甘露醇对草菇继代菌株生产性状和ROS清除能力的影响  

Effects of Mannitol on Production Characteristics and ROS Scavenging Ability of Volvariella volvacea Subcultured Strains

作　　者：赵风云[1] 程志虹 谭强飞 朱嘉宁 孙万合 张紊玮[1] 贠建民[1] ZHAO FengYun;CHENG ZhiHong;TAN QiangFei;ZHU JiaNing;SUN WanHe;ZHANG WenWei;YUN JianMin(College of Food Science and Engineering,Gansu Agricultural University,Lanzhou 730070)

机构地区：[1]甘肃农业大学食品科学与工程学院,兰州730070

出　　处：《中国农业科学》2024年第1期190-203,共14页Scientia Agricultura Sinica

基　　金：国家自然科学基金(32060708)。

摘　　要：【目的】研究甘露醇对草菇继代菌株的生产性状和活性氧(ROS)清除能力的影响,探索一种简便、有效的草菇退化菌种复壮方法。【方法】以笔者课题组前期获得的组织分离继代菌株T6、T12、T19为试验菌株,T6为连续继代6次,T12为连续继代12次,T19为连续继代19次;草菇原种V844(T0)为商业栽培种。将传统马铃薯葡萄糖琼脂培养基(PDA)中的葡萄糖替换为等质量的甘露醇,进行菌丝生理性状测定;在栽培基质中添加甘露醇,测定子实体农艺学性状;以硝基四氮唑蓝(NBT)染色、超氧阴离子(O_(2))、过氧化氢(H_(2)O_(2))含量反映活性氧(ROS)积累;采用实时荧光定量PCR(RT-qPCR)测定抗氧化酶基因表达量;利用试剂盒测定抗氧化酶活力;通过菌丝染色法测定细胞核数量和线粒体膜电位;利用高效液相色谱仪(HPLC)测定菌丝能量指标。【结果】甘露醇处理对未退化菌株T0、T6的影响不显著,但能有效恢复草菇退化菌株T12、T19的生产性状和ROS清除能力。甘露醇使T12、T19的菌丝生长速度分别提高31.46%、20.99%,菌丝生物量分别提高97.33%、76.36%;使T12的生产周期缩短12.24%,生物学效率提高17.97%,恢复至T0水平;并使退化严重、失去出菇能力的T19重新长出子实体。同时,甘露醇使T12、T19的Cu/Zn超氧化物歧化酶基因(Cu/Zn-sod)相对表达量分别上调24.64%和61.54%,Mn-sod2相对表达量分别上调19.76%和267.09%,谷胱甘肽过氧化物酶基因(gpx)相对表达量分别上调25.67%和55.82%,并使SOD活力分别提高10.79%和72.32%,GPX活力分别提高16.98%和103.85%;使T12、T19中的ROS积累量显著降低,T12、T19中的O_(2)含量分别下降35.96%和41.62%,H_(2)O_(2)含量分别下降14.44%和18.26%;并使T12、T19的细胞核数目和线粒体膜电位显著增加;使T12、T19中的ATP含量分别提高17.08%和14.55%,EC值分别提高4.52%和0.92%。【结论】甘露醇能显著提高草菇退化菌株T12、T19的抗氧化能力和线粒体【Objective】The aim of this study was to investigate the effects of mannitol on production characteristics and reactive oxygen species(ROS)scavenging ability of Volvariella volvacea subcultured strains,and to explore a simple and effective method for the rejuvenation of V.volvacea degenerated strains.【Method】The tissue isolation subcultured strains T6,T12 and T19 were obtained by previous study of our research group,and T6 was obtained after 6 successive subculture,while T12 and T19 was obtained after 12 and 19 successive subculture,respectively.The original strain(T0),referred to as V844,was a strain used in commercial agricultural cultivation.The glucose in the traditional potato dextrose agar(PDA)was replaced by mannitol of the same mass,then physiological traits were determined in mycelia.The agronomic characters of fruiting body were measured by adding mannitol to the culture medium.ROS accumulation was reflected using nitrotetrazolium blue chloride(NBT)staining of V.volvacea mycelia,superoxide anion(O_(2))and hydrogen peroxide(H_(2)O_(2))content.The expression levels of antioxidant enzyme genes were detected by real-time quantitative PCR(RT-qPCR).The activity of antioxidant enzymes was measured by the kit.The number of nuclei and mitochondrial membrane potential were determined by mycelium staining.The energy indexes were determined by high performance liquid chromatography(HPLC).【Result】Mannitol treatment had no significant effect on non-degraded strains T0 and T6,but could effectively restore the production characteristics and ROS scavenging ability of degraded strains T12 and T19.After mannitol treatment,the mycelial growth rate of T12 and T19 was increased by 31.46%and 20.99%,respectively,and the mycelial biomass was increased by 97.33%and 76.36%,respectively.The mannitol treatment shortened the production cycle of T12 by 12.24%and increased the biological efficiency by 17.97%,thus restoring it to T0 level.In addition,the mannitol treatment caused T19 to regrow its fruiting body,which had bee

关 键 词：草菇 甘露醇 菌种复壮 活性氧 抗氧化酶 线粒体 

分 类 号：S646[农业科学—蔬菜学]