Knockout of Eed/Ring1b increases expression of Hoxb genes but only Hoxb1 responds additionally to valproic acid. (A) Transcript levels of Hoxb genes were quantified by real time-quantitative (RT-q)PCR in wild-type embryonic stem (ES) cells (J1 line) and in J1 cells in which two genes encoding Polycomb proteins (Eed; Ring1b) had been knocked out (dKO cells). Light grey bars show the expression levels in wild-type J1 cells, and dark grey bars represent expression in dKO cells. (B) dKO cells were treated for 8 h with valproic acid (VPA) (1 mM). Hoxb transcript levels in treated and untreated cells were assayed by RT-qPCR. Light grey bars represent untreated dKO cells, dark grey bars show data for VPA-treated cells. For both panels error bars represent the standard error of the mean of independent experiments (n = 3). Two-tailed unpaired t-tests were used to assess the significance of differences; *P < 0.05, **P <0.01.