氧化胁迫联合UV、DES诱变选育高产、高分子量透明质酸菌株
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摘要
本实验以马疫链球菌MF003为出发菌,通过H2O2氧化胁迫培养、紫外辐照(UV)和硫酸二乙酯(DES)对菌株进行复合诱变,获得一株高产、高分子量透明质酸酶缺陷菌株ZD-25。经摇瓶发酵培养,该菌株透明质酸产量达到1.61g/L,相对分子量达到2.13×106u,分别比原始菌株提高了120.5%,57.8%。经过多次传代后,菌株透明质酸产量及相对分子量具有较高的稳定性。经5L发酵罐初步发酵,突变株ZD-25的透明质酸产量达到4.26g/L,相对分子量达到2.90×106u,分别比原始菌株提高了166.3%,93.3%。
Streptococcus equines MF003 was used as an original strain to perform H2O2 oxidative stress culture,ultraviolet radiation( UV) and diethyl sulfate( DES) compound mutation. After screening,a hyaluronidase deficient strain ZD-25 with high molecular weight and high- yield of hyaluronic acid was achieved. Based on shake flask fermentation,the yield of hyaluronic acid reached to 1.61 g / L and relative molecular weight of 2.13 × 106 u.Compared to the original strain,the yield and relative molecular weight were increased by 120.5% and 57.8%,respectively.The production and relative molecular weight of hyaluronic acid maintained high stability after several generation of reproduction.Furthermore,the yield of hyaluronic acid in mutant ZD-25 reached to 4.26 g / L and relative molecular weight of 2.90 × 106 u by 5L fermentation tank initial fermentation. Compared to the original strain,the yield and relative molecular weight increased 166.3% and 93.3%,respectively.
Streptococcus equines MF003 was used as an original strain to perform H2O2 oxidative stress culture,ultraviolet radiation( UV) and diethyl sulfate( DES) compound mutation. After screening,a hyaluronidase deficient strain ZD-25 with high molecular weight and high- yield of hyaluronic acid was achieved. Based on shake flask fermentation,the yield of hyaluronic acid reached to 1.61 g / L and relative molecular weight of 2.13 × 106 u.Compared to the original strain,the yield and relative molecular weight were increased by 120.5% and 57.8%,respectively.The production and relative molecular weight of hyaluronic acid maintained high stability after several generation of reproduction.Furthermore,the yield of hyaluronic acid in mutant ZD-25 reached to 4.26 g / L and relative molecular weight of 2.90 × 106 u by 5L fermentation tank initial fermentation. Compared to the original strain,the yield and relative molecular weight increased 166.3% and 93.3%,respectively.
引文
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[3]周晓惠,陈奕涵,周庆玮,等.溶菌酶胁迫培养提高透明质酸分子量[J].食品工业科技,2013,34(11):182-187.
[4]管世敏,荣绍丰,陈奕涵,等.响应面法优化氧化胁迫提高透明质酸分子量[J].天然产物研究与开发,2013,25:932-936.
[5]管世敏,荣绍丰,陈奕涵,等.外源氧化胁迫提高透明质酸分子量[J].中国医药工业杂志,2013,44(1):27-31.
[6]黄金明,张晶,汪江波.复合诱变选育透明质酸高产菌株[J].中国酿造,2010,1:73-75.
[7]陈奕涵,钱悦,侯永泰,等.复合诱变选育大分子量透明质酸高产菌株[J].中国酿造,2012,31(9):98-101.
[8]张元和,张宏.过氧化氢诱导三种念珠菌形成L型的初步观察[J].蚌埠医学院学报,1989(4):235-237.
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[10]邓静,吴明霞,吴华昌,等.不同诱变方法对兽疫链球菌产透明质酸的影响[J].安徽农业科学,2009,37(4):1391-1393.
[11]黄伟,刘永乐,王发祥,等.原生质诱变选育高产酸性α-淀粉酶黑曲霉菌株[J].食品工业科技,2014,35(3):160-162.
[12]Bitter T,Muir H M.A modified uronic acid carbazole reaction[J].Analytical Biochemistry,1962,4:330-334.
[13]凌沛学,张天民.透明质酸[M].北京:中国轻工业出版社,2000:42-43.
[14]Laurent T C.Biochemistry of hyaluronan[J].Acta Otolaryngologica,1987,442:7-24.
[15]Cleary P P,Larkin A.Hyaluronic acid capsule strategy for oxygen resistance in group A streptococci[J].Journal of Bacteriology,1979,140(3):1090-1097.
[16]Chong B F,Nielsen L K.Amplifying the cellular reduction potential of streptococcus zooepidemicus[J].Journal of Biotechnology,2003,100(1):153-162.
[17]丁勇,周楠,石孝勇.透明质酸产生菌的复合诱变及摇瓶发酵条件的优化[J].中国酿造,2011,12:100-103.
[2]李自刚,樊国燕,边传周,等.发酵法生产透明质酸菌种的筛选[J].安徽农业科学,2008,36(8):3307-3309.
[3]周晓惠,陈奕涵,周庆玮,等.溶菌酶胁迫培养提高透明质酸分子量[J].食品工业科技,2013,34(11):182-187.
[4]管世敏,荣绍丰,陈奕涵,等.响应面法优化氧化胁迫提高透明质酸分子量[J].天然产物研究与开发,2013,25:932-936.
[5]管世敏,荣绍丰,陈奕涵,等.外源氧化胁迫提高透明质酸分子量[J].中国医药工业杂志,2013,44(1):27-31.
[6]黄金明,张晶,汪江波.复合诱变选育透明质酸高产菌株[J].中国酿造,2010,1:73-75.
[7]陈奕涵,钱悦,侯永泰,等.复合诱变选育大分子量透明质酸高产菌株[J].中国酿造,2012,31(9):98-101.
[8]张元和,张宏.过氧化氢诱导三种念珠菌形成L型的初步观察[J].蚌埠医学院学报,1989(4):235-237.
[9]冯建成,张荣鹄,罗素兰.高产透明质酸菌种FJ-23的诱变选育[J].生物技术,2006,16(3):28-30.
[10]邓静,吴明霞,吴华昌,等.不同诱变方法对兽疫链球菌产透明质酸的影响[J].安徽农业科学,2009,37(4):1391-1393.
[11]黄伟,刘永乐,王发祥,等.原生质诱变选育高产酸性α-淀粉酶黑曲霉菌株[J].食品工业科技,2014,35(3):160-162.
[12]Bitter T,Muir H M.A modified uronic acid carbazole reaction[J].Analytical Biochemistry,1962,4:330-334.
[13]凌沛学,张天民.透明质酸[M].北京:中国轻工业出版社,2000:42-43.
[14]Laurent T C.Biochemistry of hyaluronan[J].Acta Otolaryngologica,1987,442:7-24.
[15]Cleary P P,Larkin A.Hyaluronic acid capsule strategy for oxygen resistance in group A streptococci[J].Journal of Bacteriology,1979,140(3):1090-1097.
[16]Chong B F,Nielsen L K.Amplifying the cellular reduction potential of streptococcus zooepidemicus[J].Journal of Biotechnology,2003,100(1):153-162.
[17]丁勇,周楠,石孝勇.透明质酸产生菌的复合诱变及摇瓶发酵条件的优化[J].中国酿造,2011,12:100-103.