Electronic Journal of Biotechnology
Universidad Católica de Valparaíso
Vol. 11, No. 1, 2008
Bioline Code: ej08015
Full paper language: English
Document type: Short Communication
Document available free of charge
Electronic Journal of Biotechnology, Vol. 11, No. 1, 2008
© Copyright 2008 - Pontificia Universidad Católica de Valparaíso -- Chile
Multiple pulses improve electroporation efficiency in Agrobacterium tumefaciens |
Mahmood, Tariq; Zar, Tamkina & Naqvi, S.M. Saqlan
Electroporation entails brief, high intensity pulse to create transient pores in the cell membrane to facilitate the entry of exogenous macromolecules, which may otherwise be excluded. Removal of the external field leads to the resealing of the membrane electropores permitting the survival of the electrically stimulated recipient cells. Using this technique foreign deoxyribonucleic acid (DNA) has been successfully introduced into many cell types both from prokaryotes and eukaryotes. Increase in pulse voltage and length beyond a critical limit has been reported to decrease transformation efficiency, hence in this study we have investigated another strategy i.e. increase in the number of pulses at constant high voltage and pulse duration. Commonly used Agrobacterium strains LBA4404 and EHA101 and binary vector pCAMBIA1301 were used. Transformants were selected on a combination of hygromycin and kanamycin, and confirmed by polymerase chain reaction (PCR) and restriction analysis. Increase in the number of pulses was found to show a significant and linear increase in transformation efficiency.
Agrobacterium, electroporation, pulse number, transformation efficiency.
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