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Journal of Virology, June 2003, p. 7017-7025, Vol. 77, No. 12
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.12.7017-7025.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.

Generation of Transduction-Competent Retroviral Vectors by Infection with a Single Hybrid Vaccinia Virus

Christian Konetschny, Georg W. Holzer, Carsten Urban, Thomas Hämmerle, Josef Mayrhofer, and Falko G. Falkner*

Baxter Vaccine AG, Biomedical Research Center, A-2304 Orth/Donau, Austria

Received 2 December 2002/ Accepted 21 March 2003

Recombinant vaccinia viruses that express defective retroviral vectors upon a single infection event in normal host cells were constructed. The gag-pol and envelope genes and a retroviral vector unit were inserted as vaccinia virus promoter-controlled transcription units at three separate loci. The triple recombinant virus was used to infect such diverse cell types as monkey and rabbit kidney, human lung, and primary chicken cells, resulting in the production of transduction-competent defective retroviral vectors. Infection of Chinese hamster ovary cells, which are nonpermissive for vaccinia virus replication, also resulted in production of retroviral vectors and secondary permanent transduction of the host cells. Since vaccinia virus supports the expression of cytotoxic proteins, the vesicular stomatitis virus G glycoprotein could be chosen as the envelope allowing a broad host range of transduction. Functionality of particles was monitored by expression of the green fluorescent protein in transduced 3T3 cell clones. This is the first description of a single chimeric virus encoding and releasing functional retroviral vectors, providing proof of principle of the new concept. No replication-competent retrovirus was detectable by sensitive reverse transcriptase assays. Since vaccinia virus has a broad host range, is extremely robust, and can be obtained at high titers and safe nonreplicating vaccinia virus strains are available, the hybrid system may open new perspectives for gene delivery.

* Corresponding author. Mailing address: Baxter Vaccine AG, Biomedical Research Center, Uferstrasse 15, A-2304 Orth/Donau, Austria. Phone: 43 1 20100 4232. Fax: 43 1 20100 4000. E-mail: falknef{at}baxter.com.

Journal of Virology, June 2003, p. 7017-7025, Vol. 77, No. 12
0022-538X/03/$08.00+0     DOI: 10.1128/JVI.77.12.7017-7025.2003
Copyright © 2003, American Society for Microbiology. All Rights Reserved.





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