Agrobacterium-Mediated Transformation and Targeted Mutagenesis Using SpCas12f in Rice

Area of Science:

• Molecular Biology
• Genetics
• Plant Science

Background:

• Clustered regularly interspaced short palindromic repeats (CRISPR)-Cas is a prokaryotic adaptive immune system repurposed for genome editing.
• CRISPR-Cas9 is widely used, but its size can be limiting for certain applications.
• CRISPR-Cas12f is a smaller alternative, offering potential benefits for vector construction and transformation efficiency.

Purpose of the Study:

• To describe a protocol for targeted mutagenesis in rice using Syntrophomonas palmitatica Cas12f (SpCas12f).
• To highlight the advantages of using compact CRISPR-Cas12f systems for plant genome editing.
• To summarize methods for enhancing SpCas12f-mediated targeted mutagenesis frequency.

Main Methods:

• Agrobacterium-mediated transformation of rice.
• Targeted mutagenesis using Syntrophomonas palmitatica Cas12f (SpCas12f).
• Optimization strategies for improving mutagenesis efficiency.

Main Results:

• Successful implementation of a protocol for SpCas12f-mediated targeted mutagenesis in rice.
• Demonstration of the utility of compact CRISPR-Cas12f for plant genome engineering.
• Identification of approaches to increase the frequency of targeted mutations.

Conclusions:

• The compact SpCas12f system is a viable and advantageous tool for genome editing in rice.
• This protocol facilitates efficient targeted mutagenesis in plants.
• Further optimization strategies can enhance the application of CRISPR-Cas12f in plant biotechnology.