Data quality is a concern in proteomics experimental science. In this report, we assess the intrinsic features of the data set at different levels.
Table 1. The Design of Experiment
sample | bioRep | techRep | No. of fraction |
---|---|---|---|
55 | 1 | 1 | 2 |
55 | 1 | 2 | 2 |
75 | 1 | 1 | 2 |
75 | 1 | 2 | 2 |
X!Tandem was used for analyzing the data. Parameters used in X!Tandem search were shown in Table 2. Protein identifications are inferred from peptide identifications, each protein identified having at least one associated unique peptide sequence identified at q-value equal or less than 0.01 (equivalent to a 1% FDR). The Occam's razor approach (Nesvizhskii, et al., 2003) was applied to deal with degenerate peptides by finding a minimum subset of proteins that covered all of the identified peptides.
Table 2. The database search parameters
Item | Value |
---|---|
Search engine | X!Tandem |
Enzyme | trypsin |
Fixed modifications | Carbamidomethyl (C) |
Variable modifications | Oxidation (M) |
Database | target_decoy.fasta |
Missed cleavages | 0 |
Precursor mass error | 10 ppm |
Fragment mass error | 0.6 Daltons |
This part contains the basic statistics of MS/MS data.
Table 3. Protein identification result for each fraction
sample | bioRep | techRep | fraction | spectrum_total | spectrum | peptide | protein |
---|---|---|---|---|---|---|---|
55 | 1 | 1 | 5 | 1576 | 478(30.330%) | 425 | 270 |
55 | 1 | 2 | 5 | 1562 | 456(29.193%) | 415 | 256 |
55 | 1 | 1 | 6 | 1573 | 462(29.371%) | 406 | 251 |
55 | 1 | 2 | 6 | 1558 | 487(31.258%) | 448 | 294 |
75 | 1 | 1 | 5 | 1277 | 375(29.366%) | 325 | 203 |
75 | 1 | 2 | 5 | 1256 | 350(27.866%) | 308 | 192 |
75 | 1 | 1 | 6 | 1215 | 398(32.757%) | 360 | 232 |
75 | 1 | 2 | 6 | 1208 | 361(29.884%) | 327 | 218 |
Table 4. Protein identification result for each technical replicate
sample | bioRep | techRep | spectrum_total | spectrum | peptide | protein |
---|---|---|---|---|---|---|
55 | 1 | 1 | 3149 | 940(29.851%) | 807 | 453 |
55 | 1 | 2 | 3120 | 943(30.224%) | 839 | 473 |
75 | 1 | 1 | 2492 | 773(31.019%) | 659 | 371 |
75 | 1 | 2 | 2464 | 711(28.856%) | 617 | 354 |
Table 5. Protein identification result for each biological replicate
sample | bioRep | spectrum_total | spectrum | peptide | protein |
---|---|---|---|---|---|
55 | 1 | 6269 | 1883(30.037%) | 1458 | 700 |
75 | 1 | 4956 | 1484(29.944%) | 1105 | 539 |
Summary plot for each technical replicate experiment.
Summary charts for sample: 55, biological: 1, technical: 1.
Figure 1. Missed cleavages chart for sample: 55, biological replicate: 1, technical replicate: 1.

Figure 2. Precursor ion charge chart for sample: 55, biological replicate: 1, technical replicate: 1.

Figure 3. Peptide length chart for sample: 55, biological replicate: 1, technical replicate: 1.

Figure 4. Precursor mass delta (Da) chart for sample: 55, biological replicate: 1, technical replicate: 1.

Figure 5. Precursor mass delta (ppm) chart for sample: 55, biological replicate: 1, technical replicate: 1.

Figure 6. Fragment ion mass delta (Da) chart for sample: 55, biological replicate: 1, technical replicate: 1.

Figure 7. Unique spectrum per protein chart for sample: 55, biological replicate: 1, technical replicate: 1.

Figure 8. Unique peptide per protein chart for sample: 55, biological replicate: 1, technical replicate: 1.

Figure 9. Protein mass chart for sample: 55, biological replicate: 1, technical replicate: 1.

