Figure 1.

Flow-chart of an optimised protocol for qRT-PCR in large-scale expression profiling experiments. The protocol implements multiple optimised and control steps including RNA isolation, digestion of genomic DNA (gDNA), evaluation of cDNA quality, primer design and data analysis. The absence of gDNA was confirmed by quantitative RT-PCR (qRT-PCR) with primer pairs targeting various non-coding regions. The quality of the cDNA was tested using different reference genes, as outlined in the text.