以1株能有效改善小鼠肠炎的发酵乳杆菌AR497为研究对象,在Plackett-Burman试验的基础上选取甘氨酸质量浓度、OD600、电场强度和山梨醇浓度为自变量,电转化效率为响应值,利用Box-Behnken 响应面法优化其电转化条件。最优条件为:电场强度15 kV/cm,山梨醇浓度0.6 mol/L,OD6000.2,甘氨酸质量浓度26 g/L。在此条件下,电转化效率高达2.78×105 CFU/μg DNA,比优化前提高了22倍,实现了发酵乳杆菌AR497的高效遗传转化,为后续对其进行遗传改造和解析抗炎机制奠定基础。
Lactobacillus fermentum AR497 that can evidently relieve intestinal inflammation in mice was used as the main subject. Based on the Plackett-Burman tests, glycine concentrations, OD600, electric field strength, and sorbitol concentrations were selected as independent variables, the electroporation efficiency was the response value. Further optimization was made by using Box-Behnken response surface methodology. The optimum values for electric field strength,sorbitol content,OD600, and glycine content were 15 kV/cm,0.6 mol/L,0.2 and 26 g/L, respectively. The transformants reached 2.78×105 CFU/μg DNA under the above conditions, which was improved about 22-fold compared with that of unoptimized condition. The highly frequent genetic transformation of AR497 was realized in this work, which laid the foundation for its genetic editing and elucidating its anti-inflammation mechanism.
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