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El objetivo del ARN con CRISPR-Cas13

Omar O Abudayyeh^1,2,3,4,5, Jonathan S Gootenberg^1,2,3,4,6, Patrick Essletzbichler^1,2,3,4

¹Broad Institute of MIT and Harvard, Cambridge, Massachusetts 02142, USA.

|October 5, 2017

Resumen

Los científicos diseñaron CRISPR-Cas13a para una manipulación precisa del ARN en las células de los mamíferos. Esta nueva herramienta ofrece un knockdown de ARN dirigido y un seguimiento de transcripción de células vivas con una especificidad mejorada en comparación con los métodos existentes.

Área de la Ciencia:

Biología molecular
La genética
Biotecnología

Sus antecedentes:

Las herramientas moleculares para la manipulación y medición del ARN son limitadas.
Los métodos actuales como la interferencia de ARN tienen efectos fuera del objetivo.
La visualización del ARN a menudo requiere etiquetas exógenas.

Objetivo del estudio:

Diseñar el sistema CRISPR-Cas13a para el derribo y la unión de ARN en células de mamíferos.
Evaluar la eficacia y la especificidad de LwaCas13a para la orientación del ARN.
Desarrollar una plataforma programable para el estudio del ARN en células vivas.

Principales métodos:

Se examinaron 15 ortólogos de Cas13a para identificar la variante más efectiva (LwaCas13a).
Se expresa heterológicamente en las células de mamíferos y plantas.
Utilizó LwaCas13a catalíticamente inactivo para la unión y el seguimiento del ARN.

Principales resultados:

LwaCas13a demostró una eliminación eficaz de las transcripciones reporteras y endógenas.
Se alcanzan niveles de eliminación comparables a la interferencia de ARN pero con una especificidad mejorada.
Se ha demostrado el seguimiento programable de transcripciones en células vivas utilizando LwaCas13a inactivo.

Conclusiones:

CRISPR-Cas13a es una plataforma versátil para la investigación de ARN en células de mamíferos.
LwaCas13a ofrece una herramienta específica y programable para el knockdown y la visualización de ARN.
Esta tecnología tiene aplicaciones potenciales en el desarrollo terapéutico para enfermedades relacionadas con el ARN.

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