为提高菌丝原生质体的释放量和活力,分别考查了碳源、菌龄、渗透压稳定剂、酶浓度、酶解时间及再生培养基种类对Aspergillus oryzaeHN3042和Aspergillus niger CICC2377原生质体释放和再生的影响,并研究了两个亲本原生质体的非对称灭活参数.确定了亲本原生质体的适宜制备参数.结果表明,以MPY液体培养基为菌丝培养基、0.7mol/L NaCl复合0.4mol/L山梨醇为渗透压稳定剂、以高渗豆汁固体培养基为再生培养基,原生质体有30%以上的再生率.通过非对称灭活曲线确定Aspergillus oryzae HN3042原生质体于15W紫光灯垂直距离13cm处照射15min,Aspergillus niger CICC2377原生质体于65℃水浴10min,可使亲本原生质体临界失活.

In order to improve the quantity and viability of protoplasts released from the mycelia of Aspergillus oryzae HN3042 and Aspergillus niger CICC2377,the effects of carbon resources,mycelia age,osmotic stabilizers,enzyme concentration,incubation time and regeneration media on the release and regeneration of protoplasts were investigated,and the parameters of asymmetric inactivation for the protoplasts were analyzed.Afterwards,suitable conditions for protoplast release were determined.The results indicate that,with MPY consisting of maltose,polytephone and yeast extract as the submerged medium,0.7mol/L NaCl combined with 0.4mol/L sorbitol as the osmo-tic stablizer,and with hypertonic soybean-extract solid medium as the regeneration medium,the released protoplasts show a regeneration efficiency of above 30%.Moreover,according to the asymmetric inactivation curves,it is found that both Aspergillus oryzae HN3042 and Aspergillus niger CICC2377 protoplasts are in critical inactivation stage when they are respectively under the irradiation of 15W ultraviolet from 13cm vertically away for 15min or in a 65℃ water bath for 10min.