Efficient viral delivery of Cas9 into human safe harbor

Abstract Gene editing using CRISPR/Cas9 is a promising method to cure many human genetic diseases. We have developed an efficient system to deliver Cas9 into the adeno-associated virus integration site 1 (AAVS1) locus, known as a safe harbor, using lentivirus and AAV viral vectors, as a step toward...

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Autores principales:	Hideki Hayashi, Yoshinao Kubo, Mai Izumida, Toshifumi Matsuyama
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2020
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/fce61c5bca0b424ab95f614d32a7cd92
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spelling	oai:doaj.org-article:fce61c5bca0b424ab95f614d32a7cd922021-12-02T16:18:05ZEfficient viral delivery of Cas9 into human safe harbor10.1038/s41598-020-78450-82045-2322https://doaj.org/article/fce61c5bca0b424ab95f614d32a7cd922020-12-01T00:00:00Zhttps://doi.org/10.1038/s41598-020-78450-8https://doaj.org/toc/2045-2322Abstract Gene editing using CRISPR/Cas9 is a promising method to cure many human genetic diseases. We have developed an efficient system to deliver Cas9 into the adeno-associated virus integration site 1 (AAVS1) locus, known as a safe harbor, using lentivirus and AAV viral vectors, as a step toward future in vivo transduction. First, we introduced Cas9v1 (derived from Streptococcus pyogenes) at random into the genome using a lentiviral vector. Cas9v1 activity was used when the N-terminal 1.9 kb, and C-terminal 2.3 kb fragments of another Cas9v2 (human codon-optimized) were employed sequentially with specific single-guide RNAs (sgRNAs) and homology donors carried by AAV vectors into the AAVS1 locus. Then, Cas9v1 was removed from the genome by another AAV vector containing sgRNA targeting the long terminal repeat of the lentivirus vector. The reconstituted Cas9v2 in the AAVS1 locus was functional and gene editing was efficient.Hideki HayashiYoshinao KuboMai IzumidaToshifumi MatsuyamaNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 10, Iss 1, Pp 1-14 (2020)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Hideki Hayashi Yoshinao Kubo Mai Izumida Toshifumi Matsuyama Efficient viral delivery of Cas9 into human safe harbor
description	Abstract Gene editing using CRISPR/Cas9 is a promising method to cure many human genetic diseases. We have developed an efficient system to deliver Cas9 into the adeno-associated virus integration site 1 (AAVS1) locus, known as a safe harbor, using lentivirus and AAV viral vectors, as a step toward future in vivo transduction. First, we introduced Cas9v1 (derived from Streptococcus pyogenes) at random into the genome using a lentiviral vector. Cas9v1 activity was used when the N-terminal 1.9 kb, and C-terminal 2.3 kb fragments of another Cas9v2 (human codon-optimized) were employed sequentially with specific single-guide RNAs (sgRNAs) and homology donors carried by AAV vectors into the AAVS1 locus. Then, Cas9v1 was removed from the genome by another AAV vector containing sgRNA targeting the long terminal repeat of the lentivirus vector. The reconstituted Cas9v2 in the AAVS1 locus was functional and gene editing was efficient.
format	article
author	Hideki Hayashi Yoshinao Kubo Mai Izumida Toshifumi Matsuyama
author_facet	Hideki Hayashi Yoshinao Kubo Mai Izumida Toshifumi Matsuyama
author_sort	Hideki Hayashi
title	Efficient viral delivery of Cas9 into human safe harbor
title_short	Efficient viral delivery of Cas9 into human safe harbor
title_full	Efficient viral delivery of Cas9 into human safe harbor
title_fullStr	Efficient viral delivery of Cas9 into human safe harbor
title_full_unstemmed	Efficient viral delivery of Cas9 into human safe harbor
title_sort	efficient viral delivery of cas9 into human safe harbor
publisher	Nature Portfolio
publishDate	2020
url	https://doaj.org/article/fce61c5bca0b424ab95f614d32a7cd92
work_keys_str_mv	AT hidekihayashi efficientviraldeliveryofcas9intohumansafeharbor AT yoshinaokubo efficientviraldeliveryofcas9intohumansafeharbor AT maiizumida efficientviraldeliveryofcas9intohumansafeharbor AT toshifumimatsuyama efficientviraldeliveryofcas9intohumansafeharbor
_version_	1718384210104811520

Efficient viral delivery of Cas9 into human safe harbor

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