论文部分内容阅读
目的利用响应面法对新一代必特螺旋霉素基因工程菌(Streptomyces spiramyceticus,WSJ-2),即含有整合型双拷贝4″位异戊酰基转移酶基因(4″-isovaleryltrasferase gene,ist)工程菌发酵培养基进行优化。方法以发酵效价和异戊酰螺旋霉素组分含量为指标,通过部分因子析因设计实验,筛选出影响较大的主要影响因子。其次,进行最陡爬坡实验,最后,通过中心组合设计,利用DX7trail 7.0软件进行回归分析,确定主要影响因子的最佳浓度,得到优化发酵培养基。结果综合培养基组成对发酵效价和组分含量的影响获得2个主要影响因子,即KH2PO4和NaCl,通过最陡爬坡实验和响应面法,确定了优化发酵培养基组成为(g.L-1):淀粉60、碳酸钙7.0、MgSO42.0、NH4NO36.0、酵母粉3.0、鱼粉15、MnCl20.10、NaCl 12.5、KH2PO40.645。验证实验表明在优化培养基中发酵相对效价为142%与预测值139%比较接近,效价比原配方提高了42%,而其总异戊酰基螺旋霉素组分的含量与原配方基本一致。结论对新构建的必特螺旋霉素基因工程菌采用响应面中心组合设计,可以较有效地进行发酵培养基的优化。
Objective To construct a new generation of Streptomyces spiramyceticus (WSJ-2), which contains an integrated two-copy 4 "-isovaleryltrasferase gene (ist) gene by response surface methodology Bacterial fermentation medium was optimized. Methods Fermentation titer and isovaleryl spiramycin were used as indexes. Through partial factorial factorial design experiments, the major influencing factors were screened out. Secondly, the steepest ascent experiment was carried out. Finally, through the central composite design, the regression analysis was carried out by using DX7trail 7.0 software to determine the optimal concentration of the main influencing factors and optimize the fermentation medium. Results Two main influencing factors, KH2PO4 and NaCl, were obtained by the influence of the composition of the integrated medium on the fermentation titer and the content of the components. The optimal fermentation medium was determined by the steepest ascent test and the response surface method as (gL-1 ): Starch 60, calcium carbonate 7.0, MgSO42.0, NH4NO36.0, yeast powder 3.0, fish meal 15, MnCl20.10, NaCl12.5, KH2PO40.645. The validation experiments showed that the relative titer of fermentation in optimization medium was 142% and the predicted value was 139%, which was 42% higher than that of the original formulation. However, the content of total isovaleryl spiramycin was almost the same as that of the original formulation Consistent. Conclusion The new construction of the biospirillum gene engineering bacteria response surface center combination design, can be more efficient fermentation medium optimization.