Read me file for supplementary information attached to the thesis "Thiol Signalling in Skeletal Muscle Ageing".

Samples were analysed using the following methods ultimately being exported as Excel files containing the data in its raw format as described at the end.

> An Ultimate 3000 RSLC (fast liquid chromatography) Nano system (Thermo Scientific) was coupled to a Q-Exactive mass spectrometer (Thermo Scientific) to perform the data-dependent label free analysis. 

> Raw spectra were converted to Mascot Generated Files (.mgf) using the Proteome Discoverer software (Thermo Scientific). These .mgf files were then searched against the Uniprot mouse sequence database (database last updated on 12th May 2012; 16,376 sequences were identified for Mus musculus) via an in-house Mascot database server (Matrix Science, London, UK). The search parameters were: peptide mass tolerances, 10 ppm; fragment mass tolerance, 0.01 Da, 1+, 2+ and 3+ ions; missed cleavages, 1; instrument type, ESI-TRAP. Variable modifications were included as: d0-NEM, d5-NEM, mono-, di-, and tri-oxidation of Cysteine residues and oxidation of methionine.

> Label-free relative quantification software PEAKS 7 (Bioinformatics Solutions Inc., Waterloo, Canada) was used to analyse RAW data files against the same mouse protein database for identifications with Mascot. Proteins were considered significantly changed between muscles in adult and old mice samples using the following criteria: -10 log P score of >20 (equivalent to a p value of 0.01), a fold change = 1.5 and using a quality value of 0.8. Data normalisation was performed using the total ion current.


Data is exported from Proteome Discoverer after filtering for:

> Peptides containing the amino acid: Cysteine
> Peptide confidence score of >20

Then redox peptides and proteins were exported and attached here in addition to the complete label free datasets for both quadriceps and soleus muscles of adult and old mice.

The supplementary information for the thesis is:

S1 Quadriceps - All Proteins Label Free
S2 Quadriceps - Significantly Changed Proteins
S3 Adult Quadriceps Redox Protein List
S4 Old Quadriceps Redox Protein List
S5 Soleus - All Proteins Label Free List
S6 Soleus - Significantly Changed Proteins
S7 Adult Soleus Redox Protein List
S8 Old Soleus Redox Protein List