Dynamic transition of Dnmt3b expression in mouse pre- and early post-implantation embryos
Section snippets
Results and discussion
In mammalian development, the first cell-lineage-allocation occurs in the morula stage, giving rise to the inner cell mass (ICM, embryonic lineage) and trophectoderm (TE, extra-embryonic lineage) of the blastocyst. Differential gene expression and epigenetic modifications are observed between the two lineages (Morgan et al., 2005). One such differential modification is ICM-specific de novo DNA methylation (Dean et al., 2001, Santos et al., 2002), but it is unknown which DNA methyltransferase is
Preparation of embryo
Embryos were obtained from ICR mice. Preparation of morulae (16- to 32-cell, E3.0) and blastocysts (with a middle sized blastocele, E3.5) was done as described previously (Hirasawa et al., 2008). E4.0 (hatched blastocysts) and E4.5 embryos were flushed from the uteri with PBS. E5.5 post-implantation embryos were dissected from the desidual tissues.
Antibodies
The anti-Dnmt3b monoclonal antibody (IMG-184, IMGENEX) recognized all known active isoforms of Dnmt3b. The specificity of the antibody was tested
Acknowledgements
We thank Drs. E. Li and M. Okano for providing Dnmt3b conditional knockout mice; Drs. M. Kaneda, Y. Kato, T. Sado and K. Hata for helpful discussion; H. Chiba, H. Furuumi and H. Inoue for technical assistance. This work was supported in part by Grant-in-Aid for Scientific Research on Priority Area from the Ministry of Education, Culture, Sports, Science and Technology of Japan to H.S. R.H. thanks the Japanese Society for the Promotion of Science for Young Scientists Fellowship.
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