Fig. 10. Pdgfra expression in Sox9^loxP/loxP, Sox10^lacZ/lacZ, Sox10::Cre spinal cords. In situ hybridizations with a Pdgfra-specific probe (A,B,C,D,E,F,I,J,M,N) and immunohistochemistry with antibodies specific for Pdgfra (G,H,K,L,O,P) was performed at 13.5 dpc (A-D), 14.5 dpc (E-H), 15.5 dpc (I-L) and 18.5 dpc (M-P) on transverse sections from the forelimb region of wild-type (A,E,I,M,G,K,O), Sox9^loxP/loxP, Sox10::Cre (Sox9^/) (B), Sox10^lacZ/lacZ (Sox10^-/-) (C) and Sox9^loxP/loxP, Sox10^lacZ/lacZ, Sox10::Cre (Sox9^/ Sox10^-/-) (D,F,H,J,L,N,P) embryos. The arrowhead in D indicates the Pdgfra-positive cells. (Q) Chromatin immunoprecipitation was performed on formaldehyde-fixed spinal cords from four day old wild-type mice in the absence (-) and presence of antibodies (IgG, control IgG; -Sox9, anti-Sox9 antibodies). PCR was applied on the immunoprecipitate to detect evolutionary conserved sequence elements C1, C2 and C3 in the proximal 5' flanking region of the Pdgfra gene, as well as non-conserved regions N1 and N2 in the distal 5' flanking region. These fragments were also amplified from 1/20 of the material used for immunoprecipitation (Input). H₂O: water control. (R) OLP numbers were determined in cultures of dissociated spinal cords from wild-type (white bars) and Sox9^loxP/loxP, Sox10^lacZ/lacZ, Sox10::Cre (Sox9^/ Sox10^-/-) (black bars) 18.5-day-old embryos 3 hours after seeding or after 48 hours in the absence or presence of 20 ng/ml PDGF-AA. At least 30 visual fields were counted from two independent embryos for each genotype. Data are presented as mean OLP number per visual field±s.e.m.