Custom selection of reference genes for transcriptomic analysis

We developed a R-based script to select internal control genes based solely on read counts and gene sizes. We used this method to pick custom reference genes for the differential expression analysis of three transcriptome sets from transgenic Arabidopsis plants expressing heterologous fungal effector proteins tagged with GFP (using GFP alone as the control). The custom reference genes showed lower covariance and fold change as well as a broader range of expression levels than commonly used reference genes. When analyzed with NormFinder, both typical and custom reference genes were considered suitable internal controls, but the custom selected genes were more stable. geNorm produced a similar result in which most custom selected genes ranked higher (i.e. were more stable) than commonly used reference genes.