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 β-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 β-tubulin and r-opsin genes from injected larvae identified various genomic long-fragment deletions. In situ hybridization showed gene expression patterns of β-tubulin and r-opsin were significantly altered in the mosaic mutants. Knocking out β-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.

Ruohui Li

Yue Xu

Fucun Wu

Zhangjie Peng

Jiulin Chan

Linlin Zhang

The CRISPR Journal

为表彰这一非凡的科技时代，MARY ANN LIEBERT, INC. 的出版商最近宣布创建 THE CRISPR JOURNAL——这是一份国际性、多学科的同行评审期刊，发表关于 CRISPR 无数应用和基础技术的杰出研究。 2018 年，CRISPR 期刊将以灵活的开放获取选项在线和印刷出版，为开创性研究以及生动和挑衅性的评论、分析和辩论提供一个备受瞩目的场所。 CRISPR 期刊为 MARY ANN LIEBERT, INC. 的产品组合增加了一个令人兴奋和充满活力的组成部分，其中包括 GEN（基因工程和生物技术新闻）和 80 多个领先的同行评审期刊。

To celebrate this extraordinary era of technology, the publisher MARY ANN LIEBERT, INC. recently announced the creation of The CRISPR Journal. This is an international, multidisciplinary peer-reviewed journal dedicated to publishing outstanding research on the numerous applications and foundational technologies of CRISPR. Starting in 2018, The CRISPR Journal will be published online and in print with flexible open access options, providing a prominent venue for groundbreaking research as well as vibrant and provocative commentary, analysis, and debate. The CRISPR Journal adds an exciting and dynamic component to the portfolio of MARY ANN LIEBERT, INC., which includes GEN (Genetic Engineering and Biotechnology News) and over 80 leading peer-reviewed journals.

为了庆祝这一非凡的科技时代，MARY ANN LIEBERT, INC. 的出版商最近宣布创办《CRISPR Journal》。这是一份国际性、多学科的同行评审期刊，专注于发表关于CRISPR技术及其众多应用的杰出研究。自2018年起，《CRISPR Journal》将以灵活的开放获取选项在线和印刷出版，为开创性研究以及生动且富有挑战性的评论、分析和辩论提供一个备受瞩目的平台。《CRISPR Journal》为MARY ANN LIEBERT, INC.的产品组合增添了一个令人兴奋和充满活力的组成部分，涵盖了GEN（基因工程与生物技术新闻）及80多个领先的同行评审期刊。

Category	Quartile
生物学	4
生物学, 遗传学	4

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

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Journal Info

Category	Quartile
GENETICS & HEREDITY	2