Ablations, notochord

B and E Ablation of the notochord (-Not) at the level of the anterior segmental plate, by removing neural tube plus notochord and back-grafting the neural tube alone (details in (9)) reincubation time and views as in A and D. Note that the dermomyotomal marker Pax3 (gray) and the myotomal marker MyoD (black) are correctly expressed due to the remaining expression of Shh in the floor plate of the neural tube (black, Fp). 

C and F Ablation of the posterior notochord-notochord precursors (-post Not), reincubation and views as in A and D. Note that, in the opwated region, Shh (black) and MyoD (black) are not expressed. In contrast, the expression of Pax3 (gray) is not restricted to its usual dorsal domains any more, labeling the neural tube and somite throughout (arrows). 

As the notochord is crucial for the development of the surrounding tissues but can be replaced by the floor plate of the neural tube (Fig. IB and IE), once this structure is established, the effect of notochord ablations varies depending on the type and timing of the operation. It is therefore advisable to analyze the embryos for the process of interest, but in addition for a known phenotype of the particular manipulation. This can be achieved, for instance, by double whole mount in-situ hybridization (see Fig. 1), employing molecular markers... 

Note that, when performing notochord ablations, the remarks on improving the visibility of the embryo (section 3.3.2.1) and reincubation of operated embryos (section 33.2.3) apply. 

Ablation of Posterior Notochord from Early Embryos... 

Ablation of the Notochord and Floor Plate from Late Embryos... 

To replace the ablated tissue with Shh-loaded beads, follow the instructions for notochord ablation and continne with section 3.5.2. However, the beads may be implanted anywhere in the embryo. 

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