Summary charts for sample: 55, biological: 1, technical: 2.
Figure 10. Missed cleavages chart for sample: 55, biological replicate: 1, technical replicate: 2.

Figure 11. Precursor ion charge chart for sample: 55, biological replicate: 1, technical replicate: 2.

Figure 12. Peptide length chart for sample: 55, biological replicate: 1, technical replicate: 2.

Figure 13. Precursor mass delta (Da) chart for sample: 55, biological replicate: 1, technical replicate: 2.

Figure 14. Precursor mass delta (ppm) chart for sample: 55, biological replicate: 1, technical replicate: 2.

Figure 15. Fragment ion mass delta (Da) chart for sample: 55, biological replicate: 1, technical replicate: 2.

Figure 16. Unique spectrum per protein chart for sample: 55, biological replicate: 1, technical replicate: 2.

Figure 17. Unique peptide per protein chart for sample: 55, biological replicate: 1, technical replicate: 2.

Figure 18. Protein mass chart for sample: 55, biological replicate: 1, technical replicate: 2.

Summary charts for sample: 75, biological: 1, technical: 1.
Figure 19. Missed cleavages chart for sample: 75, biological replicate: 1, technical replicate: 1.

Figure 20. Precursor ion charge chart for sample: 75, biological replicate: 1, technical replicate: 1.

Figure 21. Peptide length chart for sample: 75, biological replicate: 1, technical replicate: 1.

Figure 22. Precursor mass delta (Da) chart for sample: 75, biological replicate: 1, technical replicate: 1.

Figure 23. Precursor mass delta (ppm) chart for sample: 75, biological replicate: 1, technical replicate: 1.

Figure 24. Fragment ion mass delta (Da) chart for sample: 75, biological replicate: 1, technical replicate: 1.

Figure 25. Unique spectrum per protein chart for sample: 75, biological replicate: 1, technical replicate: 1.

Figure 26. Unique peptide per protein chart for sample: 75, biological replicate: 1, technical replicate: 1.

Figure 27. Protein mass chart for sample: 75, biological replicate: 1, technical replicate: 1.

Summary charts for sample: 75, biological: 1, technical: 2.
Figure 28. Missed cleavages chart for sample: 75, biological replicate: 1, technical replicate: 2.

Figure 29. Precursor ion charge chart for sample: 75, biological replicate: 1, technical replicate: 2.

Figure 30. Peptide length chart for sample: 75, biological replicate: 1, technical replicate: 2.

Figure 31. Precursor mass delta (Da) chart for sample: 75, biological replicate: 1, technical replicate: 2.

Figure 32. Precursor mass delta (ppm) chart for sample: 75, biological replicate: 1, technical replicate: 2.

Figure 33. Fragment ion mass delta (Da) chart for sample: 75, biological replicate: 1, technical replicate: 2.

Figure 34. Unique spectrum per protein chart for sample: 75, biological replicate: 1, technical replicate: 2.

Figure 35. Unique peptide per protein chart for sample: 75, biological replicate: 1, technical replicate: 2.

Figure 36. Protein mass chart for sample: 75, biological replicate: 1, technical replicate: 2.

