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Analysis of a Cas12a-based gene-drive system in budding yeast

The discovery and adaptation of CRISPR/Cas systems within molecular biology has provided advances across biological research, agriculture and human health. Genomic manipulation through use of a CRISPR nuclease and programmed guide RNAs has become a common and widely accessible practice. The identifi...

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Detalles Bibliográficos
Autores principales: Lewis, Isabel C., Yan, Yao, Finnigan, Gregory C.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Microbiology Society 2021
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8749140/
https://www.ncbi.nlm.nih.gov/pubmed/35024561
http://dx.doi.org/10.1099/acmi.0.000301
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author Lewis, Isabel C.
Yan, Yao
Finnigan, Gregory C.
author_facet Lewis, Isabel C.
Yan, Yao
Finnigan, Gregory C.
author_sort Lewis, Isabel C.
collection PubMed
description The discovery and adaptation of CRISPR/Cas systems within molecular biology has provided advances across biological research, agriculture and human health. Genomic manipulation through use of a CRISPR nuclease and programmed guide RNAs has become a common and widely accessible practice. The identification and introduction of new engineered variants and orthologues of Cas9 as well as alternative CRISPR systems such as the type V group have provided additional molecular options for editing. These include distinct PAM requirements, staggered DNA double-strand break formation, and the ability to multiplex guide RNAs from a single expression construct. Use of CRISPR/Cas has allowed for the construction and testing of a powerful genetic architecture known as a gene drive within eukaryotic model systems. Our previous work developed a drive within budding yeast using Streptococcus pyogenes Cas9. Here, we installed the type V Francisella novicida Cas12a (Cpf1) nuclease gene and its corresponding guide RNA to power a highly efficient artificial gene drive in diploid yeast. We examined the consequence of altering guide length or introduction of individual mutational substitutions to the crRNA sequence. Cas12a-dependent gene-drive function required a guide RNA of at least 18 bp and could not tolerate most changes within the 5′ end of the crRNA.
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spelling pubmed-87491402022-01-11 Analysis of a Cas12a-based gene-drive system in budding yeast Lewis, Isabel C. Yan, Yao Finnigan, Gregory C. Access Microbiol Research Articles The discovery and adaptation of CRISPR/Cas systems within molecular biology has provided advances across biological research, agriculture and human health. Genomic manipulation through use of a CRISPR nuclease and programmed guide RNAs has become a common and widely accessible practice. The identification and introduction of new engineered variants and orthologues of Cas9 as well as alternative CRISPR systems such as the type V group have provided additional molecular options for editing. These include distinct PAM requirements, staggered DNA double-strand break formation, and the ability to multiplex guide RNAs from a single expression construct. Use of CRISPR/Cas has allowed for the construction and testing of a powerful genetic architecture known as a gene drive within eukaryotic model systems. Our previous work developed a drive within budding yeast using Streptococcus pyogenes Cas9. Here, we installed the type V Francisella novicida Cas12a (Cpf1) nuclease gene and its corresponding guide RNA to power a highly efficient artificial gene drive in diploid yeast. We examined the consequence of altering guide length or introduction of individual mutational substitutions to the crRNA sequence. Cas12a-dependent gene-drive function required a guide RNA of at least 18 bp and could not tolerate most changes within the 5′ end of the crRNA. Microbiology Society 2021-12-17 /pmc/articles/PMC8749140/ /pubmed/35024561 http://dx.doi.org/10.1099/acmi.0.000301 Text en © 2021 The Authors https://creativecommons.org/licenses/by/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution License.
spellingShingle Research Articles
Lewis, Isabel C.
Yan, Yao
Finnigan, Gregory C.
Analysis of a Cas12a-based gene-drive system in budding yeast
title Analysis of a Cas12a-based gene-drive system in budding yeast
title_full Analysis of a Cas12a-based gene-drive system in budding yeast
title_fullStr Analysis of a Cas12a-based gene-drive system in budding yeast
title_full_unstemmed Analysis of a Cas12a-based gene-drive system in budding yeast
title_short Analysis of a Cas12a-based gene-drive system in budding yeast
title_sort analysis of a cas12a-based gene-drive system in budding yeast
topic Research Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8749140/
https://www.ncbi.nlm.nih.gov/pubmed/35024561
http://dx.doi.org/10.1099/acmi.0.000301
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