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Sample GSM2418517 Query DataSets for GSM2418517
Status Public on Jan 03, 2017
Title CB-CD34+_single culture_control 3
Sample type RNA
 
Source name Umbilical cord blood
Organism Homo sapiens
Characteristics tissue: Umbilical cord blood
Treatment protocol After 10 days of culture, cells were collected and sorted by a FACSAria II flow cytometer (Becton Dickinson). Flow cytometric data were analyzed using FACSDiva software (Becton Dickinson). MSCs were identified on the basis of CD73, CD90, CD105 co-expression and lack of CD45. CB-CD34+ cells were identified on the basis of CD45 and CD34 co-expression. At least 3x10^5 gated events were acquired for each sample. Purity of FACS-isolated cells was greater than 95%, as assessed by flow cytometric analysis of post-sorting samples.
Growth protocol BM-MSCs were thawed at 37°C and 5x10^5 cells were plated in T75 flasks. Cells were cultured in complete ISCOVE’s modified Dulbecco’s medium supplemented with 10% FBS for a week until 80% confluence. CB-CD34+ cells were seeded directly onto the MSCs layer at a final concentration of 3x10^5 cells/flask. Cells were maintained in serum-free medium (StemSpan H3000, StemCell Technologies) supplemented with 100 ng/ml Stem Cell Factor (SCF) (Peprotech), Thrombopoietin (TPO) (Peprotech) and Granulocyte Colony Stimulating Factor (G-CSF) (Italfarmaco) for 10 days at 37°C and 5% CO2.
Extracted molecule total RNA
Extraction protocol Total cellular RNA was isolated using Qiagen RNeasy kit according to manufactures’ instructions (Qiagen, Hilden, Germany).
Label Biotin
Label protocol Biotinylated aRNA were prepared according to the standard Affymetrix protocol from 50 ng total RNA using the GeneChip 3’ IVT Expression kit (Affymetrix)
 
Hybridization protocol Labelled RNA were fragmented and hybridized on Affymetrix HG-U133A PLUS 2.0 GeneChip arrays that were subsequently washed and stained using the GeneChip Hybridization, Wash and Stain Kit (Affymetrix)
Scan protocol The GeneChip was scanned and data extracted using GeneChip scanner 3000 7G (Affimetrix, SantanClara, CA).
Data processing The data were analyzed with Affymetrix® Expression Console™ Software and normalyzed with RMA algorithm. Subsequently analyses for differentially expressed genes cathegorization were performed with Partek® Genomics Suite software version 6.6 (Partek Inc., St. Louis, MO). The analysis of molecular pathways was carried out through the DAVID Functional Annotation Bioinformatics Microarray Analysis. Broad Institute Gene Set Enrichment Analysis (GSEA) software was performed to identify significant enrichments in differentially expressed genes signatures, in case of nominal P values corrected for False Discovery Rate (FDR) for enrichment scores minor than 0.05. The network analyses were generated through the use of Ingenuity Pathway Analysis® (IPA®, Qiagen).
 
Submission date Dec 07, 2016
Last update date Jan 03, 2017
Contact name Simone Perucca
E-mail(s) [email protected]
Phone +39 0303998464
Organization name University of Brescia
Department Department of Clinical and experimental sciences
Lab Chair of Hematology
Street address Viale Europa
City Brescia
ZIP/Postal code 25123
Country Italy
 
Platform ID GPL570
Series (1)
GSE90970 Mesenchymal Stromal Cells Induce Ex Vivo Proliferation and Erythroid Commitment of Cord Blood Haematopoietic Stem Cells

Data table header descriptions
ID_REF
VALUE log2-rma

Data table
ID_REF VALUE
AFFX-BioB-5_at 8.95529
AFFX-BioB-M_at 9.23806
AFFX-BioB-3_at 8.89265
AFFX-BioC-5_at 10.0371
AFFX-BioC-3_at 10.8798
AFFX-BioDn-5_at 11.699
AFFX-BioDn-3_at 12.7765
AFFX-CreX-5_at 13.3827
AFFX-CreX-3_at 13.8498
AFFX-DapX-5_at 5.79042
AFFX-DapX-M_at 7.93762
AFFX-DapX-3_at 8.84419
AFFX-LysX-5_at 4.37202
AFFX-LysX-M_at 4.57005
AFFX-LysX-3_at 5.47143
AFFX-PheX-5_at 5.02016
AFFX-PheX-M_at 5.44958
AFFX-PheX-3_at 6.09534
AFFX-ThrX-5_at 4.12262
AFFX-ThrX-M_at 4.8711

Total number of rows: 54675

Table truncated, full table size 998 Kbytes.




Supplementary file Size Download File type/resource
GSM2418517_CB-Alone_B3_HG-U133_Plus_2_.CEL.gz 4.4 Mb (ftp)(http) CEL
GSM2418517_CB-Alone_B3_HG-U133_Plus_2_.RMA.rma.chp.gz 375.7 Kb (ftp)(http) CHP
Processed data included within Sample table
Processed data provided as supplementary file

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