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Efficient Genome Editing with CRISPR/Cas9 in Pleurotus ostreatus

Summary

This research demonstrates a new genetic engineering technique for improving oyster mushrooms using CRISPR gene editing technology. The method could lead to better mushroom varieties without creating technically genetically modified organisms. Impact on everyday life: • Could lead to mushrooms with improved growth rates and yields for farmers • May result in more nutritious or better-tasting mushroom varieties for consumers • Provides a way to develop enhanced mushroom strains that avoid GMO regulations • Could reduce the time and cost needed to develop new mushroom varieties • May enable development of mushrooms with improved shelf life or disease resistance

Background

Pleurotus ostreatus is one of the most commercially produced edible mushrooms worldwide. Traditional breeding methods for improving cultivated strains are laborious and time-consuming. Molecular genetics and gene targeting methods offer potential solutions but often result in genetically modified organisms (GMOs) which face regulatory restrictions in many countries.

Objective

To develop and demonstrate efficient gene mutagenesis using plasmid-based CRISPR/Cas9 as a first step toward generating non-genetically modified P. ostreatus strains. The study aimed to test the system’s effectiveness by targeting fcy1 and pyrG genes.

Results

The system achieved high mutation frequencies of 20-94.9% depending on the target sequence. Small insertions/deletions were identified in three mutants, and plasmid insertions were found in eight mutants at the target sites. Using donor DNA templates with homology arms of 0.5-1kb length resulted in successful gene replacement via homology-directed repair with frequencies up to 83.3%.

Conclusion

The study successfully demonstrated the first application of CRISPR/Cas9 genome editing in an edible mushroom, achieving higher efficiency than previously reported in other agaricomycetes. The system enabled both random mutations and precise gene replacement, providing a foundation for future development of non-GMO mushroom strains and molecular genetics studies.
  • Published in:AMB Express,
  • Study Type:Laboratory Research,
  • Source: 10.1186/s13568-021-01193-w
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