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The enormous production of spores by the fruitbodies in the cultivation of oyster mushrooms (Pleurotus ostreatus) is develop an allergy with symptoms similar to an “extrinsic allergic alveolitis (EEA)”. the sporeless strain would noy only benefit health of mushroom workers but also reduce the risk of viral infections on the mushroom farms. For the development of a sporeless strain of P. ostreatus we used strain ASI 2069. This non-commercial strain is completely nonsporulating. We have recovered both nuclear types of strain ASI 2069 as monokaryons (hereafter referred to as neohaplonts) by protoplasting the mycelium. Crosses between neohaplonts and SSI's(single spore isolates) obtained from a sporulating commercial strain ASI 2180 yielded fruitbodies that isolated 128 strains. 13 excellent strains are selected from 30 bred strains by quality of fruitbodies and spore number. Among 13 excellent strains, G192 strain is chose finally to high yield and sporelessness.

BACKGROUND: Pleurotus ostreatus, oyster mushroom, is a popular edible mushroom cultivated world-wide and it also belongs to white-rot fungi that can degrade plant cell wall lignin effectively. To develop an efficient strategy for molecular breeding in P. ostreatus, molecular genetic tools have been developed through my 30-years investigation with wonderful colleagues from domestic and foreign countries. One of the most fruitful and newest international collaborations is that with Korean researchers. In this talk, I would like to introduce some recent results of an international collaboration with Mushroom Division, NIHHS, Rural Development Administration, Republic of Korea. RESULTS: We have successfully introduced a genome editing technology into P. ostreatus using CRISPR/Cas92. To establish a non-genetically modified (non-GM) genome editing in mushroom fungi, we set sail for developing a protocol for CRISPR/Cas9 without foreign DNA integration in the edited strain. In addition, to avoid health and environmental problems caused by spores, we selected and knocked-out candidate genes essential for spore formation using CRISPR/Cas9 technique. Three new genes essential for spore formation were found and sporeless cultivars by disrupting one of these genes were isolated. CONCLUSION: The fundamental technology of non-GM molecular breeding can be used for valuable objectives including high-value-added edible mushrooms, efficient lignin-degrading strains with high potential for bioremediation and non-existent strains su

For the development of a sporeless strain of P. ostreatus we used sporeless strain ASI 2069. We have recovered both nuclear types of strain ASI 2069 as monokaryons of nh9, nh15, nh26 and nh36 (here after referred to as neohaplonts) by protoplasting the mycelium. Crosses between neohaplonts and SSI’s(single spore isolates) obtained from a sporulating commercial strain ASI 2180. Five excellent strains are selected from 30 bred strains by quality of fruitbodies and spore number. To development of molecular markers linked to sporeless strain of P. ostreatus, we are screened helicase, recombinase(DMC1) and topoisomerase(Spo11) genes related meiosis by PCR and sequencing. Among three genes, Spo11 gene was identified into molecular marker of sporeless from neohaplonts and bred strains of P. ostreatus.