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Sample GSM132423 Query DataSets for GSM132423
Status Public on Feb 07, 2007
Title Series 5: Hyp male
Sample type RNA
 
Source name Mouse femoral shaft
Organism Mus musculus
Characteristics C57BL/6J
Treatment protocol Normal and X-linked hypophosphatemic, Hyp, mice were bred in our colony on the C57BL/6J background from stock originating from the Jackson Laboratory, Bar Harbor, ME. The breeders were fed Teklad Mouse Breeder Diet (W) No. 8626 (Harlan, Madison, WI) and tap water ad lib. Pups were weaned at three weeks of age and fed Teklad Rodent Diet (W) No. 8604 (Harlan). At five weeks of age, Hyp mice (hemizygous male (Hyp/Y) and heterozygous female (Hyp/+)) and littermate normal (wild type) male and female mice were euthanized, and both femora were harvested. The femora were cut at the metaphyseal/diaphyseal junction and the diaphyses were flushed with physiological saline to remove as much marrow as possible. The resulting shafts were frozen in liquid nitrogen and stored at 70 C.
Extracted molecule total RNA
Extraction protocol Samples were processed as described in the Affymetrix GeneChip Expression Analysis Technical Manual (copyright 2004, Affymetrix, Inc., Santa Clara, CA, Rev. 6, Part number 701025, http://www.affymetrix.com). The sample preparation is described here in brief. The femora were weighed and homogenized, and total RNA was extracted with TRIzol (Invitrogen Life Technologies, Carlsbad, CA).
Label Biotin-labeled cRNA
Label protocol Samples were processed as described in the Affymetrix GeneChip Expression Analysis Technical Manual (copyright 2004, Affymetrix, Inc., Santa Clara, CA, Rev. 6, Part number 701025, http://www.affymetrix.com). Samples of RNA were purified on RNeasy columns from Qiagen (Valencia, CA, P/N 74104) and then 8 ug purified RNA were converted to double-stranded cDNA with an Affymetrix One-Cycle cDNA Synthesis Kit (P/N 900431). The cDNA was then expressed as biotin-labeled cRNA by in vitro transcription (IVT) with the 3' Amplification Reagents for IVT Labeling (Affymetrix P/N 900449). Each sample was spiked with bioB, bioC, bioD, and cre (Affymetrix P/N 900299). The biotin-labeled cRNA was fragmented non-enzymatically.
 
Hybridization protocol Samples were processed as described in the Affymetrix GeneChip Expression Analysis Technical Manual (copyright 2004, Affymetrix, Inc., Santa Clara, CA, Rev. 6, Part number 701025, http://www.affymetrix.com). The fragmented cRNA from the 20 independent samples was hybridized to 20 mouse 430 2.0 GeneChip microarrays (Affymetrix P/N 520029) in Affymetrix hybridization buffer for 16 hours at 45 C. The hybridized arrays were washed and stained in the Affymetrix Fluidics Station 400 to attach fluorescent labels to the biotin, followed by biotin-labeled antibody, and then a second staining with fluorescent labeling of the biotin.
Scan protocol Each array was scanned once with the Affymetrix GeneArray Scanner 3000.
Description Experimental design. Equal amounts of RNA from two mice, matched for genotype and sex were pooled to create each sample for microarray analysis. Four treatment groups were done: (1) Normal male mice, (2) Hyp male mice, (3) normal female mice, and (4) Hyp female mice. Five replicates were done with each replicate containing one sample from each of the four treatment groups for a total of 20 independent samples (40 mice total). Each replicate was matched for littermates and parallel processing.
Data processing The data were analyzed with Affymetrix GeneChip Operating System (GCOS) 1.2. GCOS was used for quality control for each array and to scale the mRNA expression (signal value) of each gene so that the average of all genes is set to 500. GCOS was used to create a single pivot table with 45,101 genes for all 20 samples.
 
Submission date Aug 28, 2006
Last update date Feb 07, 2007
Contact name Ralph A. Meyer
E-mail(s) ralpham@aol.com
Organization name Carolinas Medical Center
Department Orthopaedic Research Laboratory
Lab Cannon Research Center, Rm. 304
Street address P.O. Box 32861
City Charlotte
State/province NC
ZIP/Postal code 28232-2861
Country USA
 
Platform ID GPL1261
Series (1)
GSE5657 Effect of X-linked hypophosphatemia (the Hyp mutation) on gene expression in the mid-shaft of the mouse femur

Data table header descriptions
ID_REF
VALUE Signal level
ABS_CALL Present, Absent or Marginal
DETECTION P-VALUE probability of the signal being a false positive

Data table
ID_REF VALUE ABS_CALL DETECTION P-VALUE
AFFX-BioB-5_at 1114.3 P 0.00006
AFFX-BioB-M_at 2033.3 P 0.000044
AFFX-BioB-3_at 1098.6 P 0.000044
AFFX-BioC-5_at 2579 P 0.000052
AFFX-BioC-3_at 2828.2 P 0.000044
AFFX-BioDn-5_at 8501.9 P 0.000044
AFFX-BioDn-3_at 17682.3 P 0.000044
AFFX-CreX-5_at 37064.7 P 0.000052
AFFX-CreX-3_at 44446.4 P 0.000044
AFFX-DapX-5_at 2.4 A 0.988616
AFFX-DapX-M_at 3.9 A 0.921998
AFFX-DapX-3_at 23.9 A 0.81489
AFFX-LysX-5_at 8.9 A 0.749204
AFFX-LysX-M_at 4.6 A 0.699394
AFFX-LysX-3_at 49.2 A 0.089402
AFFX-PheX-5_at 9.2 A 0.852061
AFFX-PheX-M_at 2.3 A 0.991564
AFFX-PheX-3_at 10.2 A 0.58865
AFFX-ThrX-5_at 5 A 0.949786
AFFX-ThrX-M_at 16.9 A 0.574038

Total number of rows: 45101

Table truncated, full table size 1219 Kbytes.




Supplementary file Size Download File type/resource
GSM132423.MEY.MB18.HY.04.5.CEL.gz 5.6 Mb (ftp)(http) CEL

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