Allele-specific detection of nascent transcripts by fluorescence in situ hybridization reveals temporal and culture-induced changes in lgf2 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 som e imprinted genes, such as Igf2, in pre-implantation embryos. Results: We have developed an allele-specific fluorescence in situ hybridiz ation method which involves a pair of oligonucleotide probes designed to de tect an intronic polymorphism. The method, called ASO-RNA-FISH, enabled us to distinguish allelic nascent Igf2 transcripts in the cell. nuclei of earl y mouse embryos, avoiding signals from the stored oocyte-specific transcrip ts, Igf2 transcription was first detectable in two-cell embryos, and bialle lic transcription was predominant up to the morula stage. Then, the materna l allele became silenced during the blastocyst stage. When embryos were cul tured in vitro, however, a strong bias to maternal transcription was observ ed up to the morula stage. Conclusion: ASO-RNA-FISH revealed that a transition of Igf2 from biallelic to monoallelic transcription occurs in the blastocyst stage. This developme ntal 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 techniqu e for the study of allele-specific gene expression.