To explore the optimal methods for the protoplast preparation and regeneration of Hirsutella sinensis by optimizing the limiting factors. During the treatment of enzymatic protoplast preparation, mycelium cultured for 7 days was the optimal start material. The maximum protoplast preparation rate of 4.3 × 107 protoplasts/g fresh weight (FW) was obtained after 0.5 h treatment of 1 mg/ml mixed lytic enzymes in KH2PO4-K2HPO4 buffer (pH 5.5) with 0.6 M KCl at 18 °C. As for the protoplast regeneration, the maximum protoplast regeneration rate reached 12.32% through 5 × 103 protoplasts mL−1 cultivated for 20 days in the regeneration medium with 0.6 M mannitol and 1.5% agar. The preparation and regeneration of H. sinensis protoplasts was firstly established based on process optimization and it provided a foundation for the study of H. sinensis mutagenesis.

中文翻译：

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基于工艺优化的中华被毛孢原生质体制备与再生增强

通过优化限制因素，探索中华被毛孢菌原生质体制备及再生的最佳方法。在酶促原生质体制备过程中，培养7天的菌丝体是最佳起始材料。在 18 °C 下用 0.6 M KCl 在 KH2PO4-K2HPO4 缓冲液（pH 5.5）中处理 1 mg/ml 混合裂解酶 0.5 小时后，获得了 4.3 × 107 个原生质体/g 鲜重（FW）的最大原生质体制备速率。在原生质体再生方面，在含有0.6 M甘露醇和1.5%琼脂的再生培养基中培养20天的5×103个原生质体mL-1，最大原生质体再生率达到12.32%。首次在工艺优化的基础上建立了中华鳖原生质体的制备与再生，为中华鳖的诱变研究奠定了基础。