Exercise DIS3: answers
- In hit 11, why does it say that LGGSAVISLEGKPL does not match to any component?
- Why are no quantitation results reported for hit 11?
- Try changing the quality thresholds to see whether other settings give more precise ratios
- Is normalisation appropriate for this data set?
- Select median normalisation in the format settings. Why does Distiller have to process the unassigned list?
The SILAC experiment only labels Arg, and this peptide doesn’t contain Arg
Because there are no valid peptide matches. The matches listed for this protein either have scores that are too low (method specifies matches must have scores above homology threshold) or the peptide doesn’t contain arginine
Before and after making changes, create a Proteins report, (Analysis menu; Quantitation reports; proteins), and open it in a web browser (Right click in the report window). This will make it easy to compare different settings.
The on-line help discusses this in the topic Quantitation Toolbox; In depth. For this particular data set, normalisation doesn’t make much difference because the ratios are scattered fairly symmetrically about 1 to begin with.
When Normalisation is selected, all valid peptide matches have to be processed because Distiller needs to calculate correction factors to make the median for all peptide ratios unity, not just the matches assigned to protein hits.
If you decide to use Distiller to quantitate some of your own data, make sure you read all the help topics under Quantitation Toolbox, especially Processing your own data.