|
| Status |
Public on Jan 01, 2013 |
| Title |
RB585-NN |
| Sample type |
RNA |
| |
|
| Source name |
CML CD34+
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: bone marrow
msc treatment: No MSC
imatinib treatment: No IM
|
| Treatment protocol |
CML CD34+ cells were treated with or without IM (5μM) in the presence or absence of MSC for 96 hours (n=3 each).
|
| Growth protocol |
CML CD34+ cells were cultured in the presence or absence of irradiated MSC at 37°C with 5% CO2 in serum-free expansion medium (SFEM, Stem Cell Technologies, Vancouver, BC,) supplemented with GFs at concentrations similar to those found in stroma-conditioned medium from long-term bone marrow cultures [granulocyte-macrophage colony-stimulating factor (200pg/mL), granulocyte colony-stimulating factor (1ng/mL), stem cell factor (200pg/mL), leukemia inhibitory factor (50pg/mL), macrophage inflammatory protein-1 (200pg/mL), and interleukin-6 (1ng/mL)].
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the RNAeasy Mini Kit (Qiagen, Valencia, CA), amplified, labeled and hybridized to GeneChip 1.0 arrays (Affymetrix, Santa Clara, CA).
|
| Label |
biotin
|
| Label protocol |
100ng total RNA for each sample was linear amplified and labeled with Biotin
|
| |
|
| Hybridization protocol |
~5.5ug of labeled cDNA was hybridized to Affymetrix Gene Array ST-1.0
|
| Scan protocol |
Arrays were scanned at 5um resolution with Affymetrix scanner 3000 7G
|
| Description |
Gene expression data from CML CD34+ cells
|
| Data processing |
Microarray data analysis was performed using R (version 2.9) with genomic analysis packages from Bioconductor (version 2.4). The 33297 probes represented on the microarray were filtered by cross-sample mean, and for standard deviation of greater than the 25% quantile, yielding 18624 probes representing 12553 genes. Linear regression was used to model the gene expression with the consideration of a 2x2 factorial design and matched samples. Differentially expressed genes were identified by calculating empirical Bayes moderated t-statistic, and p-values were adjusted by FDR using the “LIMMA” package. Gene Set Enrichment Analysis (GSEA) was performed using GSEA software version 2.04 [http://www.broadinstitute.org/gsea/] to detect enrichment of predetermined gene sets using t-scores from all genes for 1263 gene sets in the C2 (curated gene sets) category from the Molecular Signature Database (MsigDB)41. For significant gene sets related to the Wnt/β-catenin and Cadherin pathways, leading edge genes commonly enriched in these sets were analyzed. Heatmaps were plotted to show contrasts amongst the different treatment groups.
|
| |
|
| Submission date |
Dec 31, 2012 |
| Last update date |
Jan 01, 2013 |
| Contact name |
Ravi Bhatia |
| E-mail(s) |
rbhatia@coh.org
|
| Phone |
6263598111
|
| Fax |
6263018973
|
| Organization name |
City of Hope Beckman Research Institute
|
| Department |
Hematopoietic Stem Cell and Leukemia Research
|
| Street address |
1500 E Duarte Road
|
| City |
Duarte |
| State/province |
CA |
| ZIP/Postal code |
91010 |
| Country |
USA |
| |
|
| Platform ID |
GPL6244 |
| Series (1) |
| GSE43225 |
The effect of IM and MSC treatment on gene expression in CML CD34+ cells |
|
