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Sample GSM486555 Query DataSets for GSM486555
Status Public on Feb 01, 2010
Title F, 18de vs 1d, Replicate 5
Sample type RNA
 
Channel 1
Source name Turkey pectoralis major muscle, F, 1-day post-hatch chick
Organism Meleagris gallopavo
Characteristics tissue: pectoralis major muscle
line: F (selected from RBC2 line for increased 16wk body weight)
age: 1-day post-hatch chick
Extracted molecule total RNA
Extraction protocol Total RNA extracted using TRIReagent (Molecular Research Center, Cincinnati, OH) following manufacturer's instructions. RNA integrity and concentration was determined using an Agilent 2100 Bioanlyzer (Santa Clara, CA).
Label Cy5
Label protocol Total RNA for use in microarrays was amplified using the Amino Allyl MessageAmp™ II aRNA Amplification Kit (Ambion, Inc.) per manufacturer instructions. Briefly, 2 μg total RNA was incubated with T7 Oligo(dT) primer for 10 min at 70°C then reverse transcribed into cDNA at 42°C for 2 h. Second strand synthesis was performed at 16°C for 2 h. Double-stranded cDNA was then purified, and in vitro transcription was performed at 37°C for 4 h. Amplified RNA (aRNA) was purified and quantified using a Nanodrop ND-1000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA). For the dye coupling procedure, 10 μg aRNA was coupled to either Cy3 or Cy5 fluorescent dye (GE Healthcare, Piscataway, NJ) in the dark at room temperature for 1 hour until the reaction was quenched by the addition of 4M hydroxylamine. Dye-coupled aRNA was purified and quantified, then 5 μg was fragmented to 60-200 nucleotide segments using RNA Fragmentation Reagents (Ambion, Inc.) at 70°C for 15 min in preparation for microarray hybridization.
 
Channel 2
Source name Turkey pectoralis major muscle, F, 18-day embryo
Organism Meleagris gallopavo
Characteristics tissue: pectoralis major muscle
line: F (selected from RBC2 line for increased 16wk body weight)
age: 18-day embryo
Extracted molecule total RNA
Extraction protocol Total RNA extracted using TRIReagent (Molecular Research Center, Cincinnati, OH) following manufacturer's instructions. RNA integrity and concentration was determined using an Agilent 2100 Bioanlyzer (Santa Clara, CA).
Label Cy3
Label protocol Total RNA for use in microarrays was amplified using the Amino Allyl MessageAmp™ II aRNA Amplification Kit (Ambion, Inc.) per manufacturer instructions. Briefly, 2 μg total RNA was incubated with T7 Oligo(dT) primer for 10 min at 70°C then reverse transcribed into cDNA at 42°C for 2 h. Second strand synthesis was performed at 16°C for 2 h. Double-stranded cDNA was then purified, and in vitro transcription was performed at 37°C for 4 h. Amplified RNA (aRNA) was purified and quantified using a Nanodrop ND-1000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA). For the dye coupling procedure, 10 μg aRNA was coupled to either Cy3 or Cy5 fluorescent dye (GE Healthcare, Piscataway, NJ) in the dark at room temperature for 1 hour until the reaction was quenched by the addition of 4M hydroxylamine. Dye-coupled aRNA was purified and quantified, then 5 μg was fragmented to 60-200 nucleotide segments using RNA Fragmentation Reagents (Ambion, Inc.) at 70°C for 15 min in preparation for microarray hybridization.
 
 
Hybridization protocol Fragmented, Cy3-coupled aRNA was mixed with its Cy5-coupled partner, and the mixtures were brought to 110 μL with 68°C SlideHyb 1 (Ambion, Inc.) and incubated at 68°C for 5 min. These dye-coupled mixtures were then hybridized to oligonucleotide microarrays (described previously) for 18 h in a GeneTac Hybridization Station (Genomic Solutions, Ann Arbor, MI) using the following conditions: 54°C for 18 h, followed by a medium-stringency wash at 42°C (2X SSC, 0.1% SDS), a high-stringency wash (0.2X SSC, 0.1% SDS) at 22°C, and a wash with postwash buffer (0.2X SSC) at 22°C. Arrays were then rinsed in 2X SSC and Nanopure H2O, dried by centrifugation at 500 × g for 5 min.
Scan protocol Scanned with a GenePix 4000B (Molecular Devices, Sunnyvale, CA) scanner.
Image analysis was performed using GenePix Pro 6.0, and spot intensities were exported as GPR files.
Description F line 18d embryo versus 1d chick comparison, replicate 5 of 10.
Data processing Fluorescence intensity data was normalized for dye intensity bias using the LOESS procedure of the normalizeWithinArray function of the Bioconductor R software LIMMA (Smyth 2004).
 
Submission date Dec 16, 2009
Last update date Dec 19, 2009
Contact name Kelly R Sporer
E-mail(s) buckhamk@msu.edu
Phone 517-355-8474
Fax 517-432-0753
Organization name Michigan State University
Department Food Science and Human Nutrition
Lab Muscle Biology and Genetics Research
Street address 3365 Anthony Hall
City East Lansing
State/province MI
ZIP/Postal code 48824
Country USA
 
Platform ID GPL9788
Series (2)
GSE19531 Analysis of the turkey skeletal muscle transcriptome through development within a genetic line (Experiment 1)
GSE19585 Analysis of the turkey skeletal muscle transcriptome through development and between genetic lines

Data table header descriptions
ID_REF
Ch1_RAW Cy5 total intensity
Ch2_RAW Cy3 total intensity
VALUE LOESS-normalized Cy5 versus Cy3 log2 ratio
VALUE_A LOESS-normalized average log2 intensity

Data table
ID_REF Ch1_RAW Ch2_RAW VALUE VALUE_A
1 65 95 0.073356534 6.296111711
2 84 125 0.140824849 6.679050854
3 15324 36178 -0.908217524 14.52316517
4 1255 1115 0.843025677 10.20814982
5 14033 7140 1.50532581 13.2891221
6 6015 11835 -0.417688785 13.04256055
7 3084 5518 -0.159770979 12.0102584
8 179 203 0.573464978 7.574575847
9 1461 4359 -0.876875457 11.30126098
10 87 117 0.282782837 6.656654108
11 2079 3015 0.163831925 11.28980816
12 109 135 0.439038936 6.922499961
13 4948 1389 2.533240207 11.35623033
14 89 937 -2.673822112 8.173819334
15 29398 23848 0.604938093 14.69250552
16 111 136 0.456209356 6.940939354
17 14075 35444 -0.988654793 14.44705055
18 141 180 0.409840954 7.315702224
19 45475 8339 2.823982234 14.24922235
20 261 207 1.073333195 7.860696477

Total number of rows: 12288

Table truncated, full table size 455 Kbytes.




Supplementary file Size Download File type/resource
GSM486555.gpr.gz 1.0 Mb (ftp)(http) GPR
Processed data included within Sample table

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