Efficient Confirmation of Plant Viral Proteins and Identification of Specific Viral Strains by nanoLC-ESI-Q-TOF Using Single-Leaf-Tissue Samples.

{"sci_version": "1.2", "schema": "https://corpus.bntrg.co.uk/sci/1.0/sci.schema.json", "id": "pmid:33228257", "kind": "science-record", "revision": 6, "content_hash": "sha256:1a2a7d6b4ed33c962a63dc2abe1c1d6f20550fefe9e835b742c013b7bdf44a28", "created_at": "2026-11-01T15:41:17Z", "updated_at": "2026-11-01T21:24:09Z", "generator": {"name": "corpus-assay-engine", "run_id": "reading-sci-2026-11-01"}, "paper": {"state": "complete", "title": "Efficient Confirmation of Plant Viral Proteins and Identification of Specific Viral Strains by nanoLC-ESI-Q-TOF Using Single-Leaf-Tissue Samples.", "doi": "10.3390/pathogens9110966", "pmid": "33228257", "authors": ["Pavel Cejnar", "Štěpánka Kučková", "Jiří Šantrůček", "Miroslav Glasa", "Petr Komínek", "Daniel Mihálik", "Lucie Slavíková", "Leona Leišová-Svobodová", "Tatiana Smirnova", "Radovan Hynek", "Jiban Kumar Kundu", "Pavel Ryšánek"], "journal": "Pathogens (Basel, Switzerland)", "year": 2020, "abstract": "Plant viruses are important pathogens that cause significant crop losses. A plant protein extraction protocol that combines crushing the tissue by a pestle in liquid nitrogen with subsequent crushing by a roller-ball crusher in urea solution, followed by RuBisCO depletion, reduction, alkylation, protein digestion, and ZipTip purification allowed us to substantially simplify the sample preparation by removing any other precipitation steps and to detect viral proteins from samples, even with less than 0.2 g of leaf tissue, by a medium resolution nanoLC-ESI-Q-TOF. The presence of capsid proteins or polyproteins of fourteen important viruses from seven different families (Geminiviridae, Luteoviridae, Bromoviridae, Caulimoviridae, Virgaviridae, Potyviridae, and Secoviridae) isolated from ten different economically important plant hosts was confirmed through many identified pathogen-specific peptides from a protein database of host proteins and potential pathogen proteins assembled separately for each host and based on existing online plant virus pathogen databases. The presented extraction protocol, combined with a medium resolution LC-MS/MS, represents a cost-efficient virus protein confirmation method that proved to be effective at identifying virus strains (as demonstrated for PPV, WDV) and distinct disease species of BYDV, as well as putative new viral protein sequences from single-plant-leaf tissue samples. Data are available via ProteomeXchange with identifier PXD022456.", "claims": [{"id": "claim-method", "statement": "The peptides in raw spectra were identified and quantified by MaxQuant [11] 1.6.7.0 for Windows using reverse sequences for target-decoy database search strategy [48] and applying a 1% false discovery rate (FDR) for both the peptide spectrum match and protein group levels.", "kind": "methodological", "target": null, "evidence": [{"asset": "source:pmid:33228257", "locator": "sections[10] \"4.3. 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Plant Virus Pathogen Capsid Proteins Could Be Efficiently Confirmed in Samples of Plants with Strong Infection Using the Double Crushing Extraction Protocol Followed by nanoLC-ESI-Q-TOF", "row_count": 20, "column_count": 20}, "table": {"label": "Table 1", "caption": "Identified host and virus proteins from single-leaf-tissue samples of virus inoculated plants with strong symptoms.", "rows": [["", "Sample", "01a/barley*1", "01b/barley", "01c/barley", "02/wheat*1", "03a/Chinese cabbage", "03b/Chinese