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Sample GSM575587 Query DataSets for GSM575587
Status Public on Oct 01, 2010
Title Old_15_Basal
Sample type RNA
 
Source name Old_Basal
Organism Homo sapiens
Characteristics tissue: vastus lateralis basal skeletal muscle
gender: male
age: 66 years
Extracted molecule total RNA
Extraction protocol Tri-Reagent
Label Cy3
Label protocol Microarray assay was performed using a service provider (LC Sciences). The assay started from 4 to 8 µg total RNA sample, which was size fractionated using a YM-100 Microcon centrifugal filter (Millipore) and the small RNAs (< 300 nt) isolated were 3’-extended with a poly(A) tail using poly(A) polymerase. An oligonucleotide tag was then ligated to the poly(A) tail for later fluorescent dye staining; two different tags were used for the two RNA samples in dual-sample experiments.
 
Hybridization protocol Hybridization was performed overnight on a µParaflo microfluidic chip using a micro-circulation pump (Atactic Technologies) (1). On the microfluidic chip, each detection probe consisted of a chemically modified nucleotide coding segment complementary to target microRNA (from miRBase, http://microrna.sanger.ac.uk/sequences/) or other RNA (control or customer defined sequences) and a spacer segment of polyethylene glycol to extend the coding segment away from the substrate. The detection probes were made by in situ synthesis using PGR (photogenerated reagent) chemistry. The hybridization melting temperatures were balanced by chemical modifications of the detection probes. Hybridization used 100 uL 6xSSPE buffer (0.90 M NaCl, 60 mM Na2HPO4, 6 mM EDTA, pH 6.8) containing 25% formamide at 34 °C.
Scan protocol Fluorescence images were collected using a laser scanner (GenePix 4000B, Molecular Device) and digitized using Array-Pro image analysis software (Media Cybernetics).
Description N/A
Data processing Background is determined using a regression-based background mapping method. The regression is performed on 5% to 25% of the lowest intensity data points excluding blank spots. Raw data matrix is then subtracted by the background matrix.
Normalization is carried out using a LOWESS (Locally-weighted Regression) method on the background-subtracted data. The normalization is to remove system related variations, such as sample amount variations, different labeling dyes, and signal gain differences of scanners so that biological variations can be faithfully revealed [B. M. Bolstad, R. A. Irizarry, M. Astrandand T. P. Speed, (2003) "A comparison of normalization methods for high density oligonucleotide array data based on variance and bias", Bioinformatics, 19 (2), 185-193].
A transcript to be listed as detectable must meets at least two conditions: signal intensity higher than 3×(background standard deviation) and spot CV < 0.5. CV is calculated by (standard deviation)/(signal intensity). When repeating probes are present on an array, a transcript is listed as detectable only if the signals from at least 50% of the repeating probes are above detection level.
t-Test is performed between “control” and “test” sample groups. T-values are calculated for each miRNA, and p-values are computed from the theoretical t-distribution. miRNAs with p-values below a critical p-value (typically 0.01) are selected for cluster analysis. The clustering is done using hierarchical method and is performed with average linkage and Euclidean distance metric.
All data processes, except clustering plot, are carried out using in-house developed computer programs. The clustering plot is generated using TIGR MeV (Multiple Experimental Viewer) software from The Institute for Genomic Research.
 
Submission date Aug 06, 2010
Last update date Oct 01, 2010
Contact name micah Drummond
E-mail(s) mjdrummo@utmb.edu
Organization name University of Texas Medical Branch
Street address 301 University Boulevard
City Galveston
State/province TX
ZIP/Postal code 77555-1144
Country USA
 
Platform ID GPL10415
Series (1)
GSE23527 Aging and MicroRNA Expression in Human Skeletal Muscle

Data table header descriptions
ID_REF
VALUE LOWESS normalized signal
StDev

Data table
ID_REF VALUE StDev
hsa-let-7a 9743 720
hsa-let-7a* 13 17
hsa-let-7b 7879 455
hsa-let-7b* 91 69
hsa-let-7c 5613 741
hsa-let-7c* 8 13
hsa-let-7d 7192 499
hsa-let-7d* 254 59
hsa-let-7e 1252 80
hsa-let-7e* 12 11
hsa-let-7f 8659 661
hsa-let-7f-1* 78 15
hsa-let-7f-2* 13 16
hsa-let-7g 3926 332
hsa-let-7g* 14 21
hsa-let-7i 4256 466
hsa-let-7i* 10 16
hsa-miR-1 41845 1057
hsa-miR-100 433 104
hsa-miR-100* 6 8

Total number of rows: 924

Table truncated, full table size 16 Kbytes.




Supplementary file Size Download File type/resource
GSM575587_O7_B_435_Data.txt.gz 86.2 Kb (ftp)(http) TXT
Processed data included within Sample table

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