Liposome-mediated delivery of DNA to carrot protoplasts

Abstract

The encapsulation of DNA within liposomes and subsequent fusion of the liposomes with carrot (Daucus carota L.) protoplasts were examined to determine optimum conditions for effective liposome-mediated delivery of DNA to protoplasts. Escherichia coli [(3)H]DNA could be encapsulated with 50% efficiency using encapsulation volumes as low as 0.5 ml. Incorporation of liposome-encapsulated [(3)H]DNA by carrot protoplasts increased linearly for 2.5 h, and increasing the ratio of protoplasts to liposomes increased the total amount of radioactive label incorporated within the protoplasts. Liposome-mediated incorporation of [(3)H]DNA by protoplasts increased over a range of polyethylene glycol concentrations up to 20%, but Ca(2+) did not increase liposome-mediated incorporation when present in the liposome-protoplast incubation mixture. Optimum incorporation was observed when the pH of the liposome-protoplast incubation medium was decreased to 4.8. Encapsulation experiments using DNA of the plasmid pBR322 indicated that an average of 200-1,000 intact copies of pBR322 were sequestered within each nucleus after liposome delivery.