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dc.contributor.authorRoberts, Jennifer Anne
dc.contributor.authorMiguel-Escalada, Irene
dc.contributor.authorSlovik, Katherine Joan
dc.contributor.authorWalsh, Kathleen Theodora
dc.contributor.authorHadzhiev, Yavor
dc.contributor.authorSanges, Remo
dc.contributor.authorStupka, Elia
dc.contributor.authorMarsh, Elizabeth Kate
dc.contributor.authorBalciuniene, Jorune
dc.contributor.authorBalciunas, Darius
dc.contributor.authorMüller, Ferenc
dc.date.accessioned2019-06-12T10:27:42Z
dc.date.available2019-06-12T10:27:42Z
dc.date.issued2014-01-21
dc.identifier.citationRoberts, J.A. et al. (2014) 'Targeted transgene integration overcomes variability of position effects in zebrafish', Development, 141(3), pp.715-724. doi: 10.1242/dev.100347.en_US
dc.identifier.issn1477-9129
dc.identifier.pmid24449846
dc.identifier.doi10.1242/dev.100347
dc.identifier.urihttp://hdl.handle.net/10545/623840
dc.description.abstractZebrafish transgenesis is increasingly popular owing to the optical transparency and external development of embryos, which provide a scalable vertebrate model for in vivo experimentation. The ability to express transgenes in a tightly controlled spatio-temporal pattern is an important prerequisite for exploitation of zebrafish in a wide range of biomedical applications. However, conventional transgenesis methods are plagued by position effects: the regulatory environment of genomic integration sites leads to variation of expression patterns of transgenes driven by engineered cis-regulatory modules. This limitation represents a bottleneck when studying the precise function of cis-regulatory modules and their subtle variants or when various effector proteins are to be expressed for labelling and manipulation of defined sets of cells. Here, we provide evidence for the efficient elimination of variability of position effects by developing a PhiC31 integrase-based targeting method. To detect targeted integration events, a simple phenotype scoring of colour change in the lens of larvae is used. We compared PhiC31-based integration and Tol2 transgenesis in the analysis of the activity of a novel conserved enhancer from the developmentally regulated neural-specific esrrga gene. Reporter expression was highly variable among independent lines generated with Tol2, whereas all lines generated with PhiC31 into a single integration site displayed nearly identical, enhancer-specific reporter expression in brain nuclei. Moreover, we demonstrate that a modified integrase system can also be used for the detection of enhancer activity in transient transgenesis. These results demonstrate the power of the PhiC31-based transgene integration for the annotation and fine analysis of transcriptional regulatory elements and it promises to be a generally desirable tool for a range of applications, which rely on highly reproducible patterns of transgene activity in zebrafish.en_US
dc.description.sponsorshipThis work was funded by ‘BOLD’ Marie-Curie Initial Training Network; and ‘ZFHealth’ Integrating project in the Framework 7 programme of the European RESEARCH ARTICLE Development (2014) doi:10.1242/dev.100347Development Commission; University of Birmingham (F.M.); Temple University; and the National Institutes of Health (NIH) [HD061749 to D.B.].en_US
dc.language.isoenen_US
dc.relation.urlhttps://www.ncbi.nlm.nih.gov/pubmed/24449846en_US
dc.relation.urlhttps://dev.biologists.org/content/141/3/715en_US
dc.rightsAttribution-NonCommercial-NoDerivs 3.0 United States*
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/3.0/us/*
dc.subjectEnhanceren_US
dc.subjectIntegraseen_US
dc.subjectPosition effectsen_US
dc.subjectTol2en_US
dc.subjectTransgenesisen_US
dc.subjectZebrafishen_US
dc.titleTargeted transgene integration overcomes variability of position effects in zebrafish.en_US
dc.typeArticleen_US
dc.contributor.departmentUniversity of Birminghamen_US
dc.contributor.departmentTemple Universityen_US
dc.identifier.journalDevelopmenten_US
dc.source.journaltitleDevelopment (Cambridge, England)
dcterms.dateAccepted2013-11-10
refterms.dateFOA2019-06-12T10:27:43Z
dc.author.detail785801en_US


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