Figure 2. Oct4 is required for early embryo development prior to formation of the blastocyst.

(A) Oct4-MO-injected embryos arrested at multicell stage, while uninjected and mismatch (Oct4-MM) controls reached blastocyst stage. (B) Oct4 knockdown induced higher arrest rates at the 1- to multicell stages; p<0.01. (C) None of the Oct4-MO-injected embryos developed to blastocysts. (D) The rates of arrest at the 1- to multicell stages decreased with concentration of Oct4-MO (p<0.01). (E) There was a non-significant trend for higher rates of blastocyst development with decreasing concentrations of Oct4-MO. (F) Nuclear Oct4 expression is absent in Oct4-MO-injected embryos (top panel) but present in Oct4-MM-injected embryos and uninjected. The effect of Oct4 knockdown could not be assessed by western blot because Oct4 protein was not detectable in pooled embryos by western blot (SI Fig. 6). (G) Compared to no coinjection or mEYFP mRNA co-injection, co-injection of 36 or 3.6 ng/µL of Oct4 mRNA resulted in partial rescue of the Oct4-MO-induced phenotype by specifically decreasing the arrest rates at the 1- to multicell stages, which resulted in higher rates blastocyst development (p<0.01). (H) Overexpression of Oct4 mRNA induced higher developmental arrest in a dose-dependent manner, such that blastocyst rates are significantly lower after injection of Oct4 mRNA at 36 or 90 ng/µL, compared to overexpression of mEYFP (p<0.01). (Scale bar = 40 µm).