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Sample GSM969196 Query DataSets for GSM969196
Status Public on Feb 27, 2013
Title Subject_AF0286_SE_T5
Sample type RNA
 
Source name Blood_19.5hrsAwake_at02:40afterSleepExtension_Per3_4\4_genotype
Organism Homo sapiens
Characteristics subject: AF0286
tissue: Blood
sleepprotocol: Sleep Extension
genotype/variation: 4\4
hoursawake: 19.5
timesampletaken: 2:40
circadianphase: 0
Treatment protocol 2.5 ml of blood is drawn into the PAXGene tube at each sampling point. The tube is inverted 10 times to mix. The tube is then stored at room temperature for 4 hours in upright position. The tube is transferred to -50°C and placed lying down (never in upright position as this may cause the tube to crack) for storage at the university of Surrey Clinical Research Centre. The tube is transferred on ice to University of Surrey Faculty of Health and Medical Sciences for storage/processing
Growth protocol Blood samples were collected via an indwelling venous cannula sited in the forearm for minimum discomfort and restriction of movement. Cannulae were kept patent by the use of sterile saline. Each sample tube was coded for subject and sample collection time (participant code/sample type/sequential number)
Extracted molecule total RNA
Extraction protocol Total RNA was isolated using a PAXgene Blood RNA Kit and a QiaCube robot (Qiagen, Hilden, Germany). The RNA was quantified and the A260/280 nm and A260/230 nm ratios were determined using a NanoDrop ND1000 spectrophotometer (Wilmington, DE). RNA quality was assessed using the Bioanalyzer 2100 (Agilent, Santa Clara, CA).
Label Cy3
Label protocol cRNA was synthesized and fluorescently labelled with Cy3-CTP from 100 ng of total RNA using Agilent's Low Input Quick Amp Labeling Kit.
 
Hybridization protocol Labelled cRNA (1.65 μg) was hybridized on a Whole Human Genome 4 x 44K custom oligonucleotide microarray (G2514F, AMADID 026817; Agilent Technologies). Standard manufacturer's instructions for one-colour gene expression hybridization and washing steps were followed. The microarrays were hybridized at 65°C for 17 h in an Agilent hybridization oven with rotisserie at 10 rpm. The microarrays were washed with Agilent Wash Buffer 1, pre-warmed Wash Buffer 2 (37°C) and acetonitrile according to the manufacturer’s instructions. The last washing step was performed with Agilent Stabilization and Drying Solution for 30 sec.
Scan protocol The processed microarrays were scanned using an Agilent Microarray Scanner with a resolution of 5 μm, exploiting the extended dynamic range feature. The two images derived from each slide scanned at 10% and 100% PMT were imported into Agilent Feature Extraction software (Version 10.7.1.1) for image analysis.
Description Blood_19.5hrsAwake_at02:40afterSleepExtension_Per3_4\4genotype_Subject_AF0286
Blood taken at 02:40 following 19.5 hours awake after following a Sleep Extension protocol from a Per3 4\4 genotype subject (Subject AF0286)
Data processing Individual samples were filtered based on AgilentQC metrics of reproducibility statistics, minimum detection level estimates and feature flags. Samples with a median coefficient of variation of less than 10% in spike ins or non-control replicated probes (NCRPs) were retained and quantile-normalized using the R Bioconductor package limma. NCRPs along with their corresponding flags were averaged. Probes with more than 5 flagged samples within a subject in more than half of the total number of subjects were excluded. In addition, for time-series analyses, series with 2 consecutive or more than 2 missing time points were excluded.
 
Submission date Jul 17, 2012
Last update date Oct 15, 2014
Contact name Emma Laing
E-mail(s) e.laing@surrey.ac.uk
Organization name University of Surrey
Street address Stag Hill
City Guildford
ZIP/Postal code GU22 7XH
Country United Kingdom
 
Platform ID GPL15331
Series (1)
GSE39445 Effect of sleep restriction on the human transcriptome during extended wakefulness

Data table header descriptions
ID_REF
VALUE Quantile normalized signal intensity

Data table
ID_REF VALUE
1 17.37697094
2 2.02833535
3 2.030396845
4 2.034082287
5 2.039446846
6 2.044097268
7 2.051284593
8 2.055311924
9 2.063348231
10 2.069685401
11 2.075843918
12 4.368287436
13 8.659190785
14 2.099624545
15 5.854807808
16 3.749752954
17 9.530260368
18 4.096512566
19 7.188697791
20 8.123005749

Total number of rows: 43758

Table truncated, full table size 754 Kbytes.




Supplementary file Size Download File type/resource
GSM969196_AF0286_R_15.txt.gz 8.6 Mb (ftp)(http) TXT
Processed data included within Sample table

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