Fig. 4. The timing of the cardiac differentiation program is altered in TBX5-depleted embryos. (A) RT-PCR analysis of the expression of heart-specific isoforms of MHC, troponin and tropomyosin; and skeletal muscle-specific genes MyoD and muscle actin, throughout early and mid-tadpole stages of development in CMO (`C') and T5MO (`T') stage-matched embryos. All samples are derived from a single batch of eggs, and identical results were achieved in at least two independent sets of experiments for each marker. EF1-Alpha was used as a loading control for all RT-PCR reactions. (B-I) Images depicting embryos injected with (B-E) CMO, or (F-I) T5MO and immunostained for Tmy, showing delayed expression of Tmy in the hearts of T5MO embryos. Shown are representative sibling embryos imaged at the indicated stages. White arrows denote expression of Tmy within the heart. (J-Q) Images of living cardiac actin:GFP transgenic embryos, showing a delay in the onset of cardiac actin expression in the heart. Representative sibling embryos obtained from a single batch of embryos were injected with (J-M) CMO or (N-Q) T5MO and imaged at the indicated stages. Shown is a representative pair of embryos, while identical results were observed in over 50 embryos. White arrows denote expression of GFP within the heart field.