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Updated: Jun 5, 2025
Efficient Production and Identification of CRISPR/Cas9-generated Gene Knockouts in the Model System Danio rerio
Published on: August 28, 2018
Protocol for generating a pericyte reporter zebrafish line Ki(pdgfrb-P2A-GAL4-VP16) using a CRISPR-Cas9-mediated
Huaxing Zi1, Xiaolan Peng2, Jiulin Du3
1Institute of Neuroscience, State Key Laboratory of Neuroscience, CAS Center for Excellence in Brain Science and Intelligence Technology, Chinese Academy of Sciences, 320 Yue-Yang Road, Shanghai 200031, China; University of Chinese Academy of Sciences, 19A Yu-Quan Road, Beijing 100049, China; Dongguan Innovation Institute, Guangdong Medical University, 1 Xin-Cheng Road, Dongguan, Guangdong 523808, China.
Abstract:
Pericytes, the mural cells that envelop small blood vessels, play crucial roles in the formation of the blood-brain barrier (BBB). Here, we present a protocol for generating a pericyte reporter zebrafish line Ki(pdgfrb-P2A-GAL4-VP16) using a CRISPR-Cas9-mediated knockin technique. We describe steps for identifying efficient single guide RNA (sgRNA), constructing donor plasmid, and generating and maintaining the knockin line. We then detail procedures for in vivo imaging of brain pericytes. This protocol is adaptable for creating other knockin lines for specific cell labeling. For complete details on the use and execution of this protocol, please refer to Zi et al.1.
