Cell-free protein synthesis using cell extracts from Escherichia coli, wheat germ and rabbit reticulocytes has been used for over 40 years to produce small amounts of radiolabeled proteins for identification of gene products and other applications. In the E. coli system programmed with plasmid DNA, the cell extract contains or is supplemented with an RNA polymerase to transcribe the gene, and the mRNAs are translated by a complex mixture that contains ribosomes and a full complement of initiation, elongation and termination factors, as well as a full set of aminoacyl-tRNA synthetases and other required enzymes. The presence of molecular chaperones, protein disulfide and peptidyl-prolyl cis-trans isomerases generally ensures that proteins are correctly folded into soluble, active forms. With the eukaryotic extracts, it has additionally been possible to program protein synthesis directly with in vitro transcribed mRNA that can be produced directly from PCR products, and the presence of factors responsible for post-translational modification of proteins ensures the production of fully functional gene products.
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