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MB Sample ID: SA172721

Local Sample ID:Tugl-KD_2_IC
Subject ID:SU001927
Subject Type:Cultured cells
Subject Species:Mus musculus
Taxonomy ID:10090

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Subject:

Subject ID:SU001927
Subject Type:Cultured cells
Subject Species:Mus musculus
Taxonomy ID:10090

Factors:

Local Sample IDMB Sample IDFactor Level IDLevel ValueFactor Name
Tugl-KD_2_ICSA172721FL020830Tug1-KDTreatment

Collection:

Collection ID:CO001920
Collection Summary:Briefly, podocytes were cultured on BD BioCoat Collagen I plates (BD Biosciences, San Jose, CA) at 33°C in RPMI 1640 complete media with 20 U/ml mouse recombinant IFN-g (Thermo Fisher, Carlsbad, CA). To induce differentiation, we cultured podocytes in DMEM (5.5mM glucose and 5% FBS) at 37°C without IFN-g for 10-12 days. To rescue the expression of PGC1-a in stable Tug1-knockdown CRISPR clone (Tug1-KD)(Long et al., 2016), CMV enhancer/promoter-driven mouse Pgc1-a cDNA (Addgene, Watertown, MA) was inserted into vector Zeo-pT-MCS-GFP-T2A-Puro(Long et al., 2020), a modified PiggyBac transposon system, selected with 1ug/ml puromycin and sorted by GFP, to generate Tug1-KD/Pgc1-OE. We isolated kidney podocytes by positive selection with biotin-labelled Kirrel3 and Podocalyxin antibodies (2.5 μg/antibody/mouse, R&D Systems, Minneapolis, MN) followed by Streptavidin M-280 Dynabeads as previously described (Badal et al., 2016).
Sample Type:Epithelial cells

Treatment:

Treatment ID:TR001939
Treatment Summary:we cultured podocytes in DMEM (5.5mM glucose and 5% FBS) at 37°C without IFN-g for 10-12 days. To rescue the expression of PGC1-a in stable Tug1-knockdown CRISPR clone (Tug1-KD)(Long et al., 2016), CMV enhancer/promoter-driven mouse Pgc1-a cDNA (Addgene, Watertown, MA) was inserted into vector Zeo-pT-MCS-GFP-T2A-Puro(Long et al., 2020), a modified PiggyBac transposon system, selected with 1ug/ml puromycin and sorted by GFP, to generate Tug1-KD/Pgc1-OE.

Sample Preparation:

Sampleprep ID:SP001933
Sampleprep Summary:Metabolites from cell samples (in triplicates) grown on 10cm dishes were extracted with ice-cold 80% methanol. After centrifugation, extracts in supernatants were dried by evaporation under nitrogen.
Processing Storage Conditions:On ice
Extract Storage:-80℃

Combined analysis:

Analysis ID AN002997 AN002998
Analysis type MS MS
Chromatography type Ion exchange HILIC
Chromatography system Thermo Dionex ICS-5000+ Thermo Vanquish
Column Dionex IonPac AS11 Waters XBridge Amide (100 x 4.6mm,3.5um)
MS Type ESI ESI
MS instrument type Orbitrap Orbitrap
MS instrument name Thermo Fusion Orbitrap Thermo Fusion Orbitrap
Ion Mode NEGATIVE POSITIVE
Units AUC/ngDNA AUC/ngDNA

Chromatography:

Chromatography ID:CH002222
Chromatography Summary:ion chromatography (IC)-MS
Instrument Name:Thermo Dionex ICS-5000+
Column Name:Dionex IonPac AS11
Chromatography Type:Ion exchange
  
Chromatography ID:CH002223
Chromatography Summary:liquid chromatography (LC)-MS
Instrument Name:Thermo Vanquish
Column Name:Waters XBridge Amide (100 x 4.6mm,3.5um)
Chromatography Type:HILIC

MS:

MS ID:MS002786
Analysis ID:AN002997
Instrument Name:Thermo Fusion Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Data were acquired using a Thermo Orbitrap Fusion Tribrid Mass Spectrometer under ESI negative ionization mode. Raw data files were imported to Thermo Trace Finder software for final analysis. The relative abundance of each metabolite was normalized by DNA concentrations.
Ion Mode:NEGATIVE
  
MS ID:MS002787
Analysis ID:AN002998
Instrument Name:Thermo Fusion Orbitrap
Instrument Type:Orbitrap
MS Type:ESI
MS Comments:Data were acquired using a Thermo Orbitrap Fusion Tribrid Mass Spectrometer under ESI positive ionization mode at a resolution of 240,000.Raw data files were imported to Thermo Trace Finder software for final analysis. The relative abundance of each metabolite was normalized by DNA concentrations.
Ion Mode:POSITIVE
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