Improved Method of Expression Cloning
Summary The human genome will soon be completely sequenced, presenting a vast new array of targets against which pharmaceutical agents can potentially be designed. However, to capitalize on this new information, the cellular function and medical significance of each of these genes will need to be determined. Here we describe a new cell-free expression cloning method that can rapidly identify novel proteins based on their function. Whereas traditional expression cloning methods have been used to identify receptor proteins and secreted growth factors, both of which have proved to be excellent drug targets, this new method is especially well suited to identify intracellular components of cellular signaling or growth regulation pathways.
Applications This technology represents a significant improvement over standard methods for drug target identification, as it extends the power of expression cloning to factors that are active in the cytoplasm and nucleus. The technology is easy to use, inexpensive and versatile, and it can be used to rapidly identify the cDNA encoding almost any protein for which a biochemical assay exists. The method involves the subdivision of cDNA expression libraries into small pools, which are then used to express pools of protein in vitro.
There are multiple applications for this technology:
(1) The technology provides a rapid route to identifying new targets for pharmaceutical development. Examples include DNA-binding proteins (transcription factors), enzymes such as protein kinases or proteases, and the protein substrates of these enzymes.
(2) The technology can be used to directly identify proteins with therapeutic potential. It also facilitates the identification of new isoforms of proteins that may have novel properties.
(3) This method will facilitate the identification of enzymes that are useful in biotechnology, including restriction enzymes and other DNA modifying enzymes. It can also be used to rapidly clone the cDNAs encoding enzymes that are active at extremes of temperature or ionic strength. Such enzymes would also be of interest to manufacturers of household cleaning agents including detergents.
(4) The technology is suited to the production of a kit product for sale to research labs or pharmaceutical companies involved in target identification. This could include the sale of pooled cDNA libraries by companies which manufacture such libraries. It could also include companies that subdivide a library as a service. Finally, companies which sell the means to construct protein pools in vitro could design products especially suited for this purpose. For Further Information Please Contact the Director of Business Development Michal Preminger Email: email@example.com Telephone: (617) 432-0920
Kirschner, Marc W.
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