Efficient CRISPR /Cas9 system based on autonomously replicating plasmid with an AMA1 sequence and precisely targeted gene deletion in the edible fungus, Cordyceps militaris

can be used to improve the efficiency. Polyethylene glycol-mediated protoplast transformation was markedly more efficient than Agrobacterium tumefaciens-mediated transformation of C. militaris. To our knowledge, this is the first description of genome editing and precisely targeted gene deletion in mushrooms based on AMA1 plasmids. Our findings will enable the modification of multiple genes in both functional genomics research and strain breeding.