Expression and subcellular localization of Chd8 in cardiac development and in post-natal NCMs.

<p>A) A representation of the targets of the two sets of TaqMan probes used to measure Chd8 mRNA. Probe 1 (ABI IDRn00576005_m1) spans exon 2–3 and covers the RII binding domain. Probe 2 (ABI Rn01414467_m1) spans exons 12–13, which encode the helicase domain and detects only the two longest isoforms. B) Relative amounts of mRNA of Chd8 (Probe 1 and Probe 2) and RIIα (PKAR2A), normalized to GAPDH and calculated by the 2^−ΔΔCtt method. C) Western blot was used to detect Chd8-L1 and Chd8-L2 in NCMs. HeLa cell lysate was used as a positive control. D) NCMs were fixed at four days in culture and stained for Chd8 (green) and α-actinin (red). Arrows indicate myocytes, while arrowheads indicate fibroblasts. E) NCMs were fixed at four days in culture and stained for α-actinin (green) and RIIα/β (red). Arrows indicate myocytes, while arrowheads indicate fibroblasts. Cells were imaged with inverted fluorescence microscopy and images taken at 90X magnification. Scale bar represents 25 µm.