G7419
GC5™ Competent Cells
for generation of cDNA libraries and DNA plasmid production (96 well plate)
About This Item
grade
Molecular Biology
for molecular biology
form
suspension
availability
available only in USA
shipped in
dry ice
storage temp.
−70°C
General description
Application
Features and Benefits
- Ensures recovery of stable and high quality plasmid DNA
- Renders protection to clonal stocks from T1 and T5 bacteriophage contamination
- Allows for β-galactosidase α-complementation for blue/white screening
- Guaranteed high transformation efficiency
- Convenient pack in 96-well format (20 μL/well); each well may be used for a single transformation
Components
- GC5 chemically competent cells in one 96-well plate, 20 μL/well (G7544)
- pUC 19 control DNA (10 ng/μL), 10 μL (D2567)
Principle
Legal Information
related product
Storage Class
10 - Combustible liquids
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Are home-made competent cells as efficient at transformation as purchased cells?
They can be, depending on the technique used, the expected transformation efficiency and how the cells are handled. For special applications, competent cells prepared in-house using standard methods may not provide the efficiency you need.
Are all competent cells suitable for both plasmid production and protein expression?
No. Most strains of competent cells are suitable for producing plasmid DNA that will be used to transfect insect or mammalian cells for expression of the protein. The BL21 competent cell strains have special traits enabling protein expression in bacteria.
What are important considerations when performing bacterial transformation?
Things to consider when planning bacterial transformation:DNA impuritiesSource of DNAAmount of DNA usedStorage and handling of the competent cells
Can I store the competent cells in the -20C?
No. Competent cells should not be stored at -20C for any length of time. The cells suffer a dramatic drop in transformation efficiency when stored higher than -80C.
How should I thaw the competent cells?
Competent cells should be thawed on ice. The transformation efficiency is dependent on the proper handling of the cells (maintaining cold temperature until transformation).
Can I re-freeze the vial if I do not use the entire aliquot of competent cells?
Re-freezing competent cells will result in a decrease in their transformation efficiency. If the cells are frozen first in a dry ice / ethanol bath before placement in the -80C freezer, the loss will be about two-fold. If placed directly in the -80C freezer, the loss in transformation efficiency is about five to ten-fold.
Competent cells were left on ice overnight - can they still be used?
Competent cells left on ice, or allowed to come to room temperature slowly will suffer a dramatic loss in transformation efficiency. We recommend thawing a new aliquot of competent cells.
Can I express a recombinant protein that is toxic to the bacteria?
For expression of toxic recombinant proteins, select the BL21 strain containing the pLysS plasmid. These cells express low levels of lysozyme that will bind to T7 polymerase, and inhibit transcription.
What DNA purity do I need for use in transformation?
The DNA used should be high quality - free from phenol, proteins, detergents, and ethanol. Dissolve the DNA in sterile water or 0.5x TE buffer (5 mM Tris-HCl, 0.5 mM EDTA).
Can I grow more competent cells from the stock I purchased?
Generally, no, because the future generations of cells lose their competency.
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