Plant Gene Knockout and Knockdown by CRISPR-Cpf1 (Cas12a) Systems.
Methods in molecular biology (Clifton, N.J.), 1917: 245-256 PMID 30610641 DOI 10.1007/978-1-4939-8991-1_18
Abstract
CRISPR-Cpf1 (Cas12a) is a class II type V endonuclease, which has been used as a genome editing tool in different biological systems. Here we describe a fast, efficient, and user-friendly system for CRISPR-Cpf1 expression vector assembly. In this system, the Pol II promoter is used to drive the expression of both Cpf1 and its crRNA, with the crRNA flanked by hammerhead (HH) and hepatitis delta virus (HDV) ribozyme RNAs for precise crRNA processing. All the components of this system can be modified depending on plant species and experimental goals. Using this system, nearly 100% editing efficiency and 90% gene expression decrease were achieved in rice and Arabidopsis, respectively.