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Xcc Pan Proteome

ABSTRACT: In this project, two isolates of Xanthomonas campestris pv campestris were compared in the following conditions, 1. growth in NYG rich medium, and 2. growth in XVM1 minimal medium. Total proteins were extracted and trypsin digested. The nanoscale Liquid Chromatography was performed to separate tryptic peptides in a nanoAcquity system (Waters, USA) using 2D dilution technology. The digested peptides were analyzed in a Synapt G2 HDMS mass spectrometer (Waters, Manchester, UK) operated in the resolution mode of analysis and a resolving power of at least 20,000 full-width half-maximum (FWHM). All analyses were performed using a positive nanoelectrospray ion mode (nanoESI +). ProteinLynx Global Server (PLGS) version 2.5 (Waters, Manchester, UK) was used to process the MS data obtained from LC-MSE. For protein identification, the software embedded ion accounting algorithm was used. For the analysis of protein identification and quantification level, the observed intensity measurements were auto normalized by PLGS.

INSTRUMENT(S): Synapt G2 HDMS

ORGANISM(S): Xanthomonas Campestris Pv. Campestris (ncbitaxon:340)

SUBMITTER: Angela Mehta

PROVIDER: MSV000083181 | MassIVE | Wed Dec 05 11:16:00 GMT 2018

SECONDARY ACCESSION(S): PXD011945

REPOSITORIES: MassIVE

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Project description:<p>The sample to be tested was fully ground into powder in the grinder, 50 mg of the sample was freeze-dried, 1000 μL of the extraction solution containing the inner target was added (methanol:acetonitrile:water, 2:2:1, v/v/v, internal standard concentration 20 mg/L) and vortex mixed for 30 s; then add the steel ball to the 45 Hz grinder for 10 min, ultrasonic 10 min; the sample to be tested is obtained after filtration. When detecting metabolites, metabolite determination was performed based on the LC-MS system, which mainly consists of Waters Acquity I-Class PLUS ultra-high performance liquid tandem and Waters Xevo G2-XS QTof high-resolution mass spectrometer. Meanwhile, the Waters Acquity UPLC HSS T3 column (1.8 um 2.1 x 100 mm) was used as the chromatographic column. Positive and negative ion modes were used to determine the metabolites. Mobile phase A: 0.1% formic acid aqueous solution; Mobile phase B: 0.1% acetonitrile formate. The mobile phase conditions of liquid chromatography were as follows: the flow rate was 400 μL/min, 0.0 min: 98% flow A, 2% flow B; 0.25 min: 98% flow A, 2% flow B, 10.0 min: 2% flow A, 98% flow B; 13.0 min: 2% flow A, 98% flow B; 13.1 min: 98% flow A, 2% flow B; 15.0 min: 98% flow A, 2% flow B. MSe mode controlled by acquisition software (MassLynx v4.2, Waters) was used for primary and secondary mass spectrum data acquisition. ESI ion source parameters are as follows: Capillary voltage, 2500 V (positive ion mode) or -2000 V (negative ion mode); Cone hole voltage, 30 V; Ion source temperature, 100 °C; Desolvent temperature, 500 °C; Air flow rate, 50 L/h; Desolvent gas flow rate, 800 L/h; Plastic-nucleus ratio (m/z) collection range 50-1200 m/z. In the qualitative and quantitative analysis of metabolites, the original data collected by MassLynx v4.2 were processed by the Progenesis QI software for peak extraction, peak alignment, and other data, and the metabolites were identified based on the Progenesis QI software online METLIN database. Then, based on the results of the total score, MS2 score, and mass error (ppm), the metabolites were qualitatively determined.</p>

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Xcc Pan Proteome

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