Genomic Analysis of Sleeping Beauty Transposon Integration in Human Somatic Cells

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 t...

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Main Authors:	Turchiano, Giandomenico, Latella, Maria Carmela, Gogol-Döring, Andreas, Cattoglio, Claudia, Mavilio, Fulvio, Izsvák, Zsuzsanna, Ivics, Zoltán, Recchia, Alessandra
Format:	Online
Language:	English
Published:	Public Library of Science 2014
Online Access:	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4229213/

id	pubmed-4229213
recordtype	oai_dc
spelling	pubmed-42292132014-11-18 Genomic Analysis of Sleeping Beauty Transposon Integration in Human Somatic Cells Turchiano, Giandomenico Latella, Maria Carmela Gogol-Döring, Andreas Cattoglio, Claudia Mavilio, Fulvio Izsvák, Zsuzsanna Ivics, Zoltán Recchia, Alessandra Research Article 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. Public Library of Science 2014-11-12 /pmc/articles/PMC4229213/ /pubmed/25390293 http://dx.doi.org/10.1371/journal.pone.0112712 Text en © 2014 Turchiano et al http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.
repository_type	Open Access Journal
institution_category	Foreign Institution
institution	US National Center for Biotechnology Information
building	NCBI PubMed
collection	Online Access
language	English
format	Online
author	Turchiano, Giandomenico Latella, Maria Carmela Gogol-Döring, Andreas Cattoglio, Claudia Mavilio, Fulvio Izsvák, Zsuzsanna Ivics, Zoltán Recchia, Alessandra
spellingShingle	Turchiano, Giandomenico Latella, Maria Carmela Gogol-Döring, Andreas Cattoglio, Claudia Mavilio, Fulvio Izsvák, Zsuzsanna Ivics, Zoltán Recchia, Alessandra Genomic Analysis of Sleeping Beauty Transposon Integration in Human Somatic Cells
author_facet	Turchiano, Giandomenico Latella, Maria Carmela Gogol-Döring, Andreas Cattoglio, Claudia Mavilio, Fulvio Izsvák, Zsuzsanna Ivics, Zoltán Recchia, Alessandra
author_sort	Turchiano, Giandomenico
title	Genomic Analysis of Sleeping Beauty Transposon Integration in Human Somatic Cells
title_short	Genomic Analysis of Sleeping Beauty Transposon Integration in Human Somatic Cells
title_full	Genomic Analysis of Sleeping Beauty Transposon Integration in Human Somatic Cells
title_fullStr	Genomic Analysis of Sleeping Beauty Transposon Integration in Human Somatic Cells
title_full_unstemmed	Genomic Analysis of Sleeping Beauty Transposon Integration in Human Somatic Cells
title_sort	genomic analysis of sleeping beauty transposon integration in human somatic cells
description	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.
publisher	Public Library of Science
publishDate	2014
url	https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4229213/
_version_	1613155638093807616

Genomic Analysis of Sleeping Beauty Transposon Integration in Human Somatic Cells

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