不同培养条件对内生菌A02、A08固氮酶活力的影响及培养条件优化

Effects of Culture Conditions on Nitrogenase Activity of Endophytes A02, A08 from Cassava and Optimization of Culture Conditions

【目的】解析不同培养条件对菌株A02,A08固氮酶活力的影响,以期找到菌株的培养条件最优组合,为木薯固氮菌的培养及应用提供理论依据和技术支撑。【方法】采用单因素浓度梯度法研究5个培养条件(pH、温度、KNO3、NH4Cl、O2)对2个菌株固氮酶活性的影响;利用正交实验确定2个菌株的培养条件优化体系。【结果】2个菌株的固氮酶活性随着pH、温度和KNO3浓度的增加呈现出先升后降的规律;菌株A02和A08固氮酶活性随着NH4Cl和O2浓度升高呈现出线性下降趋势,其中A02对NH4Cl浓度增加更敏感,而A08对O2浓度升高更敏感。通过正交试验优化菌株A02的最适培养条件组合是pH 7.5,温度25℃,KNO38 g/L,O2浓度21%,此时A02固氮酶活力达935 nmol/(mL·h),各条件的影响效应为温度>O2>pH值>NH4Cl>KNO3;菌株A08的最佳培养条件组合是pH 7.5,温度25℃,KNO36 g/L,NH4Cl 8 g/L,此时固氮酶活力为768 nmol/(mL·h),各培养条件的影响效应为温度>KNO3>pH值>O2>NH4Cl。【结论】通过正交试验获得木薯内生固氮菌A02和A08的最适培养体系,在最适条件下2个菌株的固氮酶活性得到显著提高,为2种菌株在木薯生产中的应用奠定基础。

【Objective】The study aimed to analyze the effects of different culture conditions on nitrogenase activity(NA)of strains A02 and A08 and optimize the combination of them,providing evidence and technical support for culture and application of endophytic nitrogen-fixing bacteria.【Method】The effects of five culture conditions,including pH,temperature,KNO3,NH4Cl,O2 on the NAs of A02 and A08 were studied by single factor concentration gradient method,the orthogonal experiment was used to optimize the culture system for two strains.【Result】The NAs of A02 and A08 increased first and then decreased with increasing pH,temperature and the concentration of KNO3,and showed a linear downward trend with increasing concentration of NH4Cl and O2.A02 was more sensitive to the increase of NH4Cl concentration,while A08 was more sensitive to the increase of O2 concentration.Orthogonal experiments of culture conditions showed that pH 7.5,temperature 25℃,KNO38 g/L and oxygen 21%were optimal for A02,while pH 7.5,temperature 25℃,KNO36 g/L and NH4Cl 8 g/L for A08.Under these optimal culture conditions,the NAs of A02 and A08 could increase up to 935 nmol/(mL·h)and 768 nmol/(mL·h),respectively.The order of culture conditions according to their effects on NAs was:temperature>O2>pH value>NH4Cl>KNO3 for A02 and temperature>KNO3>pH value>O2>NH4Cl for A08.【Conclusion】The optimal culture conditions of A02 and A08 were obtained by orthogonal experiments,the NAs of the two strains increased significantly under the optimum conditions,which laid a foundation for the application of both strains to promote cassava production.