title: Germline-specific dgcr8 knockout in zebrafish using a BACK approach.
abstract: Zebrafish is an important model to study developmental biology and human diseases. However, an effective approach to achieve spatial and temporal gene knockout in zebrafish has not been well established. In this study, we have developed a new approach, namely bacterial artificial chromosome-rescue-based knockout (BACK), to achieve conditional gene knockout in zebrafish using the Cre/loxP system. We have successfully deleted the DiGeorge syndrome critical region gene 8 (dgcr8) in zebrafish germ line and demonstrated that the maternal-zygotic dgcr8 (MZdgcr8) embryos exhibit MZdicer-like phenotypes with morphological defects which could be rescued by miR-430, indicating that canonical microRNAs play critical role in early development. Our findings establish that Cre/loxP-mediated tissue-specific gene knockout could be achieved using this BACK strategy and that canonical microRNAs play important roles in early embryonic development in zebrafish.
authors.affiliation: State Key Laboratory of Biocontrol, Institute of Aquatic Economic Animals, and the Guangdong Province Key Laboratory for Aquatic Economic Animals, Sun Yat-Sen University, Guangzhou, China.|School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong, China.
authors.forename: Yun
authors.initials: Y
authors.lastname: Liu
authors.affiliation: School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong, China.|School of Biomedical Sciences Core Laboratory, The Chinese University of Hong Kong Shenzhen Research Institute, Shenzhen, China.
authors.forename: Zeyao
authors.initials: Z
authors.lastname: Zhu
authors.affiliation: School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong, China.|Department of Anaesthesia and Intensive Care, Prince of Wales Hospital, The Chinese University of Hong Kong, Hong Kong, China.|School of Biomedical Sciences Core Laboratory, The Chinese University of Hong Kong Shenzhen Research Institute, Shenzhen, China.
authors.forename: Idy H T
authors.initials: IHT
authors.lastname: Ho
authors.affiliation: School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong, China.|School of Biomedical Sciences Core Laboratory, The Chinese University of Hong Kong Shenzhen Research Institute, Shenzhen, China.
authors.forename: Yujian
authors.initials: Y
authors.lastname: Shi
authors.affiliation: School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong, China.|School of Biomedical Sciences Core Laboratory, The Chinese University of Hong Kong Shenzhen Research Institute, Shenzhen, China.
authors.forename: Yuxin
authors.initials: Y
authors.lastname: Xie
authors.affiliation: School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong, China.|School of Biomedical Sciences Core Laboratory, The Chinese University of Hong Kong Shenzhen Research Institute, Shenzhen, China.
authors.forename: Jianzhen
authors.initials: J
authors.lastname: Li
authors.affiliation: State Key Laboratory of Biocontrol, Institute of Aquatic Economic Animals, and the Guangdong Province Key Laboratory for Aquatic Economic Animals, Sun Yat-Sen University, Guangzhou, China.|School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong, China.|South China Sea Bio-Resource Exploitation and Utilization Collaborative Innovation Center, Guangzhou, China.
authors.forename: Yong
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Department of Anaesthesia and Intensive Care, Prince of Wales Hospital, The Chinese University of Hong Kong, Hong Kong, China.
authors.forename: Matthew T V
authors.initials: MTV
authors.lastname: Chan
authors.affiliation: School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong, China. chkcheng@cuhk.edu.hk.|School of Biomedical Sciences Core Laboratory, The Chinese University of Hong Kong Shenzhen Research Institute, Shenzhen, China. chkcheng@cuhk.edu.hk.
authors.forename: Christopher H K
authors.initials: CHK
authors.lastname: Cheng
mesh_terms: D000818:Animals; D001483:Base Sequence; D022202:Chromosomes, Artificial, Bacterial / Q000235:genetics*; D047108:Embryonic Development / Q000235:genetics; D005091:Exons / Q000235:genetics; D017353:Gene Deletion; D018507:Gene Expression Regulation, Developmental; D055786:Gene Knockout Techniques / Q000379:methods*; D018390:Gene Targeting; D005854:Germ Cells / Q000378:metabolism*; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D012323:RNA Processing, Post-Transcriptional / Q000235:genetics; D000069896:Transcription Activator-Like Effector Nucleases / Q000378:metabolism; D015027:Zebrafish / Q000196:embryology / Q000235:genetics*; D029961:Zebrafish Proteins / Q000235:genetics* / Q000378:metabolism
chemical_list: D035683:MicroRNAs; D029961:Zebrafish Proteins; C000622128:dgcr8 protein, zebrafish; D000069896:Transcription Activator-Like Effector Nucleases
keywords: Crispr-cas9; Germ layer specification; Maternal-zygotic transition; Small regulatory RNAs