Figure 1.

Patterns of 5meC and 5hmC staining in the oocyte and different stages of preimplantation embryo development. (A) z-stack projection images of staining for DNA (DAPI, blue), 5meC (green), 5hmC (red), and their merged (merge) images in the oocyte; zygote (PN1 and PN3); 2-cell, 4-cell, and 8-cell embryos; morula (ML); blastocyst (BL); and a BL stained with non-immune control IgG. (*) indicates the decondensing sperm head (in newly fertilized oocyte) or the male pronucleus (in PN3 zygote). (open star) indicates a female pronucleus. (open triangle) indicates the extruded polar body. (B) Single confocal optical sections through morulae triple stained for 5meC (green), 5hmC (red) and DNA (blue). The images show eight sequential sections taken at 0.88-μm intervals taken through morulae (1–8). (C) Single confocal optical sections through a blastocyst triple stained for 5meC (green), 5hmC (red), and DNA (blue). The images are sequential sections at 1-μm intervals at the lower pole (1–4, top row), through the equator (5–8, middle row), and upper pole (9–12, bottom row) of the embryo. Cells in the upper and lower poles are TE cells, the equatorial sections show the ICM (ICM enclosed by dotted line) encircled by the TE. (*) indicates blastocoel cavity. (D) Single confocal optical sections of staining of 5meC (green), 5hmC (red), and both merged in the individual maternal pronucleus (zygote (m)); paternal pronucleus (zygote (p)), and nuclei from two cell, four cell, eight cell embryos, and the outer and inner cells of morula (ML (o), ML (i)); and the TE and ICM of blastocysts. White arrows show representative examples of 5meC-intense staining foci; white *shows examples of nucleoli precursor bodies or nucleoli in late stage embryos. All scale bars, 10 μm.