PAXDBapplication/xmlhttp://pax-db.org/downloads/latest/datasets/bioprojects-abundance-files-v4.0.zipprimary200OK0000ProteomicsChristian von Mering4513https://pax-db.org/dataset/4513/677636284mering@imls.uzh.chUniversity of ZurichPAXDBFor the rescaling, the datasets are first parsed or processed such that the data reflect proportional abundances of whole protein molecules (i.e. proportionality to counts of complete, individual protein molecules, not to molecular weights, protein volumes, or digested peptides). In the case of spectral counting data protein. The proportional abundances are rescaled linearly to add up to one million; this means the abundance of each protein of interest is finally expressed in (parts per million,) relative to all other proteins in a sample. For a given protein abundance dataset, we then compute the absolute log abundance ratios of all pairs of proteins annotated to be functionally linked. The median of these absolute log abundance ratios represents an indirect quality metric: the closer it is to zero, the better (i.e. the more there is consistency between abundance values and functional annotations such as protein complexes or pathways). We then compute a background expectation for this metric, by permuting the abundance values in a given dataset randomly, and recomputing the median log abundance ratios. The permutation is repeated several times, yielding a distribution of medians. The actually observed median is then expressed as a Z-score distance to the random distribution ofmedians—this distance is termed the interaction consistency score.<h4>Background</h4>Sample preparation is a critical process for proteomic studies. Many efficient and reproducible sample preparation methods have been developed for mass spectrometry-based proteomic analysis of human and animal tissues or cells, but no attempt has been made to evaluate these protocols for plants. We here present an LC-MS/MS-based proteomics study of barley leaf aimed at optimization of methods to achieve efficient and unbiased trypsin digestion of proteins prior to LC-MS/MS based sequencing and quantification of peptides. We evaluated two spin filter-aided sample preparation protocols using either sodium dodecyl-sulphate or sodium deoxycholate (SDC), and three in-solution digestion (ISD) protocols using SDC or trichloroacetic acid/acetone precipitation.<h4>Results</h4>The proteomics workflow identified and quantified up to 1800 barley proteins based on sequencing of up to 6900 peptides per sample. The two spin filter-based protocols provided a 12-38% higher efficiency than the ISD protocols, including more proteins of low abundance. Among the ISD protocols, a simple one-step reduction and <i>S</i>-alkylation method (OP-ISD) was the most efficient for barley leaf sample preparation; it identified and quantified 1500 proteins and displayed higher peptide-to-protein inference ratio and higher average amino acid sequence coverage of proteins. The two spin filter-aided sample preparation protocols are compatible with TMT labelling for quantitative proteomics studies. They exhibited complementary performance as about 30% of the proteins were identified by either one or the other protocol, but also demonstrated a positive bias for membrane proteins when using SDC as detergent.