Up to 25-fold higher transformation efficiency of ligated DNA
Bruce Jerpseth Marie Callahan Alan Greener
Stratagene Cloning Systems, Inc.
Stratagenes new, high-efficiency competent E. coli cell line significantly improves the transformation efficiency of ligated DNA. Epicurian Coli XL10-Gold ultracompetent cells* yielded transformation efficiencies for ligated DNA that were up to 25-fold higher than other commercially available cell lines. XL10-Gold cells are the host cells of choice for any cloning experiment that requires a high efficiency of retrieving transformants from ligation reactions, such as constructing plasmid libraries and PCR cloning.
The ability to efficiently introduce DNA into E. coli by chemical
transformation and electroporation is a key step for many procedures, such as
generating plasmid libraries, cloning large DNA fragments and cloning PCR
products.1 The cells that have set the industry standard for these
types of procedures are Stratagenes ultracompetent cells, which yield greater
than 5 x 109 transformants/g of supercoiled pUC18 vector (2.7 kb).
However, these transformation efficiencies are typically calculated using very
small amounts of supercoiled pUC18 vector DNA. Ligated DNA molecules are known
to transform cells at a significantly lower efficiency than supercoiled
molecules, and large plasmids transform cells less efficiently than small
plasmids. This observation has particular impact for researchers making plasmid
libraries: the total number and size distribution of primary transformants is
key to finding the gene of interest and affects how easily full-length clones
can be located. The bias against transforming larger molecules influences the
ability to retrieve full-length cDNA clones. For example, many plasmid library
vectors, including two-hybrid vectors and eukaryotic expression