Minicircle DNA Immobilized in Bacterial Ghosts: In vivo Production of Safe Non-Viral DNA Delivery Vehicles

Vol. 8, No. 4, 2004

Free Abstract Article (References) Article (PDF 304 KB)

Research Article

Minicircle DNA Immobilized in Bacterial Ghosts: In vivo Production of Safe Non-Viral DNA Delivery Vehicles
Wolfgang Jechlinger^{a, b, c}, Chakameh Azimpour Tabrizi^a, Werner Lubitz^{a, b}, Peter Mayrhofer^{a, b, c}

^aInstitute of Microbiology and Genetics, Section Microbiology and Biotechnology, University of Vienna,
^bBIRD-C GmbH & CoKEG, and
^cMayrhofer and Jechlinger OEG, Vienna, Austria

Address of Corresponding Author

J Mol Microbiol Biotechnol 2004;8:222-231 (DOI: 10.1159/000086703)

Key Words

DNA delivery
Minicircle DNA
Site-specific recombination
Self-immobilization system
Bacterial ghosts

Abstract

DNA as an active agent is among the most promising technologies for vaccination and therapy. However, plasmid backbone sequences needed for the production of pDNA in bacteria are dispensable, reduce the efficiency of the DNA agent and, most importantly, represent a biological safety risk. In this report we describe a novel technique where a site-specific recombination system based on the ParA resolvase was applied to a self-immobilizing plasmid system (SIP). In addition, this system was combined with the protein E-specific lysis technology to produce non-living bacterial carrier vehicles loaded with minicircle DNA. The in vivo recombination process completely divided an origin plasmid into a minicircle and a miniplasmid. The replicative miniplasmid containing the origin of replication and the antibiotic resistance gene was lost during the subsequently induced PhiX174 gene E-mediated lysis process, which results in bacterial ghosts. The minicircle DNA was retained in these empty bacterial cell envelopes during the lysis process via the specific interaction of a membrane anchored protein with the minicircle DNA. Using this novel platform technology, a DNA delivery vehicle – consisting of a safe bacterial carrier with known adjuvant properties and minicircle DNA with an optimized safety profile – can be produced in vivo in a continuous process. Furthermore, this study provides the basis for the development of an efficient in vitro minicircle purification process.

Author Contacts

Wolfgang Jechlinger, PhD
Institute of Bacteriology, Mycology and Hygiene, Department of Pathobiology
University of Veterinary Medicine, Veterinärplatz 1, A–1210 Vienna (Austria)
Tel. +43 1 25077 2104, Fax +43 1 25077 2190
E-Mail Wolfgang.Jechlinger@vu-wien.ac.at

Article Information

Number of Print Pages : 10
Number of Figures : 2, Number of Tables : 0, Number of References : 42

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