|
| Status |
Public on Oct 25, 2012 |
| Title |
T223 |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
Peripheral blood
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: peripheral blood
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Normal blood DNA was extracted with the QIAGEN Flexigene Kit (Cat No.: 51206).
|
| Label |
Cy3
|
| Label protocol |
500ng sample and normal genomic DNA were fragmented by restriction digestion with Alu I/Rsa I (65degrees C for 20mins) and then were labeled with Cy5 and Cy3, respectively, at 85C for 30 min. Labeled DNA were purified using Microcon 30kDa spin columns. Degree of labeling was determined by measuring the absorbance at A260nm(DNA), A550nm(Cy3), and A650nm(Cy5) using a NanoDrop.
|
| |
|
| Channel 2 |
| Source name |
Colorectal tumor tissue
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: colorectal tumor
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Tumor DNA was extracted from fresh frozen material using the QIAGEN Blood & Cell Culture DNA Maxi Kit (Cat. No.: 13362).
|
| Label |
Cy5
|
| Label protocol |
500ng sample and normal genomic DNA were fragmented by restriction digestion with Alu I/Rsa I (65degrees C for 20mins) and then were labeled with Cy5 and Cy3, respectively, at 85C for 30 min. Labeled DNA were purified using Microcon 30kDa spin columns. Degree of labeling was determined by measuring the absorbance at A260nm(DNA), A550nm(Cy3), and A650nm(Cy5) using a NanoDrop.
|
| |
|
| |
| Hybridization protocol |
Appropriate equal amounts of Cy5-labeled sample and Cy3-labeled reference DNA were mixed to a 39ul final volume. 71ul Hybridization Master Mix (1mg/ml Cot-1 DNA 5ul, Agilent 10X blocking agent 11ul, and Agilent 2X HiRPM Hybridization buffer 55ul) was added and incubated at 95C for 3 min followed by 37C for 30min. 100ul of the hybridization sample mixture was applied onto the microarray gasket well and was covered by a microarray slide. The slide was incubated in a 65C rotator rack for 24 hours. After hybridization, slide was washed sequentially.
|
| Scan protocol |
Scanned on Agilent Technologies Scanner G2505C US83603551. Images were quantified using Agilent Feature Extraction Software (v10.7.3.1).
|
| Data processing |
Agilent Feature Extraction Software (v10.7.3.1) was used for background subtraction and Lowess normalization.
|
| |
|
| Submission date |
Oct 24, 2012 |
| Last update date |
Oct 25, 2012 |
| Contact name |
Ian Wallace Brock |
| E-mail(s) |
i.w.brock@sheffield.ac.uk
|
| Phone |
01142713165
|
| Fax |
447950304013
|
| Organization name |
University of Sheffield Medical School
|
| Department |
Cancer Studies
|
| Lab |
Cox
|
| Street address |
Beachhill Road
|
| City |
Sheffield |
| State/province |
South Yorkshire |
| ZIP/Postal code |
S10 2RX |
| Country |
United Kingdom |
| |
|
| Platform ID |
GPL10123 |
| Series (1) |
| GSE41813 |
Colorectal tumor DNA vs. normal control DNA |
|
