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CRISPR-based self-cleaving mechanism for controllable gene delivery in human cells

Controllable gene delivery via vector-based systems remains a formidable challenge in mammalian synthetic biology and a desirable asset in gene therapy applications. Here, we introduce a methodology to control the copies and residence time of a gene product delivered in host human cells but also sel...

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Autores principales: Moore, Richard, Spinhirne, Alec, Lai, Michael J., Preisser, Samantha, Li, Yi, Kang, Taek, Bleris, Leonidas
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4333380/
https://www.ncbi.nlm.nih.gov/pubmed/25527740
http://dx.doi.org/10.1093/nar/gku1326
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author Moore, Richard
Spinhirne, Alec
Lai, Michael J.
Preisser, Samantha
Li, Yi
Kang, Taek
Bleris, Leonidas
author_facet Moore, Richard
Spinhirne, Alec
Lai, Michael J.
Preisser, Samantha
Li, Yi
Kang, Taek
Bleris, Leonidas
author_sort Moore, Richard
collection PubMed
description Controllable gene delivery via vector-based systems remains a formidable challenge in mammalian synthetic biology and a desirable asset in gene therapy applications. Here, we introduce a methodology to control the copies and residence time of a gene product delivered in host human cells but also selectively disrupt fragments of the delivery vehicle. A crucial element of the proposed system is the CRISPR protein Cas9. Upon delivery, Cas9 guided by a custom RNA sequence cleaves the delivery vector at strategically placed targets thereby inactivating a co-expressed gene of interest. Importantly, using experiments in human embryonic kidney cells, we show that specific parameters of the system can be adjusted to fine-tune the delivery properties. We envision future applications in complex synthetic biology architectures, gene therapy and trace-free delivery.
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spelling pubmed-43333802015-02-26 CRISPR-based self-cleaving mechanism for controllable gene delivery in human cells Moore, Richard Spinhirne, Alec Lai, Michael J. Preisser, Samantha Li, Yi Kang, Taek Bleris, Leonidas Nucleic Acids Res Synthetic Biology and Bioengineering Controllable gene delivery via vector-based systems remains a formidable challenge in mammalian synthetic biology and a desirable asset in gene therapy applications. Here, we introduce a methodology to control the copies and residence time of a gene product delivered in host human cells but also selectively disrupt fragments of the delivery vehicle. A crucial element of the proposed system is the CRISPR protein Cas9. Upon delivery, Cas9 guided by a custom RNA sequence cleaves the delivery vector at strategically placed targets thereby inactivating a co-expressed gene of interest. Importantly, using experiments in human embryonic kidney cells, we show that specific parameters of the system can be adjusted to fine-tune the delivery properties. We envision future applications in complex synthetic biology architectures, gene therapy and trace-free delivery. Oxford University Press 2015-01-30 2014-12-18 /pmc/articles/PMC4333380/ /pubmed/25527740 http://dx.doi.org/10.1093/nar/gku1326 Text en © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research. http://creativecommons.org/licenses/by-nc/4.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
spellingShingle Synthetic Biology and Bioengineering
Moore, Richard
Spinhirne, Alec
Lai, Michael J.
Preisser, Samantha
Li, Yi
Kang, Taek
Bleris, Leonidas
CRISPR-based self-cleaving mechanism for controllable gene delivery in human cells
title CRISPR-based self-cleaving mechanism for controllable gene delivery in human cells
title_full CRISPR-based self-cleaving mechanism for controllable gene delivery in human cells
title_fullStr CRISPR-based self-cleaving mechanism for controllable gene delivery in human cells
title_full_unstemmed CRISPR-based self-cleaving mechanism for controllable gene delivery in human cells
title_short CRISPR-based self-cleaving mechanism for controllable gene delivery in human cells
title_sort crispr-based self-cleaving mechanism for controllable gene delivery in human cells
topic Synthetic Biology and Bioengineering
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4333380/
https://www.ncbi.nlm.nih.gov/pubmed/25527740
http://dx.doi.org/10.1093/nar/gku1326
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