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dc.contributor.authorMinskaia, Ekaterina
dc.contributor.authorNicholson, John
dc.contributor.authorRyan, Martin Denis
dc.date.accessioned2013-08-28T09:31:01Z
dc.date.available2013-08-28T09:31:01Z
dc.date.issued2013-08-22
dc.identifier.citationMinskaia , E , Nicholson , J & Ryan , M D 2013 , ' Optimisation of the foot-and-mouth disease virus 2A co-expression system for biomedical applications ' , BMC Biotechnology , vol. 13 , 67 . https://doi.org/10.1186/1472-6750-13-67en
dc.identifier.issn1472-6750
dc.identifier.otherPURE: 67129967
dc.identifier.otherPURE UUID: 2ae20532-3b5e-4b1d-9913-bd6a52fdddaf
dc.identifier.otherScopus: 84882998955
dc.identifier.otherORCID: /0000-0002-0012-0614/work/47136052
dc.identifier.urihttp://hdl.handle.net/10023/4008
dc.descriptionThis work was supported by the Medical Research Council.en
dc.description.abstractBackground Many biomedical applications require the expression or production of therapeutic hetero-multimeric proteins/protein complexes: in most cases only accomplished by co-ordinated co-expression within the same cell. Foot-and-mouth disease virus 2A (F2A) and ‘2A-like’ sequences are now widely used for this purpose. Since 2A mediates a co-translational ‘cleavage’ at its own C-terminus, sequences encoding multiple proteins (linked via 2As) can be concatenated into a single ORF: a single transgene. It has been shown that in some cases, however, the cleavage efficiency of shorter versions of F2A may be inhibited by the C-terminus of certain gene sequences immediately upstream of F2A. This paper describes further work to optimise F2A for co-expression strategies. Results We have inserted F2A of various lengths in between GFP and CherryFP ‘reporter’ proteins (in reciprocal or tandem arrangements). The co-expression of these proteins and cleavage efficiencies of F2As of various lengths were studied by in vitro coupled transcription and translation in rabbit reticulocyte lysates, western blotting of HeLa cell lysates and fluorescence microscopy. Conclusions Optimal and suboptimal lengths of F2A sequences were identified as a result of detailed ‘fine-tuning’ of the F2A sequence. Based on our data and the model according to which 2A activity is a product of its interaction with the exit tunnel of the ribosome, we suggest the length of the F2A sequence which is not ‘sensitive’ to the C-terminus of the upstream protein that can be successfully used for co-expression of two proteins for biomedical applications.
dc.language.isoeng
dc.relation.ispartofBMC Biotechnologyen
dc.rights© 2013 Minskaia et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.en
dc.subjectProtein co-expressionen
dc.subjectBicistronic vectoren
dc.subjectCo-translational ‘self-cleavage’en
dc.subjectQR355 Virologyen
dc.subject.lccQR355en
dc.titleOptimisation of the foot-and-mouth disease virus 2A co-expression system for biomedical applicationsen
dc.typeJournal articleen
dc.description.versionPublisher PDFen
dc.contributor.institutionUniversity of St Andrews.Biomedical Sciences Research Complexen
dc.contributor.institutionUniversity of St Andrews.School of Biologyen
dc.identifier.doihttps://doi.org/10.1186/1472-6750-13-67
dc.description.statusPeer revieweden


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