Transfection of mouse eggs and embryos using DNA combined to cationic liposomes.
We have used plasmid DNA in combination with cationic liposomes to transfect mouse eggs and embryos. The plasmid was rhodamine labeled, which allowed a direct visualization of the DNA uptake by the cells. Immature eggs, collected from the ovaries, were easily transfected, but once the egg was ovulated the zona pellucida (ZP) acted as a barrier and prevented transfection. Permeabilization or removal of the ZP was therefore a requirement to allow transfection. Transfected eggs were capable of being fertilized in vitro giving raise to embryos that expressed the recombinant protein. Morulae and blastocysts were also transfected when the ZP was permeabilized, but the efficiency of transfection decreased and in some cases not all the blastomeres incorporated the plasmid. Pronuclear embryos were cultured and showed expression of the transgene from the 2-cell stage. This indicates that liposome-transfection of oocytes or pronuclear embryos could be a simple and suitable method to introduce foreign genes in embryos and perhaps could be also useful to generate transgenic animals.
Carballada
R
R
Centro de Investigaciones Biológicas, CSIC, Madrid, Spain. rosac@cib.csic.es
Degefa
T
T
Esponda
P
P
D000818:Animals; D002412:Cations; D005260:Female; D005306:Fertilization; D015870:Gene Expression; D049452:Green Fluorescent Proteins; D008081:Liposomes; D008164:Luminescent Proteins / Q000235:genetics; D008297:Male; D051379:Mice / Q000196:embryology; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D010063:Ovum; D016133:Polymerase Chain Reaction / Q000379:methods; D014162:Transfection / Q000379:methods*; D019076:Transgenes
D002412:Cations; D008081:Liposomes; D008164:Luminescent Proteins; D049452:Green Fluorescent Proteins