Summary of Study ST003744

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 PR002330. The data can be accessed directly via it's Project DOI: 10.21228/M8FV76 This work is supported by NIH grant, U2C- DK119886.

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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 IDST003744
Study TitleA study exploring the epoxidation function of ODD enzymes (2-oxoglutarate/Fe(II)-dependent dioxygenases)
Study SummaryThis study identified a series of ODD (2-oxoglutarate/Fe(II)-dependent dioxygenase) genes with potential C4β-C20 epoxidation activity from the Himalayan yew (Taxus wallichiana) genome. To validate their enzymatic functions, these genes were cloned into the Ycplac22 vector and expressed in a Saccharomyces cerevisiae strain engineered to produce taxadiene, the required substrate for ODD enzymes. The fermentation process involved initial culturing of the yeast strain in test tubes, followed by inoculation into fresh medium and the addition of n-dodecane and glucose to initiate two-phase fermentation, facilitating product separation and accumulation. After 4 days of cultivation, the upper organic phase was collected and analyzed by GC-MS, confirming the production of the target compound, 4β,20-taxadiene. The obtained GC-MS data were further used to assess whether the ODD enzymes exhibit potential epoxidation activity.
Institute
Northwestern Polytechnical University
Last NameLi
First NameZhenzhu
AddressNo. 127, Youyi West Road, Xi'an City
Emaillizhenzhu@mail.nwpu.edu.cn
Phone18829012174
Submit Date2025-02-13
Raw Data AvailableYes
Raw Data File Type(s)mzML
Analysis Type DetailGC-MS
Release Date2025-02-24
Release Version1
Zhenzhu Li Zhenzhu Li
https://dx.doi.org/10.21228/M8FV76
ftp://www.metabolomicsworkbench.org/Studies/ application/zip

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Project:

Project ID:PR002330
Project DOI:doi: 10.21228/M8FV76
Project Title:A study exploring the epoxidation function of ODD enzymes (2-oxoglutarate/Fe(II)-dependent dioxygenases)
Project Type:MS Qualitative Analysis
Project Summary:This study identified a series of ODD (2-oxoglutarate/Fe(II)-dependent dioxygenase) genes with potential C4β-C20 epoxidation activity. To validate their enzymatic function, these genes were cloned into the Ycplac22 vector and introduced into Saccharomyces cerevisiae engineered to produce the necessary ODD substrate, taxadiene. The fermentation products were then analyzed using GC-MS to detect the formation of the target compound, 4β,20-taxadiene.
Institute:Northwestern Polytechnical University
Last Name:Li
First Name:Zhenzhu
Address:No. 127, Youyi West Road, Xi'an City
Email:lizhenzhu@mail.nwpu.edu.cn
Phone:18829012174

Subject:

Subject ID:SU003876
Subject Type:Yeast
Subject Species:Taxus wallichiana
Taxonomy ID:147273

Factors:

Subject type: Yeast; Subject species: Taxus wallichiana (Factor headings shown in green)

mb_sample_id local_sample_id Sample source Gene
SA408181CKSaccharomyces cerevisiae Control
SA408182GENE1Saccharomyces cerevisiae GENE1
SA408183GENE10Saccharomyces cerevisiae GENE10
SA408184GENE13Saccharomyces cerevisiae GENE13
SA408185GENE14Saccharomyces cerevisiae GENE14
SA408186GENE15Saccharomyces cerevisiae GENE15
SA408187GENE16Saccharomyces cerevisiae GENE16
SA408188GENE2Saccharomyces cerevisiae GENE2
SA408189GENE3Saccharomyces cerevisiae GENE3
SA408190GENE6Saccharomyces cerevisiae GENE6
SA408191GENE7Saccharomyces cerevisiae GENE7
SA408192GENE9Saccharomyces cerevisiae GENE9
Showing results 1 to 12 of 12

Collection:

Collection ID:CO003869
Collection Summary:GENE1-GENE16, a series of novel genes with potential epoxidation activity, were newly identified from the genome of the Himalayan yew (Taxus wallichiana), whose sequences are provided in the downloadable file "nucleotide_sequences_of_11_ODD_candidate_genes.csv". These putative ODD genes were subsequently assembled into the yeast expression vector Ycplac22. The constructed expression vector was then transformed into a pre-engineered Saccharomyces cerevisiae chassis strain, which was designed for cytoplasmic production of taxa-4(5),11(12)-diene (taxadiene), where the genes were expressed and their functions analyzed. The yeast strains were inoculated into 40 mL of fresh medium at a 1:50 ratio. A two-phase fermentation process was initiated and maintained at 30°C with agitation at 220 rpm for 4 days. After fermentation, the culture was centrifuged at 3600 rpm for 10 minutes, and the upper organic phase was collected for further analysis.
Sample Type:Yeast cells

Treatment:

Treatment ID:TR003885
Treatment Summary:After 10 hours of yeast cultivation, 5 mL of n-dodecane and 2 mL of 40% glucose were added to the culture.

Sample Preparation:

Sampleprep ID:SP003882
Sampleprep Summary:The collected upper organic phase was filtered and subsequently subjected to GC-MS analysis.

Combined analysis:

Analysis ID AN006148
Analysis type MS
Chromatography type GC-MS
Chromatography system Thermo Scientific TSQ 9000
Column ThermoFisher TraceGOLD TG-1MS (30m x 0.25mm, 0.25um)
MS Type EI
MS instrument type Triple quadrupole
MS instrument name Thermo Scientific TSQ 9000
Ion Mode POSITIVE
Units Intensity

Chromatography:

Chromatography ID:CH004669
Methods Filename:TS_Method.pdf
Instrument Name:Thermo Scientific TSQ 9000
Column Name:ThermoFisher TraceGOLD TG-1MS (30m x 0.25mm, 0.25um)
Column Temperature:300
Flow Gradient:-
Flow Rate:20mL/min
Solvent A:-
Solvent B:-
Chromatography Type:GC-MS

MS:

MS ID:MS005853
Analysis ID:AN006148
Instrument Name:Thermo Scientific TSQ 9000
Instrument Type:Triple quadrupole
MS Type:EI
MS Comments:MS acquisition was performed using an electron ionization (EI) source at 70 eV in full scan mode. Ion Source Temperature: 300 °C;Transfer Line Temperature: 290 °C;Scan Mode: Full Scan (MSFullScan); Data Type: Centroid; Mass Range: 50 amu to 600 amu; Scan Time: 0.2 seconds; Polarity: Positive Ion Mode.
Ion Mode:POSITIVE
Acquisition Parameters File:TS_Method.pdf
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