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Sample GSM849618 Query DataSets for GSM849618
Status Public on Dec 22, 2011
Title rapidly proliferating single cell derived mesenchymal cell from bone marrow 1
Sample type genomic
 
Channel 1
Source name Human Control gDNA
Organism Homo sapiens
Characteristics sample type: Promega gDNA female, G152A Lot. 25707301
Extracted molecule genomic DNA
Extraction protocol A maximum of 5x10^6 cells (RPCs, MPCs and SPCs at six weeks) were centrifuged for 5 minutes at 300 g. The pellet was resuspended in 200 μl PBS, and 20 μl proteinase K (Qiagen DNeasy Blood & Tissue Kit) and 4 μl RNase A (100 mg/ml) were added, mixed on a vortex mixer, and incubated for 2 min at room temperature. Then the suspension was spun in a microcentrifuge for 30 seconds at 6,000 g to drive the contents off the walls and lid. Genomic DNA (gDNA) was isolated using a Qiagen DNeasy Blood & Tissue Kit and Agilent’s recommended procedure (http://www.chem.agilent.com/Library/usermanuals/Public/G4410-90040 CGH Bravo Protocol 1.1.pdf). The final elution volume was 400 μl in Buffer AE. The yield and purity of the gDNA were measured using a NanoDrop ND-1000 spectrophotometer (Thermo Fisher Scientific). To assess the quality and average molecular weight of the collected gDNA, aliquots from each sample were subjected to agarose gel electrophoresis. For experimentation, high-quality gDNA samples with a 260/280 ratio of 1.8 to 2.0 (the 260/230 ratio for pure DNA is >1.8) were used.
Label Cy3
Label protocol 1.0 μg aliquots of sample DNA from RPCs, MPCs or SPCs, as well as reference DNA (Promega, female, p/n G1521), were digested with AluI and RsaI restriction enzymes and fluorescently labeled with Cy5 (test) or Cy3 (reference) using an Agilent Genomic DNA Enzymatic Labeling Kit (p/n 5190-0449).
 
Channel 2
Source name Human Mesenchymal Stem Cells
Organism Homo sapiens
Characteristics gender: female
age: 20 y
cell type: rapidly proliferating single cell derived mesenchymal cell from bone marrow
Extracted molecule genomic DNA
Extraction protocol A maximum of 5x10^6 cells (RPCs, MPCs and SPCs at six weeks) were centrifuged for 5 minutes at 300 g. The pellet was resuspended in 200 μl PBS, and 20 μl proteinase K (Qiagen DNeasy Blood & Tissue Kit) and 4 μl RNase A (100 mg/ml) were added, mixed on a vortex mixer, and incubated for 2 min at room temperature. Then the suspension was spun in a microcentrifuge for 30 seconds at 6,000 g to drive the contents off the walls and lid. Genomic DNA (gDNA) was isolated using a Qiagen DNeasy Blood & Tissue Kit and Agilent’s recommended procedure (http://www.chem.agilent.com/Library/usermanuals/Public/G4410-90040 CGH Bravo Protocol 1.1.pdf). The final elution volume was 400 μl in Buffer AE. The yield and purity of the gDNA were measured using a NanoDrop ND-1000 spectrophotometer (Thermo Fisher Scientific). To assess the quality and average molecular weight of the collected gDNA, aliquots from each sample were subjected to agarose gel electrophoresis. For experimentation, high-quality gDNA samples with a 260/280 ratio of 1.8 to 2.0 (the 260/230 ratio for pure DNA is >1.8) were used.
Label Cy5
Label protocol 1.0 μg aliquots of sample DNA from RPCs, MPCs or SPCs, as well as reference DNA (Promega, female, p/n G1521), were digested with AluI and RsaI restriction enzymes and fluorescently labeled with Cy5 (test) or Cy3 (reference) using an Agilent Genomic DNA Enzymatic Labeling Kit (p/n 5190-0449).
 
