He-Ne激光诱变姬松茸原生质体选育速生高产菌株及姬松茸液体深层发酵工艺研究

【作者】 叶勋艳；

【导师】 陈五岭；

【作者基本信息】 西北大学 ， 微生物学， 2003， 硕士

【摘要】 本文采用激光技术与微生物原生质体育种技术相结合，选育姬松茸速生高产新菌株。并研究了所选育的变异株深层发酵培养的最佳发酵工艺条件，为姬松茸工业化生产液体菌种提供微烛之光。 实验结果表明，姬松茸原生质体的最佳条件为：采用液体发酵培养第四天的菌体，2％溶壁酶，在30℃下酶解5hr，原生质体产率达9.08×10⁶个/ml；制备的原生质体，在含0.6mol/LMgSO₄的高渗培养基上培养，再生率为2.50％。 采用波长为632.8mm的He-Ne激光，以不同剂量辐照姬松茸原生质体，其变异株经酯酶同工酶谱分析，酶带条数，R_f值均发生了显著变化；斜面培养，液体培养分析表明变异株与出发菌株相比生长速率和生物量发生了明显变化，说明其遗传物质发生了改变，且不同剂量的He-Ne激光对姬松茸原生质体的诱变效果有差异，表明激光对姬松茸是一种有良好应用前景的诱变剂。 研究了不同培养基配方，不同接种量对菌丝量的影响，确定了最佳培养基配方，最佳发酵工艺条件；以及变异株在最佳培养基和最佳工艺条件下，其液体深层培养情况，包括培养过程中还原糖、氨基氮、pH等参数随菌丝生长的变化规律。更多还原

【Abstract】 In this paper, with laser-irradiation and protoplast technique, the mutants were screened whose mycelia growth speed was increased. I studied the best fermentative procedure conditions of the mutants that were cultivated in submerged culture medium, and that will offer the base of producing industrially Agaricus blazei Murrill submerged strains.The results showed that the best conditions of Agaricus blazei Murrill protoplast production and regeneration were to use the strains after cultivated in submerged culture medium for four days, while the mycelia co-treated with 2% Lywallzyme, at 32癈, for five hours, and the production rate of protoplast reached to 9.08million per milliliter, and the regeneration frequencies reached to 2.50% on hyperosmotic medium containing 0.6 M MgSO4.He-Ne laser (r=632.8nm) irradiated protoplasts for different time. The analysis of esterase isoenzgme indicated that the mutants’ esterase isenzyme patterns and Rf were of difference from the mycelidium. After solid culture and liquid culture, we can see that mutants’ mycelial growth speed and biomass were different from mycelidium. So it was concluded that the mutants’ genetic substances had changed and different doses had different effects on protoplasts. Laser is a good mutagen for Agaricus blazei Murrill.After studied the effect on wet mycelia amount of different medium, different inoculation methods, we got the best fermentative media and fermentative procedure. And we also got the law of a series of parameters about the mutants when submerged cultivated, including reducible sugar, amino nitrogen and pH.更多还原