Efficient CRISPR/Cas9-mediated Targeted Mutagenesis in Populus in the First Generation

Area of Science:

• Plant Biotechnology
• Genome Editing
• Molecular Biology

Background:

• RNA-guided genome editing using CRISPR-Cas9 is established in herbaceous plants.
• Its application in woody plants remains largely unexplored.

Purpose of the Study:

• To investigate the efficacy of the CRISPR/Cas9 system for genome editing in the woody species Populus tomentosa.
• To achieve targeted gene mutation in a specific poplar gene.

Main Methods:

• Designed four guide RNAs (gRNAs) targeting the phytoene desaturase gene 8 (PtoPDS) in Populus tomentosa.
• Utilized Agrobacterium-mediated transformation for gene delivery.
• Analyzed PCR clones for mutation events.

Main Results:

• Observed an obvious albino phenotype in transgenic poplar plants, indicative of PDS gene disruption.
• Achieved a mutation efficiency of 51.7% at the targeted PtoPDS sites.
• Identified homozygous and heterozygous mutants among the analyzed PCR clones.

Conclusions:

• The Cas9/sgRNA system is effective for precise genome editing in woody plants.
• This study validates CRISPR/Cas9 as a tool for creating targeted gene knockout mutations in Populus species.