Pre-processing for Relative Quantification (PRQ) of TMT tagged LC-MS data is an R script written to pre-process mass spectrometry data. It performs the following steps:
Median normalisation to correct for labelling and MS-run variation (Step 1) is performed within each sample and gel fraction. This involves calculating the median of the ratios of all peptide intensities from one sample versus the corresponding intensities measured in the reference sample. All intensities relating to that sample and gel fraction are then divided by the median ratio. Ratio scores for each peptide are then calculated (Step 2) by calculating the ratios of the normalised data for each peptide by dividing it by the reference intensity. Ratios corresponding to the same source protein, peptide sequence and gel fraction are then summed. Protein level data is derived from these summed peptide scores (Step 3) by taking either the mean or median of all peptide scores from the same source protein and gel fraction. This protein level data is then collected across all sixplexs (Step 4).
A paper describing and applying PRQ has been submitted.