Genetic editing and interrogation with Cpf1 and caged truncated pre-tRNA-like crRNA in mammalian cells

Cell Discovery
Xuhua ZhangYongping Song

Abstract

Cpf1, an RNA-guided DNA endonuclease that belongs to a new class II CRISPR system, has recently been harnessed for genome editing. Herein, we report an RNase-resistant caged truncated pre-tRNA-like crRNA (catRNA) that confers precise and efficient gene editing with the Lachnospiraceae bacterium Cpf1 (LbCpf1) and enables the reprogramming of catalytically dead LbCpf1 (dCpf1) lacking DNA endonuclease activity into a transcriptional modulator. Specific gene knock-outs and knock-ins were increased 3.2-fold and 4.3-fold, respectively, with catRNA compared to that induced by conventional crRNA. A much higher augmentation of gene disruption (up to 37-fold) was observed when electroporation was used. We report herein that catRNA enables efficient gene activation with dCpf1 activators. Our study reveals the potential of catRNA and a versatile application of the CRISPR/Cpf1 system, establishing a simple approach for selective gene perturbation in mammalian cells.

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Citations

May 16, 2019·Cell & Bioscience·Fatemeh SafariYounes Ghasemi
Oct 24, 2020·The CRISPR Journal·J Armando Casas-MollanoMichael J Smanski
May 1, 2021·Genes and Environment : the Official Journal of the Japanese Environmental Mutagen Society·Hussein SabitAfnan Al-Muhanaa

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Methods Mentioned

BETA
gene modification
gene
transfection
PCR
gene knock-out
transfections
gene knock-ins
flow
electrophoresis

Software Mentioned

TIDE

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