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Easy-to-Use CRISPR-Cas9 Genome Editing in the Cultured Pacific Abalone (Haliotis discus hannai)

文献类型：期刊论文

作者Li, Ruohui4,5,6,7; Xu, Yue5,6,7; Wu, Fucun1,2,4,5,6,7; Peng, Zhangjie4,5,6,7; Chan, Jiulin5,6,7; Zhang, Linlin1,2,3,4,5,6,7
刊名CRISPR JOURNAL
出版日期2024-02-01
卷号7期号:1页码:41-52
ISSN号2573-1599
DOI10.1089/crispr.2023.0070
通讯作者Wu, Fucun(wufucun@qdio.ac.cn) ; Zhang, Linlin(linlinzhang@qdio.ac.cn)
英文摘要The Pacific abalone is an important aquaculture shellfish and serves as an important model in basic biology study. However, the study of abalone is limited by lack of highly efficient and easy-to-use gene-editing tools. In this paper, we demonstrate efficient gene knockout in Pacific abalone using CRISPR-Cas9. We developed a highly effective microinjection method by nesting fertilized eggs in a low-concentration agarose gel. We identified the cilia developmental gene beta-tubulin and light-sensitive transmembrane protein r-opsin as target genes and designed highly specific sgRNAs for modifying their genomic sequences. Sanger sequencing of the genomic regions of beta-tubulin and r-opsin genes from injected larvae identified various genomic long-fragment deletions. In situ hybridization showed gene expression patterns of beta-tubulin and r-opsin were significantly altered in the mosaic mutants. Knocking out beta-tubulin in abalone embryos efficiently affected cilia development. Scanning electron microscopy and swimming behavior assay showed defecting cilia and decreased motility. Moreover, knocking out of r-opsin in abalone embryos effectively affected the expression and development of eyespots. Overall, this work developed an easy-to-use mosaic gene knockout protocol for abalone, which will allow researchers to utilize CRISPR-Cas9 approaches to study unexploited abalone biology and will lead to novel breeding methods for this aquaculture species.
WOS关键词CRISPR/CAS9 ; SPERM
资助项目National Key R&D Program of China[2022YFD2401400] ; Strategic Priority Research Program of the Chinese Academy of Sciences[XDB42000000] ; Marine S&T Fund of Shandong Province for Pilot National Laboratory for Marine Science and Technology (Qingdao)[2022QNLM030004] ; Key Research and Development Program of Shandong[2022LZGC015]
WOS研究方向Genetics & Heredity
语种英语
WOS记录号WOS:001162177200004
出版者MARY ANN LIEBERT, INC
源URL[http://ir.qdio.ac.cn/handle/337002/185106]  
专题海洋研究所_实验海洋生物学重点实验室
海洋研究所_深海极端环境与生命过程研究中心
通讯作者Wu, Fucun; Zhang, Linlin
作者单位1.Chinese Acad Sci, Inst Oceanol, Ctr Deep Sea Res, 7 Naihai Rd, Qingdao 266071, Peoples R China
2.Chinese Acad Sci, Inst Oceanol, CAS & Shandong Prov Key Lab Expt Marine Biol, 7 Naihai Rd, Qingdao 266071, Peoples R China
3.Chinese Acad Sci, Key Lab Breeding Biotechnol & Sustainable Aquacult, Wuhan, Peoples R China
4.Univ Chinese Acad Sci, Coll Marine Sci, Beijing, Peoples R China
5.Qingdao Natl Lab Marine Sci & Technol, Lab Marine Biol & Biotechnol, Qingdao, Peoples R China
6.Chinese Acad Sci, Inst Oceanol, Ctr Deep Sea Res, Qingdao, Peoples R China
7.Chinese Acad Sci, Inst Oceanol, CAS & Shandong Prov Key Lab Expt Marine Biol, Qingdao, Peoples R China
推荐引用方式
GB/T 7714		 Li, Ruohui,Xu, Yue,Wu, Fucun,et al. Easy-to-Use CRISPR-Cas9 Genome Editing in the Cultured Pacific Abalone (Haliotis discus hannai)[J]. CRISPR JOURNAL,2024,7(1):41-52.
APA Li, Ruohui,Xu, Yue,Wu, Fucun,Peng, Zhangjie,Chan, Jiulin,&Zhang, Linlin.(2024).Easy-to-Use CRISPR-Cas9 Genome Editing in the Cultured Pacific Abalone (Haliotis discus hannai).CRISPR JOURNAL,7(1),41-52.
MLA Li, Ruohui,et al."Easy-to-Use CRISPR-Cas9 Genome Editing in the Cultured Pacific Abalone (Haliotis discus hannai)".CRISPR JOURNAL 7.1(2024):41-52.

入库方式： OAI收割

来源：海洋研究所

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