<h4>Conclusions</h4>We provide detailed protocols for efficient plant protein sample preparation for LC-MS/MS-based proteomics studies. Spin filter-based protocols are the most efficient for the preparation of leaf samples for MS-based proteomics. However, a simple protocol provides comparable results although with different peptide digestion profile.Evaluation of sample preparation methods for mass spectrometry-based proteomic analysis of barley leaves.Wang Wei-Qing WQ, Jensen Ole Nørregaard ON, Møller Ian Max IM, Hebelstrup Kim H KH, Rogowska-Wrzesinska Adelina AAdd, DmelCG43443, ADD, ADD-87, Hts-RC, data, AU023367, Data Set, protein complex, supply, Proteins, Ovhts, Gene, HtsRC, CG9325, protein, neutral molecular compounds, protein-containing complex, Dmel_CG9325, Xt, Peptide, 1B1, add, Polypeptides, anon-EST:Posey9, protein polypeptide chains, native protein, peptido, htsRC, GLI3-190, natural protein, polypeptide chain, Add-hts, Protein, CG43443, Gene Products, l(2)k14523, Dmel_CG34197, l(2)00634, median, Ovhts-RC, background, supply and distribution, protein aggregate, all_pairs, molecule, Bph, molecula, Random selection by shearing, oligonucleotide random primer, proportion, HTS-R1, HTS, Hts, molecules, peptides, l(2)k06121, adducin, GLI3FL, distribution, AI854843, proportionality, add-like, HTS-RC, rate, proteins, Molekuel, Pdn, sample population, introduction, Protein Gene Products, Gene Proteins, Adducin, RANDOM, 10^[-6], ppm, sample, supply., quotient, Peptid, peptidos, Polypeptide, Attention Deficit Hyperactivity Disorder, l(2)01103, EST D, CG34197, HtsF, ratioMass Spectrum Analysis, Mass Spectrum, Procedures, Analyses, domesticated barley., Spectrometry, Methodological, procedures, Spectrum Analyses, Procedure, Methodological Study, Spectrum Analysis, Study, Spectroscopy, Barley, two-rowed barley, Techniques, Hordeum vulgare, MS, Methodological Studies, barley, Method, proteomic analysis, Studies, Mass, techniques, Analysis, Hordeum vulgare L., Technique, Horedum vulgare, Mass Spectrum Analyses, Mass Spectroscopy, methodologyexact), hoja (Spanish, 葉 (Japanese, 2018.liquid chromatography tandem mass spectroscopy, DIWS, Integral Membrane Proteins, Scientific Bias, Laundry Pods, Synthetic Detergents, Surface Proteins, SDC, Syndet, protein, Membrane-Associated Proteins, Integral, Deoxycholic Acid, trichloro-, l(2)k09905, Productivity, Alkylations, 12beta-Isomer, 2-Propanone, peptide, Polypeptides, Detergent, Techniques, protein polypeptide chains, Pod, Synthetic Detergent, Tier, sodio, exact), peptido, Method, Detergents, GRP1, Grp1, Epidemiologic Biase, Williams-Beuren syndrome chromosomal region 6 protein, 3, Ecological, Analysis, Sodium Ion, Work Flow, protein aggregate, animal, methyl ketone, Mass Spectrum Analysis, Animalia, Disodium Salt, average, DmelCG8428, Ecological Biases, Spin, me75, LCMSMS, Man (Taxonomy), [SO4](2-), peptides, Analyses, Rubidium, SULFATE ION, SUPPRESSOR OF DRM1 DRM2 