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Sample GSM1274500 Query DataSets for GSM1274500
Status Public on Dec 31, 2014
Title MCF10A_SC_R4
Sample type RNA
 
Source name MCF10A single cell
Organism Homo sapiens
Characteristics cell line: MCF10A
sample type: Amplified cDNA from single cell
Growth protocol The human epithelial cell lines MCF7 and MCF10A were grown in DMEM (Gibco, Paisley, Scotland) supplemented with 10% heat-inactivated FCS, 2.5% HEPES buffer (pH 7.2), 0.1% β-mercaptoethanol and 2mM L-glutamine (all from Sigma, UK). Both cell lines were maintained in a 5% CO2 humidified incubator at 37°C and were routinely tested for the presence of mycoplasma.
Extracted molecule total RNA
Extraction protocol RNA (1ng-25pg) or cell lysates in CLS were adjusted to 6.75ml volume and RNA-amplified using the EpiStem RNA-Amp Kit according to the manufacturers protocols (EpiStem, Manchester, UK). Briefly the samples underwent oligo-dT priming and 5’ capping prior to x35 cycles of PCR amplification using the conditions 90ºC 30 sec, 42ºC 2 min and 72ºC 6 min. Following amplification all samples were purified using a MoBio UltraClean® PCR Clean-Up Kit (Carlsbad, CA, USA) and quantified on a NanoDrop spectrophotometer.
Label Biotin
Label protocol Following amplification the cDNA was Biotin-labeled using the EpiStem RNA-Amp labeling kit (EpiStem, Manchester, UK)
 
Hybridization protocol The hybridisation cocktail containing 5mg of biotin cDNA was heated to 99°C for 5 mins. The hybridisation cocktail was then transferred to 45°C for 5 mins and centrifuged at maximum speed for 5 mins to remove insoluble material. Samples were then hybridised to Affymetrix Human HGU133 Plus 2 arrays for 16 hours overnight and then stained with SAPE using a biotin targeted antibody step and washed according to the EukGE_Ws2v4_450 fluidics protocol from Affymetrix.
Scan protocol Samples were scanned in an Affymetrix 3000 scanner.
Data processing Data were normalised and summarised using RMA, as implemented by Affymetrix Power Tools (APT) software package (http://www.affymetrix.com/estore/partners_programs/programs/developer/tools/powertools.affx) using the 'apt-probeset-summarize' default parameter setting. In subsequent analysis, gene level summaries (not included here) were computed as the geometric mean of all probesets mapping to a gene, as defined by annmap, using ENSEMBL version 70 as the source of underlying genome annotation. The empirical Bayes statistics implemented in the Bioconductor package LIMMA were used to identify protein-coding genes showing differential levels in MCF7 and MCF10A samples (FDR < 0.05; absolute fold change > 2)
 
Submission date Nov 25, 2013
Last update date Dec 31, 2014
Contact name Ged Brady
Organization name Caner Research UK Manchester Institute
Department Clinical and Experimental Pharmacology
Street address Cancer Research UK Manchester Institute, The University of Manchester, Wilmslow Rd
City Manchester
ZIP/Postal code M20 4BX
Country United Kingdom
 
Platform ID GPL570
Series (2)
GSE52712 Flexible multiplatform RNA profiling at the single cell level applied to enriched cancer initiating cells: Affymetrix array data
GSE52717 Flexible multiplatform RNA profiling at the single cell level applied to enriched cancer initiating cells

Data table header descriptions
ID_REF
VALUE Log2 RMA-normalised signal

Data table
ID_REF VALUE
AFFX-BioB-5_at 7.87079
AFFX-BioB-M_at 8.19533
AFFX-BioB-3_at 8.59847
AFFX-BioC-5_at 11.51854
AFFX-BioC-3_at 10.75317
AFFX-BioDn-5_at 12.32323
AFFX-BioDn-3_at 13.4361
AFFX-CreX-5_at 14.05034
AFFX-CreX-3_at 14.34341
AFFX-DapX-5_at 4.50043
AFFX-DapX-M_at 4.32762
AFFX-DapX-3_at 4.04259
AFFX-LysX-5_at 3.93017
AFFX-LysX-M_at 4.9249
AFFX-LysX-3_at 4.96756
AFFX-PheX-5_at 5.78283
AFFX-PheX-M_at 4.10838
AFFX-PheX-3_at 6.81286
AFFX-ThrX-5_at 5.03736
AFFX-ThrX-M_at 5.30377

Total number of rows: 54675

Table truncated, full table size 1002 Kbytes.




Supplementary file Size Download File type/resource
GSM1274500_0412_YH353_H_epi_MCF10A_1cell_r8.CEL.gz 4.8 Mb (ftp)(http) CEL
Processed data included within Sample table

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