Requirement for Pax2.1 in early MHB development. Wholemount ISH; embryos shown in one row are stained with the marker depicted on the left side. Stages and genotypes are noted; shown are lateral views, anterior to the left, except where indicated. A prime (′) indicates mutants. (A) eng3 expression is not activated in noitu29a. (B) In weak noity31a mutants expression fades away from ventral to dorsal. (C,D) Weak transient eng2 expression in the MHB primordia, dorsal views, anterior to the top; in noitu29a (C) the expression is very faint and lost at the end of epiboly; in strong noitm243a mutants (D) expression initiates normally, fades away around 2 somites, and is lost later in development. (E) Dorsal views and (F) optical cross sections through the eng2 domain of wild-type embryo and weak noitb21 mutants. Expression persists dorsally. (G,H) Expression of pax2.1 as a marker for MHB development is initiated normally in noitu29a mutants, and is gradually eliminated during somitogenesis stages. (G) pax2.1 expression in noitu29a is smaller in its a/p extent than in the wild type; later deletion is complete in the weak noitb21 allele (H). (I) At the 5-somite stage, her5 expression fades away from the MHB, except for a ventral patch in the neural tube (Q: cross section, arrowheads outline the neural keel). (J) In weak mutants, the midbrain primordium still expresses her5, also to later stages. (K) The dorsal midbrain expression of wnt1 is seen at 9 somites in noitu29a, whereas MHB expression is lost, similar to the weak noitb21 allele at later stages (L). (M) Following normal initiation, fgf8 expression is absent from the MHB except in the ventral portion (R: cross section; arrowheads outline the neural keel), that has disappeared by 22 hours (N). (O,P) At the 5-somite stage, MHB cells are present in noitu29a mutants and express fgf8 overlapping with eng3 (red) (O) and wnt1 (P). Brackets mark the MHB.