Engineering the Spatiotemporal Mosaic Self-Patterning of Pluripotent Stem Cells.
Methods Mol Biol
; 2258: 105-116, 2021.
Article
en En
| MEDLINE
| ID: mdl-33340357
ABSTRACT
Pluripotent stem cells (PSCs) possess the ability to self-organize into complex tissue-like structures; however, the genetic mechanisms and multicellular dynamics that direct such patterning are difficult to control. Here, we pair live imaging with controlled induction of gene knockdown by CRISPR interference (CRISPRi) to generate changes within subpopulations of human PSCs, allowing for control over organization and analysis of emergent behaviors. Specifically, we use forced aggregation of mixtures of cells with and without an inducible CRISPRi system to knockdown molecular regulators of tissue symmetry. We then track the resulting multicellular organization through fluorescence live imaging concurrent with the induction of knockdown. Overall, this technique allows for controlled initiation of symmetry breaking by CRISPRi to produce changes in cellular behavior that can be tracked over time within high-density pluripotent stem cell colonies.
Palabras clave
Texto completo:
1
Bases de datos:
MEDLINE
Asunto principal:
Tipificación del Cuerpo
/
Células Madre Pluripotentes
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Sistemas CRISPR-Cas
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Edición Génica
Idioma:
En
Revista:
Methods Mol Biol
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
2021
Tipo del documento:
Article
País de afiliación:
Estados Unidos