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Rapid generation of maternal mutants via oocyte transgenic expression of CRISPR-Cas9 and sgRNAs in zebrafish
Maternal products are exclusive factors to drive oogenesis and early embryonic development. As disrupting maternal gene functions is either time-consuming or technically challenging, early developmental programs regulated by maternal factors remain mostly elusive. We provide a transgenic approach to...
Autores principales: | , , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
American Association for the Advancement of Science
2021
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8346210/ https://www.ncbi.nlm.nih.gov/pubmed/34362733 http://dx.doi.org/10.1126/sciadv.abg4243 |
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author | Zhang, Chong Lu, Tong Zhang, Yizhuang Li, Jiaguang Tarique, Imran Wen, Fenfen Chen, Aijun Wang, Jiasheng Zhang, Zhuoyu Zhang, Yanjun Shi, De-Li Shao, Ming |
author_facet | Zhang, Chong Lu, Tong Zhang, Yizhuang Li, Jiaguang Tarique, Imran Wen, Fenfen Chen, Aijun Wang, Jiasheng Zhang, Zhuoyu Zhang, Yanjun Shi, De-Li Shao, Ming |
author_sort | Zhang, Chong |
collection | PubMed |
description | Maternal products are exclusive factors to drive oogenesis and early embryonic development. As disrupting maternal gene functions is either time-consuming or technically challenging, early developmental programs regulated by maternal factors remain mostly elusive. We provide a transgenic approach to inactivate maternal genes in zebrafish primary oocytes. By introducing three tandem single guide RNA (sgRNA) expression cassettes and a green fluorescent protein (GFP) reporter into Tg(zpc:zcas9) embryos, we efficiently obtained maternal nanog and ctnnb2 mutants among GFP-positive F(1) offspring. Notably, most of these maternal mutants displayed either sgRNA site–spanning genomic deletions or unintended large deletions extending distantly from the sgRNA targets, suggesting a prominent deletion-prone tendency of genome editing in the oocyte. Thus, our method allows maternal gene knockout in the absence of viable and fertile homozygous mutant adults. This approach is particularly time-saving and can be applied for functional screening of maternal factors and generating genomic deletions in zebrafish. |
format | Online Article Text |
id | pubmed-8346210 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2021 |
publisher | American Association for the Advancement of Science |
record_format | MEDLINE/PubMed |
spelling | pubmed-83462102021-08-13 Rapid generation of maternal mutants via oocyte transgenic expression of CRISPR-Cas9 and sgRNAs in zebrafish Zhang, Chong Lu, Tong Zhang, Yizhuang Li, Jiaguang Tarique, Imran Wen, Fenfen Chen, Aijun Wang, Jiasheng Zhang, Zhuoyu Zhang, Yanjun Shi, De-Li Shao, Ming Sci Adv Research Articles Maternal products are exclusive factors to drive oogenesis and early embryonic development. As disrupting maternal gene functions is either time-consuming or technically challenging, early developmental programs regulated by maternal factors remain mostly elusive. We provide a transgenic approach to inactivate maternal genes in zebrafish primary oocytes. By introducing three tandem single guide RNA (sgRNA) expression cassettes and a green fluorescent protein (GFP) reporter into Tg(zpc:zcas9) embryos, we efficiently obtained maternal nanog and ctnnb2 mutants among GFP-positive F(1) offspring. Notably, most of these maternal mutants displayed either sgRNA site–spanning genomic deletions or unintended large deletions extending distantly from the sgRNA targets, suggesting a prominent deletion-prone tendency of genome editing in the oocyte. Thus, our method allows maternal gene knockout in the absence of viable and fertile homozygous mutant adults. This approach is particularly time-saving and can be applied for functional screening of maternal factors and generating genomic deletions in zebrafish. American Association for the Advancement of Science 2021-08-06 /pmc/articles/PMC8346210/ /pubmed/34362733 http://dx.doi.org/10.1126/sciadv.abg4243 Text en Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. Distributed under a Creative Commons Attribution NonCommercial License 4.0 (CC BY-NC). https://creativecommons.org/licenses/by-nc/4.0/This is an open-access article distributed under the terms of the Creative Commons Attribution-NonCommercial license (https://creativecommons.org/licenses/by-nc/4.0/) , which permits use, distribution, and reproduction in any medium, so long as the resultant use is not for commercial advantage and provided the original work is properly cited. |
spellingShingle | Research Articles Zhang, Chong Lu, Tong Zhang, Yizhuang Li, Jiaguang Tarique, Imran Wen, Fenfen Chen, Aijun Wang, Jiasheng Zhang, Zhuoyu Zhang, Yanjun Shi, De-Li Shao, Ming Rapid generation of maternal mutants via oocyte transgenic expression of CRISPR-Cas9 and sgRNAs in zebrafish |
title | Rapid generation of maternal mutants via oocyte transgenic expression of CRISPR-Cas9 and sgRNAs in zebrafish |
title_full | Rapid generation of maternal mutants via oocyte transgenic expression of CRISPR-Cas9 and sgRNAs in zebrafish |
title_fullStr | Rapid generation of maternal mutants via oocyte transgenic expression of CRISPR-Cas9 and sgRNAs in zebrafish |
title_full_unstemmed | Rapid generation of maternal mutants via oocyte transgenic expression of CRISPR-Cas9 and sgRNAs in zebrafish |
title_short | Rapid generation of maternal mutants via oocyte transgenic expression of CRISPR-Cas9 and sgRNAs in zebrafish |
title_sort | rapid generation of maternal mutants via oocyte transgenic expression of crispr-cas9 and sgrnas in zebrafish |
topic | Research Articles |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8346210/ https://www.ncbi.nlm.nih.gov/pubmed/34362733 http://dx.doi.org/10.1126/sciadv.abg4243 |
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