|
| Status |
Public on Nov 07, 2008 |
| Title |
hAFF-4U-M-iPS-3_rep2 |
| Sample type |
RNA |
| |
|
| Source name |
iPS cell line derived from hAFF by 4F+UTF1-Myc, clone 3 (rep 2)
|
| Organism |
Homo sapiens |
| Characteristics |
Human Induced Pluripotent Stem cell
Donor: hAFF
|
| Growth protocol |
Human fibroblasts were cultured in Dulbecco’s Modified Eagle Medium (DMEM, Hyclone) containing 10% fetal bovine serum (FBS, Gibco). hES cells, iPS cells and Pre-iPS cells were maintained on Mitomycin C-treated MEFs in hES Cell culture medium consisting of 80% DMEM/F12 (Invitrogen), 20% Knockout serum replacement (KSR) (Invitrogen), 1 mM L-glutamine, 1% non-essential amino acids, 0.1 mM beta-mercaptoethanol (all from Invitrogen) and 4 ng/ml basic fibroblast growth factor bFGF (P&A Biotech).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated from cells using TRIzol (Invitrogen) according to the manufacturer’s protocol.
|
| Label |
Cy5
|
| Label protocol |
cDNA were synthesized by reverse transcription using the Amino Allyl MessageAmp II aRNA Amplification kit (Ambion) with oligo(dT) primer that contain a T7 RNA polymerase promoter sequence, according to the manufacturer’s instruction. In vitro transcription was then performed with the purified cDNA and amino allyl UTP was incorporated during transcription to produce amino allyl modified aRNA that was subsequently coupled to cy5 dye label by reacting at 25℃ for 2h.
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| |
|
| Hybridization protocol |
The Human OneArray was hybridized using 1.5× OneArrayTM Hybridization Buffer for 16 h at 60℃; then washed in Wash Buffer 1, 2 and 3 accroding to the manufacturer's instruction.
|
| Scan protocol |
Arrays were scanned using GenePix 4000B scanner (Molecular Devices) according to the manufacturer’s protocol
|
| Description |
gene expression comparation of iPS production, 4F represents OCT4/SOX2/c-Myc/KLF4
|
| Data processing |
After removing control probes, a 14/33 presence call (SNR>=5 and foreground-background>0) was used to filter probes for the 33 microarrays, resulting in 12311 probes for further quantile normalization. The median of each sample was used for hierarchical clustering. Complete-linkage hierarchical clustering was performed using cluster 3.0 with Spearman rank correlation coefficient as gene distances measurement, and Pearson correlation coefficient as sample distances measurement (Attached in supplementary data).
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| |
|
| Submission date |
Sep 24, 2008 |
| Last update date |
Nov 07, 2008 |
| Contact name |
hongkui deng |
| E-mail(s) |
hongkui_deng@pku.edu.cn
|
| Phone |
86-10-6275-6474
|
| Fax |
86-10-6275-6474
|
| Organization name |
Peking University
|
| Department |
College of Life Sciences
|
| Street address |
Yiheyuan Road 5#, Haidian District
|
| City |
Beijing |
| ZIP/Postal code |
100871 |
| Country |
China |
| |
|
| Platform ID |
GPL6254 |
| Series (1) |
| GSE12922 |
Two Supporting Factors Greatly Improve the Efficiency of Human iPS Cell Generation |
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