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| Status |
Public on Aug 17, 2021 |
| Title |
Subject_9_timepoint_2_red meat_NO_phytochemicals [IP9-2] |
| Sample type |
RNA |
| |
|
| Source name |
timepoint_2_red meat_NO_phytochemicals
|
| Organism |
Homo sapiens |
| Characteristics |
subject id: Subject_9
gender: Male
age: 29y
sample group: red meat_NO_phytochemicals
time point: 2
tissue: Colonic tissue
|
| Treatment protocol |
There are 2 study groups (1 and 2), and in each study group there are 2 meat intervention periods (A and B), separated by a wash-out period (O). During intervention period “A”, participants were asked to consume the provided red processed meat products (red processed meat group), while in period “B” red processed meat products enriched with natural compounds were provided. For study group 1, the meat products of both period ”A” and “B” had standard nitrite levels (standard nitrite PHYTOME meat), while in study group 2 the nitrite levels in period “B” were reduced (reduced nitrite PHYTOME meat). At the beginning of the study and after each intervention period of 2 weeks, colonic biopsies were collected during an endoscopic examination and analysed for gene expression changes.
|
| Growth protocol |
6 small colonic biopsies were collected, transferred to an Eppendorf container and immediately stored in liquid nitrogen.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Colon biopsies were dissolved in QIAzol® (Qiagen, Venlo, The Netherlands) using a tissue disruptor. RNA was isolated according to the manufacturer’s protocol l and followed by DNase I (Qiagen) treatment. Upon purification, RNA concentrations were measured by means of a NanoDrop® ND-1000 spectrophotometer (Thermo Scientific) at 260 and 280 nm. RNA quality and integrity were assessed by using automated gel electrophoresis on an Agilent 2100 Bioanalyzer system (Agilent Technologies) (average RNA integrity number: 7.1 ± 1.0).
|
| Label |
Cy3
|
| Label protocol |
dye-labeled cRNA (Cy3) was synthesised following the Agilent one-color Quick-Amp labeling protocol (Agilent Technologies, Amstelveen, The Netherlands).
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| |
|
| Hybridization protocol |
Microarray hybridization was performed on Agilent 8x44K whole human genome microarrays according to Agilent instructions.
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| Scan protocol |
Slides were scanned using the Agilent Technologies G2565CA DNA Microarray Scanner according to the Agilent protocol. Cy3 was excited at a wavelengths of 532 nm, respectively. Laser power was set to 100%. The photo multiplier tube gain was set to a saturation tolerance of 0.02% to minimize background and saturated spots.
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| Description |
Gene expression in colonic tissue of subject 9 after consumption of two weeks of red meat without phytochemicals
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| Data processing |
Quality control check was done in R (www.r-project.org) using Bioconductor, including local background correction, flagging of bad spots, controls and spots with too low intensity, log2 transformation and normalization. Data were normalized per batch (date of microarray hybridization) using quantile normalization. Genes with less than 30% flagged bad spots were used as a pre-processed input file. Replicates were merged using the median and missing values were imputed by k-nearest neighbours (KNN) imputation (N=15).
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| |
|
| Submission date |
Apr 02, 2020 |
| Last update date |
Aug 17, 2021 |
| Contact name |
Simone G van Breda |
| E-mail(s) |
[email protected]
|
| Phone |
0031433882127
|
| Organization name |
Maastricht University
|
| Department |
Toxicogenomics
|
| Street address |
Universiteitssingel 50
|
| City |
Maastricht |
| State/province |
Limburg |
| ZIP/Postal code |
6229 ER |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL21061 |
| Series (1) |
| GSE147996 |
Replacement of nitrite in meat products by natural bioactive compounds results in reduced exposure to N-nitroso compounds: the PHYTOME project |
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