Generation of Targeted Knockout Mutants in Arabidopsis thaliana Using CRISPR/Cas9
Florian Hahn,
Marion Eisenhut,
Otho Mantegazza,
Andreas Weber
Affiliations
Florian Hahn
Institute of Plant Biochemistry, Cluster of Excellence on Plant Science (CEPLAS), Center for Synthetic Life Sciences (CSL), Heinrich Heine University, Düsseldorf, Germany
Marion Eisenhut
Institute of Plant Biochemistry, Cluster of Excellence on Plant Science (CEPLAS), Center for Synthetic Life Sciences (CSL), Heinrich Heine University, Düsseldorf, Germany
Otho Mantegazza
Institute of Plant Biochemistry, Cluster of Excellence on Plant Science (CEPLAS), Center for Synthetic Life Sciences (CSL), Heinrich Heine University, Düsseldorf, Germany
Andreas Weber
Institute of Plant Biochemistry, Cluster of Excellence on Plant Science (CEPLAS), Center for Synthetic Life Sciences (CSL), Heinrich Heine University, Düsseldorf, Germany
The CRISPR/Cas9 system has emerged as a powerful tool for gene editing in plants and beyond. We have developed a plant vector system for targeted Cas9-dependent mutagenesis of genes in up to two different target sites in Arabidopsis thaliana. This protocol describes a simple 1-week cloning procedure for a single T-DNA vector containing the genes for Cas9 and sgRNAs, as well as the detection of induced mutations in planta. The procedure can likely be adapted for other transformable plant species.