The common Repository of Adventitious Proteins, cRAP (pronounced "cee-RAP"), is an attempt to create a list of proteins commonly found in proteomics experiments that are present either by accident or through unavoidable contamination of protein samples. The types of proteins included fall into three general classes:
-
common laboratory proteins;
-
proteins added by accident through dust or physical contact; and
-
proteins used as molecular weight or mass spectrometry quantitation standards.
We added the cRAP database.
Table 6. Identification of contaminant proteins
Accession | Peptides | Spectrum | Sample | ID | Reason | Class | Description |
---|---|---|---|---|---|---|---|
K2C1_HUMAN | 2 | 3 | 75_1_2 | 23 | Human skin | Dust/contact proteins | (P04264) Keratin, type II cytoskeletal 1 (Cytokeratin-1) (CK-1) (Keratin-1) (K1) (67 kDa cytokeratin) (Hair alpha protein) |
TRYP_PIG | 1 | 1 | 75_1_1 | 13 | Reagent | Laboratory proteins | (P00761) Trypsin precursor (EC 3.4.21.4) |
UB2E1_HUMAN | 1 | 1 | 75_1_1 | 111 | UPS | Sigma-Aldrich Universal Protein Standard | (P51965) Ubiquitin-conjugating enzyme E2 E1 [UbcH6] |
K2C1_HUMAN | 2 | 2 | 55_1_2 | 23 | Human skin | Dust/contact proteins | (P04264) Keratin, type II cytoskeletal 1 (Cytokeratin-1) (CK-1) (Keratin-1) (K1) (67 kDa cytokeratin) (Hair alpha protein) |
TAU_HUMAN | 1 | 1 | 55_1_2 | 106 | UPS | Sigma-Aldrich Universal Protein Standard | (P10636) Microtubule-associated protein tau [Tau protein] 46,810 Recombinant E. coli 6-His |
TRYP_PIG | 2 | 2 | 55_1_1 | 13 | Reagent | Laboratory proteins | (P00761) Trypsin precursor (EC 3.4.21.4) |
AMYS_HUMAN | 1 | 1 | 55_1_1 | 20 | Human saliva | Dust/contact proteins | (P04745) Salivary alpha-amylase precursor (EC 3.2.1.1) (1,4-alpha-D-glucan glucanohydrolase) |
Reproducibility of total spectra for each fraction
Figure 37. The distribution of the total spectra number

Figure 38. Error bar plot of the total spectra number for each fraction

Reproducibility of identified spectra for each fraction
Figure 39. The distribution of the identified spectra number

Figure 40. Error bar plot of the identified spectra for each fraction

Reproducibility of identified peptides for each fraction
Figure 41. The distribution of identified peptides number

Figure 42. Error bar plot of the identified peptides for each fraction

Reproducibility of identified proteins for each fraction
Figure 43. The distribution of the identified proteins number

Figure 44. Error bar plot of the identified proteins for each fraction

Figure 45. Technical replicate of peptides' overlap between sample 55 biological replicate 1.

Figure 46. Technical replicate of proteins' overlap between sample 55 biological replicate 1.

Figure 47. Technical replicate of peptides' overlap between sample 75 biological replicate 1.

Figure 48. Technical replicate of proteins' overlap between sample 75 biological replicate 1.

Figure 49. Peptides' overlap between different samples.

Figure 50. Proteins' overlap between different samples.

Figure 51. The mass error of the fragment ions

Figure 52. The mass error of the precusor

Figure 53. The mass error of the precusor

Figure 54. Sample: 55 , biological replicate: 1 , technical replicate: 1, identified spectrum accumulative effect.

Figure 55. Sample: 55 , biological replicate: 1 , technical replicate: 1, identified peptides accumulative effect.

Figure 56. Sample: 55 , biological replicate: 1 , technical replicate: 1, identified proteins accumulative effect.

Figure 57. Sample: 55 , biological replicate: 1 , technical replicate: 2, identified spectrum accumulative effect.

Figure 58. Sample: 55 , biological replicate: 1 , technical replicate: 2, identified peptides accumulative effect.

Figure 59. Sample: 55 , biological replicate: 1 , technical replicate: 2, identified proteins accumulative effect.

Figure 60. Sample: 75 , biological replicate: 1 , technical replicate: 1, identified spectrum accumulative effect.

Figure 61. Sample: 75 , biological replicate: 1 , technical replicate: 1, identified peptides accumulative effect.

Figure 62. Sample: 75 , biological replicate: 1 , technical replicate: 1, identified proteins accumulative effect.

Figure 63. Sample: 75 , biological replicate: 1 , technical replicate: 2, identified spectrum accumulative effect.

Figure 64. Sample: 75 , biological replicate: 1 , technical replicate: 2, identified peptides accumulative effect.

Figure 65. Sample: 75 , biological replicate: 1 , technical replicate: 2, identified proteins accumulative effect.
