A New ?Destination? for Protein Expression: A Lentiviral Gateway? Destination Vector for High-Level Protein Expression (pDEST-67
Posted Sep 19 2010 5:00pm
Description of Invention: A laboratory at the Science Applications International Corporation in Frederick, MD (SAIC-Frederick) has developed a lentiviral vector, pDEST-673, for high protein expression yields in cells. The pDEST-673 vector combines three features that make it optimal for protein expression in lentiviruses: the pFUGW backbone, a Gateway® vector conversion cassette, and a neomycin antibiotic resistance marker. The pFUGW portion contains a highly potent polypurine tract (PPT) that allows for the production of higher viral titers within transfected cells and a woodchuck regulatory element (WRE) to enhance protein expression. The addition of the Gateway® conversion cassette converts the vector into a Destination vector and the neomycin resistance marker allows for researchers to select for stable transfectants using antibiotic selection (a feature not possessed by many lentiviral vectors). This lentiviral Destination vector should be useful for researchers desiring to utilize neomycin resistance to select for proteins expressed in cells stably transfected with lentiviruses.
Research tool for high quantity production of a protein(s) of interest for studying the role of the protein(s) in a variety of biological processes, including pathologies such as cancers, infectious diseases, autoimmune diseases, and many other disorders
Research tool for selecting stable lentiviral transfectants following the insertion of the vector into tumor cells
Potential tool for enhancing production of proteins that are normally difficult to express in other types of bacterial, insect, or mammalian expression systems
The pFUGW backbone provides the pDEST-673 vector with optimal protein expression properties: The polypurine tract (PPT) region in the vector allows for efficient viral transcription leading to increased lentiviral production in cells. The woodchuck regulatory element acts as a posttranscriptional enhancer to promote the conversion of more mRNA transcripts into protein to yield high-levels of the protein of interest. These elements are not found in most commercially available lentiviral vectors.
The incorporation of the neomycin resistance marker facilitates selection of the transfectants of interest: Many laboratories rely on neomycin selection as a key selectable marker in their protein expression experiments. Few commercially available lentiviral vectors contain a neomycin resistance marker.
Inventors: Dominic Esposito (NCI)
Patent Status: HHS, Reference No. E-119-2009/0
Research Tool -- patent protection is not being pursued for this technology
A Ventura, et al. Cre-lox-regulated conditional RNA interference from transgenes. Proc. Natl. Acad. Sci. U S A. 2004 Jul 13;101(28):10380-10385. [ PubMed: 15240889 ]
C Lois, et al. Germline transmission and tissue-specific expression of transgenes delivered by lentiviral vectors. Science 2002 Feb 1;295(5556):868-872. [ PubMed: 11786607 ]
Licensing Status: Available for licensing under a Biological Materials License Agreement.
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