|
| Status |
Public on Oct 15, 2015 |
| Title |
mRNA_non-protein induced normal cells_rep1 |
| Sample type |
RNA |
| |
|
| Source name |
non-protein induced normal cells, 750nM for 2 days, replicate 1
|
| Organism |
Mus musculus |
| Characteristics |
cell line: C3H10T1/2
induction: none
|
| Treatment protocol |
C3H10T1/2 cells, diluted 1:1 with MEM/EBSS medium supplemented with 10% heat-inactivated fetal bovine serum in the presence of 750nM IhhN protein, were incubated for 48 hours at 37 ºC in a humidified incubator with 5% CO2.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was prepared from cells using Trizol reagent (Invitrogen) according to manufacturer's instruction.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 0.5 ug RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
|
| |
|
| Hybridization protocol |
1.5 ug of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 250 ml containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 250 ml of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole Human Genome Oligo Microarrays (G4112A) 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), then dried immediately by brief centrifugation.
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505C) using one color scan setting for 1x44k array slides (Scan Area 61x21.6 mm, Scan resolution 10um, Dye channel is set to Green and Green PMT is set to 100%).
|
| Description |
Gene expression after 48hr incubation in C3H10T1/2 cells
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 9.1 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
|
| |
|
| Submission date |
Oct 14, 2015 |
| Last update date |
Oct 15, 2015 |
| Contact name |
Lu Shen |
| E-mail(s) |
yoyomailer@sjtu.edu.cn
|
| Organization name |
Shanghai Jiao Tong University
|
| Department |
Bio-X Institutes
|
| Lab |
Pharmacogenomics
|
| Street address |
RM 218 Little White House 1954 Huashan RD
|
| City |
Shanghai |
| ZIP/Postal code |
200030 |
| Country |
China |
| |
|
| Platform ID |
GPL4134 |
| Series (2) |
| GSE74021 |
Gene expression signatures for IhhN protein induced cells |
| GSE74023 |
Gene expression and Gli1-ChIP in IhhN protein induced cells |
|
