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Fig. 4. Comparison of the localization of XMEF2C and Xscleraxis mRNA by whole-mount in situ hybridization. (A) Stage 38 (a–d). At stage 40 (e and f) XMEF2C begins to be expressed in the border of muscle masses of the head while scleraxis expression is faint and more diffuse around muscle masses. Lateral view (a and b). Ventral view (c–f). In somite region, dorsal views (g and h) indicate that XMEF2C expression is restricted to the intersomitic space while Xscleraxis expression appears faint and more diffuse. In lateral view of hypaxial region, XMEF2C mRNA begins to be expressed at stage 40 from the bottom (i) to the top (k) of connective tissue between muscle cells. No Xscleraxis expression was detected at these stages (j and l). (B) XMEF2C and Xscleraxis are largely colocalized in stage 43 embryo in the borders of muscle masses as indicated by MRF4 staining (c). (a), (b) and (c): ventral view of the head region, (d) and (e): lateral view of the head, (f) and (g): lateral view of the hypaxial region. The probe is indicated in each image. Arrows indicate Scleraxis expression in connective tissue associated with hypaxial muscles. (C) Scleraxis mRNA begins to be expressed in intersomitic space at stage 43. Embryos have been hybridized with XMEF2C, XMRF4 and Xscleraxis, and cohybridized with XMRF4 and Xscleraxis. XMRF4 mRNA accumulates around the large and centrally localized nuclei of muscle cells. At this stage, muscle cells are mononucleated and span the entire somite. The myotome presents a convex side on dorsal view. Arrow head indicates the intersomitic space between two convex myotome segments. The thin XMEF2C and Xscleraxis expression are distinct from the thick MRF4 expression and is localized between the convex myotome. Dorsal view on the top, higher magnification of dorsal view in the middle and lateral view at the bottom. The probe is indicated under each image. Arrows indicate Xscleraxis expression in intersomitic space. (D) single (a–d) or double (e–g) in situ hybridization of XMEF2C mRNA (blue) with XMRF4, XMEF2A or Xscleraxis mRNA (purple) in the ventral view of the head embryo at stage 41. XMEF2C mRNA is not colocalized with XMRF4 and XMEF2A mRNA while Xscleraxis begins to be strongly expressed in most of the XMEF2C expressing cells. |
