What happens during consensus building?
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Kristina Plate
Sep 3, 2019, 10:56:49 AM9/3/19
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Dear all,
I have a question about the consensus process. I got four spectral libraries with consensus spectra, combined them into one spectral library and applied the consensus building again on the new library. Usually I would expect that library entries which occur in only one library are untouched by the consensus building but I found some examples which are slightly changed, nonetheless.
Here two examples where some peaks changed from unknown fragments to known ones and vice versa:
Example 1:
Name: DENGNVLK/2 (old entry, original library)
LibID: 4515
MW: 889.4494
PrecursorMZ: 444.7247
Status: Normal
FullName: X.DENGNVLK.X/2 (CID-QTOF)
Comment: AvePrecursorMz=444.9799 BestRawSpectrum=file_01.29552.29552 BinaryFileOffset=23697365 ConsFracAssignedPeaks=0.333 DotConsensus=0.87,0.05;1/11 FracUnassigned=0.17,2/5;0.27,9/20;0.39,42/78 Inst=0 MassDiff=-0.0007 MassDiffCounts=1/0:11 MaxRepSN=16.7 Mods=0 NAA=8 NISTProtein=Id_01 NMC=0 NTT=1 Nreps=11/11 OrigMaxIntensity=10 Parent=444.725 Pep=Tryptic PrecursorIntensity=0 Prob=1.0000 ProbRange=1,1,1,1 Protein=1/Id_01 RepFracAssignedPeaks=0.263 RepNumPeaks=155.3/88.6 RetentionTime=1033.9,1061.1,1023.4 SN=200.0 Sample=1/sample_01,11,11 Se=1^C11:fv=-1.0692e+000/1.6865e+000,pb=0.9867/0.0281 Spec=CREATE TotalIonCurrent=1.3e+004 iRT=-21.5,-21.5,-21.5
NumPeaks: 78
305.1714 372.8 y6-35^2/0.000 9/7 0.0048|0.78
306.6166 303.6 b6-17^2/0.000,b6-18^2/0.492 8/7 0.0125|0.75
311.0964 296.5 ? 8/7 0.0066|0.38
317.0882 264.4 ? 7/7 0.0042|0.60
321.0824 206.1 ? 7/7 0.0048|0.58
322.1529 321.0 ? 7/7 0.0033|0.35
322.6899 924.5 y6^2/0.000,b3-36/-0.409 10/7 0.0018|0.58
324.2282 496.9 y3-35/0.000 7/7 0.0028|0.98
335.0982 1716.8 ? 11/7 0.0011|0.22
337.1527 217.5 ? 7/7 0.0022|0.52
339.1965 521.5 ? 9/7 0.0064|0.82
340.1598 1097.3 [b3-18/-0.949] 11/7 0.0022|0.53
341.2547 830.7 y3-18/0.000,b3-18/0.146 10/7 0.0021|1.18
345.0821 253.6 ? 7/7 0.0025|0.54
349.6770 294.1 b7-44^2/0.000,b7-45^2/0.516 7/7 0.0039|0.49
351.1283 418.3 ? 9/7 0.0054|0.70
352.1246 389.6 ? 8/7 0.0045|0.35
357.6745 572.7 a7^2/0.000 10/7 0.0031|0.28
359.1206 202.8 ? 7/7 0.0075|0.52
359.2653 786.5 y3/0.000,b3/0.146 10/7 0.0022|0.24
362.1097 241.5 ? 7/7 0.0026|0.42
