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Genomic analysis of Sleeping Beauty transposon integration in human somatic cells

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Item Type:Article
Title:Genomic analysis of Sleeping Beauty transposon integration in human somatic cells
Creators Name:Turchiano, G. and Latella, M.C. and Gogol-Döring, A. and Cattoglio, C. and Mavilio, F. and Izsvák, Z. and Ivics, Z. and Recchia, A.
Abstract:The Sleeping Beauty (SB) transposon is a non-viral integrating vector system with proven efficacy for gene transfer and functional genomics. However, integration efficiency is negatively affected by the length of the transposon. To optimize the SB transposon machinery, the inverted repeats and the transposase gene underwent several modifications, resulting in the generation of the hyperactive SB100X transposase and of the high-capacity "sandwich" (SA) transposon. In this study, we report a side-by-side comparison of the SA and the widely used T2 arrangement of transposon vectors carrying increasing DNA cargoes, up to 18 kb. Clonal analysis of SA integrants in human epithelial cells and in immortalized keratinocytes demonstrates stability and integrity of the transposon independently from the cargo size and copy number-dependent expression of the cargo cassette. A genome-wide analysis of unambiguously mapped SA integrations in keratinocytes showed an almost random distribution, with an overrepresentation in repetitive elements (satellite, LINE and small RNAs) compared to a library representing insertions of the first-generation transposon vector and to gammaretroviral and lentiviral libraries. The SA transposon/SB100X integrating system therefore shows important features as a system for delivering large gene constructs for gene therapy applications.
Keywords:Cell Line, DNA Transposable Elements, Fibroblasts, Gene Transfer Techniques, Genetic Vectors, HeLa Cells, Keratinocytes, Transposases, Animals, Mice
Source:PLoS ONE
ISSN:1932-6203
Publisher:Public Library of Science (U.S.A.)
Volume:9
Number:11
Page Range:e112712
Date:12 November 2014
Official Publication:https://doi.org/10.1371/journal.pone.0112712
PubMed:View item in PubMed

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