|
| Status |
Public on Aug 08, 2019 |
| Title |
Day 4 -2 |
| Sample type |
RNA |
| |
|
| Source name |
iPS cells during chondrocyte differentiation by the 2C method at day 4
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: iPS
day: 4
|
| Treatment protocol |
Human iPS cells were differentiated to chondrocytes by the 2C method for 9 days.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from the chondrocytes using RNeasy mini kits (Qiagen) according to manufacturer’s protocol.
|
| 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 RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-2000 Spectrophotometer.
|
| |
|
| 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 Mouse GE 8x60K Microarray (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), then dried immediately by brief centrifugation.
|
| Scan protocol |
Slides were scanned immediately after washing on the the Agilent DNA Microarray Scanner (G2565CA) 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%).
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.10.1.1 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities.
|
| |
|
| Submission date |
Mar 09, 2017 |
| Last update date |
Aug 08, 2019 |
| Contact name |
Taku Saito |
| E-mail(s) |
tasaitou-tky@umin.ac.jp
|
| Organization name |
The University of Tokyo
|
| Department |
Orthopaedic Surgery, Faculty of Medicine
|
| Street address |
7-3-1 Hongo, Bunkyo-ku
|
| City |
TOKYO |
| ZIP/Postal code |
113-8655 |
| Country |
Japan |
| |
|
| Platform ID |
GPL21185 |
| Series (1) |
| GSE96036 |
Gene expression during chondrocyte differentiation from human iPS cells by the 2C method |
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