Allele-specific detection of nascent transcripts by fluorescence in situ hybridization reveals temporal and culture-induced changes in Igf2 imprinting during pre-implantation mouse development.
BACKGROUND
Genomic imprinting causes parental-origin-specific monoallelic transcription of a subset of mammalian genes in the embryo and adult. There is conflicting evidence, however, for the monoallelic transcription of some imprinted genes, such as Igf2, in pre-implantation embryos.


RESULTS
We have developed an allele-specific fluorescence in situ hybridization method which involves a pair of oligonucleotide probes designed to detect an intronic polymorphism. The method, called ASO-RNA-FISH, enabled us to distinguish allelic nascent Igf2 transcripts in the cell nuclei of early mouse embryos, avoiding signals from the stored oocyte-specific transcripts. Igf2 transcription was first detectable in two-cell embryos, and biallelic transcription was predominant up to the morula stage. Then, the maternal allele became silenced during the blastocyst stage. When embryos were cultured in vitro, however, a strong bias to maternal transcription was observed up to the morula stage.


CONCLUSIONS
ASO-RNA-FISH revealed that a transition of Igf2 from biallelic to monoallelic transcription occurs in the blastocyst stage. This developmental regulation was modified temporarily by in vitro culture, suggesting a possible link between altered imprinting and abnormalities of the foetuses experienced in vitro culture. ASO-RNA-FISH is therefore a powerful technique for the study of allele-specific gene expression.
Ohno
M
M
Division of Disease Genes, Institute of Genetic Information, Kyushu University, Maidashi, Fukuoka 812-8582, Japan.
Aoki
N
N
Sasaki
H
H
D000483:Alleles; D000818:Animals; D046508:Culture Techniques; D004622:Embryo, Mammalian; D047108:Embryonic Development / Q000235:genetics*; D005260:Female; D005333:Fetus; D018507:Gene Expression Regulation, Developmental / Q000235:genetics; D018392:Genomic Imprinting / Q000235:genetics*; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D007335:Insulin-Like Growth Factor II / Q000235:genetics*; D007438:Introns / Q000235:genetics; D008297:Male; D051379:Mice; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D016254:Mutagenesis, Insertional / Q000235:genetics; D015345:Oligonucleotide Probes; D011110:Polymorphism, Genetic / Q000235:genetics; D011247:Pregnancy; D017384:Sequence Deletion / Q000235:genetics; D014158:Transcription, Genetic
D015345:Oligonucleotide Probes; D007335:Insulin-Like Growth Factor II