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  • NEB Turbo Competent E. coli (High Efficiency)

    Description

    Chemically competent E. coli cells suitable for high efficiency transformation and rapid colony growth.

    Highlights

    • Transformation efficiency 1-3 x 109 cfu/μg pUC19 DNA
    • Tight control of expression by laclq allows potentially toxic genes to be cloned
    • Highest growth rate on agar plates - visible colonies 6.5 hours after transformation
    • Activity nonspecific endonuclease I (endA1) eliminated for highest quality plasmid preparations
    • Resistance to phage T1 (fhuA2)
    • Suitable for blue/white screening by α-complementation of the β-galactosidase gene
    • Suitable for 5 minute transformation protocol with AmpR plasmids
    • EcoKr-m-, McrBC-
    • K12 Strain
    • Isolate DNA after 4 hours of growth from a single overnight colony
    • Free of animal products

    Genotype

    F' proA+B+ lacIq ∆lacZM15 / fhuA2  ∆(lac-proAB)  glnV galK16 galE15  R(zgb-210::Tn10)TetS  endA1 thi-1 ∆(hsdS-mcrB)5  

    Reagents Supplied

    The following reagents are supplied with this product:

    Store at (°C)Concentration
    pUC19 Transformation Control Plasmid-200.05 ng/μl
    SOC Outgrowth Medium41X

    Advantages and Features

    Features

    • Fastest growth – colonies visible after 6.5 hours
    • Plasmid preparation after 4 hours
    • Tight control/expression of toxic genes

    Applications


    DNA Effects on Transformation Efficiency and Colony Output: The optimal amount of DNA to use in a transformation reaction is lower than commonly recognized. Using clean, supercoiled pUC19, the efficiency of transformation is highest in the 100 pg-1 ng range. However, the total colonies which can be obtained from a single transformation reaction increase up to about 100 ng.
    DNA Effects on Transformation Efficiency and Colony Output: The optimal amount of DNA to use in a transformation reaction is lower than commonly recognized. Using clean, supercoiled pUC19, the efficiency of transformation is highest in the 100 pg-1 ng range. However, the total colonies which can be obtained from a single transformation reaction increase up to about 100 ng.

    Growth curve of NEB Turbo vs DH5α at 37°C in shaking flask: NEB Turbo and DH5α frozen stock (same cell density) were inoculated 1:100 into 80 ml of liquid medium (10g tryptone, 5g yeast extract, 10g NaCl, 1g MgCl2 and 1g Dextrose per liter, pH7.2) in 1 liter flask, respectively. Cells were grown at 37°C with shaking at 250 rpm. O.D.600 was measured every 30 minutes for up to 16 hours.
    Growth curve of NEB Turbo vs DH5α at 37°C in fermentor: NEB Turbo and DH5α cultures grown overnight were inoculated 1:50 into 1 liter of LB (10g tryptone, 5g yeast extract, 10g NaCl per liter, pH7.0) in 1 liter fermentor, respectively. Cells were grown at 37°C. O.D.600 was measured every 30 minutes for up to 9 hours.
    NEB Turbo cell growth and DNA yield: pUC19 was transformed into NEB Turbo competent E.coli. Six colonies grown overnight (colony size was about 2.4 mm in diameter) were inoculated into 30 ml of LB/Amp, mixed thoroughly and 5 ml was dispensed into each 25 ml test tube (equal to 1 colony in 5 ml of LB/Amp). The cells were grown at 37°C with shaking at 250 rpm. One test tube was placed on ice after 3, 4, 5, 6, 7 and 16 hours of growth, respectively and the O.D.600 was measured. Mini-prep was performed by spinning down 1.5 ml of culture 3 times (total 4.5 ml) and finally eluting the DNA into 50 μl of EB buffer.
    Comparison of DNA yield after 4 hours of growth: pUC19 was transformed into NEB Turbo competent E.coli, Invitrogen Mach1 and DH5α, respectively. One overnight grown colony (colony size for NEB Turbo was about 2.4 mm in diameter, 1.8 mm for Mach1 and 1.2 mm for DH5α) was inoculated into 5 ml of LB/Amp in a 25ml test tube and cells were grown at 37°C with shaking at 250 rpm. After 4 hours of growth, cells were measured O.D.600 and mini-prep was performed by spinning down 1.5 ml of culture 3 times (total 4.5 ml) and finally eluting the DNA into 50 μl of EB buffer. O.D.600 for NEB Turbo was 3.08, 2.36 for Mach1 and 0.75 for DH5α.

