Electroporation

* This product is for research use only. Not intended for use in the treatment or diagnosis of disease.

Electroporation is the most effective non-viral gene delivery method for introducing DNA, RNA, mRNA, RNP, proteins, and other molecules into a variety of cells (especially cells that are difficult to transfect, such as primary and stem cells). By using a precise pulse current, it can induce transient pores in the phospholipid bilayer of the cell membrane, so that the cell can absorb extracellular genetic material.

Studies have shown that during electroporation, the increased transmembrane voltage plays two roles: (a) forming pores and (b) providing local driving force.

During the opening of the pore, the nucleic acid can enter the cell and eventually enter the nucleus. Linear DNA with free ends has higher recombination capacity and is more likely to integrate into the host chromosome to produce stable transformants. Supercoiled DNA is more easily packaged into chromatin and is usually more effective for transient gene expression.

Electroporation applications

Electroporation is used in the fields of molecular biology research and medicine.

  • In vitro electroporation
  • Electroporation can be used to transfer a variety of genetic material into cells, including DNA, RNA, and oligonucleotides (including synthetic oligonucleotides with uncharged backbones), and to transfer plasmids directly between cells.

  • In vivo electroporation
  • In vivo electroporation has been shown to be effective for a variety of tissues, including tumors, skin, liver, lung, kidney, thymus, bladder, adipose tissue, vasculature, retina, cornea, ciliary muscle, brain, the umbilical cord of the spinal cord, skeletal muscle, and testis. It can be used to deliver a series of genetic material, such as DNA, RNA, and oligonucleotides (e.g., siRNA, antisense oligonucleotides).

    Now, in vivo electroporation has been used for transdermal drug delivery, cancer tumor electricity chemotherapy, and the effective delivery of gene therapy and vaccines.

Common applications of electroporation Figure 1: Common applications of electroporation

Table 1: Brief of electroporation

What can be electroporated?Typical transfectants
Bacteria
Fungus/yeast
Plants
Others, insects, fish, mold and amphibians
Mammalian
Primary explant culture
Established cell line
Human, in vitro, in vivo and ex vivo
DNA/RNA
Antibodies/proteins
Drugs
Other molecules/ions

Important tips for optimal electroporation

Optimize electroporation conditions for each cell type to ensure successful results. The following suggestions can help produce high-efficiency electroporation for most cell types.

  • Cell density
    • Perform subculture about 18–24 hours before electroporation to ensure that cells can actively divide and reach an appropriate density during transfection (i.e., 70-90% for adherent cells or 2-4×106 cells/ml for suspension cells).
  • Cell passage number
    • Using very low or very high number of passage cells may affect the experimental results. Repeat the experiment using cells with similar passage numbers.
  • Purity and concentration of nucleic acid
    • Use highly purified, sterile, and pollution free DNA for electroporation.
    • Plasmid DNA preparations that do not contain endotoxins and have an A260/280 absorbance ratio of 1.8-2.0 are ideal.
    • Use a DNA stock solution with a concentration range of 1–5 mg/ml. Too high concentration of DNA stock solution may cause uneven mixing with cells. Too low concentration of DNA stock solution may dilute the electroporation mixture and reduce the efficiency of electroporation.
  • Post-electroporation incubation time

    According to the purpose of the experiment, the incubation time is determined by the nature of the plasmid and the half-life of the expressed protein.

    • For plasmid electroporation, the optimal incubation time is usually 12-48 hours.
    • For siRNA-mediated knockout experiments, the optimal incubation time can be determined empirically by testing the stability of target mRNA and its encoded protein within 24-72 hours after electroporation.

Reference

  1. Bolhassani, A., Khavari, A., & Oraf, Z. Electroporation – Advantages and Drawbacks for Delivery of Drug, Gene and Vaccine. Application of Nanotechnology in Drug Delivery. 2014. doi:10.5772/58376.