Fig. 4. lbx1 is required for hypaxial muscle formation. RT-PCR was performed on tadpoles injected with lbx1-splice MO into one out of two cells or both cells at the two-cell stage. (A) A specific loss of the properly spliced lbx1 transcript is seen. Embryos were injected into one cell at the two-cell stage with a control MO (B-E), the lbx1-splice blocking MO (F-I) or the lbx1-translation blocking MO (J-M) along with ß-galactosidase mRNA as a lineage tracer (red). Tadpoles were stained with the 12/101 antibody (brown). The uninjected (B,D,F,H,J,L) and injected (C,E,G,I,K,M) sides of each tadpole are shown, where the uninjected sides are pictured with anterior towards the right. The control MO-injected tadpoles do not show a difference in the presence of rectus abdominus muscles on the injected side compared with the uninjected side at stage 37 (compare B with C) or stage 40 (D compared to E). Tadpoles injected with the lbx1-splice MO exhibit a loss of rectus abdominus and rectus cervicus muscles on the injected side at stage 37 (compare F with G). At stage 40, these muscles appear but are greatly reduced in the lbx1-splice MO-injected tadpoles (compare H with I). Similar results were observed for the lbx1-trans MO-injected tadpoles (J-M). (N,O) Stage 40 control injected (N) and lbx1-splice injected (O) embryos are shown from a ventral view, with the MO injected side downwards (embryos were injected into one cell at the two-cell stage). The geniohyoideus muscle is lost on the lbx1-splice injected side of the tadpole (O, arrow), whereas both are present in the control injected MO (N, arrow). Myoblast migration defects are observed in lbx1-splice injected tadpoles. (P-S) Tadpoles in P and R are stained for pax3 mRNA (purple) and then merged with the 12/101 antibody (green, Q,S). A control tadpole is pictured in P and Q, which shows the movement of pax3-positive myoblasts away from the differentiated epaxial myotome. In lbx1-splice-injected tadpoles (R,S), a large number of pax3-positive myoblasts remain adjacent to the differentiated myotome.