Electroporation is the procedure of using high intensity electric pulses to produce transient pores in the cell walls/cell membranes which enables the uptake of DNA. The efficiency of the procedure depends upon multiple factors, which could include the growth phase of the cells, the composition of the electroporation buffer, properties of the plasmid DNA including size and concentration, the voltage and time constant of the electric pulse, and the composition of the recovery medium.
Design of Experiments (DoE) is a statistics-based method that enables the efficient optimization of systems where a large number of input variables impact the outcome of the process. In this study, we identified 14 variables that could influence the electroporation efficiency for a Streptomyces species, and the experimental design space included 207,360 possible combinations of these variables.
By adopting a DoE approach, we were able to screen this large design space very efficiently with just 42 experiments. Within these 42 experiments, we found 5 sets of conditions that provided a significant improvement in the transformation efficiency compared to the baseline, with the best condition providing a 4.5X improvement over the baseline (~15000 transformants/μg DNA vs ~3300 transformants/μg DNA). The statistical analysis also revealed that 4 of the 14 variables were the most significant factors impacting the transformation efficiency, and future optimization efforts can focus on further refining this smaller variable set.
This presentation will provide an overview of our results and highlight the use of DoE as a powerful tool for the optimization of complex, multivariable processes such as the one described above.