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Fig. 4. Inhibition of PI3K-dependent signalling enhances expression of haemopoietic marker genes. (A) RT-PCR analysis of wild-type EBs formed in the absence (0 LY) or presence of 5 µM LY294002 (LY). RT-PCR analysis of EBs formed from wild-type or PDK1-/- ES cells. ES, undifferentiated ES cells; numbers indicate the day of EB differentiation. Data are representative of three (A) and two (B) individual experiments. In A the samples for Scl, Gata-1, β-H1 and β-major were all separated on the same gel but, for simplification of presentation, blank lanes were removed between ES and 0LY d3 samples and between 0LY d7 and 5 µM LY d3 samples. (C,D) Quantitative RT-PCR analysis showing relative levels of the indicated RNA normalised to β-actin. (C) The average and s.d. of three to four replicates from the same experimental RNA as in A are shown and are representative of three independent experiments. White bars, RNA from EBs generated in the absence of LY294002; black bars, RNA samples from EBs generated in the presence of 5 µM LY294002. (D) The mean and ±s.e.m. of quadruple samples from two independent experiments are shown (n=8). White bars, RNA from EBs formed from wild type EBs; black bars, RNA samples from EBs formed from PDK1-/- ES cells. *P<0.05, **P<0.01, ***P
0.005.
