Experimental procedures

Retroviral infection

Fertilized chicken eggs (SPAFAS) were incubated at 39 °C, and Noggin-'RCAS(A) virus (Pizette & Niswander 1999) injected at Hamburger—Hamilton stage (Hamburger & Hamilton 1992) 14, 17 and 20 into the limb fields, and at stage 24 or 27 in the zeugopod or autopod, respectively. For molecular studies other than alcian blue staining, embryos were injected at stage 14.

RNA in situ hybridisation andprobes

RNA in situ hybridization with digoxygenin-labelled probes was performed as referenced in Zou et al (1997). Antisense probes were generated as described: BmpRIB and colli (Francis-West et al 1999), and sox9 (Kent et al 1996).

Histology

BrdU incorporation and detection, wholemount nile blue staining, alcian blue staining on whole embryos or on sections, were executed as in (Zou et al 1997), TUNEL as in (Chen et al 1994) and PNA staining as in (Stringa & Tuan 1996).

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