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Sample GSM603580 Query DataSets for GSM603580
Status Public on Mar 09, 2011
Title muscle_48h_mouse32
Sample type RNA
 
Channel 1
Source name muscle, 48 hours fasting, mouse 32
Organism Mus musculus
Characteristics strain: FVB
mouse: 32
tissue: muscle
timepoint: 48
Extracted molecule total RNA
Extraction protocol Total RNA was extracted from each of the frozen organs of eight animals per experimental group with TRIzol reagent (Invitrogen, Breda, The Netherlands), followed by a repeated phenol-chloroform extraction, LiCl precipitation, and additional purification using the RNeasy Mini Kit (Qiagen Benelux, Venlo, The Netherlands). The RNA quality was assessed using the RNA 6000 Nano LabChip® Kit in an Agilent 2100 bioanalyzer (Agilent Technologies, Palo Alto, USA). All samples had intact bands corresponding to 18S and 28S ribosomal RNA subunits, displayed no chromosomal peaks or RNA degradation products, and had an RNA integrity number (RIN) > 8. Five out of eight animals per group were chosen based on the best RNA quality across all 5 tissues, so that the RNAs used in this study all originated from the same 5 mice per time point.
Label Cy3
Label protocol 1,5 µg of total RNA was amplified using the Amino Allyl MessageAmp aRNA kit (Ambion, Austin, Texas, USA), and labeled with Cy5 (experimental samples) and Cy3 (common reference) reactive dye according to the manufacturer’s instructions.
 
Channel 2
Source name reference pool
Organism Mus musculus
Characteristics mouse: reference pool
Extracted molecule total RNA
Extraction protocol Total RNA was extracted from each of the frozen organs of eight animals per experimental group with TRIzol reagent (Invitrogen, Breda, The Netherlands), followed by a repeated phenol-chloroform extraction, LiCl precipitation, and additional purification using the RNeasy Mini Kit (Qiagen Benelux, Venlo, The Netherlands). The RNA quality was assessed using the RNA 6000 Nano LabChip® Kit in an Agilent 2100 bioanalyzer (Agilent Technologies, Palo Alto, USA). All samples had intact bands corresponding to 18S and 28S ribosomal RNA subunits, displayed no chromosomal peaks or RNA degradation products, and had an RNA integrity number (RIN) > 8. Five out of eight animals per group were chosen based on the best RNA quality across all 5 tissues, so that the RNAs used in this study all originated from the same 5 mice per time point.
Label Cy5
Label protocol 1,5 µg of total RNA was amplified using the Amino Allyl MessageAmp aRNA kit (Ambion, Austin, Texas, USA), and labeled with Cy5 (experimental samples) and Cy3 (common reference) reactive dye according to the manufacturer’s instructions.
 
 
Hybridization protocol The microarrays were hybridized overnight with 200 µl hybridization mixture, consisting of 50 µl Cy3-and Cy5-labeled aRNA , 100 µl Formamide and 50 µl 4 x RPK0325 MicroArray Hybridization Buffer (Amersham Pharmacia Biosciences, Piscataway, NJ, USA) at 37°C and washed in an Automated Slide Processor (Amersham Pharmacia Biosciences, Piscataway, NJ, USA)
Scan protocol Scanned on an Agilent G2565AA scanner.
Spot intensities were quantified as artifact removed densities, using Array Vision software (version 6.0)
Description Common-reference sample was a pool of equal amounts of RNA from all the samples investigated, including additional samples from 5 mice that were fasted for 48 hours supplemented with vitamin B complex after 24 and 36 hours of fasting.
Data processing Estimated foreground and background signals were extracted from the ArrayVision files. Quality control was performed on the raw data; two arrays did not pass our quality criteria because of spatial artifacts and one array (from liver and clustering with brain) was excluded from further analysis. Background correction was performed by using the normexp method (Bioconductor package limma) to adjust the foreground signal without introducing negative values. Resulting log-ratios were normalized per array by using print-tip loess. Control probes were downweighted to zero in the normalization step. The oligo library was re-annotated by aligning each oligo to the Ensembl assembly using a BLAT/BLAST search. If no hit was found, oligos were blasted to a set of sequences containing Refseq RNAs (version 22) and UniGene clusters (version 163) that could not be aligned with the genome. Using information from NCBI Refseq and Unigene identifierss were linked to an Entrez gene ID. These Entrez gene IDs were used as input for the Bioconductor package AnnBuilder, with which a annotation data package was created.
 
Submission date Oct 04, 2010
Last update date Mar 09, 2011
Contact name Perry D Moerland
E-mail(s) p.d.moerland@amsterdamumc.nl
Organization name Amsterdam UMC
Department Epidemiology and Data Science
Lab Bioinformatics Laboratory
Street address Meibergdreef 9
City Amsterdam
ZIP/Postal code 1105AZ
Country Netherlands
 
Platform ID GPL11012
Series (1)
GSE24504 Interorgan coordination of the murine adaptive response to fasting

Data table header descriptions
ID_REF
VALUE normalized log2 ratio Cy3/Cy5 (test/reference)

Data table
ID_REF VALUE
1 3.889242377
2 0.006581948
3 0.011304075
4 0.015026137
5 0.005370912
6 0.034495902
7 0.000116903
8 -0.008968093
9 0.00464647
10 0.025453327
11 0.00954011
12 0.020193637
13 0.014291644
14 -0.012379291
15 -0.000150111
16 -0.000824867
17 0.198638645
18 0.112877036
19 0.026656719
20 -0.586412898

Total number of rows: 22680

Table truncated, full table size 395 Kbytes.




Supplementary file Size Download File type/resource
GSM603580.txt.gz 792.3 Kb (ftp)(http) TXT
Raw data provided as supplementary file
Processed data included within Sample table

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