Optimizing conditions for protoplasts preparation and regeneration of Agaricus bisporus Portobello

In order to improve the yield and regeneration rate of Agaricus bisporus Portabello protoplast. Mushroom strain was ZONGXIU No. 1 was selected, and two regression analysis with Design-Expert 8.0.6 software were used to optimize the preparation conditions of protoplast. The results showed that the yield of protoplast was arrived at the highest with 4.39 × 10⁷ · mL^-1 in the condition of lywallzyme concentration 1.35%, snailase concentration 7.01%, enzymolysis time 3.00 h, mycelium age 8.36 days and enzymolysis temperature 30.4℃. The highest protoplast regeneration rate with 15.1% was found in the condition of lywallzyme concentration 1.32%, snailase concentration 6.96%, enzymolysis time 2.97 h, mycelium age 9.00 days and enzymolysis temperature 31.0℃. The ultraviolet mutagenesis and chemical mutagenesis were used to mutate the protoplast growth characteristic, and eighteen strains with improved growth characteristics were obtained. The results could provide theoretical basis for protoplasts transformation, genome rearrangement and genome targeted editing for Agaricus bisporus Portobello. (authors)