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Sample GSM544728 Query DataSets for GSM544728
Status Public on May 20, 2010
Title Control_2
Sample type RNA
Source name Cultured Skin Fibroblasts, Wild-Type Control
Organism Homo sapiens
Characteristics gender: female
age: 25 yrs
Treatment protocol NA
Growth protocol Cells were grown on DMEM medium (InVitrogen, Carlsbad, CA) supplemented with 20% heat-inactivated fetal bovine serum (Hyclone, Logan, UT) and 2 mM L-glutamine, and were used before passage 10. All cultures were initiated at 7.5 x 105 cells per 75 cm2 flask. The culture medium was changed every 3-4 days.
Extracted molecule total RNA
Extraction protocol On day 16, approximately 10 days post-confluence, cells were harvested following trypsinization. Cells from triplicate flasks were pooled, washed twice with phosphate buffered saline and re-suspended in PBS. Cell numbers were determined using a Coulter Counter Model Z. The total cell suspension was divided into aliquots for RNA extraction, lipid analysis, and determination of residual glucocerebrosidase activity. The cell suspensions were centrifuged at 1000 x g for 5 min and dry cell pellets were stored at -80 C until used.
Label biotin
Label protocol Labeling was performed on a sample-by-sample basis according to manufacturer’s guidelines for use with the Affymetrix Human Genome U133A_2 GeneChip (Affymetrix, Inc). DNase treatment was included as part of isolation to remove possible contaminating DNA. Bioanalyzer nanochip (Agilent, Inc) and NanoDrop (ThermoFisher, Inc) were used to validate and quantitate the RNA prior to cRNA synthesis and labeling. For cRNA synthesis and labeling, 5ug of total RNA was used per sample in conjunction with the Affymetrix 3’ one-cycle Target labeling Kit (Affymetrix, Inc).
Hybridization protocol Labeled cRNA were hybridized to Affymetrix Human U133A_2 GeneChips (Affymetrix, Inc) in blinded interleaved fashion.
Scan protocol Affymetrix scanner 3000 was used in conjunction with Affymetrix GeneChip Operation Software to generate ~580,000 probe measurements/hybridized cRNA.
Description Cultured Skin Fibroblasts, Wild-Type Control, biological replicate 2
Data processing The “rma” function in bioconductor ( was used to summarize probe measurements and generate normalized gene fragment expression values for each hybridized cRNA. Quality of data was assured via sample-level inspection by Tukey box plot, covariance-based PCA scatter plot, and correlation-based Heat Map. Gene fragments not having at least one expression value greater then system noise were deemed non-informative and discarded. System noise was defined as the lowest expression value at which the LOWESS fit of observed CV for each gene fragment by mean expression value for each gene fragment changes from non-linear to linear. For gene fragments not discarded, expression values were floored to equal system noise if less than system noise then subject to significance testing. The one-factor ANOVA under BH FDR MCC condition (alpha criteria<0.05) was used to identify gene fragments having expression values significantly associated with differences in gender. Spearman correlation under BH FDR MCC condition (alpha criteria<0.05) was used to identify gene fragments having expression values significantly associated with differences in age. Gene fragments found to have expression values significantly associated with differences in age and/or gender were discarded before testing gene fragment expression for significant association with Type I and/or Type III GD. Testing gene fragment expression for significant association with Type I and/or Type III GD was done using the one-factor ANOVA under BH FDR MCC condition (alpha criteria<0.05). For gene fragments found to have expression significantly associated with Type I and/or Type III GD, the Tukey HSD test was used to code gene fragments having a post-hoc p-value < 0.05 between Type I GD vs. control, Type III GD vs. control, and/or Type III GD vs. Type I GD. These codes were used classify gene fragments having expression significantly association with Type I and/or Type III GD as significantly dysregulated in Type I GD only, significantly dysregulated in Type III GD only, significantly and concordantly dysregulated in both Type I GD and Type III GD, or significantly and disconcordantly dysregulated in both Type I GD and Type III GD. Gene fragment annotations and associated gene functions were obtained using IPA (Ingenuity, Inc.).
Submission date May 19, 2010
Last update date May 19, 2010
Contact name Kory R Johnson
Phone 301-402-1956
Organization name NINDS/NIH
Department DIR IT & Bioinformatics
Lab Bioinformatics Section
Street address 10/3B01, 9000 Rockville Pike
City Bethesda
State/province MD
ZIP/Postal code 20892
Country USA
Platform ID GPL571
Series (1)
GSE21899 Global Gene Expression Profiles in Cultured Skin Fibroblasts Derived from Patients with Gaucher Disease

Data table header descriptions
VALUE log (base=2) RMA signal

Data table
1007_s_at 9.044540195
1053_at 6.81242011
117_at 4.037693105
121_at 7.434929611
1255_g_at 3.151095609
1294_at 7.963359171
1316_at 5.128225171
1320_at 6.016132422
1405_i_at 3.177515862
1431_at 5.192614086
1438_at 6.323395561
1487_at 7.128653932
1494_f_at 5.355758716
1598_g_at 12.60025567
160020_at 8.032784718
1729_at 8.504669994
177_at 5.811365819
1773_at 7.077082647
179_at 8.247553923
1861_at 8.216704092

Total number of rows: 22277

Table truncated, full table size 496 Kbytes.

Supplementary file Size Download File type/resource
GSM544728.CEL.gz 1.9 Mb (ftp)(http) CEL
Processed data included within Sample table

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