MetaboLights

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ftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14382/m_MTBLS14382_LC-MS_positive_reverse-phase_metabolite_profiling_v2_maf.tsvftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14382/i_Investigation.txtftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14382/s_MTBLS14382.txtftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14382/a_MTBLS14382_LC-MS_positive_reverse-phase_metabolite_profiling.txtprimaryOK200ftp://ftp.ebi.ac.uk/pub/databases/metabolights/studies/public/MTBLS14382Serial dilutions were prepared using commercial pure ribonucleosides (0.005-150 pg, Carbosynth, Toronto Research Chemicals) in order to establish the  linear range of quantification and the limit of detection of each  compoundMetaboLightsPublicLiquid Chromatography MS - positive - reverse phaseRibonucleosides were loaded directly onto the analytical column and were separated by reversed-phase chromatography using a 50-cm column with an inner diameter of 75 μm, packed with 2 μm C18 particles (Thermo Fisher Scientific, ES903). Chromatographic gradients started at 93% buffer A and 3% buffer B with a flow rate of 250 nl/min for 5 minutes and gradually increased to 30% buffer B and 70% buffer A in 20 min. After each analysis, the column was washed for 10min with 0% buffer A and 100% buffer B. Buffer A: 0.1% formic acid in water. Buffer B: 0.1% formic acid in 80% acetonitrile. The mass spectrometer was operated in positive ionization mode with nanospray voltage set at 2.4 kV and source temperature at 305°C. For Parallel Reaction Monitoring (PRM) method the quadrupole isolation window was set to 0.7 m/z, and MSMS scans were collected over a mass range of m/z 50-450, with detection in the Orbitrap at resolution of 120,000. MSMS fragmentation of defined masses and schedule retention time (Table S15) was performed using HCD at NCE 20 (except stated differently, Table S15), the auto gain control (AGC) was set to “Standard” and a maximum injection time of 246 ms was used. In each PRM cycle, one full MS scan at resolution of 120,000 was acquired over a mass range of m/z 220-700 with detection in the Orbitrap mass analyzer. Auto gain control (AGC) was set to 1e5 and the maximum injection time was set to 50 ms. Serial dilutions were prepared using commercial pure ribonucleosides (0.005-150 pg, Carbosynth, Toronto Research Chemicals) in order to establish the  linear range of quantification and the limit of detection of each  compoundFIRST-seq: a nanopore-based cDNA sequencing platform for RNA modification and structure profiling. 10.1101/2025.04.22.649934.Center for Genomic Regulation (CRG)Eva Maria NovoaCell LineWhole Organismmass spectrometry assaySamples were digested with 1 μl of the Nucleoside Digestion Mix (New England BioLabs, M0649S) and the mixture was incubated at 37 ºC for 1 h.  Samples (20ng) were analyzed using an Orbitrap Eclipse Tribrid mass spectrometer (Thermo Fisher Scientific, San Jose, CA, USA) coupled to an EASY-nLC 1200 (Thermo Fisher Scientific (Proxeon), Odense, Denmark). Ribonucleosides were loaded directly onto the analytical column and were separated by reversed-phase chromatography using a 50-cm column with an inner diameter of 75 μm, packed with 2 μm C18 particles (Thermo Fisher Scientific, ES903). Chromatographic gradients started at 93% buffer A and 3% buffer B with a flow rate of 250 nl/min for 5 minutes and gradually increased to 30% buffer B and 70% buffer A in 20 min. After each analysis, the column was washed for 10min with 0% buffer A and 100% buffer B. Buffer A: 0.1% formic acid in water. Buffer B: 0.1% formic acid in 80% acetonitrile. The mass spectrometer was operated in positive ionization mode with nanospray voltage set at 2.4 kV and source temperature at 305°C. For Parallel Reaction Monitoring (PRM) method the quadrupole isolation window was set to 0.7 m/z, and MSMS scans were collected over a mass range of m/z 50-450, with detection in the Orbitrap at resolution of 120,000. MSMS fragmentation of defined masses and schedule retention time (Table S15) was performed using HCD at NCE 20 (except stated differently, Table S15), the auto gain control (AGC) was set to “Standard” and a maximum injection time of 246 ms was used. In each PRM cycle, one full MS scan at resolution of 120,000 was acquired over a mass range of m/z 220-700 with detection in the Orbitrap mass analyzer. Auto gain control (AGC) was set to 