title: A leader sequence capable of enhancing RNA expression and protein synthesis in mammalian cells.
abstract: Many applications in biotechnology require human proteins generated from human cells. Stable cell lines commonly used for this purpose are difficult to develop, and scaling to large numbers of proteins can be problematic. Transient expression can circumvent this problem, but protein yields are generally too low for most applications. Here we report a novel 37-nucleotide leader sequence that promotes rapid and high transgene expression in mammalian cells. This sequence was identified by in vitro selection and functions in a transient vaccinia-based cytoplasmic expression system. Vectors containing this sequence produce microgram levels of protein in just 6 h from a small-scale expression in 10(6) cells. This level of protein synthesis is ideal for high throughput production of human proteins, and could be scaled to generate milligram quantities of protein. The technology is compatible with a broad range of cell lines, accepts plasmid and linear DNA, and functions with viruses that are approved for use under BSL1 conditions. We suggest that these advantages provide a powerful method for generating human protein in mammalian cells.
authors.affiliation: Center for Evolutionary Medicine and Informatics, Arizona State University, Tempe, Arizona, 85287-5301.
authors.forename: Brian P
authors.initials: BP
authors.lastname: Wellensiek
authors.forename: Andrew C
authors.initials: AC
authors.lastname: Larsen
authors.forename: Julia
authors.initials: J
authors.lastname: Flores
authors.forename: Bertram L
authors.initials: BL
authors.lastname: Jacobs
authors.forename: John C
authors.initials: JC
authors.lastname: Chaput
mesh_terms: D020121:5' Untranslated Regions; D000818:Animals; D001709:Biotechnology / Q000379:methods*; D002460:Cell Line; D006224:Cricetinae; D015870:Gene Expression; D005822:Genetic Vectors; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000235:genetics; D014616:Vaccinia virus / Q000235:genetics*
chemical_list: D020121:5' Untranslated Regions; D012333:RNA, Messenger; D011994:Recombinant Proteins
keywords: human vaccinia virus; mammalian cell culture; recombinant protein synthesis; translation enhancing elements
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