title: TALEN- and CRISPR-enhanced DNA homologous recombination for gene editing in zebrafish.
abstract: The TALE nuclease and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated systems (Cas) have been developed as important tools for genome editing in zebrafish. Here we describe a CRISPR/Cas9-based approach for generating site-specific gene targeting in zebrafish using DNA double-strand breaks induced homologous recombination (HR)-dependent repair mechanism. Through comicroinjection of Cas9 mRNA, guide RNA targeting genomic DNA sequence corresponding to the twist2 gene and the corresponding double-strand long arm donor template with a point mutation identified in human, HR-mediated knock-in of the expected targeting sequence was obtained. To facilitate identification of germline transmission of targeted mutation, a method of screening sperms of male founder fish is designed.
authors.affiliation: Peking University Shenzhen Graduate School, Shenzhen, China; University of California Los Angeles, Los Angeles, CA, United States.
authors.forename: Y
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: University of California Los Angeles, Los Angeles, CA, United States.
authors.forename: H
authors.initials: H
authors.lastname: Huang
authors.affiliation: Peking University, Beijing, People's Republic of China.
authors.forename: B
authors.initials: B
authors.lastname: Zhang
authors.affiliation: University of California Los Angeles, Los Angeles, CA, United States.
authors.forename: S
authors.initials: S
authors.lastname: Lin
mesh_terms: D000818:Animals; D064113:CRISPR-Cas Systems / Q000235:genetics*; D004260:DNA Repair / Q000235:genetics; D000072669:Gene Editing / Q000379:methods*; D018390:Gene Targeting / Q000379:methods*; D059765:Homologous Recombination / Q000235:genetics*; D016297:Mutagenesis, Site-Directed / Q000379:methods; D000069896:Transcription Activator-Like Effector Nucleases / Q000235:genetics; D015027:Zebrafish / Q000235:genetics
chemical_list: D000069896:Transcription Activator-Like Effector Nucleases
keywords: CRISPR/Cas9; DNA double-strand breaks; Homologous recombination; In vitro fertilization; Twist2; Zebrafish