All experiments were performed in biological triplicates. Statistical comparisons of differences in biofilm formation, iTRAQ analysis, enzyme activity, histidine content and relative gene transcription level were performed using Wilcoxon test (SPSS 11.0.0 statistical software). The data of Real-time PCR were analyzed using repeated measurements in -ΔCt model (Hurtado et al., 2011 (link)). For iTRAQ analysis, MS/MS spectra were searched using MASCOT engine (Matrix Science, London, United Kingdom; version 2.2) embedded into Proteome Discoverer 1.3 (Thermo Electron, San Jose, CA, United States) against UniProt database and the decoy database. For protein identification, the following options were used. Peptide mass tolerance = 20 ppm, MS/MS tolerance = 0.1 Da, Enzyme = Trypsin, Missed cleavage = 2, Fixed modification: Carbamidomethyl (C), iTRAQ8plex (K), iTRAQ8plex (N-term), Variable modification: Oxidation (M), FDR ≤ 0.01. The protein had both a fold-change of ratio >1.2 or <0.8 (p-value < 0.05). A p < 0.05 was considered significant. The values were calculated as the mean of individual experiments in triplicate and compared with those of the control groups.
Mascot engine
Manufactured by Matrix Science
Sourced in United Kingdom, United States
The MASCOT engine is a core component of Matrix Science's laboratory equipment. Its primary function is to provide a reliable and consistent power source for various scientific instruments and experiments within the laboratory setting. The MASCOT engine is designed to deliver stable and controlled energy output, ensuring the accurate and reproducible operation of connected equipment.
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Lab products found in correlation
Proteome discoverer 1, by Thermo Fisher Scientific (46 mentions)
Q exactive mass spectrometer, by Thermo Fisher Scientific (35 mentions)
Proteome discoverer 1, by Matrix Science (29 mentions)
Easy nlc, by Thermo Fisher Scientific (19 mentions)
Proteome discoverer 2, by Matrix Science (12 mentions)
Tmt reagent, by Thermo Fisher Scientific (8 mentions)
Easy column, by Thermo Fisher Scientific (7 mentions)
Acclaim pepmap100, by Thermo Fisher Scientific (5 mentions)
Proteome discoverer 2, by Thermo Fisher Scientific (4 mentions)
Trypsin, by Promega (3 mentions)
Proteome discoverer, by Thermo Fisher Scientific (3 mentions)
Membrane and cytosol protein extraction kit, by Beyotime (3 mentions)
High ph reversed phase peptide fractionation kit, by Thermo Fisher Scientific (2 mentions)
Pierce high ph reversed phase fractionation kit, by Thermo Fisher Scientific (2 mentions)
Bca protein assay kit, by Bio-Rad (2 mentions)
Discoverer 1, by Matrix Science (2 mentions)
Ultimate 3000 rslcnano system, by Thermo Fisher Scientific (2 mentions)
Orbitrap fusion tribrid mass spectrometer, by Thermo Fisher Scientific (1 mentions)
1290 infinity lc, by Agilent Technologies (1 mentions)
Hiseq platform, by Illumina (1 mentions)
167 protocols using mascot engine
1
Comparative Multi-Omics Analysis of Bacterial Biofilm
2
Acetylation Site Analysis by Mass Spectrometry
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Protein samples were electrophoresed on an SDS/PAGE and then transferred on to a PVDF membrane for 1 h at 100 V. The membrane was blocked in BSA blocking buffer for 2 h at 25°C. We used an acetyl-lysine antibody (Cat# ICP0381, ImmuneChem Pharmaceuticals Inc., Canada) diluted to 1:15000. After incubation at 4°C for 10–12 h, the blot was washed with TBS and Tween 20 (TBST) three-times at ambient temperature. Chemiluminescence was detected using the ECL enhanced system and a luminescent image analyzer (DNR Bio Imaging Systems, Israel). To continuously monitor the acetyl-lysine sites, the acetylated Acs were isolated using the SDS/PAGE and proteins were solubilized according to previously described filter-aided sample preparation (FASP) procedures [15 (link)]. Predigested proteins were extracted by solid-phase extraction, separated using Nano-HPLC, and analyzed on an Orbitrap Fusion™ Tribrid™ Mass Spectrometer (Thermo Scientific, U.S.A.). Tandem MS (MS/MS) spectra were searched using the MASCOT engine (version 2.2; Matrix Science, United Kingdom).
