5-2.1.3. Lipofection

• Lipofection is a method of transformation first described in 1965 as a model of cellular membranes using liposomes.
• Liposomes areartificial phospholipid vesicles used for the deliveryof a variety of molecules into the cells. They may be multi-lamellar or unilamellar vesicles with a size range of0.1 to 10 micrometer or 20-25 nanometers respectively.
• They can be preloaded with DNA by two common methods- membrane-membrane fusion and endocytosisthus forming DNA- liposome complex. This complexfuses with the protoplasts to release the contents into the cell. Animal cells, plant cells, bacteria, yeast protoplasts are susceptible to lipofection method.
• Liposomes can be classified as either cationic liposome or pH-sensitive.

5-2.1.3.1. Cationic liposomes

• Cationic liposomes are positively charged liposomes which associate with the negatively charged DNA molecules by electrostatic interactions forming a stable complex.

Neutral liposomes are generally used as DNA carriers and helpers of cationic liposomes due to their non-toxic nature and high stability in serum.A positively charged lipid is often mixed with a neutral co-lipid, also called helper lipid to enhance the efficiency of gene transfer by stabilizing the liposome complex (lipoplex). Dioleoylphosphatidyl ethanolamine (DOPE) or dioleoylphosphatidyl choline (DOPC) are some commonly used neutral co-lipids.

• The negatively charged DNA molecule interacts with the positively charged groups of the DOPE or DOPC. DOPE is more efficient and useful than DOPC due to the ability of its inverted hexagonal phase to disrupt the membrane integrity.
• The overall net positive charge allows the close association of the lipoplex with the negatively charged cell membrane followed by uptake into the cell and then into nucleus.
• The lipid: DNA ratio and overall lipid concentration used in the formation of these complexes is particularly required for efficient gene transfer which varies with application.