敲除frdB基因对大肠杆菌厌氧混合酸发酵的影响

Effect of Knockouting frdB on Anaerobic Mixed Acid Fermentation for Escherichia coli

以SPUEC101(产琥珀酸)为出发菌,利用RED同源重组技术敲除延胡索酸还原酶基因frdB,得到重组菌株SPUEC103(△frdB),通过减少延胡索酸生成琥珀酸的通量,实现延胡索酸的积累。实验结果表明:敲除frdB基因后,缺陷菌株生长速率降低,利用葡萄糖的能力也有所降低,同时敲除frdB基因较大程度地改变琥珀酸、延胡索酸等的分布,在两阶段发酵中,当发酵培养基中添加30 g/L的葡萄糖时,琥珀酸和延胡索酸得率最高,对比SPUEC101,SPUEC103的琥珀酸产量产率由24.6%下降为15.4%,并有延胡索酸和少量的苹果酸生成,分别为0.182±0.002 g/L和0.023±0.002 g/L,同时丙酮酸和乙酸含量也略有升高,分别由1.87±0.02 g/L、0.012±0.002g/L上升到2.36±0.03 g/L、0.862±0.012 g/L。

A frdB defection strain, SPUEC103 （AfrdB） was constructed, by using k-RED homologous recombination technology with QLUEC001 （produced succinate） as the parent strain. The fumarate accumulated by decreasing the flux of conversion of fumarate to succinate, frdB The fermentation results showed that the mutants of Escherichia coli deficient in frdB showed slower growth, together with exploiting less glucose. At the same time, the defection of frdB could change the yields of succinate and fumarate, the yield of succinate and fumarate was the highest in LB media supplemented with 30 g/L glucosethe, the yield of succinate of dual-phase fermentation decreased from 24.6% to 15.4%. The portion of fumarate and malate in SPUEC103 increased, the final concentration of the fumarate and malate was 0. 182 ± 0. 002 g/L and 0. 023 ± 0. 002 g/L. And the concentration of pyruvate and acetate decreased from 1.87 ±0.02 g/L and 0. 012 ±0. 002 g/L to 2.36±0.03 g/ L and 0. 862±0. 012 g/L, respectively.