Transient expression of a human beta-actin promoter/lacZ gene introduced into mouse embryos correlates with a low degree of methylation.
We have analysed the correlation between expression and methylation for the human beta-actin promoter introduced into mouse embryos. The beta-actin promoter was fused to the reporter gene lacZ, and expression was analysed after pronuclear injection into fertilized mouse eggs. We analysed transient expression in in vitro cultured preimplantation embryos and expression after chromosomal integration in 5 independent lines of transgenic mice. The in vitro cultured preimplantation embryos expressed lacZ from the 2-cell to the blastocyst stages, and most abundantly at the morula stage. By increasing the amount of injected DNA, a larger proportion of embryos expressed lacZ. Embryos expressing lacZ in only a subset of the blastomeres were detected at all preimplantation stages. In contrast to the transient expression after injection, we have not detected lacZ expression in any of the 5 analysed lines of transgenic mice carrying the same construct. The lack of expression in transgenic mice correlates with hypermethylation of C residues in the vast majority of CG sequences in the integrated beta-actin/lacZ construct, whereas the injected construct was completely nonmethylated. We discuss methylation and other possible reasons for the observed differences in expression between injected and integrated copies of the beta-actin/lacZ construct and for lacZ expression in only a subset of blastomeres in preimplantation embryos.
Nilsson
E
E
Department of Molecular Genetics, Medical Nobel Institute, Karolinska Institute, Stockholm, Sweden.
Lendahl
U
U
D000199:Actins / Q000235:genetics*; D000818:Animals; D001374:Azacitidine / Q000494:pharmacology; D001483:Base Sequence; D001755:Blastocyst / Q000187:drug effects / Q000378:metabolism; D001757:Blastomeres / Q000378:metabolism; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects; D005813:Genes, Synthetic; D006801:Humans; D008297:Male; D008745:Methylation / Q000187:drug effects; D051379:Mice; D008822:Mice, Transgenic; D008845:Microinjections; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics; D001616:beta-Galactosidase / Q000096:biosynthesis* / Q000235:genetics
D000199:Actins; D011993:Recombinant Fusion Proteins; D001616:beta-Galactosidase; D001374:Azacitidine