Zona pellucida treatment before CRISPR/Cas9‐mediated genome editing of porcine zygotes
Abstract Background Increasing the permeability of the zona pellucida (ZP) of oocytes before CRISPR/Cas9 electroporation may improve the efficiency of gene editing; however, the effects of this approach on subsequent developmental processes are unclear. In this study, the effects of ZP treatment bef...
| Main Authors: | , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Wiley
2022-01-01
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| Series: | Veterinary Medicine and Science |
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| Online Access: | https://doi.org/10.1002/vms3.659 |
| _version_ | 1819243195171602432 |
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| author | Zhao Namula Maki Hirata Quynh Anh Le Qingyi Lin Koki Takebayashi Naoaki Yoshimura Fuminori Tanihara Chommanart Thongkittidilok Takeshige Otoi |
| author_facet | Zhao Namula Maki Hirata Quynh Anh Le Qingyi Lin Koki Takebayashi Naoaki Yoshimura Fuminori Tanihara Chommanart Thongkittidilok Takeshige Otoi |
| author_sort | Zhao Namula |
| collection | DOAJ |
| description | Abstract Background Increasing the permeability of the zona pellucida (ZP) of oocytes before CRISPR/Cas9 electroporation may improve the efficiency of gene editing; however, the effects of this approach on subsequent developmental processes are unclear. In this study, the effects of ZP treatment before electroporation on embryonic development and gene editing in porcine embryos were evaluated. Methods The ZP of zygotes was weakened or removed by exposure to 0.5% actinase E, followed by electroporation of the Cas9 protein with guide RNA targeting GGTA1. Results The blastocyst formation rate of ZP‐free zygotes after electroporation was significantly lower (p < 0.05) than that of ZP‐intact zygotes. The mutation rate in blastocysts from ZP‐weakened zygotes was similar to that in ZP‐intact zygotes, whereas ZP removal increased the mutation rate. The mutation efficiency in blastocysts from electroporated zygotes did not differ among ZP treatment groups. Conclusions Our results indicate that weakening the ZP does not affect the developmental competence, mutation rate, or mutation efficiency of electroporated zygotes, whereas ZP removal has a detrimental effect on embryonic development but may increase the mutation rate. |
| first_indexed | 2024-12-23T14:51:50Z |
| format | Article |
| id | doaj.art-09413eb19a4b49469c97a4af02007d69 |
| institution | Directory Open Access Journal |
| issn | 2053-1095 |
| language | English |
| last_indexed | 2024-12-23T14:51:50Z |
| publishDate | 2022-01-01 |
| publisher | Wiley |
| record_format | Article |
| series | Veterinary Medicine and Science |
| spelling | doaj.art-09413eb19a4b49469c97a4af02007d692022-12-21T17:42:54ZengWileyVeterinary Medicine and Science2053-10952022-01-018116416910.1002/vms3.659Zona pellucida treatment before CRISPR/Cas9‐mediated genome editing of porcine zygotesZhao Namula0Maki Hirata1Quynh Anh Le2Qingyi Lin3Koki Takebayashi4Naoaki Yoshimura5Fuminori Tanihara6Chommanart Thongkittidilok7Takeshige Otoi8Faculty of Veterinary Science Guangdong Ocean University Zhanjiang ChinaFaculty of Bioscience and Bioindustry Tokushima University Tokushima JapanFaculty of Bioscience and Bioindustry Tokushima University Tokushima JapanFaculty of Bioscience and Bioindustry Tokushima University Tokushima JapanFaculty of Bioscience and Bioindustry Tokushima University Tokushima JapanFaculty of Bioscience and Bioindustry Tokushima University Tokushima JapanFaculty of Bioscience and Bioindustry Tokushima University Tokushima JapanAkkhraratchakumari Veterinary College Walailak University Nakorn Sri Thammarat ThailandFaculty of Bioscience and Bioindustry Tokushima University Tokushima JapanAbstract Background Increasing the permeability of the zona pellucida (ZP) of oocytes before CRISPR/Cas9 electroporation may improve the efficiency of gene editing; however, the effects of this approach on subsequent developmental processes are unclear. In this study, the effects of ZP treatment before electroporation on embryonic development and gene editing in porcine embryos were evaluated. Methods The ZP of zygotes was weakened or removed by exposure to 0.5% actinase E, followed by electroporation of the Cas9 protein with guide RNA targeting GGTA1. Results The blastocyst formation rate of ZP‐free zygotes after electroporation was significantly lower (p < 0.05) than that of ZP‐intact zygotes. The mutation rate in blastocysts from ZP‐weakened zygotes was similar to that in ZP‐intact zygotes, whereas ZP removal increased the mutation rate. The mutation efficiency in blastocysts from electroporated zygotes did not differ among ZP treatment groups. Conclusions Our results indicate that weakening the ZP does not affect the developmental competence, mutation rate, or mutation efficiency of electroporated zygotes, whereas ZP removal has a detrimental effect on embryonic development but may increase the mutation rate.https://doi.org/10.1002/vms3.659CRISPRelectroporationgenome editingporcine zygotezona pellucida |
| spellingShingle | Zhao Namula Maki Hirata Quynh Anh Le Qingyi Lin Koki Takebayashi Naoaki Yoshimura Fuminori Tanihara Chommanart Thongkittidilok Takeshige Otoi Zona pellucida treatment before CRISPR/Cas9‐mediated genome editing of porcine zygotes Veterinary Medicine and Science CRISPR electroporation genome editing porcine zygote zona pellucida |
| title | Zona pellucida treatment before CRISPR/Cas9‐mediated genome editing of porcine zygotes |
| title_full | Zona pellucida treatment before CRISPR/Cas9‐mediated genome editing of porcine zygotes |
| title_fullStr | Zona pellucida treatment before CRISPR/Cas9‐mediated genome editing of porcine zygotes |
| title_full_unstemmed | Zona pellucida treatment before CRISPR/Cas9‐mediated genome editing of porcine zygotes |
| title_short | Zona pellucida treatment before CRISPR/Cas9‐mediated genome editing of porcine zygotes |
| title_sort | zona pellucida treatment before crispr cas9 mediated genome editing of porcine zygotes |
| topic | CRISPR electroporation genome editing porcine zygote zona pellucida |
| url | https://doi.org/10.1002/vms3.659 |
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