Chicken Hes4 (Hairy1), Jag1, Jag2, Notch1 and Lfng were kind gifts of Nicolas Daudet (University College London, UK). Fragments of mouse Jag1, Jag2 and Notch1 and of human Jag1 and Notch1 were cloned by PCR (Table 1) from, respectively, embryonic day (E)12.5 mouse cDNA (kind gift of Perrine Barraud, Dept. Physiology, Development and Neuroscience, University of Cambridge, UK) and cDNA prepared from 7-week-old dissected human foetal facial region (see previous section). Primer-BLAST software from NCBI (Ye et al., 2012 (link)) was used to design appropriate PCR primers (Table 1) and check them for specificity. The oligonucleotide properties calculator program OligoCalc (Kibbe, 2007 (link)) (http://www.basic.northwestern.edu/biotools/oligocalc.html) was used to check the melting temperature and self-complementarity of the primers. cDNA fragments were amplified by PCR and the products cloned into pDrive (Qiagen) using the Qiagen PCR cloning kit. Sequencing was performed by the Biochemistry Department DNA Sequencing Facility, University of Cambridge (Cambridge, UK). Digoxigenin-labelled antisense riboprobes were generated using standard methods (Henrique et al., 1995 (link)) and in situ hybridisation performed on cryosections as previously described (O'Neill et al., 2007 (link)) except that the slides were not treated with proteinase K.