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Genome Editing in the Mushroom-Forming Basidiomycete Coprinopsis cinerea, Optimized by a High-Throughput Transformation System

Summary

This research developed new genetic tools to modify mushroom-forming fungi more efficiently. The scientists created a system that allows them to precisely edit genes in Coprinopsis cinerea, a model mushroom species used to study fungal biology. This advancement has several real-world implications: • Improved breeding methods for edible mushrooms, potentially leading to better yields and nutritional content • Enhanced ability to study how mushrooms develop and grow, helping optimize cultivation conditions • Potential for engineering fungi to produce valuable compounds for medicine and industry • More efficient ways to study mushroom genetics, accelerating research and development • Possible applications in developing new varieties of mushrooms with desired traits

Background

Mushroom-forming basidiomycetes produce valuable metabolites and are important global natural resources. While genome engineering tools exist for other fungi, they are less developed in mushroom-forming basidiomycetes. Coprinopsis cinerea is a model species used to study fruiting body development and meiosis, but lacks efficient high-throughput transformation and gene editing systems.

Objective

To develop and optimize CRISPR/Cas9-based genome editing in C. cinerea through identification of strong promoters and establishment of an efficient transformation system using cryopreserved protoplasts.

Results

The cryopreservation method maintained protoplast viability and transformation competency for up to one year. The CcDED1pro showed seven times stronger activity than the conventional GPD2 promoter. Using basidiomycete codon-optimized Cas9 driven by CcDED1pro, successful genome editing was achieved with 21% efficiency. Two mutant lines were isolated showing clear GFP loss-of-function in both hyphae and fruiting body primordia.

Conclusion

This study established the first CRISPR/Cas9-based genome editing system in mushroom-forming basidiomycetes, along with an efficient high-throughput transformation method using cryopreserved protoplasts. These tools will accelerate functional genetics research in C. cinerea and molecular breeding of edible mushrooms.
  • Published in:Scientific Reports,
  • Study Type:Laboratory Research,
  • Source: 10.1038/s41598-017-00883-5
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