Summary of Study ST001093
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 PR000731. The data can be accessed directly via it's Project DOI: 10.21228/M8638T 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.
| Study ID | ST001093 |
| Study Title | Impact of genetic suppression (shRNA) of ALDH1A1 expression in human colon cancer cell line (COLO320) |
| Study Type | Untargeted metabolomics |
| Study Summary | Using a multi-omics approach, we have investigated the impact of genetic suppression (shRNA) of ALDH1A1 expression on transcriptomics, proteomics and untargeted metabolomics analyses in a human colon cancer cell line (COLO320). The present study (i) generates an integrative omic profile of scramble shRNA vs. ALDH1A1 shRNA COLO320 cells, and (ii) identifies possible alterations in biological pathways caused by suppression of ALDH1A1 expression. |
| Institute | Yale University |
| Last Name | Vasiliou |
| First Name | Vasilis |
| Address | 60 College str |
| georgia.charkoftaki@yale.edu; vasilis.vasiliou@yale.edu | |
| Phone | 203.737.8094 |
| Submit Date | 2018-11-12 |
| Num Groups | 2 |
| Raw Data File Type(s) | raw(Waters) |
| Analysis Type Detail | LC-MS |
| Release Date | 2019-09-23 |
| Release Version | 1 |
Select appropriate tab below to view additional metadata details:
Project:
| Project ID: | PR000731 |
| Project DOI: | doi: 10.21228/M8638T |
| Project Title: | Impact of genetic suppression (shRNA) of ALDH1A1 expression in human colon cancer cell line (COLO320) |
| Project Type: | Untargeted metabolomics analyses |
| Project Summary: | Using a multi-omics approach, we have investigated the impact of genetic suppression (shRNA) of ALDH1A1 expression on transcriptomics, proteomics and untargeted metabolomics analyses in a human colon cancer cell line (COLO320). The present study (i) generates an integrative omic profile of scramble shRNA vs. ALDH1A1 shRNA COLO320 cells, and (ii) identifies possible alterations in biological pathways caused by suppression of ALDH1A1 expression. |
| Institute: | Yale School of Public Health |
| Department: | Environmental Health Sciences |
| Last Name: | Vasiliou |
| First Name: | Vasilis |
| Address: | 60 College str, New Haven, CT, 06520, USA |
| Email: | georgia.charkoftaki@yale.edu; vasilis.vasiliou@yale.edu |
| Phone: | 2037857028 |
| Funding Source: | NIH |
Subject:
| Subject ID: | SU001137 |
| Subject Type: | Cultured cells |
| Subject Species: | Homo sapiens |
| Taxonomy ID: | 9606 |
| Genotype Strain: | ALDH1A1 |
| Cell Passage Number: | 5 |
| Cell Counts: | 4 million |
Factors:
Subject type: Cultured cells; Subject species: Homo sapiens (Factor headings shown in green)
| mb_sample_id | local_sample_id | Group name |
|---|---|---|
| SA074508 | X01222018_Colo320_20V_Lipid_POS_4 | KO ALDH1A1 |
| SA074509 | X01222018_Colo320_20V_Lipid_POS_3 | KO ALDH1A1 |
| SA074510 | X01222018_Colo320_20V_Lipid_POS_5 | KO ALDH1A1 |
| SA074511 | X01222018_Colo320_20V_Lipid_POS_7 | KO ALDH1A1 |
| SA074512 | X01222018_Colo320_20V_Lipid_POS_9 | KO ALDH1A1 |
| SA074513 | X01222018_Colo320_20V_Lipid_POS_2 | KO ALDH1A1 |
| SA074514 | X01222018_Colo320_20V_Lipid_POS_8 | KO ALDH1A1 |
| SA074515 | X01222018_Colo320_20V_Lipid_POS_QC_12 | QC |
| SA074516 | X01222018_Colo320_20V_Lipid_POS_QC_11 | QC |
| SA074517 | X01222018_Colo320_20V_Lipid_POS_QC_13 | QC |
| SA074518 | X01222018_Colo320_20V_Lipid_POS_QC_14 | QC |
| SA074519 | X01222018_Colo320_20V_Lipid_POS_QC_16 | QC |
| SA074520 | X01222018_Colo320_20V_Lipid_POS_QC_15 | QC |
| SA074521 | X01222018_Colo320_20V_Lipid_POS_QC_10 | QC |
| SA074522 | X01222018_Colo320_20V_Lipid_POS_QC_09 | QC |
| SA074523 | X01222018_Colo320_20V_Lipid_POS_QC_08 | QC |
| SA074524 | X01222018_Colo320_20V_Lipid_POS_QC_07 | QC |
| SA074525 | X01222018_Colo320_20V_Lipid_POS_15 | WT Colo320 |
| SA074526 | X01222018_Colo320_20V_Lipid_POS_11 | WT Colo320 |
| SA074527 | X01222018_Colo320_20V_Lipid_POS_14 | WT Colo320 |
| SA074528 | X01222018_Colo320_20V_Lipid_POS_10 | WT Colo320 |
| SA074529 | X01222018_Colo320_20V_Lipid_POS_12 | WT Colo320 |
| SA074530 | X01222018_Colo320_20V_Lipid_POS_16 | WT Colo320 |
| SA074531 | X01222018_Colo320_20V_Lipid_POS_13 | WT Colo320 |
| Showing results 1 to 24 of 24 |
Collection:
| Collection ID: | CO001131 |
