重离子诱变技术选育碱性蛋白酶高产菌株  

The Selection for High-yield Alkaline Protease Producing Strain by Heavy-ion Irradiation

作　　者：薛林贵[1] 景春娥[1] 赵旭[1] 张红[2] 武振华[2] 常思静[1] 

机构地区：[1]兰州交通大学化学与生物工程学院,甘肃兰州730070 [2]中国科学院近代物理研究所,甘肃兰州730000

出　　处：《微生物学通报》2010年第6期845-851,共7页Microbiology China

基　　金：国家自然科学基金项目(No30870384);甘肃省科技支撑计划项目(No090NKCA079)

摘　　要：从采集的土壤样品中分离筛选出一株碱性蛋白酶产生菌G-41,经16S rRNA分子鉴定为芽孢杆菌属菌株。该菌株在发酵培养基中能产生较高产量的胞外碱性蛋白酶(1.7×104U/mL)。以G-41为出发菌株,对其进行重离子辐照诱变处理,获得突变株G-41-68,将该突变株再次经重离子诱变,从大量突变株中筛选出碱性蛋白酶高产菌株15Gy-54,其酶活力达到6.22×104U/mL。与出发菌株相比较,突变株G-41-68和15Gy-54的酶活力分别提高了1.58倍和2.65倍。对突变株15Gy-54的发酵条件进行了优化研究,结果表明,该菌株的碱性蛋白酶活力得到进一步提高,达到7.18×104U/mL,其最适发酵条件为:培养基(g/100mL)为胰蛋白胨1、酵母膏0.5、乳糖5、Na2HPO4·12H2O0.4、KH2PO40.03、Na2CO30.1、MgSO40.0481(4×10-3mol/L)、pH8.0,培养温度41℃,振荡培养时间42-48h。实验结果表明,重离子辐照诱变技术是一种非常有效的微生物诱变育种新技术。Strain G-41 was isolated from the soil.The strain was identified as Bacillus by 16S rRNA method.Growing in fermentative medium,the strain can produce high-yield alkaline protease（1.7 × 104 U/mL）.We delt with the original strain（G-41） by the heavy-ion irradiation and obtained the mutant G-41-68.The mutant G-41-68 was again treated by the heavy-ion irradiation.We screened a high-yield alkaline protease producing strain 15Gy-54 from many of mutants and its enzyme activity reached to 6.22 × 104 U/mL.Compared with the original strain,the enzyme activity of mutant strains G-41-68 and 15Gy-54 increased by 1.58 and 2.65 times,respectivity.The fermentation conditions of the mutant strain 15Gy-54 were optimized and its enzyme activity further increased,reaching to 7.18 × 104 U/mL.The mutant＇s optimum conditions for enzyme production consisted of 1% tryptone,0.5% yeast extract,5% lactose,0.4% Na2HPO4·12H2O,0.03% KH2PO4,0.1% Na2CO3,4 × 10-3 mol/L MgSO4,the initial pH 8.0,shaking culture for 42-48 h,at 41℃.These results showed that the heavy-ion irradiation is an effective method for microbe mutagenesis.

关 键 词：重离子诱变 芽孢杆菌属 碱性蛋白酶 发酵 

分 类 号：Q933[生物学—微生物学]