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Characterization of a cell line derived from zebrafish (brachydanio rerio) embryos

  • Genetics
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Summary

During the last decade, zebrafish (Brachydanio rerio) have emerged as a novel and attractive system to study embryogenesis and organogenesis in vertebrates. The main reason is that both extensive genetic studies and detailed embryologic analysis are possible using this small tropical fresh water teleost. However, in vitro analysis using cell culture or molecular genetics are still far less advanced than in other vertebrate systems. Here we report the generation and characterization of a fibroblast like cell line, ZF4, derived from 1-day-old zebrafish embryos. The hyperploid cell line has been stable in multiple passages for more than 2 yr now and is the first zebrafish cell line that can be maintained in conventional medium containing mammalian serum. Using a series of plasmids for expression of a marker gene, we evaluate in ZF4 cells the relative strength of expression from several different viral, fish, and mammalian promoters. Stable integration can be obtained by using G418 selection. We hope that our cell line will be a useful tool for the analysis of gene regulation in zebrafish.

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References

  • Bayer, T. A.; Campos-Ortega, J. A. A transgene containing lacZ is expressed in primary sensory neurons in zebrafish. Development 115:421–426; 1992.

    PubMed  CAS  Google Scholar 

  • Bresch, H.; Beck, H.; Ehlermann, D., et al. A long-term toxicity test comprising reproduction and growth of zebrafish under 4-chloroaniline. Arch. Environ. Contam. Toxicol. 19:418–427; 1990.

    Article  Google Scholar 

  • Buono, R. J.; Linser, P. J. Transient expression of RSVCAT in transgenic zebrafish made by electroporation. Mol. Marine Biol. Biotechnol. 1:266–270; 1992.

    Google Scholar 

  • Collodi, P.; Barnes, D. W. Mitogenic activity from trout embryos. Proc. Natl. Acad. Sci. USA 87:3498–3502; 1990.

    Article  PubMed  CAS  Google Scholar 

  • Collodi, P.; Kamei, Y.; Ernst, T., et al. Culture of cells from zebrafish (Brachydanio rerio) embryo and adult tissue. Cell Biol. Toxicol. 8:43–61; 1992a.

    Article  PubMed  CAS  Google Scholar 

  • Collodi, P.; Kamei, Y.; Sharps, A., et al. Fish embryo cell cultures for derivation of stem cells and transgenic chimeras. Mol. Marine Biol. Biotechnol. 1:257–265; 1992b.

    PubMed  CAS  Google Scholar 

  • Culp, P.; Nüsslein-Volhard, C.; Hopkins, N. High-frequency germ-line transmission of plasmid DNA sequences injected into fertilized zebrafish eggs. Proc. Natl. Acad. Sci. USA 88:7953–7957; 1991.

    Article  PubMed  CAS  Google Scholar 

  • Edlund, T.; Walker, M. D.; Barr, P. J., et al. Cell specific expression of the rat insulin gene: evidence for role of two distinct 5′ flanking elements. Science 230:912–916; 1985.

    Article  PubMed  CAS  Google Scholar 

  • Ekker, N. M.; Akimenko, M.-A.; Bremiller, R., et al. Regional expression of three homeobox transcripts in the inner ear of the zebrafish embryo. Neuron 9:27–35; 1992.

    Article  PubMed  CAS  Google Scholar 

  • Endo, A.; Ingalls, T. Chromosomes of the zebrafish. J. Hered. 59:382–384; 1968.

    PubMed  CAS  Google Scholar 

  • Evans, M. H.; Kaufman, M. H. Establishment in culture of pluripotent cells from mouse embryos. Nature 292:154–156; 1981.

    Article  PubMed  CAS  Google Scholar 

  • Friedenreich, H.; Schartl, M. Transient expression directed by homologous and heterologous promoter and enhancer sequences in fish cells. Nucleic Acids Res. 18:3299–3305; 1990.

    Article  PubMed  CAS  Google Scholar 

  • Friedrich, G.; Soriano, P. Promoter traps in embryonic stem cells: a genetic screen to identify and mutate developmental genes in mice. Genes & Dev. 5:1513–1523; 1991.

