METHODS OF GENE TRANSFER - ELECTROPORATION
Electroporation is a mechanical method used for the introduction of polar molecules into a host cell
through the cell membrane.
• This method was first demonstrated by Wong and Neumann in 1982 to study gene transfer in
mouse cells.
• It is now a widely used method for the introduction of transgene either stably or transiently into
bacterial, fungal, plant and animal cells.
• It involves use of a large electric pulse that temporarily disturbs the phospholipid bilayer, allowing
the passage of molecules such as DNA.
The basis of electroporation is the relatively weak hydrophobic/hydrophilic interaction of the
phospholipids bilayer and ability to spontaneously reassemble after disturbance. A quick voltage
shock may cause the temporary disruption of areas of the membrane and allow the passage of polar
molecules. The membrane reseals leaving the cell intact soon afterwards.
Procedure
The host cells and the DNA molecules to be transported into the cells are suspended in a solution.
The basic process inside an electroporation apparatus is represented in a schematic diagram (Figure
5-3.1.1(a).).
When the first switch is closed, the capacitor charges up and stores a high voltage which gets
discharged on closing the second switch.
• Typically, 10,000-100,000 V/cm in a pulse lasting a few microseconds to a millisecond is essential
for electroporation which varies with the cell size.
• This electric pulse disrupts the phospholipid bilayer of the membrane causing the formation of
temporary aqueous pores.
• When the electric potential across the cell membrane is increased by about 0.5-1.0 V, the charged
molecules e.g. DNA migrate across the membrane through the pores in a similar manner to
electrophoresis.
• The initiation of electroporation generally occurs when the transmembrane voltage reaches at 0.5-
1.5 V. The cell membrane discharges with the subsequent
flow of the charged ions and molecules and the pores of the membrane quickly close reassembling
the phospholipid bilayer.
Applications
Electroporation is widely used in many areas of molecular biology and in medical field. Some
applications of electroporation include:
• DNA transfection or transformation
Electroporation is mainly used in DNA transfection/transformation which involves introduction of
foreign DNA into the host cell (animal, bacterial or plant cell).
• Direct transfer of plasmids between cells
It involves the incubation of bacterial cells containing a plasmid with another strain lacking plasmids
but containing some other desirable features. The voltage of electroporation creates pores, allowing
the transfer of plasmids from one cell to another. This type of transfer may also be performed
between species. As a result, a large number of plasmids may be grown in rapidly dividing bacterial
colonies and transferred to yeast cells by electroporation.
• Gene transfer to a wide range of tissues
Electroporation can be performed in vivo for more efficient gene transfer in a wide range of tissues
like skin, muscle, lung, kidney, liver, artery, brain, cornea etc. It avoids the vector-specific immune-
responses that are achieved with recombinant viral vectors and thus are promising in clinical
applications.
Advantages
• It is highly versatile and effective for nearly all cell types and species.
• It is highly efficient method as majority of cells take in the target DNA molecule.
• It can be performed at a small scale and only a small amount of DNA is required as compared to
other methods
Disadvantages
• Cell damage is one of the limitations of this method caused by irregular intensity pulses resulting in
too large pores which fail to close after membrane discharge.
• Another limitation is the non-specific transport which may result in an ion imbalance causing
improper cell function and cell death.
