Optimization of electroporation method of Streptococcus thermophilus AR333 by response surface methodology

Abstract:

[Background] Streptococcus thermophilus AR333, isolated from fermented dairy products, could produce a high-yield active exopolysaccharide. [Objective] To establish a high-efficiency electroporation method of S. thermophilus AR333. [Methods] Electroporation conditions were optimized by single factor experiment and Box-Behnken response surface methodology. [Results] The optimum condition was glycine content 8.3 g/L, cell density of 0.8 (OD600), buffer composed of 10% glycerol (V/V) and 0.5 mol/L sucrose, 80 ng pIB184 plasmid, electric field strength 14 kV/cm, the recovery LM17 media composed of 0.4 mol/L sorbitol, 2 mmol/L CaCl2 and 20 mmol/L MgCl2 and recovery time of 5 h. [Conclusion] The transformants reached 3.68×105 CFU/μg-DNA under the optimal condition, which was improved about 14-fold in comparison with that of initial condition. The optimized condition was used to achieve a high electro-transformation efficiency, which lays a foundation for genetic engineering of AR333.

Key words:Response surface methodology, Streptococcus thermophilus, Electroporation, Transformation efficiency