cabbage", "04a/tobacco", "04b/tobacco", "04c/tobacco", "04d/tobacco", "04e/tobacco", "05/apricot", "06a/plum", "06b/plum", "07/bean", "08/goosefoot", "09/sorghum", "10/maize"], ["PREPARATION", "Sample weight (g)", "0.14", "0.34", "0.46", "0.34", "0.52", "0.47", "0.47", "0.46", "0.45", "0.55", "0.63", "0.60", "0.60", "0.97", "0.48", "0.63", "0.32", "0.13"], ["Host taxonomic classification", "Hordeum vulgare", "Hordeum vulgare", "Hordeum vulgare", "Triticum aestivum", "Brassica rapa subsp. pekinensis", "Brassica rapa subsp. pekinensis", "Nicotiana benthamiana", "Nicotiana benthamiana", "Nicotiana benthamiana", "Nicotiana clevelandia x N. glutinosa", "Nicotiana tabacum", "Prunus armeniaca", "Prunus domestica", "Prunus Pdomestica", "Phaseolus vulgaris", "Chenopodium amaranticolor", "Sorghum bicolor", "Zea mays"], ["UniProt reference proteome used", "UP000011116", "UP000019116", "UP000011750", "UP000084051 *2", "*3", "UP000000226", "*4", "UP000000768", "UP000007305"], ["Reference genome proteins", "189,799", "130,673", "40,809", "73,605 + 53,411 + 74,802 *2", "155,017 *3", "30,501", "1063 *4", "41,380", "99,254"], ["Known host viruses in online databases", "66", "63", "29", "369", "17", "227", "378", "22", "69"], ["Tested viruses for sample (UniProt protein sequences available)", "47", "45", "24", "232", "13", "151", "232", "18", "52"], ["RESULTS", "Identified peptides in sample", "3233", "2871", "3963", "3888", "1844", "1094", "3176", "2772", "3412", "3426", "1270", "2344", "1648", "1047", "4364", "346", "1942", "1956"], ["Identified host proteins *", "886", "810", "1082", "1076", "537", "339", "756", "668", "865", "865", "287", "593", "435", "362", "1163", "71", "570", "549"], ["Identified virus", "Wheatdwarf virus(WDV)", "Barley yellow dwarf virus(BYDV)", "Bromemosaic virus(BMV)", "Wheatdwarf virus(WDV)", "Cauliflower mosaic virus (CaMV)", "Turnip vein clearing virus(TVCV)", "Plum pox virus(PPV)", "Turnipmosaic virus(TuMV)", "Tobaccoringspot virus(TRSV)", "Tomatoaspermy virus(TAV)", "Tobaccomosaic virus(TMV)", "Plum pox virus(PPV)", "Plum pox virus(PPV)", "Plum pox virus(PPV)", "Bean common mosaic virus(BCMV)", "Broad BeanWilt Virus 2(BBWV-2)", "Sorghummosaic virus(SrMV)", "Sugarcanemosaic virus(SCMV)"], ["Identified virus family", "Geminiviridae (DNA)", "Luteoviridae (RNA)", "Bromoviridae (RNA)", "Geminiviridae (DNA)", "Caulimoviridae (DNA)", "Virgaviridae (RNA)", "Potyviridae(RNA)", "Potyviridae(RNA)", "Secoviridae(RNA)", "Bromoviridae (RNA)", "Virgaviridae (RNA)", "Potyviridae(RNA)", "Potyviridae(RNA)", "Potyviridae(RNA)", "Potyviridae(RNA)", "Secoviridae(RNA)", "Potyviridae(RNA)", "Potyviridae(RNA)"], ["Identified capsid protein peptides **", "10", "2", "11", "10", "10", "6", "11", "14", "8", "12", "12", "6", "6", "5", "4", "17", "9", "4"], ["Capsid protein sequence coverage (AA) **", "147 of 260", "36 of 199", "147 of 189", "147 of 260", "110 of 489", "82 of 157", "119 of 330", "154 of 286", "104 of 513", "134 of 218", "92 of 159", "86 of 330", "65 of 330", "57 of 330", "71 of 287", "247 of 599", "95 of 320", "59 of 313"], ["(56.5%)", "(18.1%)", "(77.8%)", "(56.5%)", "(22.5%)", "(52.2%)", "(36.1%)", "(53.8%)", "(20.3%)", "(61.5%)", "(57.9%)", "(26.0%)", "(19.7%)", "(17.2%)", "(24.7%)", "(41.2%)", "(29.7%)", "(18.8%)"], ["Capsid protein sequence coverage on ≥ 7 amino acid sequences (AA) **", "147 of 189", "36 of 148", "147 of 152", "147 of 189", "110 of 333", "82 of 142", "119 of 278", "154 of 210", "104 of 437", "134 of 187", "92 of 148", "86 of 278", "65 of 278", "57 of 278", "71 of 232", "247 of 506", "95 of 211", "59 of 247"], ["(77.8%)", "(24.3%)", "(96.7%)", "(77.8%)", "(33.0%)", "(57.7%)", "(42.8%)", "(73.3%)", "(23.8%)", "(71.7%)", "(62.2%)", "(30.1%)", "(23.4%)", "(20.5%)", "(30.6%)", "(48.8%)", "(45.0%)", "(23.9%)"], ["Other