|
| Status |
Public on Jan 01, 2015 |
| Title |
P00322 Non-failing Left Ventricle |
| Sample type |
RNA |
| |
|
| Source name |
heart left ventricle, non-failing
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: heart left ventricle
heart failure: no
disease status: non-failing
gender: male
age: 75
|
| Treatment protocol |
Explanted myocardium snap frozen in liquid nitrogen.
|
| Growth protocol |
None
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol extraction of total RNA was performed according to the manufacturer's instructions.
|
| Label |
biotin
|
| Label protocol |
100ng of total RNA was converted to first-strand cDNA using reverse transcriptase primed by a poly(T) oligomer that incorporated the T7 RNA polymerase promoter. Second-strand cDNA synthesis was followed by in vitro transcription (Affymetrix One-Cycle Target Labeling Kit) for linear amplification and biotinylation of each transcript.
|
| |
|
| Hybridization protocol |
cDNAs were added to Affymetrix hybridization cocktails, heated at 99ºC for 2 min and hybridized for 16 h at 45ºC to Affymetrix Human Gene 1.1 ST Array GeneChips (Affymetrix Inc., Santa Clara, CA). The microarrays were then washed at low (6X SSPE) and high (100mM MES, 0.1M NaCl) stringency and stained with streptavidin-phycoerythrin. Fluorescence was amplified by adding biotinylated anti-streptavidin and an additional aliquot of streptavidin-phycoerythrin stain.
|
| Scan protocol |
A confocal scanner was used to collect fluorescence signal after excitation at 570 nm.
|
| Description |
P00322
PolyA mRNA
|
| Data processing |
Microarray data were analyzed using the Oligo package available at the Bioconductor website (www.bioconductor.org). The raw data were first background-corrected by the Robust Multichip Average (RMA) method and then normalized by an invariant set method. Microarray Processing batch were controlled for using the R package Combat. Differential gene expression was analyzed by the Limma package available at the Bioconductor website. P-values obtained from the multiple comparison tests were corrected by false discovery rates.
|
| |
|
| Submission date |
May 06, 2014 |
| Last update date |
Jan 01, 2015 |
| Contact name |
Michael Patrick Morley |
| E-mail(s) |
mmorley@pennmedicine.upenn.edu
|
| Phone |
215-898-2026
|
| Organization name |
Perelman School of Medicine at the University of Pennsylvania
|
| Department |
Penn Cardiovascular Institute
|
| Street address |
3400 Civic Center Blvd, Bldg 421
|
| City |
Philadelphia |
| State/province |
PA |
| ZIP/Postal code |
19104 |
| Country |
USA |
| |
|
| Platform ID |
GPL11532 |
| Series (2) |
| GSE57338 |
RNA-Seq Identifies Novel Myocardial Gene Expression Signatures of Heart Failure [microarray] |
| GSE57345 |
RNA-Seq Identifies Novel Myocardial Gene Expression Signatures of Heart Failure |
|
