|
| Status |
Public on Jun 25, 2020 |
| Title |
60weeks_1 |
| Sample type |
RNA |
| |
|
| Source name |
Mouse colon organoids treated with inflammatory reagents for 60 weeks
|
| Organism |
Mus musculus |
| Characteristics |
tissue: colon organoid
treatment: inflammatory reagents for 60 weeks
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using the RNeasy Micro Kit (Qiagen, Hilden, Germany), according to the manufacturer’s instructions. The protocol includes on-column DNase digestion. RNA was quantified using a NanoDrop 1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 50 ng RNA using Low Input Quick Amp Labeling Kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy mini spin columns purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with Agilent 2100 BioAnalyzer and the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
600 ng of Cy3-labelled cRNA was fragmented at 60°C for 30 minutes in a reaction volume of 25 ul containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 25 ul of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Microarrays (G4852A) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with Gene Expression Wash Buffer 1 (Agilent) and 1 minute with 37°C Gene Expression Wash buffer 2 (Agilent), then dried immediately by air-drying.
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent Microarray Scanner using one color scan setting (Scan resolution 3um, TIFF file dynamic range 20bit).
|
| Description |
60 weeks treatment with inflammatory reagents
|
| Data processing |
Agilent Feature Extraction was used to quantify each spot. The output RAW data was imported into GeneSpring (Agilent) and normalized with 75th percentile shift.
|
| |
|
| Submission date |
Jun 24, 2020 |
| Last update date |
Jun 25, 2020 |
| Contact name |
Shuji Hibiya |
| Organization name |
Tokyo Medical and Dental University
|
| Department |
Gastroenterology and Hepatology
|
| Street address |
1-5-45, Yushima, Bunkyo-ku
|
| City |
Tokyo |
| ZIP/Postal code |
1138519 |
| Country |
Japan |
| |
|
| Platform ID |
GPL10787 |
| Series (2) |
| GSE153176 |
Gene expression of mouse colon organoids treated with inflammatory reagents. |
| GSE153178 |
Mouse colon organoids treated with inflammatory reagents. |
|
