Abstract
As an integral part of modern cell biology, fluorescence microscopy enables quantification of the stability and dynamics of fluorescence-labeled biomolecules inside cultured cells. However, obtaining time-resolved data from individual cells within a live vertebrate organism remains challenging. Here we demonstrate a customized pipeline that integrates meganuclease-mediated mosaic transformation with fluorescence-detected temperature-jump microscopy to probe dynamics and stability of endogenously expressed proteins in different tissues of living multicellular organisms.
Publication types
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Research Support, Non-U.S. Gov't
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Research Support, U.S. Gov't, Non-P.H.S.
MeSH terms
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Animals
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Cell Line, Tumor
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Embryo, Nonmammalian
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Endodeoxyribonucleases / metabolism
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Fluorescence Resonance Energy Transfer / instrumentation
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Fluorescence Resonance Energy Transfer / methods
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Fungal Proteins / chemistry
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Fungal Proteins / genetics
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Fungal Proteins / metabolism*
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Humans
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Intravital Microscopy / instrumentation
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Intravital Microscopy / methods*
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Kinetics
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Microscopy, Fluorescence / instrumentation
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Microscopy, Fluorescence / methods
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Phosphoglycerate Kinase / chemistry
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Phosphoglycerate Kinase / genetics
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Phosphoglycerate Kinase / metabolism*
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Protein Folding
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Protein Stability
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Recombinant Fusion Proteins / chemistry
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Recombinant Fusion Proteins / genetics
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Recombinant Fusion Proteins / metabolism
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Temperature
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Zebrafish
Substances
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Fungal Proteins
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Recombinant Fusion Proteins
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Phosphoglycerate Kinase
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Endodeoxyribonucleases