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Escherichia coli BL21(DE3) is a commonly used protein production strain of the E. coli bacterium. This strain combines several features that allow for excessive expression of heterologous proteins . It is derived from the B lineage of E. coli .
The expression vector is transformed into one of several relevant strains of E. coli, most frequently BL21(DE3). The E. coli cell also has its own chromosome, which possesses a gene that is expressed to produce T7 RNA polymerase.
The DE3 prophage found in BL21(DE3) provides T7 RNA polymerase (driven by the LacUV5 promoter), allowing for vectors with the T7 promoter to be used instead. [15]
Agilent Technologies, Inc. is an American global company headquartered in Santa Clara, California, that provides instruments, software, services, and consumables for laboratories. Agilent was established in 1999 as a spin-off from Hewlett-Packard. The resulting IPO of Agilent stock was the largest in the history of Silicon Valley at the time.
Super Optimal Broth (SOB medium) is a nutrient-rich bacterial growth medium used for microbiological culture, generally of Escherichia coli.This nutrient-rich microbial broth contains peptides, amino acids, water soluble vitamins and glucose in a low-salt formulation.
In the case of deletions, a single transformation protocol is more efficient and shows a 100% deletion rate following colony PCR. [1] Further work should be pursued to determine causes for the different transformation efficiencies for various mutation types. Short sequence insertions are also possible using the SCAR-less method.
This device is able to transform almost any type of cell and is not limited to the transformation of the nucleus; it can also transform organelles, including plastids and mitochondria. [3] A gene gun is used for delivery of exogenous DNA to cells. This method is known as 'biolistics'.
Protocols, however, exist for making supercompetent cells that may yield a transformation efficiency of over 10 9. [61] The chemical method, however, usually does not work well for linear DNA, such as fragments of chromosomal DNA, probably because the cell's native exonuclease enzymes rapidly degrade linear DNA.