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Episomal Transgene Expression in Pluripotent Stem Cells

  • Michele M. P. Lufino
  • Anna R. Popplestone
  • Sally A. Cowley
  • Pauline A. H. Edser
  • William S. James
  • Richard Wade-Martins
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 767)

Abstract

Herpes simplex type 1 (HSV-1) amplicon vectors possess a number of features that make them excellent vectors for the delivery of transgenes into stem cells. HSV-1 amplicon vectors are capable of efficiently transducing both dividing and nondividing cells and since the virus is quite large, 152 kb, it is of sufficient size to allow for incorporation of entire genomic DNA loci with native promoters. HSV-1 amplicon vectors can also be used to incorporate and deliver to cells a variety of sequences that allow extrachromosomal retention. These elements offer advantages over integrating vectors as they avoid transgene silencing and insertional mutagenesis. The construction of amplicon vectors carrying extrachromosomal retention elements, their packaging into HSV-1 viral particles, and the use of HSV-1 amplicons for stem cell transduction will be described.

Key words

HSV-1 amplicon iBAC Extrachromosomal vector Stem cells Gene expression vector 

Notes

Acknowledgments

This work was supported by the Parkinson’s UK Monument Trust Discovery Award; the Friedreich’s Ataxia Research Alliance, Ataxia UK and the National Ataxia Foundation; the Medical Research Council and the Biotechnology and Biological Sciences Research Council. M.M.P.L. is an Ataxia UK Research Fellow, A.R.P. is a Medical Research Council student and S.A.C. is a Wellcome Trust Research Fellow.

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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  • Michele M. P. Lufino
    • 1
  • Anna R. Popplestone
    • 1
  • Sally A. Cowley
    • 2
  • Pauline A. H. Edser
    • 1
  • William S. James
    • 2
  • Richard Wade-Martins
    • 1
  1. 1.Molecular Neurodegeneration and Gene Therapy Research Group, Department of Physiology, Anatomy and GeneticsUniversity of OxfordOxfordUK
  2. 2.Sir William Dunn School of PathologyUniversity of OxfordOxfordUK

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