    CAS  Google Scholar 

  • Gatchalian, C. L.; Eisen, J. S. Pathway selection by ectopic motoneurons in embryonic zebrafish. Neuron 9:105–112; 1992.

    Article  PubMed  CAS  Google Scholar 

  • Gillis, D. M.; Kramer, D. Ideal interference distributions: population density and patch use by zebrafish. Anim. Behav. 35:1875–1882; 1987.

    Article  Google Scholar 

  • Grunwald, D. J.; Streisinger, G. Induction of recessive lethal and specific locus mutations in the zebrafish with ethyl nitrosourea. Genet. Res. 59:103–116; 1992.

    Article  PubMed  CAS  Google Scholar 

  • Gruss, P.; Khoury, G. Rescue of a splicing defective mutant by insertion of an heterologous intron. Nature 286:634–637; 1980.

    Article  PubMed  CAS  Google Scholar 

  • Hatta, K.; Kimmel, C. B.; Ho, R. K., et al. The cyclops mutation blocks specification of the floor plate of the zebrafish central nervous system. Nature 350:339–341; 1991.

    Article  PubMed  CAS  Google Scholar 

  • Hightower, L. E.; Renfro, J. L. Recent applications of fish cell culture in biomedical research. J. Exp. Zool. 248:290–302; 1988.

    Article  PubMed  CAS  Google Scholar 

  • Ho, R. H.; Kane, D. Cell-autonomous action of zebrafishspt-1 mutation in specific mesodermal precursors. Nature 348:728–730; 1990.

    Article  PubMed  CAS  Google Scholar 

  • Inoue, K. Expression of reporter genes introduced by microinjection and electroporation in fish embryos and fry. Mol. Marine Biol. Biotechnol. 1:266–270; 1992.

    PubMed  CAS  Google Scholar 

  • Kimmel, C. B.; Warga, R. M. Tissue specific cell lineages originate in the gastrula of the zebrafish. Science 231:365–368; 1986.

    Article  PubMed  Google Scholar 

  • Kimmel, C. B.; Kane, D. A.; Walker, C., et al. A mutation that changes cell movement and cell fate in the zebrafish embryo. Nature 337:358–362; 1989.

    Article  PubMed  CAS  Google Scholar 

  • Kirschbaum, F. Untersuchungen über das Farbmuster der ZebrabarbeBrachydanio rerio (Cyprinidae, Teleostei). Wilhelm Roux Arch. 177:129–152; 1975.

    Article  Google Scholar 

  • Krauss, S.; Maden, M.; Holder, N., et al. Zebrafish pax[b] is involved in the formation of the midbrain-hindbrain boundary. Nature 360:87–89; 1992.

    Article  PubMed  CAS  Google Scholar 

  • Lin, S.; Long, W.; Chen, J., et al. Production of germ line chimeras in zebrafish by cell transplants from genetically pigmented to albino embryos. Proc. Natl. Acad. Sci. USA 89:4519–4523; 1992.

    Article  PubMed  CAS  Google Scholar 

  • Lindl, T.; Bauer, J. Zell- and Gewebekultur. Stuttgart/New York: Gustav Fischer Verlag; 1989.

    Google Scholar 

  • Liu, Z.; Moav, B.; Faras, A. J., et al. Development of expression vectors for transgenic fish. Bio/Technology 8:1268–1272; 1990.

    Article  PubMed  CAS  Google Scholar 

  • Liu, Z.; Zhu, Z.; Roberg, K., et al. Isolation and characterization of theβ-actin gene of carp (Cyprinus carpio). DNA Sequence J. 1:125–136; 1990.

    CAS  Google Scholar 

  • MacGregor, G. R.; Caskey, C. T. Construction of plasmids that expressE. coli β-galactosidase in mammalian cells. Nucleic Acids. Res. 17:2365; 1989.

    Article  PubMed  CAS  Google Scholar 

  • Maniatis, T.; Fritsch, E. F.; Sambrook, J. Molecular cloning, a laboratory manual. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press; 1989.

