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Type: Journal article
Title: Co-expression of a scFv antibody fragment and a reporter protein using lentiviral shuttle plasmid containing a self-processing furin-2A sequence
Author: Appleby, S.
Irani, Y.
Mortimer, L.
Brereton, H.
Klebe, S.
Keane, M.
Cowan, P.
Williams, K.
Citation: Journal of Immunological Methods, 2013; 397(1-2):61-65
Publisher: Elsevier
Issue Date: 2013
ISSN: 0022-1759
Statement of
Sarah L.Appleby, Yazad Irani, Lauren A.Mortimer, Helen M.Brereton, Sonja Klebe, Miriam C.Keane, Peter J.Cowan, Keryn A.Williams
Abstract: It is often desirable to co-express a reporter protein with a potential therapeutic protein, to verify correct targeting of an expression strategy. Vectors containing a viral self-processing 2A sequence have been reported to drive equimolar expression of two or more transgenes from a single promoter. Here, we report on the co-expression of a secreted antibody fragment and an intracellular reporter protein, enhanced yellow fluorescent protein from lentiviral shuttle plasmids by inserting a furin-2A (F2A) sequence between the two cDNAs, in two different orientations, in the expression cassette. We show that the order of these two transgenes relative to the F2A sequence affects expression levels. Reduced expression of each transgene positioned downstream of F2A, compared with upstream of F2A, was observed (p<0.05). Moreover, protein expression from double-cDNA plasmids was significantly lower than from their corresponding single transgene counterparts (p<0.05).
Keywords: scFv; reporter protein; co-expression; 2A self-processing sequence; furin cleavage site; lentiviral vector
Rights: © 2013 Elsevier B.V. All rights reserved.
DOI: 10.1016/j.jim.2013.08.012
Appears in Collections:Aurora harvest 8
Opthalmology & Visual Sciences publications

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