Efficient encapsulation of plasmid DNA in anionic liposomes by a freeze/thaw extrusion procedure

In this study we investigated whether intact plasmid DNA can be efficiently encapsulated in anionic liposomes prepared by freeze/thaw and extrusion te chniques. There is controversy about this method of DNA encapsulation, espe cially as to whether DNA remains intact and retains its biological activity during extrusion. A solution containing supercoiled plasmid pCMV beta (716 4 base pairs) was added to dry lipid films, and after freezing and thawing, the suspension was extruded through a filter with 0.2 mu m pores. About 20 % of the DNA became encapsulated, as evidenced by protection from degradati on by endonuclease added externally. Plasmid isolated from the liposomes wa s structurally intact, and had essentially the same transfection activity a s untreated DNA. These results show that plasmid DNA can be reliably and ef ficiently encapsulated in anionic liposomes by freeze/thaw and extrusion.