Identification of genes expressed in the mouse limb using a novel ZPA microarray approach

One well-studied signaling center in the developing vertebrate limb, the zone of polarizing activity (ZPA), produces the morphogen sonic hedgehog that is necessary for normal growth and pattern formation. To identify additional factors expressed in the ZPA of the mouse limb bud, the Shhgfpcre allele was used to purify ZPA cells using fluorescence-activated cell sorting. Microarray technology was then used to identify genes whose expression was elevated in the ZPA compared to the rest of the limb. In situ hybridization confirmed the expression of two known transcription factors, Hlxb9 and Tcfap2b, an uncharacterized EST, and a transmembrane protein of unknown function in domains overlapping the ZPA. The expression of two other genes was confirmed by rtPCR. The methods described in this report will be applicable for identifying genes enriched in Shh-expressing cells throughout development.