Summary
Recombinant retroviruses enabling transfer of genes into a broad range of cell lines are used for different purposes. High virus titers and efficient expression of the integrated genes or both are usually required.
In testing different retroviral vectors, we were unable to find high-yield expression of integrated genes. Expression from promoters is significantly reduced upon integration into retroviral vectors. We show that a DNA fragment from the commonly used Tn5 transposon conferring G418 resistance is able to repress a heterologous promoter in cis. This DNA fragment is regarded to be responsible for inefficient expression of genes from internal promoters in retroviral vectors carrying the neomycin resistance gene. In contrast, LTRs do not influence downstream located SV40 promoters in cis or by transcriptional interference.
In order to facilitate the screening of high-yield virus producing subclones, we have adapted a previously established method for fast and simple isolation of over-expressing cell clones. This method allows the direct isolation of ψ2 transformants with high virus titers. Transformants derived by this procedure produce approximately 10-fold more recombinant virus as compared to pools of conventionally selected ψ2 transfectants.
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© 1989 Springer-Verlag Berlin Heidelberg
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Artelt, P., Bartsch, J., Hauser, H., Wirth, M. (1989). Improvement of Gene Expression and Virus Production in the Use of Retroviral Vectors for Gene Transfer. In: Lother, H., Dernick, R., Ostertag, W. (eds) Vectors as Tools for the Study of Normal and Abnormal Growth and Differentiation. NATO ASI Series, vol 34. Springer, Berlin, Heidelberg. https://doi.org/10.1007/978-3-642-74197-5_15
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DOI: https://doi.org/10.1007/978-3-642-74197-5_15
Publisher Name: Springer, Berlin, Heidelberg
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