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SHuffle® Express Competent E. coli

Catalog # Concentration Size List Price Quantity Your Price
C3028J 12 x 0.05 ml $383.00
$344.70
Catalog # Size List Price Your Price
C3028J 12 x 0.05 ml $383.00
$344.70
Catalog #
Qty:
 
*On-line ordering is for Canadian customers only. Web pricing is applicable only to orders placed online at www.neb.ca

Chemically competent E. coli cells suitable for non-T7 protein expression with enhanced capacity to correctly fold proteins with multiple disulfide bonds in the cytoplasm.

  • Non-T7 expression
  • Engineered E. coli B strain to promote disulfide bond formation in the cytoplasm
  • Deficient in proteases Lon and OmpT
  • Resistant to phage T1 (fhuA2)
  • Free of animal products
  • No dry ice surcharge on competent cell shipments
Featured Videos
View Video Library
  • Monarch_MiniprepProtocol_VideoThumb
    Stages of SHUFFLE® Optimization
  • Monarch_MiniprepProtocol_VideoThumb
    Fast transformation plating with glass beads
  • Monarch_MiniprepProtocol_VideoThumb
    What is a Disulfide Bond (1 of 4)

Chemically competent E. coli B cells engineered to form proteins containing disulfide bonds  in the cytoplasm.

Highlights
  • Engineered E. coli B strain to promote disulfide bond formation in the cytoplasm
  • Constitutively expresses a chromosomal copy of the disulfide bond isomerase DsbC
  • DsbC promotes the correction of mis-oxidized proteins into their correct form (1,3)
  • The cytoplasmic DsbC is also a chaperone that can assist in the folding of proteins that do not require disulfide bonds (4)
  • Enhanced BL21 derivative
  • Activity of nonspecific endonuclease I (endA1) eliminated for highest quality plasmid preparations
  • Deficient in proteases Lon and OmpT
  • Resistance to phage T1 (fhuA2), Nit, Spec, Str*
    *Resistance to low levels of streptomycin may be observed.
Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  SHuffle® Express Competent E. coli C3028JVIAL -80 12 x 0.05 ml
Features
  • Protease deficient/B strain
  • Enhanced capacity to correctly fold proteins with multiple disulfide bonds in the cytoplasm
Application Features

Figure 1, vtPA activity assayed from crude lysates: Figure 1, vtPA activity assayed from crude lysates:
Truncated tissue plasminogen activator (vtPA), which contains nine disulfide bonds when folded and oxidized correctly, was expressed from a pTrc99a plasmid in the cytoplasm of E. coli cells. After induction, cells were harvested and crude cell lysates were prepared. vtPA was assayed using a chromogenic substrate Chromozym t-PA (Roche #11093037001) and standardized to protein concentration using Bradford reagent. E. coli wt+ cells are DHB4, which is the parent of FÅ113 (Origami™).


Figure 2, PfCHT1 chitinase activity assayed from crude lysates: Figure 2, PfCHT1 chitinase activity assayed from crude lysates:
Plasmodium falciparum chitinase (PfCHT1) with three cysteines were expressed from a plasmid under the regulation of T7 promoter. After induction, cells were harvested and crude cell lysates were prepared. PfCHT1 was assayed using a chromogenic substrate (CalBioChem #474550) and standardized to protein concentration using Bradford reagent.

Properties & Usage

Antibiotic for Plasmid Selection

Antibiotics for Plasmid Selection Working Concentration
Ampicillin 100 µg/ml
Carbenicillin 100 µg/ml
Chloramphenicol 33 µg/ml
Kanamycin 30 µg/ml
Tetracycline 15 µg/ml

Shipping Notes

  • Ships on dry ice

Antibiotic Resistance

  • str (resistance to low levels of streptomycin may be observed)
  • nit
  • spec


Companion Products
Notes
  • STORAGE AND HANDLING: Competent cells should be stored at -80°C. Storage at -20°C will result in a significant decrease in transformation efficiency. Cells lose efficiency whenever they are warmed above -80°C, even if they do not thaw.
References
  • Bessette, P.H. et al. (1999). Proc. Natl. Acad. Sci. USA. 96, 13703-13708.
  • Qiu, J., Swartz, J.R. and Georgiou, G. (1998). Appl. Environ. Microbiol. 64, 4891-4896.
  • Levy, R. et al. (2001). Protein Expr. Purif. 20, 338-347.
  • Chen, J. et al. (1999). J. Biol. Chem. 274, 19601-19605.
  • Boyd, D. et al. (2000). J. Bacteriol. 182, 842-847.
  • de Marco, A. (2009). Microbial Cell Factories. 8, 26.
Quality Control Assay
Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.
Specifications
The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]
Certificate of Analysis
The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]
Safety Data Sheets
Legal And Disclaimer

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This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.

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