| Collection Summary: | the cell culture medium was removed from the culture dishes, and the cells were rinsed three times with ice-cold DPBS. For quenching and metabolite extraction, 1 mL ice-cold methanol was added to the plate and the cells were immediately removed from the culture dish by scraping. The cell suspension was collected into 2 mL Protein Lobind Eppendorf tubes that were placed on ice. After cell counting (ViaStain AOPI Staining Solution,cridine orange/propidium iodide Nexcelom Bioscience, Lawrence, MA) using an automated cellometer (Cellometer K2, Nexcelom Bioscience, Lawrence, MA), the cell number was adjusted to 4x106 cells/tube. The cells were lysed by the application of a freeze-thaw cycle (freezing in liquid nitrogen for 5 min, thawing in warm water for 5 min), followed by sonication for 2 min. This process was repeated three times. The samples were then centrifuged at 15,000 rpm for 10 min at 4°C, and the supernatant was transferred to 2 mL tubes and dried using a Speed Vac (SPD111V Savant, Thermo Fisher Scientific). The samples were then reconstituted in isopropanol:acetonitrile:water (2:1:1 %v/v) and 2.5 μL Splash® Lipidomix® (Avanti Polar Lipids Inc.) was added to each sample as an internal standard |
| Sample Type: | Cultured cells |
Treatment:
| Treatment ID: | TR001151 |
| Treatment Summary: | Cellular ALDH1A1 expression was suppressed using MISSION shRNA lentiviral transduction particles containing validated ALDH1A1 shRNA in a pLKO plasmid vector. Scramble control shRNA in pLKO plasmid vector was used to generate control cells (Sigma-Aldrich, St. Louis, MO). The shRNA transfection was carried out as described previously [1] and individual stable clones were selected using puromycin (10 μg/mL). Western blot analysis was used to verify changes in ALDH1A1 expression using methods described previously [2]. References [1] S. Guo, H. Bai, C.M. Megyola, S. Halene, D.S. Krause, D.T. Scadden, J. Lu, Complex oncogene dependence in microRNA-125a-induced myeloproliferative neoplasms, Proc Natl Acad Sci U S A, 109 (2012) 16636-16641. [2] Y. Chen, D.C. Thompson, V. Koppaka, J.V. Jester, V. Vasiliou, Ocular aldehyde dehydrogenases: protection against ultraviolet damage and maintenance of transparency for vision, Prog Retin Eye Res, 33 (2013) 28-39. |
Sample Preparation:
| Sampleprep ID: | SP001144 |
| Sampleprep Summary: | Briefly, the cell culture medium was removed from the culture dishes, and the cells were rinsed three times with ice-cold DPBS. For quenching and metabolite extraction, 1 mL ice-cold methanol was added to the plate and the cells were immediately removed from the culture dish by scraping. The cell suspension was collected into 2 mL Protein Lobind Eppendorf tubes that were placed on ice. After cell counting (ViaStain AOPI Staining Solution,cridine orange/propidium iodide Nexcelom Bioscience, Lawrence, MA) using an automated cellometer (Cellometer K2, Nexcelom Bioscience, Lawrence, MA), the cell number was adjusted to 4x106 cells/tube. The cells were lysed by the application of a freeze-thaw cycle (freezing in liquid nitrogen for 5 min, thawing in warm water for 5 min), followed by sonication for 2 min. This process was repeated three times. The samples were then centrifuged at 15,000 rpm for 10 min at 4°C, and the supernatant was transferred to 2 mL tubes and dried using a Speed Vac (SPD111V Savant, Thermo Fisher Scientific). The samples were then reconstituted in isopropanol:acetonitrile:water (2:1:1 %v/v) and 2.5 μL Splash® Lipidomix® (Avanti Polar Lipids Inc.) was added to each sample as an internal standard. |
Combined analysis:
| Analysis ID | AN001779 |
|---|---|
| Analysis type | MS |
| Chromatography type | Reversed phase |
| Chromatography system | Waters |
| Column | Waters Acquity BEH C18 (100 x 2.1mm,1.7um) |
| MS Type | ESI |
| MS instrument type | QTOF |
| MS instrument name | Waters Synapt G2 XS |
| Ion Mode | POSITIVE |
| Units | abundance corresponding to m/z |
Chromatography:
| Chromatography ID: | CH001257 |
| Chromatography Summary: | The mobile phase involved a gradient comprising varying amounts of solutions A (10 mM ammonium formate in 60% v/v acetonitrile) and B (10 mM ammonium formate in 95% isopropanol and 5% acetonitrile). The linear gradient elution was: 40–43% B (0–2 min), 43–50% B (over 0.1 min), 50–54% B (over 10 min), 54–70% B (over 0.1 min), 70–99% B (over 5.9 min), 99-40% B (over 0.1 min), and 40% B (for 2 min), run at 300 μl/min in positive Electrospray Ionization mode and the column temperature was maintained at 55 °C |
| Instrument Name: | Waters |
| Column Name: | Waters Acquity BEH C18 (100 x 2.1mm,1.7um) |
| Flow Rate: | 300 μl/min |
| Injection Temperature: | 4 |
| Internal Standard: | SPLASH™ Lipidomix® Mass Spec Standard (Avanti Polar Lipids, Inc.) |
| Solvent A: | 60% acetonitrile/40% water; 10 mM ammonium formate |
| Solvent B: | 95% isopropanol/5% acetonitrile; 10 mM ammonium formate |
| Chromatography Type: | Reversed phase |
MS:
| MS ID: | MS001643 |
| Analysis ID: | AN001779 |
| Instrument Name: | Waters Synapt G2 XS |
| Instrument Type: | QTOF |
| MS Type: | ESI |
| Ion Mode: | POSITIVE |
