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Fig. 1. Dissections and recombinations. (A) Diagram of the tissue
manipulations used for these experiments. The top arrows indicate separation
of tips and stalk, and the culture of the amputated stalk with fresh
mesenchyme, the middle arrows indicate injury to mesenchyme (mes) alone, or to
stalk (ub) and mesenchyme, and the bottom arrows indicate culture of the
`wrong' end of the stalk with fresh mesenchyme. (B-D) Discarded
portions stained with an in situ probe show the complete
Wnt11-expressing tip regions (B), in addition to short regions of
Wnt11-negative stalks; those stained with the stalk-specific stain
DBA again show that the tips and a short length of stalk are present in the
discarded region (C). Staining the same specimen with
calbindin-D28K, which stains both tips and stalks (D), shows where
the tips are. Scale bar: 100 µm. (E) Dilution PCR analysis of
Wnt11 expression in kidneys, including tips, and in de-tipped stalks.
The numbers below the dilutions are the number of tip cells represented in the
PCR. The signal in de-tipped stalks is far dimmer than the 1/100 dilution of
kidney (with 0.81 tip cells). The actin bands demonstrate that the undiluted
samples of kidney and stalk cDNA represent equal amounts of total cells, as
intended.
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(C,D) Tips formed from the `wrong end' of the ureter, as in A,
induce the formation of nephrons in the surrounding mesenchyme. These are not
detectable in the ureteric bud-specific anti-calbindin stain (C) but are
visible in the anti-laminin stain (D, arrow).