Breeding and Gene Manipulation for the Sawdust-Based Cultivation of Lentinula edodes

研究了木屑培养香菇结实过程中漆酶和纤维素酶mRNA的转录调控。（材料与方法）本研究共检测了12株菌种。KS-3、-4、-16、-23、-46、-56、-58、-60、-67和-76（广天菌株），KS-12和-24（寒天菌株）。按照标准方法进行木屑基质培养。测定了7个不同生长阶段漆酶和纤维素酶的活性。生长阶段分为菌丝生长阶段（90日龄）、聚集体阶段（94日龄）、针状阶段（98日龄）、扣状阶段（100日龄）、破纱阶段（102日龄）、衰老阶段（104日龄）和二次同花顺阶段（140日龄）。对木屑基基材的水分条件进行了调整浸泡处理。用水势（φ）值评价水分状况。RT-PCR法检测漆酶和纤维素酶基因的转录调控。（结果）以木屑为底物，研究了香菇不同生长阶段漆酶和纤维素酶的活性及其转录调控。漆酶和纤维素酶活性的变化与果体发育阶段有明显的关系。漆酶和纤维素酶活性在基因转录水平上受到调控。漆酶mRNA水平在完全定植期最高，在果体发育期间下降。纤维素酶mRNA在小果体形成阶段开始积累。纤维素酶mRNA转录物在果体发育的破纱期表达量最大。这种趋势在广域气候菌株的结果培养中比在非冷气候菌株的结果培养中更为明显。漆酶和纤维素酶基因的转录也受到木屑基质水分条件的影响。当培养基的水势高时，原基起始发生，以便菌丝快速转录mRNA。少

Transcriptional regulation of laccase and cellulase mRNA were investigated for fruiting on the sawdust-based cultivation of Lentinula edodes.(Materials and Methods)1. Twelve spawn strains were examined in this study. KS-3, -4, -16, -23, -46, -56, -58, -60, -67, and -76 (wide-weather strains), KS-12 and -24 (cold weather strains).2. The sawdust-based substrate cultivation was done according to the standard methods. Activities of laccase and cellulase were measured at the 7 different growth stages. The growth stages were defined to mycelial growth (90-days old), aggregate (94-days old), pin (98-days old), button (100-days old), veil break (102-days old), senescent (104-days old), and second flush (140-days old).3. Moisture condition of the sawdust-based substrate was adjusted soaking treatment. The moisture condition was evaluated by the water potential (φ) values.4. Transcriptional regulation of laccase and cellulase genes was measured by the RT-PCR method.(Results)Extracellular enzyme … More activities of laccase and cellulase and their transcriptional regulation were investigated at various growth stages in a sawdust-based substrate for Lentinula edodes. Changes of laccase and cellulase activities revealed a clear relationship with fruit body development stages. Laccase and cellulase activities were regulated at the level of gene transcription. The level of laccase mRNA was maximal at the fully colonized stage and declined during fruit body development. Cellulase mRNA began to accumulate at the pin (miniature fruit bodies) formation stage. Cellulase mRNA transcripts were maximally expressed at the veil-break stage of fruit body development. This tendency was clearer in the fruiting cultures with the wide-range-weather strains than in non-fruiting cold-weather strains. Transcription of laccase and cellulase genes was also affected by the water conditions of the sawdust-based substrate. Primordia initiation occurred when the water potential of the medium was high for rapid mRNA transcription by the mycelium. Less