optional steps might prolong the overall timeline slightly.
For any large-scale protein production requirements, GenScript offers fully customizable E. coli fermentation services.
Service feature - Soluble protein strategies
Solution 1: Various tag screening
- Purpose: Produce fusion proteins in soluble form with ease.
- Feature: Around 11 different fusion tag modifications from High Expression_pET-28 a (+) are available in stock functionalized vectors with an N-terminal tag.
- Advantages: Saves time without requiring a drawn-out dilution or dialysis procedure.
Image Credit: GenScript
Solution 2: Standard denaturation
- Purpose: Preserve the protein's soluble state.
- Feature: Protein may not be active but can be re-dissoluble in detergent buffer and used for dialysis.
- Advantages: Antigen (generally produces antibody to western blot)
- Workflow: Evaluate expression > Scale up expression > Wash IBs > Solubilize IBs > Purify > Dialysis and QC
Image Credit: GenScript
Solution 3: Professional inclusion body refolding services
- Purpose: Eagle was supposed to get active protein but did not work with the supernatant or other cells.
- Feature: A lot of work has been done on refolding condition screening, IB treatment, purification, and analysis techniques that can be more similar to how refolding occurs naturally with activity.
- Application: For refolding and large-scale expression.
- Technical work:
- Around 17 different dilution refolding buffer screening methods
- Examine the results of refolding by reduced and non-reduced LC-MS, SDS-PAGE, and SEC-HPLC