3
Quantitative Proteomics of Streptococcus suis
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MS/MS spectra were searched using MASCOT engine (Matrix Science, London, UK; version 2.2) against 83,725 S. suis protein-coding sequences deposited in the Uniprot database and embedded into Proteome Discoverer 1.4. The search was conducted with trypsin applied as a specific enzyme and parameters used for normal peptides as follows: peptide mass tolerance, 20 ppm; fragment mass tolerance, 0.1 Da; max missed cleavages, 2; fixed modifications, carbamidomethyl (C), TMT 10plex (N-term), TMT 10plex (K); variable modifications: oxidation (M), TMT 10plex (Y); database pattern, decoy; and false-discovery rate ≤0.01. Each of the identified proteins involved at least one unique peptide. Protein quantification was accomplished by correlating the relative intensities of reporter ions extracted from tandem mass spectra to that of the peptides selected for MS/MS fragmentation. Fold change (ΔgidAΔmnmE/SC19) > 1.2 and < 0.83, and a p-value of <0.05 were used to represent up- or down-regulation, respectively. The mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier PXD012716.
4
Protein Identification and Quantification
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MS/MS spectra were searched against the UniProtKB database (www.uniprot.org ) using the MASCOT engine (Matrix Science, London, UK; version 2.2) embedded into Proteome Discoverer 1.4. Relative parameters was set as follows: trypsin was chosen as the enzyme, and Carbamidomethyl (C), iTRAQ 4/8 plex (N-term) and iTRAQ 4/8 plex (K) as fixed modifications; Oxidation (M) and iTRAQ 4/8plex (Y) as variable modifications; peptide mass tolerance: ±20 mg/l and fragment mass tolerance: 0.1 Da. To reduce the probability of false peptide identification (false discovery rate (FDR)), only peptides with FDR of 1% at the protein level were counted as the identified protein and each identified protein had at least one unique peptide. For protein quantification, the protein ratios are calculated as the median of only unique peptides of the protein. The thresholds of unique peptide were determined by FDR <0.01, and protein was considered as a differentially expressed protein (DEP) if its fold change was at least 1.2 and its P value <0.05 (Student’s t-test).
5
Multiomics Analysis of Breast Cancer
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The HNCH-BC database was used to acquire RNA-seq, proteomic, and metabolomic data from 24 BC patients (24 normal tissue samples and 24 BC samples). SSR scores were determined at the mRNA and protein levels using the BC sample's RNA-seq and proteomic data. The levels of nine EAAs were obtained from metabolomic data of 24 BC samples (Supplementary Table 2 and 3 ). Correlation between the level of nine EAAs and SSR scores in BC tissues was evaluated by using Pearson's correlation analysis.
Multi-omics analysis was performed at Applied Protein Technology Co., Ltd (Shanghai, China). Sequencing was performed using the Next-Generation Sequencing (NGS) technology based on an Illumina Hiseq platform. The study of free amino acids and their derivatives was performed using a liquid chromatography-tandem mass spectrometry (LC-MS/MS) system. A UHPLC (1290 Infinity LC, Agilent Technologies) coupled to a QTRAP (AB SCIEX 5500) was used to separate amino acids with greater efficiency. A quantitative proteomic analysis was performed via tandem mass tag (TMT) technology. Tandem mass spectrometry (TMS) spectral data were obtained from the MASCOT engine (Matrix Science, London, UK; version 2.2) embedded into Proteome Discoverer 1.4 software.
