|
| Status |
Public on Dec 10, 2009 |
| Title |
3 NCEP 011408 2-20-08 |
| Sample type |
RNA |
| |
|
| Source name |
colon
|
| Organism |
Homo sapiens |
| Characteristics |
gender: male
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Using a random hexamer incorporating a T7 promoter, double-stranded cDNA was synthesized from 500 ng total RNA from which the majority of the ribosomal RNA had been removed using a RiboMinus Human/Mouse Transcriptome Isolation Kit (Invitrogen, Carlsbad, CA). cRNA was generated from the double-stranded cDNA template though an in-vitro transcription reaction and purified using the Affymetrix sample cleanup module. cDNA was regenerated through a random-primed reverse transcription using a dNTP mix containing dUTP. The RNA was hydrolyzed with RNase H and the cDNA purified. The cDNA was then fragmented by incubation with a mixture of UDG and APE 1 restriction endonucleases; and end-labeled via a terminal transferase reaction incorporating a biotinylated dideoxynucleotide. 5.5 ug of the fragmented, biotinylated cDNA was added to a hybridization cocktail,
|
| Label |
biotin
|
| Label protocol |
The cDNA was then fragmented by incubation with a mixture of UDG and APE 1 restriction endonucleases; and end-labeled via a terminal transferase reaction incorporating a biotinylated dideoxynucleotide. 5.5 ug of the fragmented, biotinylated cDNA was added to a hybridization cocktail,
|
| |
|
| Hybridization protocol |
The hybridization cocktail was loaded on the Human Exon 1.0 ST GeneChip and hybridized for 16 hours at 45oC and 60 rpm
|
| Scan protocol |
The stained array was scanned at 532 nm using an Affymetrix GeneChip Scanner 3000, generating CEL files for each array.
|
| Description |
Comparion data--colon tumor and normal were available at (http://www.affymetrix.com/support/technical/sample_data/exon_array_data.affx).
|
| Data processing |
PARTEK background correction, quantile normalization and probe summarization was performed using RMA
|
| |
|
| Submission date |
Nov 24, 2009 |
| Last update date |
Nov 24, 2009 |
| Contact name |
Lesleyann Hawthorn |
| E-mail(s) |
[email protected]
|
| Phone |
7067214384
|
| Fax |
7067211660
|
| Organization name |
Medical College of Georgia Cancer Center
|
| Department |
Molecular Oncology
|
| Street address |
1120 15th St
|
| City |
Augusta |
| State/province |
GA |
| ZIP/Postal code |
30912 |
| Country |
USA |
| |
|
| Platform ID |
GPL5175 |
| Series (1) |
|