 
Hybridization protocol Experimental and reference targets for each hybridization were purified using a Microcon YM-30 column (Millipore) and validated using a NanoDrop ND-1000 to ensure the yield and specific incorporation of the labeled gDNA. Labeled test and reference DNAs were combined, denatured, pre-annealed with Cot-1 DNA (Invitrogen, Carlsbad, CA) and blocking reagent (Agilent), and then hybridized to Agilent Human 4x180K CGH arrays (Design ID = 022060) for 24 hours in a rotating oven (Agilent Technologies) at 65°C and 20 rpm.
Scan protocol After the hybridization, the array slides were washed and then scanned at 3 μm resolution with an Agilent G2565CA Scanner. Data were extracted from the microarray images using Agilent Feature Extraction Software v10.5.1.1 with the Feature Extraction protocol CGH-105_Dec09. These ratio data, along with the associated error values and flagged features, were imported into the DNA Analytics Software v5.0.14 (Agilent).
Data processing To make aberration calls, an aberration detection algorithm, ADM-2, was used with a threshold of 5.5 and an aberration filter set at 2 for the minimum number of probe regions and 0.25 for the minimum absolute average log2 ratio for regions in the DNA Analytics to reduce false positives.
 
Submission date Dec 15, 2011
Last update date Dec 22, 2011
Contact name Yo Mabuchi
E-mail(s) yomabuchi@2009.jukuin.keio.ac.jp
Organization name Keio University School of Medicine
Department Physiology
Street address 35 Shinanomachi, Shinjuku-ku
City Tokyo
ZIP/Postal code 160-8582
Country Japan
 
Platform ID GPL10123
Series (1)
GSE34484 LNGFR- and Thy-1-based prospective isolation of human mesenchymal stem cells reveals functionally distinct subpopulations.

Data table header descriptions
ID_REF
VALUE Agilent Feature Extract default normalized log ratio test/ref
NormCh1 gProcessedSignal
NormCh2 rProcessedSignal
QUALITY1 gIsSaturated
QUALITY2 rIsSaturated
QUALITY3 gIsFeatNonUnifOL
QUALITY4 rIsFeatNonUnifOL
QUALITY5 gIsPosAndSignif
QUALITY6 rIsPosAndSignif

Data table
ID_REF VALUE NormCh1 NormCh2 QUALITY1 QUALITY2 QUALITY3 QUALITY4 QUALITY5 QUALITY6
1 -6.32E-02 3.58E+03 3.10E+03 0 0 0 0 1 1
2 0.00E+00 7.27E+00 1.54E+01 0 0 0 0 0 0
3 0.00E+00 7.28E+00 1.54E+01 0 0 0 0 0 0
4 0.00E+00 7.28E+00 1.53E+01 0 0 0 0 0 0
5 0.00E+00 7.29E+00 1.53E+01 0 0 0 0 0 0
6 0.00E+00 7.29E+00 1.53E+01 0 0 0 0 0 0
7 0.00E+00 7.30E+00 1.52E+01 0 0 0 0 0 0
8 0.00E+00 7.30E+00 1.52E+01 0 0 0 0 0 0
9 5.59E-01 7.31E+00 2.65E+01 0 0 0 0 0 1
10 0.00E+00 7.31E+00 1.51E+01 0 0 0 0 0 0
11 0.00E+00 7.31E+00 1.51E+01 0 0 0 0 0 0
12 0.00E+00 7.32E+00 1.51E+01 0 0 0 0 0 0
13 0.00E+00 1.50E+01 1.51E+01 0 0 0 0 1 0
14 0.00E+00 7.33E+00 1.50E+01 0 0 0 0 0 0
15 0.00E+00 7.33E+00 1.50E+01 0 0 0 0 0 0
16 0.00E+00 7.33E+00 1.50E+01 0 0 0 0 0 0
17 0.00E+00 7.34E+00 1.49E+01 0 0 0 0 0 0
18 0.00E+00 7.34E+00 1.49E+01 0 0 0 0 0 0
19 0.00E+00 7.34E+00 1.49E+01 0 0 0 0 0 0
20 0.00E+00 7.35E+00 1.49E+01 0 0 0 0 0 0

Total number of rows: 180880

Table truncated, full table size 8105 Kbytes.




Supplementary file Size Download File type/resource
GSM849618_RPC_1.txt.gz 18.7 Mb (ftp)(http) TXT
Processed data included within Sample table

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