Accedi

University of Cantabria, Santander y Torrelavega, Spain

1 ARTICLES PUBLISHED IN JoVE

Bioengineering

Construction of CRISPR Plasmids and Detection of Knockout Efficiency in Mammalian Cells through a Dual Luciferase Reporter System

Hegang Li^*1, Huaiyuan Qin^*1, Ning Zhang^*1, Jinshan Zhao¹, Jingjing Xin¹, Flor M. Perez-Campo², Huawei Liu¹

¹Qingdao Agricultural University, Qingdao, China, ²University of Cantabria, Santander y Torrelavega, Spain

Here, we present a protocol describing a streamlined method for the efficient generation of plasmids expressing both the CRISPR enzyme and associated single guide RNA (sgRNAs). Co-transfection of mammalian cells with this sgRNA/CRISPR vector and a dual luciferase reporter vector that examines double-strand break repair allows evaluation of knockout efficiency.