CMT3, PTPSTEP, Surface, Tissue, plants, proteins, procedures, Structures, D17Mit170, Trichloroacetic, T1, SPIN, Methodological Studies, Propanon, Statistical Biases, BTKAP1, Diws1t, l(2)k08110, sample, 3beta-Isomer, Ecological Bias, Sodium Trichloroacetate, Surface Protein, Membrane Associated Proteins, Systematic, Hordeum vulgare L., SDC1, SIMPLE, Epidemiologic Biases, Membrane Protein, GTFII-I, Magnesium (2:1) Salt, T17A5.18, GPH, LC-MS2, 6030441I21Rik, Plant Protein, Primary Protein Structure, sulfate, Detergent Pod, Sulfate dianion, anatomical protrusion, Scientific, animalia, dimethylketone, Epidemiologic, Modern, land plants, LC-MS/MS, Ecological Fallacies, Procedure, BAP135, Tl3, Tl2, Spectrum Analysis, Aceton, Spectroscopy, Outcome Measurement Errors, organism, Sodium-23, TPSG1, Workflows, S-adenosyl-L-methionine:thiol S-methyltransferase activity, Dihydroxycholanoic Acid, Experimental, Pyroacetic ether, tetraoxosulfate(2-), PIG7, 3.1.3.48, SRF-Phox1-interacting protein, sodium, Laundry Detergent Pod, Cleansing Agents, BAP-135, Sulfate anion(2-), Truncation Bias, Detergent Pods, natrium, Acetic acid, 5beta, l(2R)W5, Acid, Amino Acid, Cell Membrane Protein, Bias, whole organism, Pods, Step, CG11628, Membrane-Associated, Spectrometry, Plant, Striatum-enriched protein-tyrosine phosphatase, Cell Membrane Proteins, Methodological, Outcome Measurement, GtfII-I, study protocol, Methodological Study, beta Trypsin, beta-Ketopropane, 葉 (Japanese, human, Experimental Bias, 12beta-Isomer Deoxycholic Acid, Outcome Measurement Error, higher plants, Sodium Salt, l(2)W5, 5alpha-Isomer, Agents, WBS, spine, Acide, STEP, liquid chromatography-tandem mass spectroscopy, Koerper, PRSS31, CG8428, Polypeptide, TMT, Work Flows, Cholan-24-oic acid, Amino Acid Sequences, Integral Membrane, 5alpha Isomer, Acide trichloracetique, human being, multi-cellular organism, GRP1/cytohesin 1, Procedures, Peptidomics, Laundry Detergent Pods, Ecological Fallacy, Truncation, Lagodeoxycholic Acid, Gene, Systematic Bias, Spectrum Analyses, protein-containing complex, TFII-I, SO4(2-), Synd-1, LC-MS-MS, protrusion, Human, Bruton tyrosine kinase-associated protein 135, Ion Level, CG11633, Agent, cytohesin/GRP1, two-rowed barley, method, propanone, Homo sapiens, polypeptide chain, 12-dihydroxy-, Membrane-Associated Protein, method used in an experiment, LC-MSMS, dimethylcetone, Studies, Mass, Gene Products, Sodium Deoxycholate, IB291, tetraoxosulfate(VI), Low, Animal, 12alpha)-, AA673430, trichloracetique, Level, Technique, Man, Mass Spectroscopy, Fallacies, Cell Membrane, study, Synthetic, 12beta Isomer, Aggregation, beta-Trypsin, Monosodium Salt, 3beta Isomer, l(2)SH2 0323, metazoa, labeling, Desoxycholic Acid, man, Structure, Laundry Detergent, Study, Barley, Hordeum vulgare, Neural-specific protein-tyrosine phosphatase, Protein Structure, Kybella, peptidos, Laundry Pod, Choleic Acid, AA553322, BTK-associated protein 135, Azeton, Primary Protein, Dimethyl ketone, thiol methyltransferase activity, Laundry, TP53I7, Biase, WBSCR6, Monoammonium Salt, Fallacy, T17A5_18, Dimethylketon, cou, body, protein complex, 5alpha-Isomer Deoxycholic Acid, Proteins, Cell Surface, whole body, Deoxycholate, stepk, Truncation