Abstract

Electroporation is a mechanical method used to introduce polar molecules into a host cell through the cell membrane. In this procedure, a large electric pulse temporarily disturbs the phospholipid bilayer allowing molecules like DNA to pass into the cell. Application of statistical methods to determine the appropriate processes have been suggested for genetic engineering and biotechnology technique such as electroporation. This study explains the use of Taguchi statistical method to optimize the conditions for efficient plasmid transformation into Escherichia coli via electroporation. In order to improve electroporation, optical density of bacteria, recovery time and electrical parameter (field strength and capacitance) were optimized using the Taguchi statistical method. ANOVA of obtained data indicated that the optimal conditions of electrotransformation of pET-28a (+) plasmid intoEscherichia coli BL21(DE3)pLysS was 0.7, 120 min, 12 kV/cm and 50 µF, for optical density of cell culture, recovery time, field strength and capacitance, respectively. The most significant alterations are decrease in field strength and increase in optical density in comparison with common electroporation protocol. The maximum level of plasmid transformation obtained under optimal condition was 8.7×10⁸ transformants/μg DNA plasmid, which was 6.7 fold higher than the control condition.

Kay words: Electroporation, Taguchi statistical method, Plasmid.

Abbreviation

Abbreviation: ANOVA, Analysis of variance.