Generation of cells with a loss-of-function mutation in a gene (knockout cells) is a valuable technique for studying the function of a given gene product. However, if the product of the target gene is essential for cell viability, conditional knockout cell lines must be generated. Recently, as gene editing technology using CRISPR/Cas9 has developed, it has become possible to produce conditional knockout cell lines using this technique. However, to obtain final conditional knockout cell lines, it is necessary to perform several experiments with multiple complicated steps. In this paper, we introduce an easy and efficient method to generate conditional knockout cell lines based on combining auxin-inducible degron (AID) technology with CRISPR/Cas9 gene editing. Our method only requires performing a single transfection and is therefore an easy and rapid method to obtain a conditional knockout cell line.
Aid CRISPR/Cas9 Rapid degradation
Clustered regularly interspaced short palindromic repeat
CRISPR-associated protein 9
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We are very grateful to R. Fukuoka and Y. Fukagawa for their technical assistance. This work was supported by MEXT KAKENHI Grant Numbers 25221106 and 15H05972 to TF and MEXT KAKENHI Grant Number 17K15041 to KN.