Biases, Cell Surface Protein, sulphate, Primary, CD138, Cell, Peptide, bnch, l(2)k02511, Sulfate(2-), polypeptide, Sequences, LC/MS/MS, MS, Lr, native protein, natural protein, Sequence, Metazoa, Protein, SYND1, proteomic analysis, 3beta-Isomer Deoxycholic Acid, Trichloroacetate, Membrane Proteins, Errors, Sodium, 25/11, Rubidium Trichloroacetate, techniques, Protein Structures, (3alpha, l(2)SH0323, Mass Spectrum Analyses, Cell Surface Proteins, domesticated barley, Membrane Associated Protein, Sodium Ion Level, Cleansing, CYH1, Mass Spectrum, PRSS, tetraoxidosulfate(2-), Tripcellim, Biases, Na, Natrium, Monopotassium Salt, Membrane, hoja (Spanish, Sulfuric acid ion(2-), sample population, DmelCG11628, Protein Gene Products, l(2)10403, plan specification, Trypure, Gene Proteins, 11Na, Integral Membrane Protein, Cleansing Agent, Error, barley, Aggregation Bias, Modern Man, Statistical Bias, Bra, liquid chromatography tandem mass spectrometry, Peptid, Primary Protein Structures, Statistical, Sodium 23, Horedum vulgare, Sulfate, Peptide., methodology, sulphate ion0falseHvulgare - Leaf, SC (Wangetal,plantmethods.2018)abundance based on Spectral counting, Interaction consistency score: 8.6, Coverage: 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VVA0A287TBY9_HORVVA0A287FZQ3_HORVVA0A287K2L4_HORVVF2DSV5_HORVVM0W5F0_HORVVM0Y9J4_HORVVM0XG50_HORVVA0A287UC51_HORVVF2CQ05_HORVV4513.MLOC_29257.1M0UF93_HORVVA0A287Y2G1_HORVVA0A287NJB9_HORVVA0A287J7G9_HORVVS4Z8A5_HORVVA0A287TFZ0_HORVVM0YDJ4_HORVVM0ZBG0_HORVV4513.MLOC_42860.1Q70DK2_HORVVA0A287RWW7_HORVVA0A287SML4_HORVVA0A287NK41_HORVVA0A287EF53_HORVVS4Z1Z4_HORVVF2CX23_HORVVA0A287LTV2_HORVVA0A287QQN6_HORVVA0A287LPR3_HORVVA0A287S5U2_HORVVA0A287XF72_HORVVF2D6Z3_HORVV4513.MLOC_39970.1A0A287LJL4_HORVVA0A287HKS9_HORVVA0A287NWJ5_HORVVA0A287IZU9_HORVVA0A287T973_HORVVF2D587_HORVVA0A287QR99_HORVVA0A287S6Q1_HORVVA0A287IB96_HORVVA0A287L3J7_HORVVM0WUN8_HORVVA0A287UQX0_HORVV4513.MLOC_12540.14513.MLOC_64667.1A0A287LL35_HORVVA0A287WTF4_HORVVM0VIJ4_HORVVM0Z170_HORVVA0A287TFH8_HORVVA0A287HP43_HORVVA0A287HLK2_HORVVF2D426_HORVVA0A287KR61_HORVVA0A287WBY9_HORVVA0A287RZJ7_HORVVA0A287SNR6_HORVVA0A287LUR0_HORVVF2CYS4_HORVVM0VU12_HORVVA0A287LZU9_HORVVA0A287NZS0_HORVVA0A287QE28_HORVVA0A287V720_HORVVM0WC86_HORVVA0A287PHL1_HORVVA0A287INL5_HORVVQ5DVL6_HORVVA0A287LM58_HORVVA0A287TUE1_HORVVF2D6E2_HORVVA0A287N4X8_HORVVA5CFY2_HORVVA0A287RJY1_HORVVA0A287H8R8_HORVVA0A287LTY0_HORVVM0XNN4_HORVVA0A287WLY2_HORVVM0XFC6_HORVVF2E390_HORVVM0YQ06_HORVVA0A287K2S5_HORVVM0YQX1_HORVVM0V284_HORVVA0A287PRL3_HORVVA0A287TE56_HORVVA0A287RG91_HORVVA0A287JRE8_HORVVA0A287UD10_HORVV4513.MLOC_11877.1A0A287W312_HORVVA0A287F300_HORVVA0A287HY59_HORVVF2EEU3_HORVVA0A287ITA6_HORVVA0A287LB65_HORVVA0A287T4G8_HORVVM0YWT6_HORVVM0Y1W3_HORVVM0WA37_HORVVM0WJK0_HORVVM0WK00_HORVV4513.MLOC_67147.1A0A287PIH5_HORVVM0WVZ7_HORVVM0VGX7_HORVVA0A287U6Y1_HORVVF2DQX5_HORVVA0A287RJ48_HORVVF2DTY8_HORVVA0A287HK87_HORVVA0A287WQ77_HORVVA0A287GXX6_HORVVF2CQR8_HORVVIPYR_HORVVA0A287JCS1_HORVVA0A287HU17_HORVVA0A287TKB7_HORVVA0A287V3C0_HORVVA0A287HPE1_HORVVA0A287FEE6_HORVVM0URR9_HORVVA0A287MU28_HORVVA0A287K5X8_HORVVA0A287GXU4_HORVVF2EJQ4_HORVVM0XTF9_HORVVA0A287INP0_HORVVM0YB37_HORVVM0Y1I9_HORVVA0A287GF91_HORVVF2CTM5_HORVVM0X1J7_HORVVA0A287XZN2_HORVVA0A287FZG6_HORVV4513.MLOC_75016.2A0A287X4M9_HORVVM0XWK0_HORVVA0A287PXX3_HORVVA0A287VMH0_HORVVA0A287L1V8_HORVVM0YNW0_HORVVM0XJ27_HORVVA0A287QW66_HORVVF2CSC5_HORVVA0A287WSC5_HORVVF2CS91_HORVV4513.MLOC_69436.4M0UPI3_HORVVF2CZL7_HORVVA0A287NB42_HORVVA0A287JBI1_HORVVM0WMW0_HORVVA0A287S858_HORVVF2D5Z1_HORVVM0XMY2_HORVV4513.MLOC_62162.1A0A287GFM8_HORVVF2E797_HORVVM0YMJ5_HORVVA0A287RRS8_HORVVA0A287I770_HORVVA0A287T433_HORVVA0A287RRF8_HORVVA0A287IXV3_HORVVA0A287T3H0_HORVVA0A287PE35_HORVVF2E7J2_HORVV4513.MLOC_59320.1M0W0Z0_HORVVF2D4G2_HORVVF2CSG2_HORVVS4Z0F2_HORVVM0Y7B5_HORVVM0YB72_HORVVA0A287XGC8_HORVVM0XNU3_HORVVM0USD7_HORVVF2CPV6_HORVVA0A287SQZ2_HORVVA0A287M8W6_HORVVM0Y9X7_HORVVA0A287T9K2_HORVVA0A287SV12_HORVVA0A287IZ01_HORVVF2DDF2_HORVVF2DLQ0_HORVVM0YEB2_HORVVF2DX89_HORVVF2CRK1_HORVVS4YYM1_HORVVA0A287J7C6_HORVVA0A287H1C9_HORVVB5AAU3_HORVV4513.MLOC_73559.1A0A287HSM4_HORVVA0A287JYJ6_HORVVM0Z2K3_HORVVA0A287HB47_HORVVM0WNB1_HORVVM0XRA3_HORVVF2DAK4_HORVVM0Z3J1_HORVVA0A287LWZ3_HORVVA0A287R3G1_HORVVA0A287LMC2_HORVVA0A287J1U9_HORVVM0YVN1_HORVVA0A287SZW7_HORVVA0A287X2N9_HORVVA0A287FFR4_HORVVM0UNQ9_HORVVM0YF40_HORVVA0A287UW89_HORVVA0A287G1X9_HORVVA0A287GWH7_HORVVM0WYN7_HORVVA0A287PNA7_HORVVA0A287X055_HORVVA0A287T875_HORVVM0W5L2_HORVVA0A287JS74_HORVVA0A287XPQ7_HORVVA0A287FDB7_HORVVM0WEK4_HORVV4513.MLOC_33777.1M0YZA4_HORVVM0UGS6_HORVV4513.MLOC_64988.1A0A287PRE6_HORVVF2E3F5_HORVVF2DBD4_HORVVA0A287SND3_HORVVA0A287W654_HORVVA0A287VSA1_HORVVM0XDX5_HORVVF2CRK3_HORVVM0YB06_HORVV4513.MLOC_12188.3M0UYT8_HORVVM0VE87_HORVVM0XYC9_HORVV4513.MLOC_7610.2M0YES9_HORVVA0A287IIL7_HORVVM0W4W4_HORVVA0A287L4J4_HORVVA0A287M799_HORVVA0A287I5I6_HORVVA0A287EE27_HORVVA0A287SU56_HORVVM0WYK6_HORVVF2D961_HORVVM0WNE6_HORVVA0A287XDV5_HORVVA0A287SSU5_HORVVA0A287MXU9_HORVVA0A287RB60_HORVVA0A287TQ09_HORVVA0A287Y2J0_HORVVA0A287G041_HORVVM0V6F5_HORVV4513.MLOC_56244.1M0YJS4_HORVVF2E4I7_HORVV4513.MLOC_78332.1A0A287R6E0_HORVVF2CQH5_HORVVA0A287H5G8_HORVVA0A287LCT1_HORVVA0A287NYS4_HORVVM0XR76_HORVV4513.MLOC_58510.2A0A287NHD3_HORVV4513.MLOC_12843.2A0A287KFL8_HORVVM0XQB0_HORVVM0VJI6_HORVVA0A287UQJ1_HORVVQ6RYF4_HORVVM0Y1P2_HORVVA0A287QAB4_HORVVF2D5W1_HORVVA0A287PV82_HORVVF2DC29_HORVV4513.MLOC_42305.14513.MLOC_10422.1A0A287NXT3_HORVVM0WGC3_HORVVM0XZZ4_HORVVA0A287FZ34_HORVVF2EC14_HORVVF2CT57_HORVVS4Z1V8_HORVVA0A287T069_HORVVM0WFD2_HORVVA0A287RL62_HORVVA0A287LHF7_HORVVF2D8N3_HORVVA0A287IG64_HORVVM0YT83_HORVVA0A287EP81_HORVVA0A287PV87_HORVVM0V0J0_HORVVA0A287VLS1_HORVVA0A287M7J7_HORVVA0A287SVB8_HORVVM0YYV4_HORVVA0A287N905_HORVVA0A287QBH7_HORVVM0VJI5_HORVVA0A287U5I1_HORVVA0A287IQT8_HORVVA0A287KTZ1_HORVVM0YQD9_HORVVF2DYY1_HORVVF2E600_HORVVA0A287U9C4_HORVVM0YG07_HORVVA0A287M5Y3_HORVVA0A287ILG2_HORVVF2CWD2_HORVVA0A287VDK5_HORVVA0A287IM25_HORVVA0A287N846_HORVVF2E7Q2_HORVVA0A287TCG9_HORVVA0A287P854_HORVVF2DQG1_HORVV4513.MLOC_38928.1M0XEJ4_HORVVF2DYH7_HORVVA0A287PU20_HORVVA0A287UWZ2_HORVVA0A287GYG2_HORVVA0A287HDJ7_HORVVS4Z0F5_HORVV4513.MLOC_57988.5A0A287E8E2_HORVVM0YLD7_HORVVM0VJ84_HORVVF2DTH9_HORVVM0XK55_HORVVF2D9J3_HORVVA0A287SYQ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