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The Development of an AAV-Based CRISPR SaCas9 Genome Editing System That Can Be Delivered to Neurons in vivo and Regulated via Doxycycline and Cre-Recombinase
The RNA-guided Cas9 nuclease, from the type II prokaryotic clustered regularly interspersed short palindromic repeats (CRISPR) adaptive immune system, has been adapted by scientists to enable site specific genome editing of eukaryotic cells both in vitro and in vivo. Previously, we reported the deve...
Autores principales: | , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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Frontiers Media S.A.
2018
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Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6243075/ https://www.ncbi.nlm.nih.gov/pubmed/30483052 http://dx.doi.org/10.3389/fnmol.2018.00413 |
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author | Kumar, Namrata Stanford, William de Solis, Christopher Aradhana, Abraham, Nigel D. Dao, Trieu-Mi J. Thaseen, Sadiqa Sairavi, Anusha Gonzalez, Cuauhtemoc Ulises Ploski, Jonathan E. |
author_facet | Kumar, Namrata Stanford, William de Solis, Christopher Aradhana, Abraham, Nigel D. Dao, Trieu-Mi J. Thaseen, Sadiqa Sairavi, Anusha Gonzalez, Cuauhtemoc Ulises Ploski, Jonathan E. |
author_sort | Kumar, Namrata |
collection | PubMed |
description | The RNA-guided Cas9 nuclease, from the type II prokaryotic clustered regularly interspersed short palindromic repeats (CRISPR) adaptive immune system, has been adapted by scientists to enable site specific genome editing of eukaryotic cells both in vitro and in vivo. Previously, we reported the development of an adeno-associated virus (AAV)-mediated CRISPR Streptococcus pyogenes (Sp) Cas9 system, in which the genome editing function can be regulated by controlling the expression of the guide RNA (sgRNA) in a doxycycline (Dox)-dependent manner. Here, we report the development of an AAV vector tool kit utilizing the Cas9 from Staphylococcus aureus (SaCas9). We demonstrate in vitro genome editing in human derived 293FT cells and mouse derived Neuro2A (N2A) cells and in vivo in neurons of the mouse brain. We also demonstrate the ability to regulate the induction of genome editing temporally with Dox and spatially with Cre-recombinase. The combination of these systems enables AAV-mediated CRISPR/Cas9 genome editing to be regulated both spatially and temporally. |
format | Online Article Text |
id | pubmed-6243075 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2018 |
publisher | Frontiers Media S.A. |
record_format | MEDLINE/PubMed |
spelling | pubmed-62430752018-11-27 The Development of an AAV-Based CRISPR SaCas9 Genome Editing System That Can Be Delivered to Neurons in vivo and Regulated via Doxycycline and Cre-Recombinase Kumar, Namrata Stanford, William de Solis, Christopher Aradhana, Abraham, Nigel D. Dao, Trieu-Mi J. Thaseen, Sadiqa Sairavi, Anusha Gonzalez, Cuauhtemoc Ulises Ploski, Jonathan E. Front Mol Neurosci Neuroscience The RNA-guided Cas9 nuclease, from the type II prokaryotic clustered regularly interspersed short palindromic repeats (CRISPR) adaptive immune system, has been adapted by scientists to enable site specific genome editing of eukaryotic cells both in vitro and in vivo. Previously, we reported the development of an adeno-associated virus (AAV)-mediated CRISPR Streptococcus pyogenes (Sp) Cas9 system, in which the genome editing function can be regulated by controlling the expression of the guide RNA (sgRNA) in a doxycycline (Dox)-dependent manner. Here, we report the development of an AAV vector tool kit utilizing the Cas9 from Staphylococcus aureus (SaCas9). We demonstrate in vitro genome editing in human derived 293FT cells and mouse derived Neuro2A (N2A) cells and in vivo in neurons of the mouse brain. We also demonstrate the ability to regulate the induction of genome editing temporally with Dox and spatially with Cre-recombinase. The combination of these systems enables AAV-mediated CRISPR/Cas9 genome editing to be regulated both spatially and temporally. Frontiers Media S.A. 2018-11-13 /pmc/articles/PMC6243075/ /pubmed/30483052 http://dx.doi.org/10.3389/fnmol.2018.00413 Text en Copyright © 2018 Kumar, Stanford, de Solis, Aradhana, Abraham, Dao, Thaseen, Sairavi, Gonzalez and Ploski. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms. |
spellingShingle | Neuroscience Kumar, Namrata Stanford, William de Solis, Christopher Aradhana, Abraham, Nigel D. Dao, Trieu-Mi J. Thaseen, Sadiqa Sairavi, Anusha Gonzalez, Cuauhtemoc Ulises Ploski, Jonathan E. The Development of an AAV-Based CRISPR SaCas9 Genome Editing System That Can Be Delivered to Neurons in vivo and Regulated via Doxycycline and Cre-Recombinase |
title | The Development of an AAV-Based CRISPR SaCas9 Genome Editing System That Can Be Delivered to Neurons in vivo and Regulated via Doxycycline and Cre-Recombinase |
title_full | The Development of an AAV-Based CRISPR SaCas9 Genome Editing System That Can Be Delivered to Neurons in vivo and Regulated via Doxycycline and Cre-Recombinase |
title_fullStr | The Development of an AAV-Based CRISPR SaCas9 Genome Editing System That Can Be Delivered to Neurons in vivo and Regulated via Doxycycline and Cre-Recombinase |
title_full_unstemmed | The Development of an AAV-Based CRISPR SaCas9 Genome Editing System That Can Be Delivered to Neurons in vivo and Regulated via Doxycycline and Cre-Recombinase |
title_short | The Development of an AAV-Based CRISPR SaCas9 Genome Editing System That Can Be Delivered to Neurons in vivo and Regulated via Doxycycline and Cre-Recombinase |
title_sort | development of an aav-based crispr sacas9 genome editing system that can be delivered to neurons in vivo and regulated via doxycycline and cre-recombinase |
topic | Neuroscience |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6243075/ https://www.ncbi.nlm.nih.gov/pubmed/30483052 http://dx.doi.org/10.3389/fnmol.2018.00413 |
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