PUBLICATION

Cellular differentiation in primary cell cultures from single zebrafish embryos as a model for the study of myogenesis

Authors
Myhre, J.L., and Pilgrim, D.B.
ID
ZDB-PUB-101018-18
Date
2010
Source
Zebrafish   7(3): 255-266 (Journal)
Registered Authors
Myhre, Layne, Pilgrim, David
Keywords
none
MeSH Terms
  • Zebrafish/embryology*
  • Zebrafish/genetics
  • Zebrafish/metabolism
  • Animals
  • Gene Expression Regulation, Developmental
  • Heart/embryology*
  • Cell Differentiation*
  • Myocardium/cytology*
  • Myocardium/metabolism
  • Models, Biological*
  • Cell Culture Techniques/methods*
  • Muscle Development*
(all 12)
PubMed
20936983 Full text @ Zebrafish
Abstract
Culturing cells in vitro can produce a uniform population for the study of cellular differentiation, which is especially useful for the quantification of gene expression or the observation of subcellular structures. In zebrafish, a handful of immortalized cell lines have been used for these purposes, despite being heavily selected by passaging. Methods for primary cell culture of zebrafish embryonic blastomeres have been previously reported, but require combining a large number of genetically heterogeneous embryos, meaning that subsequent cell cultures are not clonal. Without genetically uniform cultures, this model system cannot exploit the wealth of available embryonic lethal mutants in zebrafish. We therefore describe methods for the generation of zebrafish embryonic blastomere cell cultures from single genetically characterized embryos. We examined myogenic differentiation and gene expression in single-embryo cultures from early wild-type embryos, as well as embryos containing an embryonic lethal mutation of unc45b, a myosin chaperone known to be required for sarcomere organization during myogenesis. We also demonstrated the practical usefulness of this technique by experimentally manipulating expression of specific genes in individual embryos before cell culture using standard tools of zebrafish biology such as morpholino-oligonucleotide gene knockdown and transgene-mediated gene expression.
Genes / Markers
Figures
Figure Gallery (7 images)
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Expression
No data available
Phenotype
No data available
Mutations / Transgenics
Allele Construct Type Affected Genomic Region
sb60
    Point Mutation
    ua1TgTransgenic Insertion
      y1TgTransgenic Insertion
        1 - 3 of 3
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        Human Disease / Model
        No data available
        Sequence Targeting Reagents
        Target Reagent Reagent Type
        tp53MO4-tp53MRPHLNO
        unc45bMO1-unc45bMRPHLNO
        1 - 2 of 2
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        Fish
        1 - 3 of 3
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        Antibodies
        Orthology
        No data available
        Engineered Foreign Genes
        Marker Marker Type Name
        EGFPEFGEGFP
        1 - 1 of 1
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        Mapping
        No data available