利福霉素高产菌株快速筛选方法的建立及应用  

Establishment and Its Application of a Fast Screening Method for Rifamycin SV-Producing Strain

作　　者：杜吉泉[1] 王军峰[1] 储炬[1] 王永红[1] 庄英萍[1] 张嗣良[1] 

机构地区：[1]华东理工大学生物反应器工程国家重点实验室

出　　处：《华东理工大学学报（自然科学版）》2006年第12期1400-1403,1408,共5页Journal of East China University of Science and Technology

基　　金：上海市科委重点科技攻关项目(034319220)

摘　　要：利福霉素发酵过程中pH变化规律能够反映菌体对不同营养物质的利用情况及菌体生理特性的变化,并与最终放瓶效价有一定的关联。据此建立了一种以发酵前期(±50 h)pH变化为判断依据的新的快速筛选菌种的方法,即发酵前期pH下降到谷底较早的菌株高产的可能性很大(概率接近70%),并在双亲株灭活种间融合法选育利福霉素SV菌株中进行了应用,筛选到了高产融合菌株F-3及F-16,两菌株分别比出发菌株效价提高了11%和17%。同时,得到了两个亲株(利福霉素SV产生菌U-32、利福霉素B产生菌X#-1)的酶解条件(U-32:10 m g/mL、34°C、2 h;X#-1:10 m g/mL、34°C、3 h)及原生质体双灭活标记条件(U-32:UV 20 m in;X#-1:60°C、50 m in),双灭活原生质体用500 g/L PEG融合,融合频率约1.04×10-5。In order to obtain a high yield rifamycin SV-producing strain, the protoplasts fusion between two Amycolatopsis mediterranei strains U-32 and X^#-1 which produce rifamycin SV and rifamycin B respectively was studied. The protoplasts of U-32 and X^#-1 were inactivated by ultraviolet radiation（15 W, 25 cm, 20 min） and by heating （60 ℃, 50 min） respectively before fusion. The variation of pH during rifamycin SV fermentation in flask could reflect the consumption of substrates, which is relevant to the final titer of production. Accordingly, a method of fast selection of rifamycin SV-producing strain was established depending on the variation of pH during the initial 50 h of fermentation using the pH-controlled fed-batch shaker （pH-shaker）. Two fusion strains F-3 and F-16 with high potency of rifamycin SV were gained by this method, which was increased by 11 % and 17 % respectively.

关 键 词：地中海拟无枝酸菌 利福霉素SV pH检测控制 菌种快速筛选 双亲株灭活 原生 质体融合 

分 类 号：TQ920.6[轻工技术与工程—发酵工程]