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Expanding plant genome editing scope and profiles with CRISPR‐FrCas9 systems targeting palindromic TA sites
Expanding plant genome editing using CRISPR-FrCas9 systems that target palindrome TA DNA sites
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Abstract
FrCas9 recognizes a concise 5'-NNTA-3' , enabling targeted mutagenesis in rice.
- FrCas9 targets more abundant palindromic TA sites in plant genomes compared to SpCas9's PAM requirements.
- It demonstrates cleavage activities at all tested 5'-NNTA-3' PAM sites, producing editing outcomes typical of conventional CRISPR systems.
- FrCas9 enables high-efficiency targeted mutagenesis and generates biallelic mutants in stable rice lines.
- The fusion of FrCas9 with TREX2 enhances its capacity to produce larger deletions while maintaining editing efficiency.
- FrCas9-based cytosine and adenine base editors can create targeted C-to-T and A-to-G edits in rice plants.
- Whole-genome sequencing indicates that while FrCas9 is highly specific, TREX2-FrCas9 may lead to off-target mutations, primarily single nucleotide variants.
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Key numbers
2.7% to 53.9%
Editing Efficiency Range
Measured at 16 endogenous sites in rice protoplasts.
6–20 bp
Larger Deletions Achieved
Deletion sizes generated by TREX2-FrCas9 compared to FrCas9.
59.5%
C-to-T Conversion Rate
Achieved by CBE V2.1 at four target sites.