|
| Status |
Public on Nov 01, 2018 |
| Title |
PBEC_Dex-and-TNFa_ANR273_rep3 |
| Sample type |
RNA |
| |
|
| Source name |
NuLi-1 human airway epithelial cell line, Dex+TNFa treated
|
| Organism |
Homo sapiens |
| Characteristics |
glucocorticoid treatment: Dex
cytokine treatment: TNFa
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted using the TRIzol method
|
| Label |
Oyster-550
|
| Label protocol |
The total RNA was DNAse digested and low-molecular weight (LMW) RNA was isolated by ultrafiltration through YM-100 columns (Millipore) and subsequent purification using the RNeasy MinElute Clean-Up Kit (Qiagen). The LMW RNA samples were 3’-end labeled with Oyster-550 fluorescent dye using the Flash Tag RNA labeling Kit (Genisphere).
|
| |
|
| Hybridization protocol |
Labeled LMW RNA samples were hybridized to the MicroRNA microarrays according to conditions recommended in the Flash Tag RNA labeling Kit manual.
|
| Scan protocol |
The microarrays were scanned on an Axon Genepix 4000B scanner, and data was extracted from images using GenePix V4.1 software.
|
| Description |
Array D, blocks 25-32 of raw data file
2013-03-1_19717741_650PMT_5um_0532.txt
|
| Data processing |
Data for 4 arrays were extracted from a single microarray image into a single text file using GenePix version 4.1 software. The raw text files were split in to four subfiles based on block assignment, namely blocks 1-8 contained data for array A, blocks 9-16 contained data for array B, blocks 17-24 contained data for array C, and blocks 25-32 contained data for array D. Spot intensities were obtained for the 10336 features on each microarray by subtracting the median local background from the median local foreground for each spot. Detection Thresholds for each array were determined by calculating the 10% trim mean intensity of the negative controls spots and adding 5X the standard deviation of the background (non-spot area). The spot intensities and the Threshold (T) were transformed by taking the log (base 2) of each value. The mean probe intensities for each of the 2017 human probes on each of the 12 arrays were then determined by averaging the triplicate spot intensities. Spots flagged as poor quality during data extraction were omitted prior to averaging and spots that have an average saturated signal were removed prior to normalization, which yielded 1995 human non-saturated probes intensities. The normalization factor (N) for each microarray was determined by obtaining the 20% trim mean of the mouse probes intensities above Threshold in > 90% of samples. The log2-transformed spot intensities for all 10,336 features were normalized, by subtracting N from each spot intensity, and scaled by adding the grand mean of N across all microarrays. The 1995 human non-control non-saturated log2-transformed, normalized, and averaged probe intensities were filtered to obtain a list of 606 human microRNA probes showing probe intensity above Threshold in at least 10% of samples. The data was untransformed using the formula 2^N.
|
| |
|
| Submission date |
Oct 02, 2018 |
| Last update date |
Nov 01, 2018 |
| Contact name |
Faoud T Ishmael |
| E-mail(s) |
Faoud.Ishmael@mountnittany.org
|
| Phone |
717-531-0003
|
| Organization name |
Penn State College of Medicine
|
| Department |
Division of Pulmonary and Critical Care Medicine
|
| Lab |
Section of Allergy and Immunology
|
| Street address |
500 University Dr.
|
| City |
Hershey |
| State/province |
PA |
| ZIP/Postal code |
17033 |
| Country |
USA |
| |
|
| Platform ID |
GPL25635 |
| Series (1) |
| GSE120758 |
MicroRNA-146a and glucocorticoids in inflamed airway epithelial cells |
|
