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Sample GSM497551 Query DataSets for GSM497551
Status Public on Jan 12, 2011
Title T3_CMMEF-2, biological rep2
Sample type RNA
 
Source name hES-T3 cells grown on feeder-free Matrigel in MEF-conditioned medium
Organism Homo sapiens
Characteristics growth conditions: grown on feeder-free Matrigel in MEF-conditioned medium
gender: female
cell line: hES-T3
Treatment protocol No
Growth protocol The MEF cells were cultured in MEF medium overnight, and the mitotically inactivated MEF cells were maintained in hES medium containing 4 ng/ml bFGF. After 24 h, the MEF-conditioned medium was collected and filtered through 0.2 um membrane (PN4612, Pall Life Sciences) . The culture dish was coated with Matrigel diluted with DMEM/F12 (1:30) overnight at 4oC. The cryopreserved stock of hES-T3 cells (36 passages) were continuously maintained on feeder-free Matrigel-coated dish in MEF-conditioned medium (with additional 4 ng/ml bFGF) for 12 passages, and these hES-T3 cells were designated as T3/CMMEF.
Extracted molecule total RNA
Extraction protocol Trizol extraction of total RNA was performed according to the manufacturer's instructions. Total RNAs extracted using TRIZOL reagent were quantified spectrophotometrically. The cDNA synthesis was carried out by using SUPERSCRIPTTM one-step RT-PCR kit (Invitrogen). For the reverse transcription step, the whole 5 ul of the resuspended RNAs were incubated for 60 min at 42oC, then 15 min at 72oC in 50 ul of reaction mixture containing 25 ul of 2x Reaction Mix (Invitrogen) and 1 ul of RT/PLANTINUM Taq Mix (Invitrogen). 28.5 ul of the cDNAs present in the 50 ul RT reaction mixture purified by Microarray Target Purification Kit (Roche Applied Science, Indianapolis, ID, USA, http://www.roche-applies-science.com) were used as templates to amplify cDNA by Microarray Target Amplification Kit. The amplified cDNAs were purified by Microarray Target Purification Kit, and complementary RNAs (cRNAs) were synthesized from 200 ng cDNA with Microarray RNA Target Synthesis Kit (Roche Applied Science).
Label biotin
Label protocol Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 5 micrograms purified cRNA (Expression Analysis Technical Manual, 2004, Affymetrix).
 
Hybridization protocol Following fragmentation, 10 micrograms of cRNA were hybridized for 16 hr at 45oC on Affymetrix Human Genome U133 plus 2.0 GeneChip. GeneChips were washed and stained in the Affymetrix Fluidics Station 400.
Scan protocol GeneChips from the hybridization experiments were read by the Affymetrix GeneChip scanner 3000.
Description The mRNA profiles from T3/CMHDF, T3/CMMEF, T3/HDF and T3/MEF were quantitatively determined.
Data processing The data were processed by GeneSpring software version 7.3 GC-RMA preprocessor to use on the CEL files
 
Submission date Jan 14, 2010
Last update date Jan 12, 2011
Contact name Sung-Liang Yu
E-mail(s) slyu@ntu.edu.tw
Phone 886-2-23958341
Organization name National Taiwan University
Department Clinical Laboratory Sciences and Medical Biotechnology
Lab Microarray Core Facility
Street address Jen Ai Road Section1
City Taipei
ZIP/Postal code 100
Country Taiwan
 
Platform ID GPL570
Series (1)
GSE19902 Comparative expression profiles of human embryonic stem cells among different feeders and conditioned media

Data table header descriptions
ID_REF Affymetrix Synthetic ID
VALUE GC-RMA-calculated Signal intensity

Data table
ID_REF VALUE
1007_s_at 0.924
1053_at 1.104
117_at 1.015
121_at 1.579
1255_g_at 30.03
1294_at 0.265
1316_at 1.507
1320_at 0.577
1405_i_at 1.296
1431_at 1.002
1438_at 1.057
1487_at 0.907
1494_f_at 0.695
1552256_a_at 0.646
1552257_a_at 0.972
1552258_at 0.684
1552261_at 1.655
1552263_at 1.228
1552264_a_at 0.936
1552266_at 1.223

Total number of rows: 54675

Table truncated, full table size 888 Kbytes.




Supplementary file Size Download File type/resource
GSM497551_T3_CMMEF_2.CEL.gz 4.2 Mb (ftp)(http) CEL
Processed data included within Sample table

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