Antibody staining (Lecture 3, p.4) In situ hybridization

Antibody staining (Lecture 3, p.4) In situ hybridization

Antibody staining (Lecture 3, p.4) In situ hybridization (Lecture 4 p.4) mRNA depletion (knockdown) methods (Lecture 3, p.4) Wnt11 protein

Outside cell Inside cell -catenincatenin mRNA Axin

-catenincatenin builds up -catenincatenin Wnt is absent. -catenin mRNA is continuouslycatenin mRNA is continuously

translated and the protein is continuously degraded. Wnt is present. -catenin mRNA is continuouslycatenin protein is not degraded. It builds up, binds to Tcf/LEF and enters the nucleus.

Wnt, beta-catenin, and organizer formation * * * *

* * * Endo-cateninmesoderm induction. Pieter Nieuwkoop 1968 (Nieuwkoop center)

(Nieuwkoop center) Nodal signaling pathway (belongs to TGF-beta family of pathways) phospho-catenin Smad2

A dominant negative form of the Nodal (or Bmp) receptor can block Nodal (or Bmp) signaling Deplete and replace vegT mRNA vegT depleted No mesoderm or endoderm,

No blastopore vegT depleted and restored vegT mRNA restored by injection into vegetal hemisphere VegT depleted embryos: rescue with

mRNA for xnr1, 5, 6, or derriere, injected into vegetal pole cells xnr1 Normal late blastula: xnr1 expressed in vegetal cells xnr1 rescue

derriere rescue xnr5, xnr6 rescue

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