Simple and effective method of electroporation for introduction of plasmid and cosmid DNAs to mammalian cells.
We have established a simple and efficient method of electroporation applicable to gene transfer in mammalian cells. It uses a single decaying pulse of around 1 ms at room temperature in the medium such as Saline G appropriate for repair of pulse-induced pores in the plasma membrane. Many types of cells (both floating and adherent) could be transformed efficiently by the electric field strengths between 1-2 kV/cm. For instance P3U1, mouse myeloma cell, could be transformed by a pulse at 1.2 kV/cm with the frequency of 10(-2) per viable cells and with survivals of 90%. We have applied these conditions to transform tsBN2 cell line of BHK21/13 by a cosmid clone (approximately 45 kb) carrying the human gene complementing to tsBN2 mutation. Significant levels of transformation were observed for this gene. Since this gene can only work as a whole size (approximately 30 kb), the results show that electroporation is useful to introduce cosmid or possibly genomic DNA to mammalian cells.
Hama-Inaba
H
H
Division of Genetics, National Institute of Radiological Sciences, Chiba, Japan.
Nishimoto
T
T
Ohtsubo
M
M
Sato
K
K
Kasai
M
M
D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000502:physiology; D002463:Cell Membrane Permeability; D003360:Cosmids; D006801:Humans; D010957:Plasmids; D014162:Transfection