Electronic Journal of Biotechnology
Universidad Católica de Valparaíso
Vol. 13, No. 5, 2010
Bioline Code: ej10065
Full paper language: English
Document type: Note
Document available free of charge
Electronic Journal of Biotechnology, Vol. 13, No. 5, 2010
© Copyright 2010 - Pontificia Universidad Católica de Valparaíso -- Chile
Enhancing DNA electrotransformation efficiency in Escherichia coli DH10B electrocompetent cells|
Wu, Ning; Matand, Kanyand; Kebede, Bizuayehu; Acquaah, George & Williams, Sonya
Electrotransformation also known as electroporation is the most reliable and efficient tool for plasmid DNA uptake. Electrotransformation efficiency is function of many factors which include (1) number of cell washes prior to electroporation, (2) electroporation cell number, (3) electroporation DNA amount, and (4) cell growth phase. Those factors have limitedly been concomitantly investigated in E. coli DH10B strain. This study is aimed to explore above key factors to define the optimal conditions for high electrotransformation efficiency. The results showed that electrotransformation efficiency of E. coli DH10B was enhanced to 1.5 x 109 cfu/μg by washing cells three times with 15 ml of 10% glycerol. This washed off extra salts from cell suspension and enhanced electrotransformation by preventing arcing and enhancing cell resistance while ensuring minimal level of conductivity. Early exponential phase at 0.15 OD600 was the best growth phase for enhancing electrotransformation of E. coli DH10B. The results also showed that higher electrotransformation efficiency was similarly achieved when 0.5 x 1010 and 0.6 x 1010 cell numbers were electroporated with DNA amount ranging from 10 to 40 pg. This study confirmed the optimal conditions for electro competent cell preparation and plasmid DNA electrotransformation, which can result highest transformation efficiency.
cell competency, DNA, E. coli DH10B, electroporation.
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