    Google Scholar 

  • Marcey, D.; Nüsslein-Volhard, C. Embryology goes fishing. Nature 321:380–381; 1986.

    Article  Google Scholar 

  • Miller, J. Experiments in molecular genetics. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory Press; 1972.

    Google Scholar 

  • Moav, B.; Liu, Z.; Groll, Y., et al. Selection of promoters for gene transfer into fish. Mol. Marine Biol. Biotechnol. 1:338–345; 1992.

    PubMed  CAS  Google Scholar 

  • Molven, A.; Njolstad, P. R.; Fjose, A. Genomic structure and restricted neural expression of the zebrafishwnt-1 (int-1) gene. EMBO J. 10:799–807; 1991.

    PubMed  CAS  Google Scholar 

  • Ono, H.; Klein, D.; Vincek, V., et al. Major histocompatibility complex class II genes of zebrafish. Proc. Natl. Acad. Sci. USA 89:11886–11890; 1992.

    Article  PubMed  CAS  Google Scholar 

  • Ozato, K.; Wakamatsu, Y.; Inoue, K. Medaka as a model of transgenic fish. Mol. Marine Biol. Biotechnol. 1:346–354; 1992.

    PubMed  CAS  Google Scholar 

  • Powers, D.; Hereford, L.; Cole, T., et al. Electroporation: a method for transferring genes into the gametes of zebrafish (Brachydanio rerio), channel catfish (Ictalurus punctatus) and common carp (Cyprinus carpio). Mol. Marine Biol. Biotechnol. 1:301–308; 1992.

    PubMed  CAS  Google Scholar 

  • Roosen-Runge, E. C. Furchung und Primitiventwicklung vonBrachydanio rerio. Anat. Anz. 81:297–301; 1936.

    Google Scholar 

  • Schulte-Merker, S.; Ho, R. K.; Herrmann, B. G., et al. The protein product of the zebrafish homologue of the mouse T gene is expressed in nuclei of the germ ring and the notochord of the early embryo. Development 116:1021–1032; 1992.

    PubMed  CAS  Google Scholar 

  • St. Johnston, D.; Nüsslein-Volhard, C. The origin of pattern and polarity in theDrosophila embryo. Cell 68:201–219; 1992.

    Article  Google Scholar 

  • Streisinger, G. F.; Walker, C.; Dower, D., et al. Production of clones of homozygous diploid zebrafish (Brachydanio rerio). Nature 291:293–296; 1981.

    Article  PubMed  CAS  Google Scholar 

  • Stuart, G. W.; McMurray, J. V.; Westerfield, M. Replication, integration, and stable germ line transmission of foreign sequences injected into early zebrafish embryos. Development 103:403–412; 1988.

    PubMed  CAS  Google Scholar 

  • Stuart, G. W.; Vielkind, J. R.; McMurray, J. V., et al. Stable lines of transgenic zebrafish exhibit reproducible patterns of transgenic expression. Development 109:577–584; 1990.

    PubMed  CAS  Google Scholar 

  • Westerfield, M. The zebrafish book. Eugene: University of Oregon Press; 1989.

    Google Scholar 

  • Wilkins, A. S. Genetic analysis of animal development. New York: Wiley & Sons; 1993.

    Google Scholar 

  • Winkler, C.; Hong, Y.; Wittbrodt, J., et al. Analysis of heterologous and homologous promoters and enhancers in vitro and in vivo by gene transfer into Japanese Medaka (Oryzias latipes) andXiphophorus. Mol. Marine Biol. Biotechnol. 1:326–337; 1992.

    PubMed  CAS  Google Scholar 

  • Wolf, K.; Quimby, M. C. Established eurythermic line of fish cells in vitro. Science 137:1065–1066; 1962.

    Article  Google Scholar 

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Driever, W., Rangini, Z. Characterization of a cell line derived from zebrafish (brachydanio rerio) embryos. In Vitro Cell Dev Biol - Animal 29, 749–754 (1993). https://doi.org/10.1007/BF02631432

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