|
| Status |
Public on May 10, 2011 |
| Title |
heart_hibernating_N05-035 |
| Sample type |
RNA |
| |
|
| Source name |
heart,hibernating
|
| Organism |
Ursus americanus |
| Characteristics |
tissue type: heart
gender: male
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from frozen tissues stored at -80 C by grinding in liquid nitrogen with mortar and pestle and using RNeasy Kit (Qiagen). Skeletal muscle tissue was treated by proteinase K and RNA was extracted by using RNeasy Fibrous Tissue Kit (Qiagen). All RNA samples were processed by DNase I (Qiagen) treatment. For cDNA library construction, mRNA was selected from total RNA with the oligo(dT) cellulose by the use of Poly(A) Purist Kit (Ambion).
|
| Label |
33P
|
| Label protocol |
Samples of total RNA were linearly amplified with Illumina TotalPrep RNA Amplification Kit (Ambion), and 1.6 μg of the amplified RNA was labeled with 65 μCi of [33P]dCTP. All RNA samples were amplified, labeled and hybridized in the same batch.
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| |
|
| Hybridization protocol |
The hybridization was carried out for 18 hours at 42°C in 4 ml of MicroHyb buffer (Invitrogen). Filters were rinsed at room temperature with 2× SSC/1% SDS to remove residual probe and MicroHyb solution and then transferred to preheated wash solutions in a temperature-controlled shaking water bath. Filters were washed twice for 30 min in 1.5 l of 2× SSC/1% SDS at 50°C and then once for 30 min in 1.5 l of 0.5× SSC/1% SDS at 55°C.
|
| Scan protocol |
Filters were then exposed to phosphorimager screens for four days and scanned at 50-µm resolution in a Storm Phosphorimager.
|
| Description |
heart tissue from hibernating black bear
|
| Data processing |
Image analysis was performed with the ImaGene program (Biodiscovery). Only nonempty spots with signal median density exceeding double background median in all individuals were included in the analysis. Background corrected signal was obtained by subtracting local background median density from signal median density. Background corrected signals were divided by their median on the array to obtain the normalized median densities representing the normalized expression values.
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| |
|
| Submission date |
Mar 10, 2011 |
| Last update date |
May 10, 2011 |
| Contact name |
Haifang Wang |
| Organization name |
Shanghai institute for biology science
|
| Department |
CAS-MPG Partner institute for Computational Biology
|
| Street address |
320 yueyang road
|
| City |
Shanghai |
| ZIP/Postal code |
200031 |
| Country |
China |
| |
|
| Platform ID |
GPL8249 |
| Series (1) |
| GSE27875 |
Modulation of gene expression in heart and liver of hibernating black bears (Ursus americanus) |
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