|
| Status |
Public on Dec 25, 2010 |
| Title |
HMEC/GHRD-5 |
| Sample type |
RNA |
| |
|
| Source name |
HMEC/GHRD
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Primary Human Mammalian Epithelial Cells (HMECs)
|
| Biomaterial provider |
ScienCell Research Laboratories (Carlsbad, CA)
|
| Treatment protocol |
Treatment consisted of HMEC cells being stimulated with HMEC basal medium containing 15% GHRD serum for 6 hours then cells were harvested and processed for RNA extraction.
|
| Growth protocol |
Cells were cultured in HMEC medium (ScienCell) at 37oC and 5% CO2 in Poly-L-Lysine coated culture dishes (Sigma).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted from the serum treated cell cultures using TRI Reagent® (Ambion). Quality and quantity of the total RNA was checked with the Agilent 2100 bioanalyzer using RNA 6000 Nano chips.
|
| Label |
Streptavidin-Cy3 bound to biotin-labeled cRNA.
|
| Label protocol |
standard Illumina protocol using Illumina TotalPrep RNA Amplification Kit (Ambion; Austin, TX, cat # IL1791) In short, 0.5g of total RNA was first converted into single-stranded cDNA with reverse transcriptase using an oligo-dT primer containing the T7 RNA polymerase promoter site and then copied to produce double-stranded cDNA molecules. The double stranded cDNA was cleaned and concentrated with the supplied columns and used in an overnight in-vitro transcription reaction where single-stranded RNA (cRNA) was generated and labeled by incorporation of biotin-16-UTP.
|
| |
|
| Hybridization protocol |
Standard Illumina protocol. In short, a total of 0.75ug of biotin-labeled cRNA was hybridized at 58 degrees C for 16 hours to Illumina's Sentrix Human HT-12, v3 Expression BeadChips (Illumina, San Diego, CA). Each BeadChip targets more than 25,000 annotated genes with more than 48,000 probes. Probes were designed using the RefSeq (Build 36.2, Rel 22) and the UniGene (Build 199) databases with an average of 15-fold redundancy. The arrays were washed, blocked and the biotin labeled cRNA detected by staining with streptavidin-Cy3.
|
| Scan protocol |
Arrays were scanned at a resolution of 0.8um using the Beadstation 500 X from Illumina.
|
| Description |
Primary Human Mammalian Epithelial Cells (HMECs) were cultured in HMEC medium in the prescence of 15% GHRD serum for 6 hours
|
| Data processing |
Data was extracted using the Illumina BeadStudio software(v3.0). Any spots at or below the background were filtered out using an Illumina detection p-value of 0.02 and above. The natural log of all remaining scores was used to find the avg and std of each array and the z-score normalization was calculated and presented below. Z-score = (raw value - avg)/std. Complete data including detection scores will be included in the supplemental file.
|
| |
|
| Submission date |
May 24, 2010 |
| Last update date |
Jun 22, 2020 |
| Contact name |
Supriyo De |
| Organization name |
NIA-IRP, NIH
|
| Department |
Laboratory of Genetics and Genomics
|
| Lab |
Computational Biology & Genomics Core
|
| Street address |
251 Bayview Blvd
|
| City |
Baltimore |
| State/province |
Maryland |
| ZIP/Postal code |
21224 |
| Country |
USA |
| |
|
| Platform ID |
GPL6947 |
| Series (1) |
| GSE21980 |
Growth hormone receptor deficiency is associated with a major reduction in pro-aging signaling, cancer, and diabetes in humans |
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