FIG. 9.

Perinucleolar association present in two- and four-cell embryos is lost during the 8-cell stage. (A) Cot-1 and Xist RNA FISH with subsequent DNA FISH using the Xic probe, Sx7, in the wild-type four-cell embryo. (B) Cot-1 RNA FISH with subsequent DNA FISH in the Xist mutant four-cell embryo using a combination of Sx7 and πXE9 probes to distinguish X^M (wild-type) from X^P (Xist deficient). For panels A and B, two z sections (top and bottom) are shown for each blastomere to capture the X^M and X^P planes. (C) Xic nucleolar association of X^P versus that of X^M in 4- and 8-cell embryos. Cot-1 RNA FISH was performed on wild-type and Xist-deficient embryos as described for Fig. 7C with the following exception: for 4- and 8-cell embryos, nucleolar association correlated with a Cot-1⁻ state of X^P in 100% of blastomeres; however, for the wild-type embryos, a lack of nucleolar association was correlated with silencing in a fraction of blastomeres. P values were calculated using the unpaired student t test. WT, wild-type. Xist−, X^MX^P;Xist−. (D) X chromosome painting of a two-cell embryo of an Xist mutant reveals large X territories (circled) at this stage. (E) Cot-1 RNA FISH and subsequent X chromosome painting show no compaction of X^P in the Xist mutant embryo at the blastocyst stage. (F) Pictorial representation of the deduced X^P and X^M structures in the early embryo. Repeat elements of X^P lie in the silent perinucleolar compartment, while X^M and active genic loci of X^P reside in Cot-1⁺ regions.