bioRxivCell BiologyPreprint
Collagen assembly and turnover imaged with a CRISPR-Cas9 engineered Dendra2 tag
Adam PickardiD, Antony AdamsoniD, Yinhui Lu, Joan ChangiD, Richa GarvaiD, Nigel HodsoniD, Karl E. KadleriD
Abstract
Electron microscopy has been the \"gold standard\" for studying collagen networks but dynamic information on how cells synthesise the networks has been lacking. Live imaging methods have been unable to distinguish newly-synthesised fibrils from pre-existing fibrils and intracellular collagen. Here, we tagged endogenous collagen-I using CRISPR-Cas9 with photoswitchable Dendra2 and demonstrate live cells synthesising, migrating on, and interacting with, collagen fibrils. This strategy is applicable for other long half-life proteins.
- organism: human
- assay: imaging
- assay: fluorescence / live-cell microscopy
- research focus: cell signaling & trafficking
- research focus: cell type annotation
- approach: experimental assay
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