Breeding of Bacterial Cellulose Strain Induced by Ultra-High Pressure and Optimization of Fermentation Medium
-
摘要: 为了获得高产细菌纤维素菌株,对初选的细菌纤维素菌株J2进行超高压诱变,运用Plackett-Burman设计对影响高压诱变菌株生产细菌纤维素的因素效应进行评价,采用Box-Behnken试验优化发酵培养基组成。试验结果表明,超高压诱变压力、时间对细菌纤维素菌株有显著或极显著影响。细菌纤维素菌株高压诱变条件为压力250 MPa、时间15 min、温度25 ℃。经超高压诱变,获得产纤维素能力高、遗传稳定性好的诱变菌株M438。影响诱变菌株M438发酵生产细菌纤维素的关键因子是酵母浸出汁、MgSO4和无水乙醇。优化的发酵培养基为碳源5%(葡萄糖∶蔗糖为4∶1)、酵母浸出汁1.25%、CaCl20 15%、ZnSO4 0.20%、K2HPO4 0.20%、MgSO4 0.93%、富马酸0.30%、无水乙醇0.50%。利用此培养基培养诱变细菌纤维素菌株M438,其纤维素产量是优化前的1.84倍,是超高压诱变之前的2.69倍。超高压技术用于细菌纤维素菌株的诱变育种是可行的。发酵培养基的优化可显著提高菌株M438发酵生产细菌纤维素的能力。Abstract: In order to obtain high-yield strains of bacterial cellulose (BC), the primary bacterial cellulose strain was induced by ultra-high pressure. Plackett-Burman design was employed to evaluate variables which were relevant to yield of BC which was produced by strain induced by ultra-high pressure. Fermentation medium composition was optimized through Box-Behnken experiment. The results show that mutagenic pressure and time on the strain of bacterial cellulose have significant or extremely significant influence. The mutagenic conditions of BC strain are pressure of 250 MPa, time of 15 min, and temperature of 25 ℃. The mutagenic strain M438 with high cellulose production capacity and genetic stability is obtained through the ultra-high pressure treatment. The key factors affected bacterial cellulose production of mutation strain M438 are yeast juice, MgSO4, and anhydrous ethanol. The optimistic fermentation medium is as follows, carbon source 5% (glucose∶sucrose=4∶1), yeast juice 1.25%, CaCl2 0.15%, ZnSO4 0.2%, K2HPO4 0.2%, MgSO4 0.93%, fumaric acid 0.3%, and ethanol 0.5%. BC yield of M438 cultured with optimized fermentation medium is as 1.84 times as that cultured with the initial fermentation medium, and is as 2.69 times as that of the unmutant strain. Ultra-high pressure technology for mutation breeding of bacterial cellulose strain is feasible. BC producing capacity of M438 can be significantly improved through fermentation medium optimization.
-
Ross P, Mayer R, Benziman M. Cellulose Biosynthesis and Function in Bacteria [J]. Microbiol Mol Biol Rev, 1991, 55(1): 35-58. Ma C Z, Gu Z R. Biological and Physicochemical Properties and Commercial Application of Bacterial Cellulose Produced by Acetobacter xylinum [J]. Acta Agriculture Shanghai, 2001, 17(4): 93-98. (in Chinese) 马承铸, 顾真荣. 细菌纤维素生物理化特性和商业用途 [J]. 上海农业学报, 2001, 17(4): 93-98. Tan Y J, Hong F, Shao Z Y. Bacterial Cellulose Application in Foods [J]. China Biotechnology, 2007, 27(4): 126-131. (in Chinese) 谭玉静, 洪枫, 邵志宇. 细菌纤维素在食品中的应用 [J]. 中国生物工程杂志, 2007, 27(4): 126-131. Pu H Y, Chen Y Q. Bacterial Cellulose and Its Application in Food Industry [J]. Journal of Cereals and Oils, 2005, (9): 16-17. (in Chinese) 蒲海燕, 陈宇前. 细菌纤维素及其在食品工业中的应用 [J]. 粮食与油脂, 2005, (9): 16-17. Song H N, Zhang Y Q, Guo H Q. Bacterial Cellulose the Applications and Prospect on Papermaking [J]. Journal of Guangxi University, 2004, 29(1): 73-76. (in Chinese) 宋海农, 张远秋, 郭华清. 细菌纤维素在造纸工业中的应用和展望 [J]. 广西大学学报(自然科学版), 2004, 29(1): 73-76. Sanchavanakit N, Sangrungraungroj W, Aomongkolgit R, et al. Growth of Human Keratinocytes and Fibroblasts on Bacterial Cellulose Film [J]. Biotechnology Prog, 2006, 22(4): 1194-1199. Ma X, Chen S W, Wang R M, et al. Nanometer Biomaterial Cellulose for Accelerating Skin Wound Healing in Rats [J]. Chinese Journal of Clinical Rehabilitation, 2006, 10(37): 45-48. (in Chinese) 马霞, 陈世文, 王瑞明, 等. 