viral proteins identified **", "−", "2 fragments of movement protein", "−", "−", "more than 10 fragments of other viral proteins - movement protein, reverse transcriptase, aphid transmission protein, etc.", "−", "other more than 10 fragments of genome polyprotein", "other more than 10 fragments of genome polyprotein", "−", "−", "more than 10 fragments of replication protein", "other more than 10 fragments of genome polyprotein", "other 6 fragments of genome polyprotein", "−", "other more than 10 fragments of genome polyprotein", "one other fragment of genome polyprotein from RNA 1 component", "other more than 10 fragments of genome polyprotein", "other more than 10 fragments of genome polyprotein"], ["Sequence covered (aa) **", "147", "87", "147", "147", "794", "82", "936", "695", "104", "134", "405", "291", "126", "57", "380", "275", "486", "343"], ["Putative sequence covered (aa) ***", "147", "87", "147", "147", "807", "82", "1073", "734", "104", "142", "405", "291", "126", "57", "407", "327", "522", "398"], ["Virus presence also confirmed by ****", "DAS-ELISAqPCR", "DAS-ELISART-PCRRFLP", "DAS-ELISA", "DAS-ELISAqPCR", "DAS-ELISA", "DAS-ELISA", "DAS-ELISAqPCR", "DAS-ELISAqPCR", "DAS-ELISA", "DAS-ELISA", "DAS-ELISA", "DAS-ELISAqPCR", "DAS-ELISAqPCR", "DAS-ELISAqPCR", "DAS-ELISA", "electron microscopyRT-PCR", "DAS-ELISA", "DAS-ELISA"]], "row_count": 20, "column_count": 20, "numeric_cells": 247, "header_rows": 0, "locator": "section[4]/table-wrap[1]#pathogens-09-00966-t001"}}], "gaps": ["asset:data:geo:PXD022456 not downloaded", "asset:data:geo:PXD022456 licence not read"]}, "analysis": {"state": "partial", "methods": [{"id": "method-1", "family": null, "specific": null, "label": "Target-decoy database search strategy with a 1% false discovery rate (FDR) applied at both peptide-spectrum-match and protein-group levels", "dsl_operations": [], "missing_operations": [], "expressible": null, "parameters": {}, "data_shapes": [], "evidence": {"score": 1.0, "band": "verbatim", "quote": "The peptides in raw spectra were identified and quantified by MaxQuant [11] 1.6.7.0 for Windows using reverse sequences for target-decoy database search strategy [48] and applying a 1% false discovery rate (FDR) for both the peptide spectrum match and protein group levels."}}], "targets": [], "gaps": ["method.reader_named_unmapped"]}, "capability": {"state": "complete", "requirements": [{"capability_id": "capability:computational_analysis", "label": "Computational analysis platform", "assay_families": ["meta_analysis", "machine_learning", "survival_analysis", "statistical_regression", "pca", "molecular_docking", "network_analysis", "differential_expression", "gwas"], "availability_state": "owned", "tier_unlocked": "R1", "candidate": "Automated Assays Laravel application and simulated adapters"}], "gaps": []}, "readiness": {"state": "blocked", "tier": "R4", "blocking_axis": "method", "blocked_on": "method:not_determinable", "axes": {"method": {"tier": "R4", "expressible": false, "required_operations": [], "missing_operations": [], "basis": "The reader quoted the method the paper names (Target-decoy database search strategy with a 1% false discovery rate (FDR) applied at both peptide-spectrum-match and protein-group levels), but the corpus's method extractor could not map it onto a DSL operation, so the method is published but not expressible."}, "capability": {"tier": "R1", "capability_id": "capability:computational_analysis", "availability_state": "owned", "basis": "The computational analysis is owned. 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