Fig. 7

Measurement of erythropoietic and vasculogenic gene transcript levels after MO injection by real-time RT-PCR. Wild-type embryos were injected with antisense MO (Zebb302), a 5-bp mismatch MO (Zebb302e), or Danieau′s solution as described under Materials and methods. Total RNA was collected from pooled embryos (7 per treatment) at each time point, and transcript levels for β-globin, klfd, klf2a, and scl were determined by the comparative threshold cycle method. For β-globin, the mRNA level for antisense-MO-treated embryos was suppressed significantly at 18 h with respect to the Danieau′s treatment (P = 0.002) but not with respect to the 5-bp mismatch MO. At 24 h, β-globin mRNA levels observed for the antisense MO were significantly different from those observed for the 5-bp mismatch MO and Danieau′s solution controls [P = 0.0016 and 0.00011, standard error of the difference (SED)], whereas the levels for the two controls were not significantly different. klfd expression was reduced significantly at 18 h (P = 0.020 and 0.0042, respectively, SED) and at 24 h (P = 0.011 and 0.019, respectively, SED) with respect to the two controls. By contrast, there were no significant differences among the three treatment conditions for klf2a and scl mRNA levels (range of P values for all time points = 0.09–0.86, ANOVA).