NDLI: Retroviral Vectors Produced in the Cytoplasmic Vaccinia Virus System Transduce Intron-Containing Genes

Content Provider	PubMed Central
Author	Konetschny, C. Holzer, G. W. Falkner, F. G.
Copyright Year	2002
Abstract	Introns and polyadenylation (pA) sites are known to improve transcript stability and nuclear-cytoplasmic transport and are normally present in efficient gene expression vectors. Standard retroviral vectors, however, do not allow the inclusion of such sequence elements, as mRNA processing at internal splice and pA sites interferes with the production of functional full-length vector genomes. In this report we examined the capability of hybrid vaccinia/retroviral vectors to transduce complex gene cassettes with nuclear RNA processing signals within the retroviral genome. A retroviral vector was constructed that contains a gene of interest (the human coagulation factor IX [FIX] cDNA), including an intron and an internal pA site. The modified proviral vector genome was cloned downstream of a vaccinia virus promoter and was inserted into the vaccinia virus genome. Infection of a packaging cell line with the recombinant vaccinia virus vector resulted in secretion of retroviral particles at average titers of 105 CFU per ml of cell culture supernatant. Due to the cytoplasmic transcription and the nonrecognition of nuclear transcription signals in the vaccinia virus system, full-length transcripts were obtained that still contained the intron. In the retrovirally transduced cell lines the FIX transcripts were terminated at the internal pA site. The transcripts were quantitatively spliced, and FIX was secreted. Recombinant cell lines with stable single-copy inserts containing sequence elements necessary for efficient gene function could be generated. Thus, a relatively simple cytoplasmic system for the generation of complex retroviral vectors is described. Retroviral vectors transducing intron-containing gene cassettes may play a further role in gene therapy applications.
Related Links	http://dx.doi.org/10.1128/jvi.76.3.1236-1243.2002
Ending Page	1243
Page Count	8
Starting Page	1236
File Format	PDF
ISSN	0022538X
e-ISSN	10985514
Journal	Journal of Virology
Issue Number	3
Volume Number	76
Language	English
Publisher	American Society for Microbiology
Publisher Date	2002-02-01
Access Restriction	Open
Rights Holder	American Society for Microbiology
Subject Keyword	Immunology Virology Research in Higher Education
Content Type	Text
Resource Type	Article
Subject	Virology Immunology Microbiology Insect Science

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