High-Yield E. coli Cell-Free Protein Synthesis (CFPS) System Service

The Bacterial Cell-Free Protein Synthesis (CFPS) System , predominantly utilizing optimized E. coli lysates, is a powerful platform that recreates the essential components of the translation machinery in vitro . By decoupling protein synthesis from the cellular environment, this system offers unparalleled speed, control, and flexibility for expressing challenging proteins, toxic gene products, and non-canonical amino acid incorporated variants.

CD Biosynsis provides a premium, optimized E. coli CFPS System Service tailored for R&D and industrial applications. Our system is designed for high yields and features proprietary lysate preparation and energy regeneration components, ensuring efficient expression of milligram quantities of protein in just a few hours. The open nature of our bacterial CFPS system makes it the preferred choice for applications requiring high-throughput screening (HTS), rapid prototyping, and sophisticated labeling (e.g., isotope or non-natural amino acid incorporation) , offering a fast, robust, and cost-effective alternative to traditional cell-based expression.

Highlights

Key advantages of using our E. coli CFPS system for protein production:

• High Yield Performance: Optimized lysate preparation and energy mix deliver superior protein yields, often reaching hundreds of micrograms per milliliter of reaction.
• Fast Turnaround: Protein synthesis is typically completed within 4–8 hours , drastically accelerating the time from gene to functional protein.
• Unrestricted Protein Expression: Capable of producing proteins that are toxic, insoluble, or aggregation-prone when expressed in vivo .
• Open & Flexible Platform: Allows for direct modification of the reaction environment, including the addition of inhibitors, detergents, cofactors, and labeling precursors.

Applications

The bacterial CFPS system is a versatile tool for diverse biochemical and engineering applications:

High-Throughput Screening (HTS)

           

Enables the rapid, parallel synthesis and screening of protein libraries or mutants in 96-well plate format for enzyme activity or ligand binding.

Non-natural Amino Acid (nnAA) Incorporation

The preferred system for site-specifically incorporating nnAAs, allowing for protein functionalization via Click Chemistry or the introduction of novel probes.

Isotope Labeling for NMR/Structure

Cost-effective and high-fidelity incorporation of stable isotopes (^15N, ^13C, ^2H) required for structural analysis of challenging proteins.

Synthesis of Toxic/Membrane Proteins

Ideal for expressing gene products that poison living cells or require controlled environments (e.g., detergents, liposomes) for proper folding.

System Features & Options

Key components and customizable features of our E. coli CFPS Service:

Optimized Lysate Preparation

Ribosome-rich E. coli S30 extract prepared under proprietary conditions to minimize endogenous nuclease activity and maximize protein yield.

High-Efficiency Energy Mix

Regeneration system (e.g., phosphoenolpyruvate) formulated for long-lasting protein synthesis and sustained high-yield performance in batch or fed-batch mode.

Linear vs. Plasmid DNA Templates

Supports both linear PCR products (for speed and HTS) and circular plasmid DNA (for maximum yield and long-term stability).

Ribosome Recycling Capability

Engineered systems that promote efficient ribosome release and re-initiation, increasing the total number of proteins synthesized per ribosome unit.

Tag and Purification Options

Immediate purification upon synthesis using integrated tags (e.g., His-tag, Strep-tag) and optimized protocols for rapid downstream processing.

Workflow

Our streamlined process for efficient bacterial CFPS protein production:

• Gene Optimization & Template Preparation: The customer provides the gene sequence, which is optimized for CFPS expression and converted into a high-quality linear or plasmid DNA template.
• CFPS Reaction Setup: The DNA template, the lyophilized energy mix, and the specialized E. coli lysate are combined in the reaction vessel.
• In Vitro Synthesis: The mixture is incubated at 30℃-37℃ for the designated time (e.g., 6 hours) to maximize protein expression.
• Purification: The crude lysate containing the target protein is harvested and purified using affinity chromatography (e.g., Ni-NTA for His-tag).
• Quality Control (QC): Protein yield is determined by SDS-PAGE and protein quantification assays. Functional activity assays and MS verification are available upon request.
• Delivery: Purified protein is delivered in the requested buffer, often lyophilized or flash-frozen, along with a detailed COA.

We provide essential assurance for high-quality CFPS outcomes:

• Guaranteed High Yield: We provide a guaranteed minimum yield (typically based on the protein size and complexity) for standard targets.
• Nuclease-Free Environment: Strict control measures are in place to minimize endogenous nuclease activity, protecting the DNA and mRNA templates and maximizing protein synthesis duration.
• Custom Additive Testing: Capability to test the impact of various additives (e.g., chaperones, specific ions, non-detergents) to optimize the folding and solubility of challenging proteins.
• Scalability: Expertise in scaling up the reaction from muL-scale analytical scale to multi-milliliter preparatory scale required for structural and therapeutic studies.