|
| Status |
Public on Nov 22, 2007 |
| Title |
NHBE_Chip 02 rep1_rep2_rep3 |
| Sample type |
RNA |
| |
|
| Source name |
normal human bronchial/tracheal epithelial cells
|
| Organism |
Homo sapiens |
| Characteristics |
normal human bronchial/tracheal epithelial cells
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using Gentra Systems Versagene Total RNA Cell Kit (included their DNAse Kit) was carried out according to manufacture's instructions
|
| Label |
biotin
|
| Label protocol |
GeneChip® Whole Transcript (WT) Double-Stranded DNA Terminal Labeling Kit and GeneChip® WT Amplified Double-Stranded cDNA Synthesis Kit from Affymetrix (Santa Clara, CA). This protocol entails first and second strand cDNA synthesis using random hexamers, RNA removal and cDNA purification, a quality control cDNA step, a cDNA fragmentation step, TdT labeling, prehybridization of the chips, and the final hybridization of the labeled cDNA onto the GeneChip® Human 35 bp Tiling Array 1.0R Set. Detailed procedure is found in the GeneChip® WT Double-Stranded Target Assay Manual.
|
| |
|
| Hybridization protocol |
Used GeneChip® Hybridization, Wash, and Stain Kit. Followed the procedure from the GeneChip® WT Double-Stranded Target Assay Manual.
|
| Scan protocol |
Each array was scanned using the Affymetrix GeneChip® 300 G7 scanner.
|
| Description |
Ribominus untreated: rep1, rep2
Ribominus treated: rep3
|
| Data processing |
Data was analyzed using Tiling Analysis Software (TAS) and Integrated Genome Browser (IGB). Tiling Analysis Software v1.1 User Guide (Affymetrix) provides detailed description of its analysis capabilities including various uses of quality control measures. Briefly, the analyses provided within the TAS application included analyzing feature intensity data stored in CEL files to produce signal and p-values for each genomic position, computation of genomic intervals based on those computed signal and p-values, and computation of summary statistics and visualizations for assessing the quality of the array data. The results of this analysis were imported into applications such as IGB or the UCSC Human Genome Browser. With IGB, annotation variations were compared in different data sets as described in the User's Guide. Specifically, our data was normalized using quantile normalization plus scaling (scaled to the default target intensity of 500 for each ship). Array types were verified against library files. Replicates were not normalized together. BAR files were created in TAS and opened in IGB for signal visualization. Once a threshold of 99.5th percentile was selected, a "track" was made (which mark regions over the threshold level). Annotations from these "tracks" were saved into a bed file that can be opened in Excel for further analysis.
|
| |
|
| Submission date |
Nov 21, 2007 |
| Last update date |
Aug 14, 2011 |
| Contact name |
Damon Scott Perez |
| E-mail(s) |
perez.damon@mayo.edu
|
| Phone |
507-266-0311
|
| Organization name |
Mayo Clinic
|
| Department |
Experimental Pathology & Lab Medicine
|
| Lab |
David I Smith; Stabile 2-50
|
| Street address |
200 Fist Street SW
|
| City |
Rochester |
| State/province |
MN |
| ZIP/Postal code |
55905 |
| Country |
USA |
| |
|
| Platform ID |
GPL6157 |
| Series (1) |
| GSE9655 |
Whole-genome tiling array analysis in normal human bronchial/tracheal epithelial cells (NHBE) |
|
