|
| Status |
Public on Sep 02, 2009 |
| Title |
Breast cell line BT549 |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
genomic DNA isolated from breast cell line
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: BT549
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
blocks were trimmed with a razor blade to remove normal tissue and cryosections were obtained from either side of the block to ascertain that tumor cells comprised greater than 70% of the specimen. DNA was extracted using the QUIamp tissue kits (29304, Qiagen)
|
| Label |
Cy3
|
| Label protocol |
The tumor genomic DNA and normal male reference DNA (300 ng each) were labeled by random priming in separate 50 ul reactions to incorporate Cy3 and Cy5, respectively.
|
| |
|
| Channel 2 |
| Source name |
normal male genomic DNA
|
| Organism |
Homo sapiens |
| Characteristics |
gender: male
tissue: blood
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
blocks were trimmed with a razor blade to remove normal tissue and cryosections were obtained from either side of the block to ascertain that tumor cells comprised greater than 70% of the specimen. DNA was extracted using the QUIamp tissue kits (29304, Qiagen)
|
| Label |
Cy5
|
| Label protocol |
The tumor genomic DNA and normal male reference DNA (300 ng each) were labeled by random priming in separate 50 ul reactions to incorporate Cy3 and Cy5, respectively.
|
| |
|
| |
| Hybridization protocol |
The test and reference DNAs together with 100 ug human Cot-1 DNA were hybridized to the BAC arrays for ~48 hrs at 37ºC. After post-hybridization washes, the arrays were mounted in a solution containing 90% glycerol, 10% PBS and 1 uM DAPI and sealed with a cover slip.
|
| Scan protocol |
A custom built CCD camera system was used to acquire 16 bit 1024x1024 pixel DAPI, Cy3 and Cy5 images (Pinkel et al., 1998 and Hamilton et al. 2006). Image analysis was carried out using UCSF SPOT software (Jain et al., 2002).
|
| Description |
n/a
|
| Data processing |
The log2ratio of the total integrated Cy3 and Cy5 intensities for each spot after background subtraction was calculated, normalized to the median log2ratio of all the clones on the array and the average of the triplicates calculated using a second custom program, SPROC. Automatic data filtering to reject data points based on low DAPI intensity, low correlation between Cy3 and Cy5 within each segmented spot and low reference/DAPI signal intensity was also carried out using SPROC. Data files were subsequently manually edited by rejecting clones for which only one spot of the triplicate survived after SPROC analysis and for which the standard deviation of the log2ratio of the triplicate was >0.2. For each tumor, the data are plotted as the mean log2ratio of the triplicate spots for each clone normalized to the genome median log2ratio. The clones are ordered by position in the genome (UCSC draft genome sequence, July 2003 freeze) beginning at 1p and ending with Xq.
|
| |
|
| Submission date |
Sep 12, 2008 |
| Last update date |
Sep 02, 2009 |
| Contact name |
Serena S kwek |
| E-mail(s) |
[email protected]
|
| Phone |
415-514-9301
|
| Organization name |
UCSF
|
| Department |
Cancer Research Institute
|
| Lab |
Donna Albertson
|
| Street address |
2340 Sutter St.
|
| City |
San Francsico |
| State/province |
CA |
| ZIP/Postal code |
94143-0808 |
| Country |
USA |
| |
|
| Platform ID |
GPL7298 |
| Series (1) |
| GSE12761 |
Co-amplification at 8p and 11q in breast tumors and cell lines with 8p11q array Genomic Comparative Hybridization |
|