    Effect of heat shock time on NEB Turbo competent E.coli transformation efficiency: 50 μl of competent cells were transformed with 100 pg of pUC19 control DNA following the provided High Efficiency Transformation Protocol except heat shock time varied from 0 to 80 seconds.
    Effect of DNA incubation time on NEB Turbo competent E.coli transformation efficiency: 50 μl of competent cells were transformed with 100 pg of pUC19 control DNA following the provided High Efficiency Transformation Protocol except DNA incubation time varied from 0 to 40 minutes.
    Effect of outgrowth medium on transformation efficiency: 50 μl of NEB Turbo competent E.coli was transformed with 100 pg of pUC19 control DNA following the provided High Efficiency Transformation Protocol with the exception of varying the outgrowth medium. NEB SOC outgrowth medium delivers the highest transformation efficiency.
    Take advantage of the low cost per transformation with NEB Turbo. Calculations were based on list price and recommended transformation volumes.

    Properties and Usage

    Antibiotics for Plasmid SelectionWorking Concentration
    Ampicillin100 μg/ml
    Carbenicillin100 μg/ml
    Chloramphenicol33 μg/ml
    Kanamycin30 μg/ml
    Streptomycin25 μg/ml
    Tetracycline15 μg/ml

    Storage Temperature

    -80°C

    Shipping Notes

    • Ships on dry ice

    Antibiotic Resistance

    • nit

    Notes

    1. CAUTION: This product contains DMSO, a hazardous material. Review the MSDS before handling.
    2. 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.

    Supporting Documents

    Material Safety Datasheets

    The following is a list of Material Safety Data Sheets (MSDS) that apply to this product to help you use it safely. The following file naming structure is used to name these document files: [Product Name] MSDS. For international versions please contact us at info@neb.com.

    Datacards

    The Product Summary Sheet, or Data Card, includes details for how to use the product, as well as details of its formulation and quality controls. The following file naming structure is used to name the majority of these document files: [Catalog Number]Datasheet-Lot[Lot Number]. For those product lots not listed below, please contact NEB at info@neb.com or fill out the Technical Support Form for appropriate document.
    1. Does plasmid size affect colony size (C2984)?
    2. How long can I purify plasmid after the inoculation from a single colony using NEB Turbo competent E.coli (C2984)?
    3. How long should I incubate cells on ice after DNA has been added (NEB #C2984H and NEB #C2984I)?
    4. What are the strain properties (C2984)?
    5. What does the growth curve of NEB Turbo cells look like compared to DH5α?
    6. What is the difference between NEB #C2984H and NEB #C2984I?
    7. What is the optimal heat shock time for this strain (NEB #C2984H and NEB #C2984I)?
    8. Which strain of Competent E. coli should I use for general cloning?
    9. How should I calculate the transformation efficiency (C2984)?
    10. What are the solutions/recipes (C2984)?
    11. Can I store competent cells at -20°C instead of -80°C?
    12. Which kind of transformation tubes should be used?
    13. What volume of DNA can be added into competent cells?
    14. What is the shelf life for this strain (NEB #C2984H and NEB #C2984I)?
    15. Are NEB's competent cells compatible with the "Plate and Go" protocol?
    1. High Efficiency Transformation Protocol (C2984)
    2. 5 Minute Transformation Protocol (C2984)

    Selection Tools

    Usage Guidelines & Tips

    Application Notes