Making your own competent cells?
Cloning or subcloning is often just a preliminary step in research programs. It should be simple and consistent. That is why many researchers choose to get their competent cells from a commercial supplier - quality-controlled lots, that are tested to meet clear specifications for transformation efficiency. This alleviates concerns about batch-to-batch variability and helps to make transformation of your DNA constructs routine and reliable.
Important factors to consider when choosing competent cells include transformation efficiency, the presence of genetic markers in the bacterial strain, and the convenience of the packaging format. These factors will directly impact the time and effort required for your project, as well as its success. That’s why Life Technologies offers a wide selection of Invitrogen™ competent cell types and formats, so that you can make the right choice to help you achieve your cloning goals.
The transformation efficiency you need will be largely determined by your application. To meet different cloning requirements, competent cells are available in a wide range of transformation efficiencies - from >1 × 106 for routine cloning to >3 × 1010 cfu/μg pUC DNA for library construction or working with limited amounts of DNA.
Figure 1. A range of transformation efficiencies are available to meet your application needs. One Shot® cells are provided at >5 x 109, 1 x 109, or >1 x 108 cfu/µg.
Determining Transformation Efficiency
Usually the transformation efficiency of competent cells is measured by transforming them with subsaturating amounts of supercoiled pUC DNA (~10–500 pg). The results are expressed in number of transformants (or colony forming units) per microgram of plasmid DNA (cfu/μg).
Transformation Efficiency Calculations
|# of colonies||x||106 pg||x||300 µl total
|=||# of transformants|
|100 pg transformed DNA||µg||? µl plated||µg plasmid DNA|
Single-use One Shot® aliquots for unparalleled convenience
The unique One Shot® format provides 50 μl aliquots of chemically competent cells in single-use tubes. Transformation is performed directly in the tube, saving you time and effort. There are no extra pipetting steps, no extra tubes, and no loss of efficiency due to repeated freeze-thaw cycles. These kits come with SOC medium and pUC19 positive control (Figure 1).
Our DH5α™ competent cells are designed for general cloning & subcloning, and are available in various transformation efficiencies (from >106 to 109 cfu/µg) and packing formats. DH5α™ Cells are a well-known, versatile strain that can be used in many everyday cloning applications. In addition to supporting blue/white screening, recA1 and endA1 mutations in DH5α™ Cells increase insert stability and improve the quality of plasmid DNA prepared from minipreps. DH5α™ Cells offer the following benefits:
- A wide range of efficiencies from >106 to >109 cfu/µg
- Greatly increased plasmid yield and quality due to endA1 mutation
- Blue/white screening of recombinant clones due to lacZΔM15
- Superior insert stability due to recA1 mutation