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林产化学与工业

林产化学与工业 ›› 2013, Vol. 33 ›› Issue (5): 30-34.doi: 10.3969/j.issn.0253-2417.2013.05.006

• 1研究报告 • 上一篇    下一篇

脂肪酶产生菌微波-亚硝基胍复合诱变及培养条件优化

苗长林1, 罗文1, 吕鹏梅1, 李惠文1, 杨玲梅1, 袁振宏1, 蒋剑春2   

  1. 1. 中国科学院 广州能源研究所, 中国科学院可再生能源与天然气水合物重点实验室, 广东 广州 510640;2. 中国林业科学研究院 林产化学工业研究所, 江苏 南京 210042
  • 收稿日期:2012-08-20 修回日期:1900-01-01 出版日期:2013-10-30 发布日期:2013-10-30
  • 通讯作者: 吕鹏梅(1973-),女,山东烟台人,研究员,博士生导师,从事生物质能研究;E-mail:lvpm@ms.giec.ac.cn。

Screening of Lipase Production Strains by Microwave-nitrosoguanidine Mutagenesis and Optimizing of Lipase Producing Condition

MIAO Chang-lin1, LUO Wen1, LŰ Peng-mei1, LI Hui-wen1, YANG Ling-mei1, YUAN Zhen-hong1, JIANG Jian-chun2   

  1. 1. Key Laboratory of Renewable Energy and Gas Hydrate, Guangzhou Institute of Energy Conversion, CAS, Guangzhou 510640, China;2. Institute of Chemical Industry of Forest Products, CAF, Nanjing 210042, China
  • Received:2012-08-20 Revised:1900-01-01 Online:2013-10-30 Published:2013-10-30

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摘要： 采用微波-亚硝基胍诱变方法对粗酶活为 10 U/mL 的米黑根毛霉进行诱变处理以获得高产酶能力的菌株,实验获得最佳诱变条件为：功率 500 W 微波处理时间为 100 s,亚硝基胍处理时间为 45 min。通过荧光圈初筛和摇瓶复筛,筛选出一株酶活可达 20.50 U/mL 突变菌株,该菌株比原始出发菌株脂肪酶活力提高了 105%,将MN4连续传代5次,酶活稳定。对MN4突变菌株的产酶条件进行研究,结果表明,MN4突变菌株最适产酶条件为：发酵温度 30℃,起始pH值为7.0,发酵时间 4 d。在此条件下,该突变菌株所产脂肪酶活最高,达 22.90 U/mL,产酶活力相对原始出发菌株提高了 129.0%。试验结果表明:微波、亚硝基胍对米黑根毛霉菌体细胞的致死作用表现出相似的趋势,在一定范围内都与诱变时间呈线性正相关。微波-亚硝基胍复合处理还呈现一定的协同效应,可以减弱单一诱变剂反复诱变产生的诱变抗性和饱和性,能够实现优势互补,提高诱变效率。

关键词: 脂肪酶, 微波, 亚硝基胍, 诱变, 米黑根毛霉

Abstract: In order to obtain mutants with high lipase production, the Rhizomucor miehei with 10 U/mL enzyme activity was selected as the original strain, and was treated by combining microwave irradiation and nitrosoguanidine(NTG). The optimum treating conditions were got by primary screening. A mutant named MN 4 with higher activity of lipase was obtained as the original strain was treated by a power of microwave irradiation 500 W for 100 s and then a NTG treatment for 60 min. The activity of lipase of MN 4 was 20.50 U/mL, which was 105% higher than that of the original strain. Then an experiment for studying the genetic stability was taken by means of passaging 5 times and its results showed that MN4 had a better genetic stability. The Lipase-producing conditions of such mutant were investigated and the optimum conditions were summed up as follows: fermentation temperature was at 30℃, pH value was 7.0, and fermentation time was 4 d. Under these conditions, the produced Lipase had the highest activity of 22.90 U/mL, which was 129.0% higher than that of the original strain. The experimental results showed that microwave and NTG mutation agents possessed analogical mortal ratio trend. It positively related to the mutation agent's concentration microwave-NTG composite mutation presents some synergies. This could weaken repellence and saturation by single mutation agent.Microwave-NTG composite mutation could realize the complementary advantages and inerease mutant efficiency.

Key words: lipase, microwave, nitrosoguanidine, mutation, Rhizomucor miehei

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