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Highly selective transgene expression through the flip-excision switch system by using a unilateral spacer sequence
The flip-excision switch (FLEX) system with an adeno-associated viral (AAV) vector allows expression of transgenes in specific cell populations having Cre recombinase. A significant issue with this system is non-specific expression of transgenes in tissues after vector injection. We show here that C...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Elsevier
2023
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10014282/ https://www.ncbi.nlm.nih.gov/pubmed/36936079 http://dx.doi.org/10.1016/j.crmeth.2022.100393 |
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author | Matsushita, Natsuki Kato, Shigeki Nishizawa, Kayo Sugawara, Masateru Takeuchi, Kosei Miyasaka, Yoshiki Mashimo, Tomoji Kobayashi, Kazuto |
author_facet | Matsushita, Natsuki Kato, Shigeki Nishizawa, Kayo Sugawara, Masateru Takeuchi, Kosei Miyasaka, Yoshiki Mashimo, Tomoji Kobayashi, Kazuto |
author_sort | Matsushita, Natsuki |
collection | PubMed |
description | The flip-excision switch (FLEX) system with an adeno-associated viral (AAV) vector allows expression of transgenes in specific cell populations having Cre recombinase. A significant issue with this system is non-specific expression of transgenes in tissues after vector injection. We show here that Cre-independent recombination events in the AAV genome carrying the FLEX sequence occur mainly during the production of viral vectors in packaging cells, which results in transgene expression in off-target populations. Introduction of a relatively longer nucleotide sequence between two recognition sites at the unilateral side of the transgene cassette, termed a unilateral spacer sequence (USS), is useful to suppress the recombination in the viral genome, leading to the protection of non-specific transgene expression with enhanced gene expression selectivity. Our FLEX/USS system offers a powerful strategy for highly specific Cre-dependent transgene expression, aiming at various applications for structural and functional analyses of target cell populations. |
format | Online Article Text |
id | pubmed-10014282 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2023 |
publisher | Elsevier |
record_format | MEDLINE/PubMed |
spelling | pubmed-100142822023-03-16 Highly selective transgene expression through the flip-excision switch system by using a unilateral spacer sequence Matsushita, Natsuki Kato, Shigeki Nishizawa, Kayo Sugawara, Masateru Takeuchi, Kosei Miyasaka, Yoshiki Mashimo, Tomoji Kobayashi, Kazuto Cell Rep Methods Article The flip-excision switch (FLEX) system with an adeno-associated viral (AAV) vector allows expression of transgenes in specific cell populations having Cre recombinase. A significant issue with this system is non-specific expression of transgenes in tissues after vector injection. We show here that Cre-independent recombination events in the AAV genome carrying the FLEX sequence occur mainly during the production of viral vectors in packaging cells, which results in transgene expression in off-target populations. Introduction of a relatively longer nucleotide sequence between two recognition sites at the unilateral side of the transgene cassette, termed a unilateral spacer sequence (USS), is useful to suppress the recombination in the viral genome, leading to the protection of non-specific transgene expression with enhanced gene expression selectivity. Our FLEX/USS system offers a powerful strategy for highly specific Cre-dependent transgene expression, aiming at various applications for structural and functional analyses of target cell populations. Elsevier 2023-01-18 /pmc/articles/PMC10014282/ /pubmed/36936079 http://dx.doi.org/10.1016/j.crmeth.2022.100393 Text en © 2022 The Author(s) https://creativecommons.org/licenses/by-nc-nd/4.0/This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/). |
spellingShingle | Article Matsushita, Natsuki Kato, Shigeki Nishizawa, Kayo Sugawara, Masateru Takeuchi, Kosei Miyasaka, Yoshiki Mashimo, Tomoji Kobayashi, Kazuto Highly selective transgene expression through the flip-excision switch system by using a unilateral spacer sequence |
title | Highly selective transgene expression through the flip-excision switch system by using a unilateral spacer sequence |
title_full | Highly selective transgene expression through the flip-excision switch system by using a unilateral spacer sequence |
title_fullStr | Highly selective transgene expression through the flip-excision switch system by using a unilateral spacer sequence |
title_full_unstemmed | Highly selective transgene expression through the flip-excision switch system by using a unilateral spacer sequence |
title_short | Highly selective transgene expression through the flip-excision switch system by using a unilateral spacer sequence |
title_sort | highly selective transgene expression through the flip-excision switch system by using a unilateral spacer sequence |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10014282/ https://www.ncbi.nlm.nih.gov/pubmed/36936079 http://dx.doi.org/10.1016/j.crmeth.2022.100393 |
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