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Blood, 15 January 2006, Vol. 107, No. 2, pp. 483-491.
Prepublished online as a Blood First Edition Paper on September 27, 2005; DOI 10.1182/blood-2005-05-2133.
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Submitted May 26, 2005
Accepted September 10, 2005
Stable gene transfer and expression in human primary T-cells by the Sleeping Beauty transposon system
Xin Huang, Andrew C Wilber, Lei Bao, Dong Tuong, Jakub Tolar, Paul J Orchard, Bruce L Levine, Carl H June, R S McIvor, Bruce R Blazar, and Xianzheng Zhou*
Division of Blood and Marrow Transplantation, Department of Pediatrics, University of Minnesota, Minneapolis, MN, USA; The Cancer Center, University of Minnesota, Minneapolis, MN, USA
Department of Genetics, Cell Biology and Development, University of Minnesota, Minneapolis, MN, USA; The Cancer Center, University of Minnesota, Minneapolis, MN, USA
Department of Pathology and Laboratory Medicine, the Abramson Family Cancer Research Institute, University of Pennsylvania, Philadelphia, PA, USA
* Corresponding author; email: zhoux058{at}umn.edu.
The Sleeping Beauty (SB) transposon system is a non-viral DNA delivery system in which a transposase directs integration of an SB transposon into TA-dinucleotide sites in the genome. To determine whether the SB transposon system can mediate stable gene expression in human T-cells, primary peripheral blood lymphocytes (PBLs) were nucleofected with SB vectors carrying a DsRed reporter gene. Plasmids containing the SB transposase on the same (cis) or separate molecule (trans) as the SB transposon mediated long-term and stable reporter gene expression in human primary T-cells. Sequencing of transposon:chromosome junctions confirmed that stable gene expression was due to SB-mediated transposition. In other studies, PBLs were successfully transfected using the SB transposon system and shown to stably express a fusion protein consisting of (i) a surface receptor useful for positive T-cell selection and (ii) a "suicide" gene useful for elimination of transfected T-cells after chemotherapy. This study is the first report demonstrating that the SB transposon system can mediate stable gene transfer in human primary PBLs, which may be advantageous for T-cell based gene therapies.
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