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Updated: Sep 27, 2026
High-Efficiency Transduction of Liver Cancer Cells by Recombinant Adeno-Associated Virus Serotype 3 Vectors
Published on: March 22, 2011
Comparison of the EF-1 alpha and the CMV promoter for engineering stable tumor cell lines using recombinant
Christian Teschendorf1, Kenneth H Warrington, Dietmar W Siemann
1Department of Molecular Genetics and Microbiology, College of Medicine, University of Florida, P.O. Box 100266, JHMHSC, Gainesville, FL 32610, USA. Christian.Teschendorf@ruhr-uni-bochum.de
Background:
Silencing of the viral CMV immediate early enhancer promoter can be a problem in certain cell types when engineering stable cell lines.
Materials And Methods:
We compared the efficacy of the CMV promoter to the promoter of the elongation factor-1 alpha (EF-1 alpha) for the generation of stable colon carcinoma cell lines (HT-29). Green fluorescent protein (GFP) expression cassettes were delivered by recombinant adeno-associated virus (AAV) which is known for its ability to stably transduce cells. Stable cell lines were characterized in vitro by FACS and in vivo after HT-29 clones were grown as xenografts in nude mice.
Results:
Stable HT-29 clones with > 97% of all cells homogeneously expressing GFP were generated with the EF-1 alpha promoter. In contrast in clones carrying the CMV promoter, only up to 60% of the cells were GFP-positive with expression levels varying widely between cells. Superinfection with wild-type adenovirus induced GFP expression in more than 90% of the cells indicating that the CMV promoter was silenced. In vivo the tumors carrying the EF-1 alpha promoter were homogeneously GFP-positive, whereas the CMV promoter gave rise to a scattered pattern of GFP expression.
Conclusion:
This study underlines the importance of the promoter for the generation of stable cell lines. In addition it demonstrates that recombinant AAV can effectively be used as a gene delivery system for this purpose.
