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CRISPR-Cas9 knockin ratones para la edición del genoma y el modelado del cáncer

Randall J Platt¹, Sidi Chen², Yang Zhou³

¹Broad Institute of MIT and Harvard, Cambridge, MA 02142, USA; McGovern Institute for Brain Research, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Brain and Cognitive Sciences, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Department of Biological Engineering, Massachusetts Institute of Technology, Cambridge, MA 02139, USA.

|September 30, 2014

Resumen

Hemos desarrollado un ratón knockin Cas9 Cre-dependiente para la edición del genoma in vivo. Esta herramienta permite modelar enfermedades complejas como el adenocarcinoma pulmonar mediante la alteración simultánea de genes clave.

Área de la Ciencia:

Genética La genética.
Biología Molecular Biología Molecular
Investigación del cáncer Investigación del cáncer.

Sus antecedentes:

CRISPR-Cas9 es una poderosa herramienta para la investigación genética.
Las aplicaciones in vivo de CRISPR-Cas9 se están expandiendo.
Se necesitan sistemas eficientes de edición del genoma para el modelado de enfermedades.

Objetivo del estudio:

Para crear un versátil modelo de ratón knockin Cas9 dependiente de Cre.
Para demostrar las capacidades de edición del genoma in vivo y ex vivo.
Para modelar el adenocarcinoma pulmonar mediante la orientación de los genes mutados clave.

Principales métodos:

Generación de un ratón knockin Cas9 dependiente del Cre.
Entrega de ARN guía a través de métodos AAV, lentivirus o de partículas.
Modelado simultáneo de las mutaciones genéticas KRAS, p53 y LKB1 in vivo.

Principales resultados:

Edición exitosa del genoma in vivo y ex vivo en varios tipos de células (neuronas, inmunes, endoteliales).
Inducción de mutaciones de pérdida de función en p53 y Lkb1 utilizando la administración de AAV.
Generación de mutaciones Kras ((G12D) a través de la reparación dirigida a la homología, lo que lleva al adenocarcinoma pulmonar.
Demostración del modelado simultáneo de múltiples mutaciones genéticas.

Conclusiones:

Los ratones Cas9 desarrollados facilitan amplias aplicaciones de edición del genoma in vivo.
Esta tecnología permite el modelado rápido y preciso de enfermedades genéticas complejas.
Los ratones Cas9 son herramientas valiosas para la investigación biológica y el modelado de enfermedades.

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