363.1587 466.0 b7-17^2/0.000,b7-18^2/0.492 10/7 0.0158|0.57
367.1761 357.9 ? 9/7 0.0048|0.28
368.1553 902.2 ? 10/7 0.0022|0.35
380.6772 652.1 b7+18^2/0.000,b4-35/-0.427,b4-36/0.557 11/7 0.0038|0.39
381.1772 323.6 b7+18^2i/0.000 8/7 0.0150|0.48
382.1715 322.1 ? 7/7 0.0057|0.63
385.1814 1855.7 ? 11/7 0.0010|0.19
390.1026 454.8 ? 10/7 0.0033|0.22
391.1944 368.2 ? 8/7 0.0044|0.71
397.1561 364.0 ? 10/7 0.0190|0.47
399.1147 380.4 b4-17/0.000,b4-18/0.984 9/7 0.0095|0.34
407.1278 194.0 ? 8/7 0.0063|0.44
408.1134 591.3 ? 11/7 0.0034|0.27
415.0899 350.6 ? 7/7 0.0033|0.66
426.1283 301.0 ? 7/7 0.0103|0.28
427.2062 302.1 p-35^2/0.000,p-36^2/0.492,y4-46/-0.097 8/7 0.0024|0.51
432.1130 818.0 ? 11/7 0.0021|0.57
434.1669 630.8 ? 10/7 0.0018|0.21
436.2114 527.0 p-17^2/0.000,p-18^2/0.492 10/7 0.0112|0.18
444.1509 209.8 ? 7/7 0.0091|0.40
444.7247 372.3 p^2/0.000 9/7 0.0038|0.99
450.1291 332.0 ? 7/7 0.0095|0.28
451.1907 307.9 ? 7/7 0.0084|0.19
453.2455 470.7 ? 9/7 0.0019|0.30
456.2817 281.1 y4-17/0.000,y4-18/0.984 7/7 0.0066|0.34
464.2124 444.1 ? 9/7 0.0144|0.77
473.3082 786.0 y4/0.000 10/7 0.0035|0.88
478.1221 300.7 ? 8/7 0.0056|0.30
479.1876 634.6 ? 9/7 0.0029|0.34
481.2389 523.7 ? 10/7 0.0054|0.27
489.1725 308.4 ? 7/7 0.0028|0.36
495.1470 277.1 b5-35/0.000,y5-35/-0.146 7/7 0.0058|0.70
496.1470 671.3 b5-35i/0.000 10/7 0.0038|0.25
497.1470 359.0 b5-35i/0.000 8/7 0.0083|0.41
498.2665 363.6 ? 7/7 0.0034|0.28
506.1980 244.5 ? 7/7 0.0078|0.46
507.1826 368.6 ? 9/7 0.0073|0.47
512.3191 320.0 y5-18/0.000,b5-18/0.146 7/7 0.0039|0.32
513.3031 337.1 y5-17/0.000,b5-17/0.146 7/7 0.0050|0.68
524.2086 352.5 ? 9/7 0.0039|0.27
530.3297 3591.3 y5/0.000,b5/0.146 11/7 0.0020|0.17
531.3297 519.0 y5i/0.000 9/7 0.0066|1.19
540.3092 304.2 ? 9/7 0.0058|0.33
594.2154 301.1 b6-35/0.000,b6-36/0.984 7/7 0.0097|0.32
609.3355 591.4 y6-35/0.000 10/7 0.0124|0.34
611.2420 591.0 b6-18/0.000 10/7 0.0106|0.47
626.3620 213.7 y6-18/0.000 7/7 0.0055|0.35
627.3460 2553.2 y6-17/0.000 11/7 0.0038|0.33
629.2525 442.2 b6/0.000 8/7 0.0062|0.56
644.3726 10000.0 y6/0.000 11/7 0.0029|0.00
645.3726 452.3 y6i/0.000 7/7 0.0054|0.34
655.3404 466.5 ? 9/7 0.0051|0.25
738.3781 365.6 y7-35/0.000,y7-36/0.984 8/7 0.0062|0.27
755.4046 551.9 y7-18/0.000 10/7 0.0162|0.83
756.3886 431.0 y7-17/0.000 8/7 0.0095|0.48
783.3949 1519.3 ? 11/7 0.0048|0.15
784.3822 1186.5 ? 10/7 0.0066|0.22
