African Journal of
Biotechnology

  • Abbreviation: Afr. J. Biotechnol.
  • Language: English
  • ISSN: 1684-5315
  • DOI: 10.5897/AJB
  • Start Year: 2002
  • Published Articles: 12401

Full Length Research Paper

Optimization of plasmid electrotransformation into Escherichia coli using Taguchi statistical method

  Mohamad Heiat1, Hossein Aghamollaei1*, Seeyed mostafa Hoseinei2, Reza Abbasi Larki3 and Kheirollah Yari4  
  1Applied Biotechnology Research Center, Baqiyatallah University of Medical Science, Tehran, Iran. 2Young Researchers Club, Science and Research Branch, Islamic Azad university, Tehran, Iran. 3Department of microbiology, Islamic Azad university, Tonekabon Branch, Tonekabon, Iran. 4Medical Biology Research Center, Kermanshah University of Medical Sciences, Kermanshah, Iran.
Email: [email protected]

  •  Accepted: 16 February 2012
  •  Published: 12 April 2012

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.