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Use of Minicircle Plasmids for Gene Therapy

Protocol
First Online:
Part of the Methods in Molecular Biology™ book series (MIMB, volume 542)

Summary

A large number of cancer gene therapy clinical trials are currently being performed that are attempting to evaluate novel approaches to eliminate tumor cells by the introduction of genetic material into patients. One of the most important objectives in gene therapy is the development of highly safe and efficient vector systems for gene transfer in eukaryotic cells. Currently, viral and nonviral vector systems are used, both having their advantages and limitations. Minicircles are novel supercoiled minimal expression cassettes, derived from conventional plasmid DNA by site-specific recombination in vivo in Escherichia coli for the use in nonviral gene therapy and vaccination. Minicircle DNA lacks the bacterial backbone sequence consisting of an antibiotic resistance gene, an origin of replication, and inflammatory sequences intrinsic to bacterial DNA. In addition to their improved safety profile, minicircles have been shown to greatly increase the efficiency of transgene expression in various in vitro and in vivo studies. In this chapter, we describe the production, purification, and application of minicircle DNA and discuss the rationale of the improved gene transfer efficiencies compared to conventional plasmid DNA.

Keywords

Bacterial backbone sequences biosafety minicircle DNA nonviral gene therapy persistent transgene expression plasmid site-specific recombination 
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Copyright information

© Humana Press, a part of Springer Science+Business Media, LLC 2009

Authors and Affiliations

  1. 1.Mayrhofer & Jechlinger OEGViennaAustria
  2. 2.PlasmidFactory GmbH & Co. KGBielefeldGermany
  3. 3.Institute of Bacteriology, Mycology and Hygiene, Department of PathobiologyUniversity of Veterinary Medicine ViennaViennaAustria

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