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High-level expression by tissue/cancer-specific promoter with strict specificity using a single-adenoviral vector

Tissue-/cancer-specific promoters for use in adenovirus vectors (AdVs) are valuable for elucidating specific gene functions and for use in gene therapy. However, low activity, non-specific expression and size limitations in the vector are always problems. Here, we developed a ‘double-unit’ AdV conta...

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Autores principales: Kanegae, Yumi, Terashima, Miho, Kondo, Saki, Fukuda, Hiromitsu, Maekawa, Aya, Pei, Zheng, Saito, Izumu
Formato: Texto
Lenguaje:English
Publicado: Oxford University Press 2011
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3025582/
https://www.ncbi.nlm.nih.gov/pubmed/21051352
http://dx.doi.org/10.1093/nar/gkq966
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author Kanegae, Yumi
Terashima, Miho
Kondo, Saki
Fukuda, Hiromitsu
Maekawa, Aya
Pei, Zheng
Saito, Izumu
author_facet Kanegae, Yumi
Terashima, Miho
Kondo, Saki
Fukuda, Hiromitsu
Maekawa, Aya
Pei, Zheng
Saito, Izumu
author_sort Kanegae, Yumi
collection PubMed
description Tissue-/cancer-specific promoters for use in adenovirus vectors (AdVs) are valuable for elucidating specific gene functions and for use in gene therapy. However, low activity, non-specific expression and size limitations in the vector are always problems. Here, we developed a ‘double-unit’ AdV containing the Cre gene under the control of an α-fetoprotein promoter near the right end of its genome and bearing a compact ‘excisional-expression’ unit consisting of a target cDNA ‘upstream’ of a potent promoter between two loxPs near the left end of its genome. When Cre was expressed, the expression unit was excised as a circular molecule and strongly expressed. Undesired leak expression of Cre during virus preparation was completely suppressed by a dominant-negative Cre and a short-hairpin RNA against Cre. Using this novel construct, a very strict specificity was maintained while achieving a 40- to 90-fold higher expression level, compared with that attainable using a direct specific promoter. Therefore, the ‘double-unit’ AdV enabled us to produce a tissue-/cancer-specific promoter in an AdV with a high expression level and strict specificity.
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spelling pubmed-30255822011-01-24 High-level expression by tissue/cancer-specific promoter with strict specificity using a single-adenoviral vector Kanegae, Yumi Terashima, Miho Kondo, Saki Fukuda, Hiromitsu Maekawa, Aya Pei, Zheng Saito, Izumu Nucleic Acids Res Methods Online Tissue-/cancer-specific promoters for use in adenovirus vectors (AdVs) are valuable for elucidating specific gene functions and for use in gene therapy. However, low activity, non-specific expression and size limitations in the vector are always problems. Here, we developed a ‘double-unit’ AdV containing the Cre gene under the control of an α-fetoprotein promoter near the right end of its genome and bearing a compact ‘excisional-expression’ unit consisting of a target cDNA ‘upstream’ of a potent promoter between two loxPs near the left end of its genome. When Cre was expressed, the expression unit was excised as a circular molecule and strongly expressed. Undesired leak expression of Cre during virus preparation was completely suppressed by a dominant-negative Cre and a short-hairpin RNA against Cre. Using this novel construct, a very strict specificity was maintained while achieving a 40- to 90-fold higher expression level, compared with that attainable using a direct specific promoter. Therefore, the ‘double-unit’ AdV enabled us to produce a tissue-/cancer-specific promoter in an AdV with a high expression level and strict specificity. Oxford University Press 2011-01 2010-11-04 /pmc/articles/PMC3025582/ /pubmed/21051352 http://dx.doi.org/10.1093/nar/gkq966 Text en © The Author(s) 2010. Published by Oxford University Press. http://creativecommons.org/licenses/by-nc/2.5 This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.5), which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
spellingShingle Methods Online
Kanegae, Yumi
Terashima, Miho
Kondo, Saki
Fukuda, Hiromitsu
Maekawa, Aya
Pei, Zheng
Saito, Izumu
High-level expression by tissue/cancer-specific promoter with strict specificity using a single-adenoviral vector
title High-level expression by tissue/cancer-specific promoter with strict specificity using a single-adenoviral vector
title_full High-level expression by tissue/cancer-specific promoter with strict specificity using a single-adenoviral vector
title_fullStr High-level expression by tissue/cancer-specific promoter with strict specificity using a single-adenoviral vector
title_full_unstemmed High-level expression by tissue/cancer-specific promoter with strict specificity using a single-adenoviral vector
title_short High-level expression by tissue/cancer-specific promoter with strict specificity using a single-adenoviral vector
title_sort high-level expression by tissue/cancer-specific promoter with strict specificity using a single-adenoviral vector
topic Methods Online
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3025582/
https://www.ncbi.nlm.nih.gov/pubmed/21051352
http://dx.doi.org/10.1093/nar/gkq966
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