Name: DENGNVLK/2 (new entry, combined library)
LibID: 4018
MW: 889.4494
PrecursorMZ: 444.7247
Status: Normal
FullName: X.DENGNVLK.X/2 (CID-QTOF)
Comment: AvePrecursorMz=444.9799 BestRawSpectrum=file_01.29552.29552 BinaryFileOffset=17834540 ConsFracAssignedPeaks=0.333 DotConsensus=0.87,0.05;1/11 FracUnassigned=0.17,2/5;0.40,14/20;0.48,52/78 Inst=0 MassDiff=-0.0007 MassDiffCounts=1/0:11 MaxRepSN=16.7 Mods=0 NAA=8 NISTProtein=Id_01 NMC=0 NTT=1 Nreps=11/11 OrigMaxIntensity=10 Parent=444.725 Pep=Tryptic PepContext=1/DTNK_DNLS PrecursorIntensity=0 Prob=1.0000 ProbRange=1,1,1,1 Protein=1/Id_01 RepFracAssignedPeaks=0.263 RepNumPeaks=155.3/88.6 RetentionTime=1033.9,1061.1,1023.4 SN=200.0 Sample=1/sample_01,11,11 Se=1^C11:fv=-1.0692e+000/1.6865e+000,pb=0.9867/0.0281 Spec=CREATE TotalIonCurrent=1.3e+004 iRT=-21.5,-21.5,-21.5
NumPeaks: 78
305.1228 372.8 ? 9/7 0.0048|0.78
306.1029 303.6 ? 8/7 0.0125|0.75
311.0964 296.5 ? 8/7 0.0066|0.38
317.0882 264.4 ? 7/7 0.0042|0.60
321.0824 206.1 ? 7/7 0.0048|0.58
322.1529 321.0 ? 7/7 0.0033|0.35
323.1336 924.5 ? 10/7 0.0018|0.58
324.9859 496.9 ? 7/7 0.0028|0.98
335.0982 1716.8 ? 11/7 0.0011|0.22
337.1527 217.5 ? 7/7 0.0022|0.52
339.1965 521.5 ? 9/7 0.0064|0.82
340.1598 1097.3 ? 11/7 0.0022|0.53
341.0172 830.7 ? 10/7 0.0021|1.18
345.0821 253.6 ? 7/7 0.0025|0.54
350.1439 294.1 ? 7/7 0.0039|0.49
351.1283 418.3 ? 9/7 0.0054|0.70
352.1246 389.6 ? 8/7 0.0045|0.35
357.1872 572.7 ? 10/7 0.0031|0.28
359.1197 202.8 b3/0.000 7/7 0.0075|0.52
359.2653 786.5 y3/0.000 10/7 0.0022|0.24
362.1097 241.5 ? 7/7 0.0026|0.42
363.0953 466.0 ? 10/7 0.0158|0.57
367.1761 357.9 ? 9/7 0.0048|0.28
368.1553 902.2 ? 10/7 0.0022|0.35
380.1201 652.1 b4-36/0.000 11/7 0.0038|0.39
381.1041 323.6 b4-35/0.000 8/7 0.0150|0.48
382.1715 322.1 ? 7/7 0.0057|0.63
385.1830 1855.7 m3:6/0.000 11/7 0.0010|0.19
390.1026 454.8 ? 10/7 0.0033|0.22
391.1944 368.2 ? 8/7 0.0044|0.71
397.1561 364.0 ? 10/7 0.0190|0.47
398.1306 380.4 b4-18/0.000 9/7 0.0095|0.34
407.1278 194.0 ? 8/7 0.0063|0.44
408.1134 591.3 ? 11/7 0.0034|0.27
415.0899 350.6 ? 7/7 0.0033|0.66
426.1283 301.0 ? 7/7 0.0103|0.28
427.2062 302.1 p-35^2/0.000 8/7 0.0024|0.51
432.1130 818.0 ? 11/7 0.0021|0.57
434.1669 630.8 ? 10/7 0.0018|0.21
436.2114 527.0 p-17^2/0.000 10/7 0.0112|0.18
444.1509 209.8 ? 7/7 0.0091|0.40
444.7247 372.3 p^2/0.000 9/7 0.0038|0.99
450.1291 332.0 ? 7/7 0.0095|0.28
