|
| Status |
Public on Jun 30, 2019 |
| Title |
DMSO_treatment_2 as ctrl for Ban Ex |
| Sample type |
RNA |
| |
|
| Source name |
DMSO_treated_hfNSC_rep2
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: hfNSCs
treatment: DMSO
time: 2 days
|
| Treatment protocol |
hfNSCs were treated by 10ng/ml Bangle extract and 10μM cis-Banglene for two days in neuronal differentiated medium (neurobasal medium containing 2% B27 supplement and 0.5 mM L-glutamine).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted by using TRIZOL reagent.
|
| Label |
Cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 0.1ug Total RNA using the Low Input Quick Amp Labeling Kit (Agilent) according to the manufacturer's instructions, followed by RNeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-2000 Spectrophotometer.
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| |
|
| Hybridization protocol |
0.6ug 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 manufacturer’s instructions. On completion of the fragmentation reaction, 25 ul of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to SurePrint G3 Human GE 8x60K Microarray Ver3.0 (Agilent) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent).
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent SureScan Microarray Scanner (G2600D) using one color scan setting for 8x60k array slides (Scan Area 61x21.6 mm, Scan resolution 3um, Dye channel is set to Green PMT is set to 100%).
|
| Description |
AR-2955_02
Gene expression after 2 days in DMSO treated hfNSCs.
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 12.0.3.1 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities. Processed signal intensities were normalized by the global scaling method. A trimmed mean probe intensity was determined by removing 2% of the lower and the higher end of the probe intensities in order to calculate the scaling factor. Normalized signal intensities were then calculated from the target intensity on each array using the scaling factor, so that the trimmed mean target intensity of each array was arbitrarily set to 2500.
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| |
|
| Submission date |
Apr 05, 2019 |
| Last update date |
Mar 25, 2020 |
| Contact name |
Kazumi Hirano |
| E-mail(s) |
kazumi-hirano@aist.go.jp
|
| Phone |
+81-80-4336-7711
|
| Organization name |
AIST
|
| Department |
Biomedical
|
| Street address |
1-1-1 Higashi
|
| City |
Tsukuba |
| State/province |
Ibaraki |
| ZIP/Postal code |
3058566 |
| Country |
Japan |
| |
|
| Platform ID |
GPL20844 |
| Series (1) |
| GSE129415 |
Effect of Bangle extract and cis-Banglene on human NSC neuronal differentiation |
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