Temporal and spatial expression of lipospermine-compacted genes transferred into chick embryos in vivo.
We have optimized a lipospermine-based transfection method for introducing genes into intact vertebrate embryos in vivo. The method employs small amounts of the cationic lipid Transfectam (DOGS), in a concentrated (40 mM) ethanolic solution, to compact and to transfer exogenous genes into chick embryos during the early stages of development (< 36 h of incubation). Plasmid vectors containing the reporter gene luciferase were used to follow the time course of expression. Luciferase activity was detected as early as 12 h post-transfection and was highest at this time. Enzyme activity then decreased over the next two days and was usually undetectable by 72-h post-transfection. To follow the spatial expression of the exogenous genes, a Rous sarcoma virus (RSV)-beta-galactosidase vector was used. When the transfection complex was applied externally around the developing embryo, the main site of expression was the cardiac tissue. Expression could be targeted to the nervous system by micro-injecting the DNA/DOGS (DNA/dioctadecylamidoglycylspermine) complex into the developing brain. The results show that reporter genes can be efficiently expressed in both the developing central nervous system and heart. This raises the possibility that lipospermines can be used to transfer functional genes into embryos during defined periods of development and also to deliver genes in other species and in other in vivo contexts.
Demeneix
B A
BA
Muséum Nationale d'Histoire Naturelle, Paris, France.
Abdel-Taweb
H
H
Benoist
C
C
Seugnet
I
I
Behr
J P
JP
D000818:Animals; D001358:Avian Sarcoma Viruses / Q000235:genetics; D001921:Brain / Q000196:embryology / Q000378:metabolism; D002642:Chick Embryo; D004247:DNA / Q000008:administration & dosage; D015870:Gene Expression; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D005998:Glycine / Q000008:administration & dosage / Q000031:analogs & derivatives*; D006321:Heart / Q000196:embryology; D007700:Kinetics; D008156:Luciferases / Q000235:genetics; D008845:Microinjections; D009206:Myocardium / Q000378:metabolism; D010957:Plasmids; D011401:Promoter Regions, Genetic; D013096:Spermine / Q000008:administration & dosage / Q000031:analogs & derivatives*; D001616:beta-Galactosidase / Q000235:genetics
C061134:dioctadecylamidoglycylspermine; D013096:Spermine; D004247:DNA; D008156:Luciferases; D001616:beta-Galactosidase; D005998:Glycine