纳米材料细菌纤维素对大鼠皮肤创伤的促愈作用 [J]. 中国临床康复, 2006, 10(37): 45-48. Nguyen V T, Gidley M J, Dykes G A, et al. Potential of a Nisin-Containing Bacterial Cellulose Film to Inhibit Listeria monocytogenes on Processed Meats [J]. Food Microbiology, 2008, 25(3): 471-478. Wang S L, Wu X Z, Hao L H, et al. Mutation Effect of Ultra High Pressure on Industrialized Microbe [J]. China Biotechnology, 2005, 25(6): 7-9. (in Chinese) 王岁楼, 吴晓宗, 郝莉花, 等. 超高压在工业微生物诱变选育中的应用初探 [J]. 中国生物工程杂志, 2005, 25(6): 7-9. Gao X, Li J, Ruan K C. Barotolerant E. coli Induced by High Hydrostatic Pressure [J]. Acta Biochimica et Biophysica Sinica, 2001, 33(1): 79-84. (in Chinese) 高翔, 李炯, 阮康成. 高压力诱变的耐压大肠杆菌 [J]. 生物化学与生物物理学报, 2001, 33(1): 77-81. Silva J L, Luan P, Glaser M, et al. Effects of Hydrostatic Pressure on a Membrane-Enveloped Virus: High Immunogenicity of the Pressure-Inactivated Virus [J]. J Virology, 1992, 66(4): 2111-2117. Wang S L, Wu X Z, Hao L H, et al. Mutation Effect of Ultra High Pressure on Microbe [J]. Acta Microbiologica Sinica, 2005, 45(6): 970-973. (in Chinese) 王岁楼, 吴晓宗, 郝利花, 等. (超)高压对微生物的影响及其诱变效应探讨 [J]. 微生物学报, 2005, 45(6): 970-973. Wang S L, Li G F, Wang Q B, et al. Mutagenic Effect of UV Radiation and Ultra High Pressure on Strains Producing Laccase [J]. Journal of Zhengzhou University of Light Industry, 2005, 20(3): 39-42. (in Chinese) 王岁楼, 李国富, 王琼波, 等. 紫外和超高压诱导漆酶产生菌变异的对比研究 [J]. 郑州轻工业学院学报(自然科学版), 2005, 20(3): 39-42. Wang H, Shen S L, Li Z H, et al. High Pressure-Induced Changes of Physico-Chemical Properties and Mutagenesis in Aspergillus Oryza [J]. Chinese Journal of High Pressure Physics, 2008, 22(3): 259-263. (in Chinese) 王华, 申斯乐, 李志慧, 等. 高压对米曲霉理化性质影响及诱变的研究 [J]. 高压物理学报, 2008, 22(3): 259-263. Wu R Q, Li Z X, Shao D Y, et al. Screening and Primary Identification of Bacterial Cellulose Producing Strain [J]. China Brewing, 2008, (10): 37-38. (in Chinese) 毋锐琴, 李志西, 邵东燕, 等. 高产细菌纤维素菌株的分离及初步鉴定 [J]. 中国酿造, 2008, (10): 37-38. Wu R Q, Du S K, Li Z X, et al. Optimization of Bacterial Cellulose Fermentation Medium and Observation of Bacterial Cellulose Ultra-Micro-Structure [J]. Chinese Journal of Biotechnology, 2008, 24(6): 1068-1074. (in Chinese) 毋锐琴, 杜双奎, 李志西, 等. 细菌纤维素发酵培养基的优化及超微观结构分析 [J]. 生物工程学报, 2008, 24(6): 1068-1074. Guo W Q, Ren N Q, Wang X J, et al. Optimization of Culture Conditions for Hydrogen Production by Ethanoligenens Harbinense B49 Using Response Surface Methodology [J]. Bioresource Technology, 2008, 100(3): 1192-1196. Banik R M, Santhiagu A, Upadhyay S N, et al. Optimization of Nutrients for Gellan Gum Production by Sphingomonas Paucimobilis ATCC-31461 in Molasses Based Medium Using Response Surface Methology [J]. Bioresource Technology, 2007, 98(4): 792-797. Sheu F, Wng C L, Shyu Y T. Fermentation of Monascus purpureus on Bacterial Cellulose-nata and the Color Stability of Monascus-nata Complex [J]. J Food Sci, 2000, 65(2): 342-345. Dalian Polytechnic University, Zhengzhou University of Light Industry. Food Analysis [M]. Beijing: China Light Industry Press, 2006: 100-216. (in Chinese) 大连轻工学院, 郑州轻工学院. 食品分析 [M]. 北京: 中国轻工业出版社, 2006: 100-216. Yang G. Microbiology Experiment [M]. Beijing: Science Press, 2004: 159-170. (in Chinese) 杨革. 微生物学试验教程 [M]. 北京: 科学出版社, 2004: 159-170.
点击查看大图
计量
- 文章访问数: 7942
- HTML全文浏览量: 258
- PDF下载量: 605