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Targeted Mutagenesis Using FnCpf1 in Tobacco
Akira Endo1, Seiichi Toki2,3
1Plant Genome Engineering Research Unit, Institute of Agrobiological Sciences, National Agriculture and Food Research Organization, Tsukuba, Ibaraki, Japan.
Methods in Molecular Biology (Clifton, N.J.)
|January 6, 2019
Summary
Francisella novicida Cpf1 (FnCpf1) enables targeted mutagenesis in plants. Its shortest PAM (TTN) and sticky DNA ends offer advantages for plant genome engineering and molecular breeding.
Area of Science:
- Molecular Biology
- Genetics
- Biotechnology
Background:
- CRISPR/Cas9 systems are widely used for targeted mutagenesis.
- CRISPR from Prevotella and Francisella 1 (Cpf1) is an RNA-directed endonuclease with unique properties.
- Cpf1 offers versatility in plant genome engineering.
Purpose of the Study:
- To describe procedures for targeted mutagenesis in tobacco using FnCpf1.
- To highlight FnCpf1 as a practical tool for plant genome engineering.
Main Methods:
- Utilizing Francisella novicida Cpf1 (FnCpf1) for targeted mutagenesis.
- Employing FnCpf1's recognition of the TTN protospacer adjacent motif (PAM).
Main Results:
- FnCpf1 recognizes the shortest known PAM (TTN), expanding targetable sequences.
- Cpf1 generates cohesive DNA ends, facilitating in vivo ligation compared to Cas9's blunt ends.
Conclusions:
- FnCpf1 is a practical and versatile tool for targeted mutagenesis in plants.
- FnCpf1-mediated mutagenesis can accelerate molecular breeding of crops.
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