高压电晕电场诱变纳豆芽孢杆菌合成γ-聚谷氨酸产量提高的研究

发布时间：2018-01-22 11:53

本文关键词： 纳豆芽孢杆菌针板电晕电场 γ-PGA 稀土元素响应面分析 γ-PGA合成酶　出处：《内蒙古大学》2017年硕士论文　论文类型：学位论文

【摘要】：为了提高纳豆芽孢杆菌产γ-聚谷氨酸能力,本试验采用针板高压电晕电场和稀土元素来处理纳豆芽孢杆菌N-0,处理结果显示:3kV-30min-0.8cm的针板处理电场,可筛选到γ-PGA产量为21.2g·L-1的菌株N-E5.2,是菌株N-0 y-PGA产量的8倍;稀土元素二次处理,筛选出γ-PGA产量为22.8g.L-1的N-E5.2b,产量又提高了 7.5%,且连续7次传代,发酵性能稳定。以纳豆芽孢杆菌N-E5.2b为研究对象,进行培养基响应面优化研究构建响应方程。获得的优化培养基(g·L-1):细菌学蛋白胨15、蔗糖46.38、谷氨酸钠45.85、酵母提取粉 2.5、NaCI5、MgSO4·7H2O0.5、MnSO4·4H2O0.4、CaCI20.25、K2HP04·3H20 5.2和KH2PO42。摇瓶发酵γ-PGA产量达38.94 g.L-1,较优化前提高了70.8%。提取和扩增6株不同处理菌株的pgsBCA合成酶基因,对比这6株菌的pgsB、pgsC和pgsA的基因序列及合成酶蛋白PgsB、PgsC和PgsA的氨基酸序列差异。结果显示:pgsB、pgsC 和和pgsA基因分别约含11 70个、450个和1140个核苷酸,分别编码约390个、150个和380个氨基酸。高压电晕电场和稀土元素促使pgsBCA基因序列发生了碱基的置换和颠换、插入和缺失,合成酶PgsBCA氨基酸序列出现多处氨基酸间的替换、插入和缺失,且对序列的开端和尾端影响较大,PgsA在30～40个氨基酸区间变化显著。
[Abstract]:In order to improve the ability of producing 纬 -polyglutamic acid from Bacillus natto, N-0 of Bacillus natto was treated with high electric field and rare earth elements. The results showed that the strain N-E5.2 with 纬 -PGA yield of 21.2 g 路L ~ (-1) could be screened by the needle plate treatment electric field of 30 min -0.8 cm. The yield of N-0y-PGA was 8 times of that of strain N-0y-PGA. The yield of 纬 -PGA was 22.8g 路L ~ (-1) N-E 5.2b, the yield was increased by 7.5g 路L ~ (-1). The fermentation performance was stable. Bacillus natto N-E5.2b was used as the research object. The response equation was established by optimizing the response surface of culture medium. The optimized medium was: bacteriological peptone 15, sucrose 46.38, sodium glutamate 45.85. The yeast extract was 2.5% NaCI5, MgSO4 路7H2O 0.5MNO _ 4 路4H _ 2O _ (0.4) and CaCI _ (20.25). K2HP04 路3H20 5.2 and KH2PO422.The yield of 纬 -PGA reached 38.94 g 路L ~ (-1). The pgsBCA synthase genes of 6 strains treated with different treatments were extracted and amplified, and the pgsB of the 6 strains were compared. The gene sequence of pgsC and pgsA and the amino acid sequence of PgsBU PgsC and PgsA were different. PgsC and pgsA genes contain about 11 70 nucleotides, 450 nucleotides and 1 140 nucleotides respectively, encoding 390 nucleotides, respectively. 150 amino acids and 380 amino acids. High voltage halo electric field and rare earth elements lead to the replacement, transversion, insertion and deletion of pgsBCA gene sequence. Substitution, insertion and deletion of multiple amino acids were observed in the amino acid sequence of the synthase PgsBCA, and significant changes of PgsA were observed in the range of 30-40 amino acids, which had a great influence on the beginning and end of the sequence.
【学位授予单位】：内蒙古大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q936

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