TABLE 3.
An iTRAQ Experiment with a 20-Fold Range of Protein Quantities among Four Samples Compared
| 5/100 μg (114/117) | 25/100 μg (115/117) | 50/100 μg (116/117) | SwissProt entry name | prot_mass | emPAI | emPAI* protein mass | 180 μg total | 114 protein (μg) | 115 protein (μg) | 116 protein (μg) | 117 protein (μg) | R2 | Normalized to 100 μg total/label | sd/average CV % | |||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
| A | B | C | D | E | F | G | H | I | J | K | L | M | N | O | P | Q | R |
| 0.06 | 0.28 | 0.55 | GRCA_ECOBW | 15485 | 6.5 | 1.00E+05 | 1.5 | 0.04 | 0.22 | 0.44 | 0.79 | 1.00 | 0.81 | 0.83 | 0.85 | 0.83 | 1.9 |
| 0.06 | 0.28 | 0.58 | RS10_ECO24 | 12593 | 4.2 | 5.25E+04 | 0.8 | 0.03 | 0.11 | 0.24 | 0.41 | 1.00 | 0.47 | 0.43 | 0.46 | 0.42 | 5.0 |
| 0.06 | 0.31 | 0.59 | RL11_ECO24 | 17229 | 4.1 | 7.03E+04 | 1.0 | 0.03 | 0.16 | 0.32 | 0.54 | 1.00 | 0.58 | 0.61 | 0.61 | 0.56 | 4.7 |
| 0.05 | 0.25 | 0.50 | HNS_ECOLI | 17460 | 4.0 | 6.95E+04 | 1.0 | 0.03 | 0.15 | 0.29 | 0.58 | 1.00 | 0.51 | 0.54 | 0.56 | 0.60 | 6.2 |
| 0.06 | 0.28 | 0.50 | CSPC_ECOLI | 8551 | 3.8 | 3.22E+04 | 0.5 | 0.02 | 0.07 | 0.13 | 0.26 | 1.00 | 0.29 | 0.27 | 0.25 | 0.27 | 5.1 |
| 0.06 | 0.29 | 0.55 | EFTU1_ECO24 | 46886 | 3.6 | 1.71E+05 | 2.5 | 0.08 | 0.39 | 0.74 | 1.34 | 1.00 | 1.37 | 1.46 | 1.44 | 1.39 | 2.9 |
| 0.05 | 0.25 | 0.58 | RL7_ECO24 | 14305 | 3.3 | 4.68E+04 | 0.7 | 0.02 | 0.09 | 0.22 | 0.37 | 0.99 | 0.34 | 0.34 | 0.42 | 0.39 | 10.2 |
| 0.07 | 0.28 | 0.55 | TALB_ECOLI | 38683 | 3.0 | 1.17E+05 | 1.7 | 0.06 | 0.26 | 0.50 | 0.92 | 1.00 | 1.16 | 0.95 | 0.98 | 0.96 | 9.8 |
| 0.07 | 0.26 | 0.53 | RL1_ECO24 | 28173 | 3.0 | 8.48E+04 | 1.3 | 0.05 | 0.18 | 0.36 | 0.68 | 1.00 | 0.86 | 0.67 | 0.70 | 0.71 | 11.7 |
| 0.05 | 0.27 | 0.52 | TNAA_ECOBW | 57462 | 3.0 | 1.71E+05 | 2.6 | 0.07 | 0.37 | 0.72 | 1.39 | 1.00 | 1.37 | 1.37 | 1.41 | 1.44 | 2.5 |
| 0.05 | 0.24 | 0.52 | CSRA_ECOBW | 7428 | 2.8 | 2.06E+04 | 0.3 | 0.01 | 0.04 | 0.09 | 0.17 | 1.00 | 0.16 | 0.15 | 0.17 | 0.18 | 6.6 |
| 0.05 | 0.32 | 0.58 | AHPC_ECOLI | 23024 | 2.4 | 5.57E+04 | 0.8 | 0.02 | 0.14 | 0.25 | 0.43 | 1.00 | 0.40 | 0.51 | 0.48 | 0.44 | 10.2 |
| 0.05 | 0.27 | 0.53 | DBHA_ECOLI | 11258 | 2.4 | 2.68E+04 | 0.4 | 0.01 | 0.06 | 0.11 | 0.22 | 1.00 | 0.19 | 0.22 | 0.22 | 0.23 | 8.5 |
| 0.06 | 0.27 | 0.52 | RS4_ECO24 | 26538 | 2.3 | 6.18E+04 | 0.9 | 0.03 | 0.13 | 0.26 | 0.50 | 1.00 | 0.50 | 0.50 | 0.50 | 0.52 | 2.2 |
| 0.07 | 0.28 | 0.51 | RL9_ECO24 | 17489 | 2.3 | 4.04E+04 | 0.6 | 0.02 | 0.09 | 0.17 | 0.33 | 1.00 | 0.39 | 0.34 | 0.32 | 0.34 | 8.1 |
| 0.06 | 0.28 | 0.54 | RS1_ECO57 | 67575 | 2.2 | 1.51E+05 | 2.3 | 0.07 | 0.33 | 0.65 | 1.21 | 1.00 | 1.21 | 1.24 | 1.27 | 1.26 | 1.8 |
