PUBLICATION

An Efficient Vector-Based CRISPR/Cas9 System in Zebrafish Cell Line

Authors
Ye, X., Lin, J., Chen, Q., Lv, J., Liu, C., Wang, Y., Wang, S., Wen, X., Lin, F.
ID
ZDB-PUB-240424-6
Date
2024
Source
Marine biotechnology (New York, N.Y.)   26(3): 588-598 (Journal)
Registered Authors
Keywords
CRISPR/Cas9, Fish cell, U6 promoter, Zebrafish
MeSH Terms
  • Animals
  • Cell Line
  • Zebrafish*/genetics
  • Monophenol Monooxygenase/genetics
  • Monophenol Monooxygenase/metabolism
  • Zebrafish Proteins/genetics
  • Zebrafish Proteins/metabolism
  • Humans
  • Promoter Regions, Genetic*
  • Gene Editing*/methods
  • Genetic Vectors*
  • CRISPR-Cas Systems*
  • HEK293 Cells
(all 13)
PubMed
38652190 Full text @ Mar. Biotechnol.
Abstract
The clustered regularly interspaced short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) system has been widely applied in animals as an efficient genome editing tool. However, the technique is difficult to implement in fish cell lines partially due to the lack of efficient promoters to drive the expression of both sgRNA and the Cas9 protein within a single vector. In this study, it was indicated that the zebrafish U6 RNA polymerase III (ZFU6) promoter could efficiently induce tyrosinase (tyr) gene editing and lead to loss of retinal pigments when co-injection with Cas9 mRNA in zebrafish embryo. Furthermore, an optimized all-in-one vector for expression of the CRISPR/Cas9 system in the zebrafish fibroblast cell line (PAC2) was constructed by replacing the human U6 promoter with ZFU6 promoter, basing on the lentiCRISPRV2 system that widely applied in mammal cells. This new vector could successfully target the cellular communication network factor 2a (ctgfa) gene and demonstrated its function in the PAC2 cell. Notably, the vector could also be used to edit the endogenous EMX1 gene in the mammal 293 T cell line, implying its wide application potential. In conclusion, we established a new gene editing tool for zebrafish cell line, which could be a useful in vitro platform for high-throughput analyzing gene function in fish.
Genes / Markers
Figures
Figure Gallery (6 images)
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Expression
Phenotype
No data available
Mutations / Transgenics
No data available
Human Disease / Model
No data available
Sequence Targeting Reagents
Target Reagent Reagent Type
ccn2aCRISPR9-ccn2aCRISPR
tyrCRISPR1-tyrCRISPR
1 - 2 of 2
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Fish
No data available
Antibodies
No data available
Orthology
No data available
Engineered Foreign Genes
No data available
Mapping
No data available
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Citation
Ye, X., Lin, J., Chen, Q., Lv, J., Liu, C., Wang, Y., Wang, S., Wen, X., Lin, F. (2024) An Efficient Vector-Based CRISPR/Cas9 System in Zebrafish Cell Line. Marine biotechnology (New York, N.Y.). 26(3):588-598.