This triple-knockout strain is a "maximally unstable" model designed for extreme membrane permeability and total secretion failure. It combines the loss of structural integrity (TolAB) with the loss of the primary export channel (TolC).
For Research Use Only. Not intended for use in food manufacturing or medical procedures (diagnostics or therapeutics). Do Not Use in Humans.
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Product Information
Product Overview
This triple-knockout strain is a "maximally unstable" model designed for extreme membrane permeability and total secretion failure. It combines the loss of structural integrity (TolAB) with the loss of the primary export channel (TolC).
Target
Escherichia coli Nissle 1917
Applications
This strain is often used in specialized laboratory settings to study extreme bacterial stress responses or to create a "leaky" host that releases almost all synthesized periplasmic content into the medium. The main advantage is the near-total lack of a protective barrier.
Product Identity & Gene Details
Chassis Strain
Escherichia coli Nissle 1917
Target Gene(s)
tolA, tolB and tolC
Gene Function
tolA / tolB: Essential components of the Tol-Pal complex that maintain the integrity of the cell envelope.
tolC: The multi-functional outer membrane pore used for drug efflux and protein secretion.
Modification Type
Triple gene knockout (Structural and transport collapse)
Genetic Architecture
Promoter System
Native promoter
Selection Marker
Standard Antibiotic Marker or Markerless/Scarless (No resistance genes)
Delivery & Service Standards
Deliverables
Glycerol Stocks
Shipping Conditions
Dry Ice Shipping
Lead Time
4-8 weeks
Documents
Certificate of Analysis (COA) or Project Report
Verification
PCR Sequencing
Engineered Strain Recovery & Handling
Immediate Post-Delivery Steps
1. Transfer to Storage: Immediately transfer the cryovials to a -80°C ultra-low temperature freezer. 2. Avoid Fluctuations: Do not store the vials in a "frost-free" freezer, as the temperature cycles can damage the cell membranes of the engineered strains.
Strain Recovery Protocol
To ensure maximum viability, follow this standard recovery procedure: 1. Preparation: Prepare the appropriate selective agar medium (e.g., LB agar + specific antibiotic as listed in the COA). 2. Partial Thawing (The "Scratch" Method): * Place the cryovial on dry ice. Do not thaw the entire vial. Use a sterile inoculation loop or pipette tip to "scratch" a small amount of ice from the surface of the frozen stock. 3. Streaking: Streak the cells onto the selective agar plate to obtain single colonies. 4. Incubation: Invert the plate and incubate at the specified temperature (e.g., 37°C for E. coli or 30°C for S. cerevisiae) for 16-24 hours. 5. Subculturing: Pick a single, well-isolated colony to inoculate into liquid media for your experiments.
Usage Precautions & Best Practices
Antibiotic Pressure
1. Maintenance: For strains containing plasmids or specific resistance markers, always use the correct concentration of antibiotics in both solid and liquid media to prevent plasmid loss or contamination.
2. Markerless Strains: If the strain is a "scarless" integrated knockout, antibiotic pressure is not required for maintenance, but periodic genotypic verification is recommended.
Induction Conditions
1. If your strain uses an inducible promoter (e.g., T7/IPTG or Arabinose), ensure the culture reaches the optimal OD600 (typically 0.6-0.8) before adding the inducer. 2. Verify the optimal induction temperature, as engineered strains often produce better results at lower temperatures (e.g., 16-25°C) to prevent inclusion body formation.
Disposal
All waste (vials, tips, plates) must be autoclaved at 121°C for at least 20 minutes or chemically disinfected before disposal.