| 0.05 | 0.30 | 0.56 | OMPA_ECOLI | 40174 | 2.2 | 8.80E+04 | 1.3 | 0.04 | 0.20 | 0.38 | 0.69 | 1.00 | 0.67 | 0.76 | 0.75 | 0.72 | 5.6 |
| 0.06 | 0.24 | 0.56 | ALKH_ECOLI | 24458 | 2.2 | 5.36E+04 | 0.8 | 0.02 | 0.10 | 0.24 | 0.43 | 1.00 | 0.45 | 0.39 | 0.47 | 0.45 | 7.6 |
| 0.06 | 0.27 | 0.54 | ATPB_ECOBW | 53377 | 2.2 | 1.15E+05 | 1.7 | 0.06 | 0.25 | 0.49 | 0.91 | 1.00 | 1.07 | 0.92 | 0.96 | 0.95 | 6.6 |
| 0.05 | 0.27 | 0.57 | RL3_ECO24 | 25112 | 2.1 | 5.27E+04 | 0.8 | 0.02 | 0.11 | 0.24 | 0.42 | 1.00 | 0.40 | 0.41 | 0.46 | 0.43 | 6.1 |
| 0.06 | 0.25 | 0.52 | RBSB_ECOLI | 34822 | 2.1 | 7.28E+04 | 1.1 | 0.03 | 0.15 | 0.31 | 0.60 | 1.00 | 0.60 | 0.55 | 0.60 | 0.62 | 4.6 |
| 0.06 | 0.27 | 0.56 | RS3_ECO24 | 29426 | 2.0 | 5.80E+04 | 0.9 | 0.03 | 0.12 | 0.25 | 0.46 | 1.00 | 0.50 | 0.46 | 0.50 | 0.48 | 3.7 |
| 0.06 | 0.31 | 0.55 | RL5_ECO24 | 22795 | 2.0 | 4.47E+04 | 0.7 | 0.02 | 0.11 | 0.19 | 0.35 | 1.00 | 0.37 | 0.40 | 0.37 | 0.36 | 4.3 |
| 0.06 | 0.27 | 0.53 | G3P1_ECO57 | 39716 | 2.0 | 7.74E+04 | 1.2 | 0.03 | 0.17 | 0.33 | 0.63 | 1.00 | 0.63 | 0.62 | 0.64 | 0.65 | 2.2 |
| 0.06 | 0.43 | 0.56 | RS6_ECO24 | 16186 | 1.9 | 3.12E+04 | 0.5 | 0.01 | 0.10 | 0.13 | 0.23 | 0.97 | 0.23 | 0.36 | 0.25 | 0.24 | 23.0 |
| 0.07 | 0.29 | 0.51 | RL10_ECO24 | 19631 | 1.9 | 3.77E+04 | 0.6 | 0.02 | 0.09 | 0.15 | 0.30 | 1.00 | 0.39 | 0.32 | 0.30 | 0.31 | 12.3 |
| 0.06 | 0.26 | 0.51 | ATPF_ECODH | 19616 | 1.9 | 3.77E+04 | 0.6 | 0.02 | 0.08 | 0.16 | 0.31 | 1.00 | 0.32 | 0.30 | 0.30 | 0.32 | 3.6 |
| 0.05 | 0.26 | 0.53 | EFTS_ECOBW | 34697 | 1.8 | 6.25E+04 | 0.9 | 0.03 | 0.13 | 0.27 | 0.50 | 1.00 | 0.50 | 0.49 | 0.52 | 0.52 | 3.2 |
| 0.05 | 0.26 | 0.50 | THIO_ECOLI | 13498 | 1.8 | 2.39E+04 | 0.4 | 0.01 | 0.05 | 0.10 | 0.20 | 1.00 | 0.19 | 0.19 | 0.19 | 0.20 | 3.4 |
| 0.05 | 0.27 | 0.53 | TIG_ECOBW | 53351 | 1.8 | 9.34E+04 | 1.4 | 0.04 | 0.20 | 0.40 | 0.75 | 1.00 | 0.73 | 0.74 | 0.78 | 0.78 | 3.4 |
| Sum | 1.21E+07 | 180 | |||||||||||||||
| Only bold red and ion score >20 | iTRAQ-emPAI calculated protein amounts* | 5.5 | 26.8 | 51.5 | 96.2 | ||||||||||||
| Original planned amounts** | 5 | 25 | 50 | 100 | |||||||||||||
Proteins (522) of the E. coli lysate were compared by iTRAQ labeling (Supplemental Data file 1), but only the top 30 are shown in this table. The original amounts of protein planned for iTRAQ 114, 115, 116, and 117 were 5, 25, 50, and 100 μg, respectively.
*The calculated protein amounts reflect the relative efficiencies of the iTRAQ labeling reaction under the different reagent composition designs.
**The original amounts of protein planned for each iTRAQ labeling were 5 g. Only peptides identified by Mascot with bold red and ion score >20 were used in this analysis. R2, R-squared value of the trendline; CV, coefficient of variation.