Multi-omics analysis was performed at Applied Protein Technology Co., Ltd (Shanghai, China). Sequencing was performed using the Next-Generation Sequencing (NGS) technology based on an Illumina Hiseq platform. The study of free amino acids and their derivatives was performed using a liquid chromatography-tandem mass spectrometry (LC-MS/MS) system. A UHPLC (1290 Infinity LC, Agilent Technologies) coupled to a QTRAP (AB SCIEX 5500) was used to separate amino acids with greater efficiency. A quantitative proteomic analysis was performed via tandem mass tag (TMT) technology. Tandem mass spectrometry (TMS) spectral data were obtained from the MASCOT engine (Matrix Science, London, UK; version 2.2) embedded into Proteome Discoverer 1.4 software.
6
Protein Identification via MASCOT MS/MS Analysis
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MS/MS spectra were searched using MASCOT engine (Matrix Science, London, UK; version 2.2) embedded into Proteome Discoverer 1.4.
7
Differential Proteomics Analysis Pipeline
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MS/MS raw files were processed using MASCOT engine (Matrix Science, London, United Kingdom; version 2.6) embedded into Proteome Discoverer 2.2, and searched against the NCBI database (Accession: NZ_CP006961.1 and NZ_CP006962.1 ). The search parameters included trypsin as the enzyme used to generate peptides with a maximum of two missed cleavages permitted. A precursor mass tolerance of 10 ppm was specified and 0.05 Da tolerance for MS2 fragments. Except for TMT labels, carbamidomethyl (C) was set as a fixed modification. Variable modifications were Oxidation (M) and Acetyl (Protein N-term). A peptide and protein false discovery rate of 1% was enforced using a reverse database search strategy. Proteins with fold change (FC) >1.5, p-value < 0.05 (by Student’s t-test), and the false discovery rate (FDR) < 0.05 were considered differentially expressed.
8
Comprehensive LC-MS/MS Evaluation Protocol
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For 60 min, an Easy nLC linked to a Q Exactive mass spectrometer (Thermo Scientific, Waltham, MA, USA) was used to evaluate liquid chromatography–mass spectrometry (LC-MS/MS). The mass spectrometer was operated in positive ion mode. The MASCOT engine (Matrix Science, London, UK; version 2.2) incorporated in Proteome Discoverer 1.4 was used to search for tandem mass spectrometry (MS/MS) spectra.
9
Ovine and Cetartiodactyla Proteome Analysis
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Raw data files were submitted to MASCOT engine (Matrix Science, London, UK; version 2.2) through Proteome Discoverer 1.4 (Thermo Electron, San Jose, CA.). The MS/MS spectra were searched against UniProt ovis aries (27065 sequences, download at 09, 10, 2015) and UniProt cetartiodactyla (712254 sequences, download at 15, 10, 2015). For protein identification, the following options were used: Peptide mass tolerance=20 ppm, MS/MS tolerance=0.1 Da, Enzyme=Trypsin, Missed cleavage=2, Fixed modification: Carbamidomethyl (C), iTRAQ 8 plex (K), iTRAQ 8 plex (N-term), Variable modification: Oxidation (M), FDR ≤ 0.01.
10
Quantitative Proteomics of M. oryzae
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The proteins from the 8 h and 24 h collections were mixed in a 1:1 (w/w total protein) ratio followed by digestion through the FASP procedure, as described previously [60 (link)]. LC-MS/MS analysis was performed on a Q Exactive mass spectrometer equipped with an EASY-Spray ion source (Thermo Fisher Scientific, Waltham, MA, USA), coupled to an Easy-nLC (Thermo Fisher Scientific) for 1 h. MS data acquired using a data-dependent top10 method dynamically choosing the most abundant precursor ions from the survey scan (300–1800 m/z) for HCD fragmentation were analyzed using MASCOT engine (Matrix Science, London, UK; version 2.4) against the UniProtKB M. oryzae database. For protein identification, the following options were used: trypsin cleavage, double missed cleavage, peptide mass tolerance set to 20 ppm, MS/MS tolerance set to 0.1 Da, carbamido methylation set as fixed modification, FDR ≤ 0.01.
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