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Biological Chemistry

Editor-in-Chief: Brüne, Bernhard

Editorial Board Member: Buchner, Johannes / Ludwig, Stephan / Sies, Helmut / Turk, Boris / Wittinghofer, Alfred

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Improved lentiviral gene transfer into human embryonic stem cells grown in co-culture with murine feeder and stroma cells

Melanie Wurm1, 2 / Benjamin Groß3, 4 / Malte Sgodda4 / Ludger Ständker5 / Thomas Müller1 / Wolf-Georg Forssmann5 / Peter A. Horn6 / Rainer Blasczyk1 / 4

1Institute for Transfusion Medicine, Hannover Medical School, D-30625 Hannover, Germany

2Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA

3Department of Paediatric Haematology and Oncology, Hannover Medical School, D-30625 Hannover, Germany

4JRG Stem Cell Biology, Cluster of Excellence REBIRTH, Hannover Medical School, D-30625 Hannover, Germany

5Department of Immunology and Rheumatology, Hannover Medical School, D-30625 Hannover, Germany

6Institute for Transfusion Medicine, University Hospital Essen, D-45147 Essen, Germany

Corresponding author

Citation Information: Biological Chemistry. Volume 392, Issue 10, Pages 887–895, ISSN (Online) 1437-4315, ISSN (Print) 1431-6730, DOI: 10.1515/BC.2011.085, August 2011

Publication History

Received:
2011-04-19
Accepted:
2011-06-27
Published Online:
2011-08-04

Abstract

Genetic modification of human embryonic stem cells (hESCs) using biophysical DNA transfection methods are hampered by the very low single cell survival rate and cloning efficiency of hESCs. Lentiviral gene transfer strategies are widely used to genetically modify hESCs but limited transduction efficiencies in the presence of feeder or stroma cells present problems, particularly if vesicular stomatitis virus glycoprotein (VSV-G) pseudotyped viral particles are applied. Here, we investigated whether the recently described semen derived enhancer of virus infection (SEVI) and alternative viral envelope proteins derived from either Gibbon ape leukaemia virus (GALV) or feline leukaemia virus (RD114) are applicable for transducing hESCs during co-culture with feeder or stroma cells. Our first set of experiments demonstrates that SEVI has no toxic effect on murine or hESCs and that exposure to SEVI does not interfere with the pluripotency-associated phenotype. Focusing on hESCs, we were able to further demonstrate that SEVI increases the transduction efficiencies of GALV and RD114 pseudotyped lentiviral vectors. More importantly, aiming at targeted differentiation of hESCs into functional somatic cell types, GALV pseudotyped lentiviral particles could efficiently and exclusively transduce hESCs grown in co-culture with OP9-GFP stroma cells (which were often used to induce differentiation into haematopoietic derivatives).

Keywords: embryonic stem cells; lentiviral gene transfer; SEVI; transduction efficiency

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