1e5 and the maximum injection time was set to 50 ms. Serial dilutions were prepared using commercial pure ribonucleosides (0.005-150 pg, Carbosynth, Toronto Research Chemicals) in order to establish the  linear range of quantification and the limit of detection of each  compound [62] A mix of commercial ribonucleosides was injected before and after each batch of samples to assess instrument stability and to be used as external standard to calibrate the retention time of each ribonucleoside. Acquired data were analyzed with the Skyline-daily software (v24.1.1.284) and extracted precursor areas of the ribonucleosides were used for quantification. Escherichia coliHomo sapienshttps://www.ebi.ac.uk/metabolights/MTBLS14382Eva Maria Novoa. Centre for Genomic Regulation. eva.novoa@crg.eu.Eva Maria Novoa. Center for Genomic Regulation (CRG). evamaria.novoa@gmail.com.Acquired data were analyzed with the Skyline-daily software (v24.1.1.284) and extracted precursor areas of the ribonucleosides were used for quantification. ReplicateDoseevamaria.novoa@gmail.comSamples were digested with 1 μl of the Nucleoside Digestion Mix (New England BioLabs, M0649S) and the mixture was incubated at 37 ºC for 1 h.  Samples (20ng) were analyzed using an Orbitrap Eclipse Tribrid mass spectrometer (Thermo Fisher Scientific, San Jose, CA, USA) coupled to an EASY-nLC 1200 (Thermo Fisher Scientific (Proxeon), Odense, Denmark). Ribonucleosides were loaded directly onto the analytical column and were separated by reversed-phase chromatography using a 50-cm column with an inner diameter of 75 μm, packed with 2 μm C18 particles (Thermo Fisher Scientific, ES903). Chromatographic gradients started at 93% buffer A and 3% buffer B with a flow rate of 250 nl/min for 5 minutes and gradually increased to 30% buffer B and 70% buffer A in 20 min. After each analysis, the column was washed for 10min with 0% buffer A and 100% buffer B. Buffer A: 0.1% formic acid in water. Buffer B: 0.1% formic acid in 80% acetonitrile. The mass spectrometer was operated in positive ionization mode with nanospray voltage set at 2.4 kV and source temperature at 305°C. For Parallel Reaction Monitoring (PRM) method the quadrupole isolation window was set to 0.7 m/z, and MSMS scans were collected over a mass range of m/z 50-450, with detection in the Orbitrap at resolution of 120,000. MSMS fragmentation of defined masses and schedule retention time (Table S15) was performed using HCD at NCE 20 (except stated differently, Table S15), the auto gain control (AGC) was set to “Standard” and a maximum injection time of 246 ms was used. In each PRM cycle, one full MS scan at resolution of 120,000 was acquired over a mass range of m/z 220-700 with detection in the Orbitrap mass analyzer. Auto gain control (AGC) was set to 1e5 and the maximum injection time was set to 50 ms. Serial dilutions were prepared using commercial pure ribonucleosides (0.005-150 pg, Carbosynth, Toronto Research Chemicals) in order to establish the  linear range of quantification and the limit of detection of each  compound [62] A mix of commercial ribonucleosides was injected before and after each batch of samples to assess instrument stability and to be used as external standard to calibrate the retention time of each ribonucleoside. Acquired data were analyzed with the Skyline-daily software (v24.1.1.284) and extracted precursor areas of the ribonucleosides were used for quantification. MetabolomicsCell LineMetabolomicsRNA modificationThermo Scientific LTQ Orbitraptargeted analysisrnaEscherichia coliHomo sapiensexperimental blankThermo Scientific EASY-nLC 1200 System (UPLC)RNA structureWhole OrganismCell LineMetabolomicsRNA modificationThermo Scientific LTQ Orbitraprnatargeted analysisEscherichia coliHomo sapiensexperimental blankThermo Scientific EASY-nLC 1200 System (UPLC)RNA structureWhole OrganismSamples (20ng) were analyzed using an Orbitrap Eclipse Tribrid mass spectrometer (Thermo Fisher Scientific, San Jose, CA, USA) coupled to an EASY-nLC 1200 (Thermo Fisher Scientific (Proxeon), Odense, Denmark).1-methyladenosine7-Methylguanosine3-methylcytidinefalseFIRST-seq: a nanopore-based cDNA sequencing platform for RNA modification and structure profilingThis repository contains the LC-MS/MS raw data for the RNA quantifications obtained from in vivo DMS probed RNAs from E.coli and MDA-MB231 cell lines.2026-04-262026-04-26MTBLS14382MTBLC20129MTBLC16020MTBLC20794CHEBI:20129CHEBI:16020CHEBI:20794