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Avoidance of reporter assay distortions from fused dual reporters
Loughran, Gary; Howard, Michael T.; Firth, Andrew E.; Atkins, John F.
Positioning test sequences between fused reporters permits monitoring of both translation levels and framing, before and after the test sequence. Many studies, including those on recoding such as productive ribosomal frameshifting and stop codon readthrough, use distinguishable luciferases or fluorescent proteins as reporters. Occasional distortions, due to test sequence product interference with the individual reporter activities or stabilities, are here shown to be avoidable by the introduction of tandem StopGo sequences (2A) flanking the test sequence. Using this new vector system (pSGDluc), we provide evidence for the use of a 3' stem-loop stimulator for ACP2 readthrough, but failed to detect the reported VEGFA readthrough.
Keyword(s): Recoding; Frameshifting; Readthrough; Dual luciferase; StopGo; VEGF-Ax; Cleavage; Stop; 2A; Codon; Translation; Polyprotein; Aphthovirus; Fusion
Publication Date:
Type: Journal article
Peer-Reviewed: Yes
Language(s): English
Institution: University College Cork
Funder(s): Science Foundation Ireland
Citation(s): Loughran, G., Howard, M. T., Firth, A. E. and Atkins, J. F. (2017) 'Avoidance of reporter assay distortions from fused dual reporters', RNA, 23(8) (6pp). doi: 10.1261/rna.061051.117
Publisher(s): Cold Spring Harbor Lab Press
File Format(s): application/pdf
Related Link(s):
First Indexed: 2017-09-27 06:38:20 Last Updated: 2017-09-27 06:38:20