Figure 6

mRNA and protein validation of differential expression identified by microarray analysis. A) Real-time qRT-PCR analysis of six genes with increased expression in unfused sutures (RBP4, GPC3, C1QTNF3 short isoform, FMOD, PRELP, and PTN) and three genes with increased expression in fused sutures (WIF1, ANXA3, and CYFIP2) for unfused, fusing and fused suture tissue isolated from sagittal, coronal, lambdoid and metopic sutures. Significant differential expression (P < 0.05, *; P < 0.01, **) was analysed for fusing and fused sutures compared to unfused sutures. Mean expression + SEM is shown; n = 3 for all comparisons, except fused sagittal (n = 5), fused metopic (n = 4), and fused lambdoid (n = 2). Absolute expression values represent molecules per ng cDNA. B) Western blot analysis of individual protein samples in the order seen in (C), for collagen type 1 (COL1), GPC3, C1QTNF3 and RBP4. C) Densitometry analysis of western blots normalised to COL1 expression.