Summary of study ST000258

This data is available at the NIH Common Fund's National Metabolomics Data Repository (NMDR) website, the Metabolomics Workbench, https://www.metabolomicsworkbench.org, where it has been assigned Project ID PR000210. The data can be accessed directly via it's Project DOI: 10.21228/M87W2S This work is supported by NIH grant, U2C- DK119886.

See: https://www.metabolomicsworkbench.org/about/howtocite.php

This study contains a large results data set and is not available in the mwTab file. It is only available for download via FTP as data file(s) here.

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Study IDST000258
Study TitleMetabolic contribution of pSymA and pSymB megaplasmid/chromid for multipartite Sinorhizobium meliloti cultured in minimal M9 medium
Study Typemegaplasmid deletion
Study SummaryTo understand the contribution of pSymA and pSymB to the metabolism of S. meliloti, the intracellular metabolome was analyzed at five time points (exponential and stationary growth phases) across the growth curve of strains with or without pSymA and/or pSymB grown in a defined, minimal medium (M9).
Institute
McMaster University
DepartmentDepartment of Biology
Last NameFinan
First NameTurlough
AddressDepartment of Biology, McMaster University, Hamilton, Canada L8S4K1
Emailfinan@mcmaster.ca
Phone(+1)905-525-9140 ext 22932
Submit Date2015-09-17
Num Groups20
Total Subjects115
Raw Data AvailableYes
Raw Data File Type(s).mcf, .ami, .hdx, .txt, .xml
Analysis Type DetailLC-MS
Release Date2015-10-26
Release Version1
Turlough Finan Turlough Finan
https://dx.doi.org/10.21228/M87W2S
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

Select appropriate tab below to view additional metadata details:


Project:

Project ID:PR000210
Project DOI:doi: 10.21228/M87W2S
Project Title:Effects of synthetic large-scale genome reduction on metabolism and metabolic preferences in a nutritionally complex environment
Project Summary:The soil bacterium Sinorhizobium meliloti forms nodules on the roots of leguminous plants, where N2 is reduced to ammonia. Its genome includes a 3.65 Mb chromosome, a 1.35 Mb pSymA megaplasmid, and a 1.68 Mb pSymB chromid. pSymA and pSymB constitute ~45% of the genome and here a non-targeted approach was used to identify the metabolic consequences of the removal of these replicons. Polar and non-polar metabolites from wild-type, ∆pSymA, ∆pSymB, and ∆pSymAB cells and supernatants across a growth curve were analyzed by LC-HILIC-TOF-MS. 2008 metabolite features were identified in the extracellular metabolome of cells grown in LBmc containing yeast extract and casein hydrolysate. 1474 features were found from the intracellular metabolites of cells grown in minimal M9-sucrose medium. Analysis revealed both time and genotype influenced the metabolome, with the removal of pSymB having a much greater effect than the loss of pSymA. Strains lacking pSymB showed an increase in sugar, amino acid, and nucleotide metabolites in the intracellular metabolome, and the loss of pSymB clearly impaired the cell’s ability to catabolize exogenous amino acids. We conclude that despite the ability of wild-type, ∆pSymA, ∆pSymB, and ∆pSymAB strains to grow in both M9-sucrose and LBmc media, the removal of pSymA, and particularly pSymB, had clear and dramatic effects on the S. meliloti metabolome. The larger effect associated with the pSymB chromid is consistent with the large number of metabolic genes on this replicon and the greater genetic and metabolic integration of this replicon with the S. meliloti chromosome.
Institute:McMaster University
Department:Department of Biology
Last Name:Finan
First Name:Turlough
Address:Department of Biology, McMaster University, Hamilton, Canada L8S4K1
Email:finan@mcmaster.ca
Phone:(+1)905-525-9140 ext 22932

Subject:

Subject ID:SU000278
Subject Type:Bacterial cells
Subject Species:Sinorhizobium meliloti
Taxonomy ID:382
Genotype Strain:Rm2011 (SU47 str-3)|SmA818|RmP3009|RmP2917
Human Ethnicity:wild type|Rm2011 lack pSymA|Rm2011 lack pSymB|Rm2011 lack pSymA and pSymB | a tRNAarg and the smb20996-engA operon from pSymB were integrated to chromosome in strains RmP3009 and RmP2917
Species Group:Microorganism