451.1907 307.9 ? 7/7 0.0084|0.19
453.2455 470.7 ? 9/7 0.0019|0.30
456.2817 281.1 y4-17/0.000 7/7 0.0066|0.34
464.2124 444.1 ? 9/7 0.0144|0.77
473.3082 786.0 y4/0.000 10/7 0.0035|0.88
478.1221 300.7 ? 8/7 0.0056|0.30
479.1876 634.6 ? 9/7 0.0029|0.34
481.2389 523.7 ? 10/7 0.0054|0.27
489.1725 308.4 ? 7/7 0.0028|0.36
496.1315 277.1 ? 7/7 0.0058|0.70
496.2130 671.3 ? 10/7 0.0038|0.25
497.2055 359.0 ? 8/7 0.0083|0.41
498.2671 363.6 m3:7/0.000 7/7 0.0034|0.28
506.1980 244.5 ? 7/7 0.0078|0.46
507.1826 368.6 ? 9/7 0.0073|0.47
513.1576 320.0 b5-17/0.000 7/7 0.0039|0.32
513.3031 337.1 y5-17/0.000 7/7 0.0050|0.68
524.2086 352.5 ? 9/7 0.0039|0.27
530.3297 3591.3 y5/0.000 11/7 0.0020|0.17
531.1792 519.0 ? 9/7 0.0066|1.19
540.3092 304.2 ? 9/7 0.0058|0.33
594.2154 301.1 b6-35/0.000 7/7 0.0097|0.32
609.3355 591.4 y6-35/0.000 10/7 0.0124|0.34
610.3355 591.0 y6-35i/0.000 10/7 0.0106|0.47
626.3620 213.7 y6-18/0.000 7/7 0.0055|0.35
627.3460 2553.2 y6-17/0.000 11/7 0.0038|0.33
629.2525 442.2 b6/0.000 8/7 0.0062|0.56
644.3726 10000.0 y6/0.000 11/7 0.0029|0.00
645.3726 452.3 y6i/0.000 7/7 0.0054|0.34
655.3404 466.5 ? 9/7 0.0051|0.25
738.3781 365.6 y7-35/0.000 8/7 0.0062|0.27
755.4046 551.9 y7-18/0.000 10/7 0.0162|0.83
756.3886 431.0 y7-17/0.000 8/7 0.0095|0.48
783.3949 1519.3 ? 11/7 0.0048|0.15
784.3822 1186.5 ? 10/7 0.0066|0.22
-----------------------------------------------------------------------------------
Example 2:
Name: DIETIIQK/2 (old entry, original library)
LibID: 4160
MW: 960.5481
PrecursorMZ: 480.2740
Status: Normal
FullName: X.DIETIIQK.X/2 (CID-QTOF)
Comment: AvePrecursorMz=480.5611 BinaryFileOffset=23385298 FracUnassigned=0.09,1/5;0.23,11/20;0.06,7/31 MassDiff=-0.0014 Mods=0 NAA=8 NISTProtein=Id_02 NMC=0 NTT=1 Nreps=1/1 OrigMaxIntensity=11 Parent=480.274 Pep=Tryptic PrecursorIntensity=0 Prob=1.0000 Protein=1/Id_02 RawSpectrum=file_02_04.69683.69683 RetentionTime=2745.2,2745.2,2745.2 Sample=1/sample_02,1,1 Se=1^C1:pb=1.0000/0,fv=2.7040/0 Spec=CREATE TotalIonCurrent=8.9e+003 iRT=33.4,33.4,33.4
NumPeaks: 31
308.1581 226.5 ?
309.0826 359.4 ?
325.6734 1596.8 b6-35^2/0.000,b6-36^2/0.492
329.1945 141.2 a6^2/0.000
343.1920 668.4 b6^2/0.000,y3-45/-0.042,y6-45^2/-0.516,y6-46^2/-0.024
353.2183 967.7 y3-35/0.000
354.2183 480.9 y3-35i/0.000
355.2183 481.6 y3-35i/0.000
358.1609 442.5 b3/0.000,y6-17^2/0.456,y6-18^2/0.948
358.1970 96.7 ?
