NEB® 10-beta Electrocompetent E. coli
Supplied with outgrowth medium optimized for NEB 10-beta & NEB Stable Competent E.coli ; please do not use SOC outgrowth medium.
NEB 10-beta Competent E. coli is a derivative of the popular DH10B. It is T1 phage resistant and endonuclease I (endA1) deficient for high- quality plasmid preparations.
- High efficiency strain ideal for cloning large plasmids and BACs
- Available in chemically-competent format (single-use and 200 μl vials)
- No dry ice surcharge on competent cell shipments
- Outgrowth medium included
- Free of animal products
- Refer to our Electroporation Tips
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Product Information
NEB 10-beta electrocompetent E. coli cells are optimized for high efficiency transformation by electroporation. These cells are ideal for DNA library constructions and all cloning purposes.
Highlights
- DH10B™ derivative
- Accomodation of large plasmids including BAC and cosmid constructs
- Efficient transformation of methylated DNA derived from eukaryotic sources and unmethylated DNA derived from PCR, cDNA and other sources
- Activity of nonspecific endonuclease I eliminated for highest quality plasmid preparations (endA1)
- Suitable for blue/white screening by α-complementation of the β-galactosidase gene (Φ80ΔlacZM15)
- Reduced recombination of cloned DNA (recA)
- Resistance to phage T1 (fhuA)
- K-12 Strain
- Free of animal products
Transformation Efficiency
> 2 x 1010 cfu/μg pUC19Genotype
Δ(ara-leu) 7697 araD139 fhuA ΔlacX74 galK16 galE15 e14- Φ80dlacZΔM15 recA1 relA1 endA1 nupG rpsL (StrR) rph spoT1 Δ(mrr-hsdRMS-mcrBC)
- This product is related to the following categories:
- Cloning Competent Cell Strains
- This product can be used in the following applications:
- Transformation
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Features
Electroporation Tips:
- Electroporation cuvettes and microcentrifuge tubes should be pre-chilled on ice.
- Electrocompetent cells should be thawed on ice and suspended well by carefully flicking the tubes.
- Once DNA is added to the cells, electroporation can be carried out immediately. It is not necessary to incubate DNA with cells. The maximum recommended volume of a DNA solution to be added is 2.5 µl. Addition of a large volume of DNA decreases transformation efficiency.
- Contaminants such as salts and proteins can lower electroporation efficiency. Ideally, DNA for transformation should be purified and suspended in water or TE. Transformation efficiency is more than10-fold lower for ligation mixtures than the control pUC19 plasmid due to the presence of ligase and salts. If used directly, ligation reactions should be heat-inactivated at 65°C for 20 min and then diluted 10-fold. For optimal results, spin columns (NEB #T1030) are recommended for cleanup of ligation reactions.
- Electroporation conditions vary with different cuvettes and electroporators. If you are using electroporators or cuvettes not specified in the protocol, you may need to optimize the electroporation conditions. Cuvettes with 1mm gap are recommended (e.g. BTX Model 610/613 and Bio-Rad #165-2089). Higher voltage is required for cuvettes with 2 mm gap.
- Arcing may occur due to high concentration of salts or air bubbles.
- It is essential to add recovery medium to the cells immediately after electroporation. One minute delay can cause a 3-fold reduction in efficiency.
- Cold and dry selection plates lead to lower transformation efficiency. Pre-warm plates at 37°C for 1 hour. Using 37°C pre-warmed recovery medium increases the efficiency by about 20%.
- Refreeze unused cells in a dry ice/ethanol bath for 5 min and then store at -80°C. Do not use liquid nitrogen. Additional freeze-thaw cycles result in lower transformation efficiency.
Application Features
DNA Effects on Transformation Efficiency and Colony Output: Electroporation efficiency remains extremely high up to about 10 ng of input DNA, then decreases at higher DNA concentrations. Total colony output continues to increase with increasing DNA input up to at least 1 μg of puc19.
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