Factors:

Subject type: Bacterial cells; Subject species: Sinorhizobium meliloti (Factor headings shown in green)

mb_sample_id local_sample_id strains
SA011773A_M2dΔpSymA
SA011774A_M2cΔpSymA
SA011775A_M2eΔpSymA
SA011776A_M2fΔpSymA
SA011777A_M3bΔpSymA
SA011778A_M3aΔpSymA
SA011779A_M2bΔpSymA
SA011780A_M2aΔpSymA
SA011781A_M1cΔpSymA
SA011782A_M1bΔpSymA
SA011783A_M1dΔpSymA
SA011784A_M1eΔpSymA
SA011785A_M1fΔpSymA
SA011786A_M3cΔpSymA
SA011787A_M3dΔpSymA
SA011788A_M5aΔpSymA
SA011789A_M4fΔpSymA
SA011790A_M5bΔpSymA
SA011791A_M5cΔpSymA
SA011792A_M5fΔpSymA
SA011793A_M5eΔpSymA
SA011794A_M4eΔpSymA
SA011795A_M4dΔpSymA
SA011796A_M3fΔpSymA
SA011797A_M3eΔpSymA
SA011798A_M4aΔpSymA
SA011799A_M4bΔpSymA
SA011800A_M4cΔpSymA
SA011801A_M1aΔpSymA
SA011802A_M5dΔpSymA
SA011803AB_M2eΔpSymAB
SA011804AB_M2dΔpSymAB
SA011805AB_M2fΔpSymAB
SA011806AB_M3aΔpSymAB
SA011807AB_M3bΔpSymAB
SA011808AB_M2cΔpSymAB
SA011809AB_M2aΔpSymAB
SA011810AB_M1bΔpSymAB
SA011811AB_M1aΔpSymAB
SA011812AB_M1cΔpSymAB
SA011813AB_M1dΔpSymAB
SA011814AB_M1eΔpSymAB
SA011815AB_M3cΔpSymAB
SA011816AB_M2bΔpSymAB
SA011817AB_M5cΔpSymAB
SA011818AB_M5aΔpSymAB
SA011819AB_M5dΔpSymAB
SA011820AB_M5eΔpSymAB
SA011821AB_M3dΔpSymAB
SA011822AB_M4eΔpSymAB
SA011823AB_M5bΔpSymAB
SA011824AB_M4bΔpSymAB
SA011825AB_M4aΔpSymAB
SA011826AB_M4cΔpSymAB
SA011827AB_M3eΔpSymAB
SA011828AB_M4dΔpSymAB
SA011829B_M4aΔpSymB
SA011830B_M4bΔpSymB
SA011831B_M4cΔpSymB
SA011832B_M3fΔpSymB
SA011833B_M3eΔpSymB
SA011834B_M3cΔpSymB
SA011835B_M3dΔpSymB
SA011836B_M4dΔpSymB
SA011837B_M5eΔpSymB
SA011838B_M5dΔpSymB
SA011839B_M3bΔpSymB
SA011840B_M5cΔpSymB
SA011841B_M5bΔpSymB
SA011842B_M4fΔpSymB
SA011843B_M5aΔpSymB
SA011844B_M4eΔpSymB
SA011845B_M3aΔpSymB
SA011846B_M1eΔpSymB
SA011847B_M1fΔpSymB
SA011848B_M1cΔpSymB
SA011849B_M1bΔpSymB
SA011850B_M1aΔpSymB
SA011851B_M2aΔpSymB
SA011852B_M1dΔpSymB
SA011853B_M2eΔpSymB
SA011854B_M2bΔpSymB
SA011855B_M2dΔpSymB
SA011856B_M2fΔpSymB
SA011857B_M2cΔpSymB
SA011858WT_M4bwild type
SA011859WT_M4cwild type
SA011860WT_M4awild type
SA011861WT_M3ewild type
SA011862WT_M3dwild type
SA011863WT_M4dwild type
SA011864WT_M3fwild type
SA011865WT_M5ewild type
SA011866WT_M5dwild type
SA011867WT_M3cwild type
SA011868WT_M5cwild type
SA011869WT_M5bwild type
SA011870WT_M4fwild type
SA011871WT_M5awild type
SA011872WT_M4ewild type
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Collection:

Collection ID:CO000271
Sample Type:cells
Volumeoramount Collected:6 x 108 cells
Storage Conditions:-80ºC freezer
Collection Vials:eppendorf tube
Tissue Cell Identification:6 x 108 cells

Treatment:

Treatment ID:TR000291
Animal Vet Treatments:-80ºC freezer
Animal Anesthesia:test tube
Animal Endp Tissue Proc Method:M9 miminal medium plus 10 mM sucrose
Animal Endp Clinical Signs:n/a
Human Fasting:30ºC incubator
Human Endp Clinical Signs:OD600 values of approximately 0.3, 0.5, 0.9, 1.6, and 3.5 (except for Rm2011 which reached ~4.8)

Sample Preparation:

Sampleprep ID:SP000285
Sampleprep Protocol Filename:Fei,_F.,_Bowdish,_D._M._E.,_&_McCarry,_B._E._(2014)._Analytical_and_Bioanalytical_Chemistry,_406,_3723–3733.
Extraction Method:cell pellet extracted with 2:2:1 MeOH/EtOH/H2O with beadbeating
Extract Storage:-80ºC freezer
Sample Resuspension:cell extracts suspended 60%v/v acetonitrile/water
Sample Spiking:L-methionine-d3, L-tryptophan-d5 as standards for recovery determination ; L-phenylalanine-d8, diphenylalanine and glycine-phenylalanine as internal standards for peak area normalization
Cell Type:Gram negative bacterium

Combined analysis:

Analysis ID AN000409 AN000410
Analysis type MS MS
Chromatography type HILIC HILIC
Chromatography system Agilent 1200 RR Series II Agilent 1200 RR Series II
Column Phenomenex Kinetex HILIC 100A (50 x 2.1 mm, 2.6um) Phenomenex Kinetex HILIC 100A (50 x 2.1 mm, 2.6um)
MS Type ESI ESI
MS instrument type QTOF QTOF
MS instrument name Bruker MicrOTOF II Bruker MicrOTOF II
Ion Mode POSITIVE NEGATIVE
Units normalized peak area normalized peak area

Chromatography:

Chromatography ID:CH000290
Methods Filename:Opt_silicaHILIC.m
Instrument Name:Agilent 1200 RR Series II
Column Name:Phenomenex Kinetex HILIC 100A (50 x 2.1 mm, 2.6um)
Column Temperature:40C
Flow Gradient:0-0.5 min, 95%A; 0.5-12.5 min, 35%A; 12.5-13.0 min, 35%A; 13.0-14.0 min, 95%A; 14.0-24.0 min, 95%A
Flow Rate:200 ?L/min
Internal Standard:L-phenylalanine-d8, diphenylalanine and glycine-phenylalanine
Sample Injection:2 ?L
Solvent A:HPLC grade acetonitrile
Solvent B:10 mM ammonium acetate in HPLC grade water adjusted to pH 3 with formic acid
Analytical Time:24 min
Chromatography Type:HILIC

MS:

MS ID:MS000351
Analysis ID:AN000409
Instrument Name:Bruker MicrOTOF II
Instrument Type:QTOF
MS Type:ESI
Ion Mode:POSITIVE
Capillary Voltage:-3800 V
Dry Gas Flow:8.0 L/min
Dry Gas Temp:250ºC
Scan Range Moverz:50-1000 m/z
  
MS ID:MS000352
Analysis ID:AN000410
Instrument Name:Bruker MicrOTOF II
Instrument Type:QTOF
MS Type:ESI
Ion Mode:NEGATIVE
Capillary Voltage:4500 V
Dry Gas Flow:8.0 L/min
Dry Gas Temp:250ºC
Scan Range Moverz:50-1000 m/z
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