365.2030 293.0 ?
373.2367 296.2 ?
388.2554 1744.6 y3/0.000,b7-36^2/-0.955
424.1714 315.8 b4-35/0.000,b4-36/0.984
442.1820 644.1 b4-17/0.000,b4-18/0.984
457.7633 546.4 p-45^2/0.000,p-46^2/0.492,y4-44/0.414
458.2791 109.5 p-44^2/0.000,b4/-0.929
471.7608 110.9 p-17^2/0.000,p-18^2/0.492
472.2608 111.1 p-17^2i/0.000
477.1321 111.6 ?
480.2740 336.0 p^2/0.000
484.3130 224.9 y4-17/0.000,y4-18/0.984
501.3395 523.5 y4/0.000
553.1674 240.4 ?
567.3501 487.3 y5-35/0.000
584.3766 247.1 y5-18/0.000
602.3872 2935.1 y5/0.000
696.3927 205.5 y6-35/0.000,y6-36/0.984
714.4032 2211.4 y6-17/0.000,y6-18/0.984
731.4298 10000.0 y6/0.000
732.4298 414.9 y6i/0.000
Name: DIETIIQK/2 (new entry, combined library)
LibID: 4673
MW: 960.5481
PrecursorMZ: 480.2740
Status: Normal
FullName: X.DIETIIQK.X/2 (CID-QTOF)
Comment: AvePrecursorMz=480.5611 BinaryFileOffset=20613807 FracUnassigned=0.09,1/5;0.25,13/20;0.17,13/31 MassDiff=-0.0014 Mods=0 NAA=8 NISTProtein=Id_02 NMC=0 NTT=1 Nreps=1/1 OrigMaxIntensity=11 Parent=480.274 Pep=Tryptic PepContext=1/NTPR_DIHE PrecursorIntensity=0 Prob=1.0000 Protein=1/Id_02 RawSpectrum=file_02_04.69683.69683 RetentionTime=2745.2,2745.2,2745.2 Sample=1/sample_02,1,1 Se=1^C1:pb=1.0000/0,fv=2.7040/0 Spec=CREATE TotalIonCurrent=8.9e+003 iRT=33.4,33.4,33.4
NumPeaks: 31
308.1581 226.5 ?
309.0826 359.4 ?
326.1680 1596.8 ?
328.2231 141.2 m4:6/0.000
344.1816 668.4 m3:5/0.000
353.2183 967.7 y3-35/0.000
354.2183 480.9 y3-35i/0.000
355.2183 481.6 y3-35i/0.000,m5:7/-0.016
358.1609 442.5 b3/0.000
358.1970 96.7 ?
365.2030 293.0 ?
373.2367 296.2 ?
388.2554 1744.6 y3/0.000
424.1714 315.8 b4-35/0.000
441.1980 644.1 b4-18/0.000
457.2713 546.4 p-46^2/0.000,m3:6/0.006
459.2086 109.5 b4/0.000
471.2688 110.9 p-18^2/0.000
472.2512 111.1 ?
477.1321 111.6 ?
480.2410 336.0 ?
484.3130 224.9 y4-17/0.000
501.3395 523.5 y4/0.000
553.1674 240.4 ?
568.3124 487.3 ?
584.2763 247.1 ?
602.3872 2935.1 y5/0.000
695.4087 205.5 y6-36/0.000
713.4192 2211.4 y6-18/0.000
731.4298 10000.0 y6/0.000
732.4675 414.9 ?
What happened during the consensus building? Does anyone have a clue?
I'm looking forward to your help und thank you in advance!
Kind regards,
Kristina
Kristina Plate
Sep 16, 2019, 8:56:32 AM9/16/19
to spctools-discuss
Hello again,
I already figured out that this problem is not caused by the consensus building but a reimport by spectrast in mrm format (-cM parameter) which I did for the subsequent assay library generation.
So, is the difference between the spectra a normal process during the MRM file import? What is happening there?
Kind regards and thank you,
Kristina
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