|
| Status |
Public on Mar 01, 2017 |
| Title |
genomic DNA of mouse brain, GZH021 |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
Mouse brain
|
| Organism |
Mus musculus |
| Characteristics |
background strain: FVB/C57BL/6
gender: F
genotype/varation: Pten;p53 double knockout
tissue: Medulloblastoma
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Pulverized tissue was lysed in 200mM NaCl, 5mM EDTA, 0.2% SDS, 100mM Tris pH8.5, 100ug/ml proteinase K overnight at 55C, then extracted with phenol/chloroform.
|
| Label |
cy5
|
| Label protocol |
Genomic DNA Labeling Kit (Enzo Life Sciences, Farmingdale, NY) was used as per manufacturer's protocol.
|
| |
|
| Channel 2 |
| Source name |
normal genomic DNA with opposite sex to ch1
|
| Organism |
Mus musculus |
| Characteristics |
tissue: normal brain
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Pulverized tissue was lysed in 200mM NaCl, 5mM EDTA, 0.2% SDS, 100mM Tris pH8.5, 100ug/ml proteinase K overnight at 55C, then extracted with phenol/chloroform.
|
| Label |
cy3
|
| Label protocol |
Genomic DNA Labeling Kit (Enzo Life Sciences, Farmingdale, NY) was used as per manufacturer's protocol.
|
| |
|
| |
| Hybridization protocol |
Prior to hybridization, the test and reference probes were combined with 100 ug mouse fluorimetric Cot-1 (Invitrogen) and ethanol precipitated. Following centrifugation, the probes were resuspended in 110ul SlideHyb Buffer #3 (Ambion) containing 5ul of 100µg/ul Yeast tRNA (Invitrogen), heated to 95ºC for 5 minutes and incubated for 30 minutes at 37ºC to block repetitive elements on the probe. Hybridization was performed for 16 hours at 55ºC using a GeneMachine hybridization station (Genomic Solutions, Inc.) as described (Cowell, J. K. and N. J. Nowak. 2003. High-resolution analysis of genetic events in cancer cells using bacterial artificial chromosome arrays and comparative genome hybridization. Adv Cancer Res 90:91-125). After hybridization, the slides were automatically washed in the GeneMachines station with reducing concentrations of SSC and SDS. The final wash was 30 seconds in 0.1X SSC, followed by a two second 100 % ethanol dip. The slides were spun dry and scanned immediately at 5µm resolution using a GenePix 4000B laser scanner (Axon Inc.)
|
| Scan protocol |
Slides were scanned on a GenePix 4000B laser Scanner (Axon Inc.) using GenePix Pro V6.0/1. Both lasers were set to 100% and the PMT was set to 500 for cy3 and 450 for cy5
|
| Description |
tissue from mouse brain with Medulloblastoma
|
| Data processing |
Images were analyzed using Imagene 6.0 (BioDiscovery Inc.). The output of Image analysis was further processed by in-house developed programs. The log2 ratios of the test/control (values are not background subtracted) were normalized using a subgrid loess, with all flagged spots and clones on the sex chromosome are given a weight of zero. The mean of the replicate measures was taken and the final normalized log2 ratio was plotted against its genomic location.
|
| |
|
| Submission date |
Aug 14, 2012 |
| Last update date |
Mar 01, 2017 |
| Contact name |
David Finkelstein |
| E-mail(s) |
david.finkelstein@stjude.org
|
| Phone |
9014953931
|
| Organization name |
St Jude Children's Research Hospital
|
| Department |
Computational Biology
|
| Street address |
332 N. Lauderdale St.
|
| City |
Memphis |
| State/province |
TN |
| ZIP/Postal code |
38105 |
| Country |
USA |
| |
|
| Platform ID |
GPL4714 |
| Series (1) |
| GSE40106 |
Pten deletion in neonatal brain induces an abnormal neural progenitor niche that can synergize with Trp53 loss to generate medulloblastoma |
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