Gene Expression Profiling using Fluidigm Biomark

Fluidigm 48.48 Biomark gene expression assays were performed, as described previously^{20 (link),40 (link)}. Briefly, the integrated fluidic chip (IFC) (Fluidigm) was primed using the IFC Controller MX, according to the manufacturer’s instructions. cDNA was diluted 1:1 with DEPC-treated water and 20× GE Sample Loading reagent was diluted 1:9 in TaqMan PCR Master Mix. Diluted cDNA and loading reagent were combined 1:1 and loaded onto the IFC chip. Primer assays were diluted 1:1 with 2X GE Assay Loading reagent and loaded into the IFC. The sample and assay were subsequently distributed on the chip via the IFC Controller MX loading program and the gene expression assay was performed and analysed using the GE 48.48 Standard.pcl on the Fluidim Biomark. Ct values were acquired from the Fluidigm Biomark, being normalised to ROX.