|
| Status |
Public on Sep 01, 2007 |
| Title |
D1 NPF_cyclin D1_2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
NPF Control
|
| Organism |
Homo sapiens |
| Characteristics |
NPF Control
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated from NPFs, CAFs, and NPF cyclin D1 cells using total RNA isolation kit (Qiagen).
|
| Label |
Cy5
|
| Label protocol |
Total RNA was amplified through one round of linear amplification using the MessageAmp aRNA kit (Ambion, Austin, TX). Sample quality and quantification was assessed by agarose gel electrophoresis and absorbance at A260. Cy3 and Cy5 labeled cDNA probes were made from 4 µg of amplified RNA.
|
| |
|
| Channel 2 |
| Source name |
Human normal prostate fibroblasts overexpressing cyclin D1 NPF_cyclin D1_2
|
| Organism |
Homo sapiens |
| Characteristics |
Human normal prostate fibroblasts overexpressing cyclin D1 NPF_cyclin D1_2. prostate cancer status: N; Specimen ID: NPF_cyclin D1_2; origin: primary prostate stromal cells from benign peripheral prostate from patient with bladder cancer; Tissue: normal prostate fibroblasts overexpressing cyclin D1;
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was isolated from NPFs, CAFs, and NPF cyclin D1 cells using total RNA isolation kit (Qiagen).
|
| Label |
Cy3
|
| Label protocol |
Total RNA was amplified through one round of linear amplification using the MessageAmp aRNA kit (Ambion, Austin, TX). Sample quality and quantification was assessed by agarose gel electrophoresis and absorbance at A260. Cy3 and Cy5 labeled cDNA probes were made from 4 µg of amplified RNA.
|
| |
|
| |
| Hybridization protocol |
Two NPF cyclin D1 and two CAF samples (labeled with Cy3) were hybridized head-to-head with an NPF control sample labeled with Cy5. Probes were hybridized competitively to microarrays under a coverslip for 16 h at 63oC.
|
| Scan protocol |
Fluorescent array images were collected for both Cy3 and Cy5 by using a GenePix 4000B fluorescent scanner, and image intensity data were gridded and extracted using GenePix Pro 4.1 software.
|
| Description |
Human normal prostate fibroblasts overexpressing cyclin D1 NPF_cyclin D1_2 from Human hybridized against NPF Control
|
| Data processing |
Differences in gene expression between NPF cyclin D1/NPF and CAF/NPF groups were determined using a two-sample t-test with Significance Analysis of Microarrays (SAM) software (http://www-stat.stanford.edu/_tibs/SAM/) with a False Discovery Rate (FDR) of +/- 10% considered significant (Tusher, PNAS, 2001). Similarities in gene expression between NPF cyclin D1/NPF and CAF/NPF groups were determined using a one-sample t-test in SAM with an FDR of +/- 0.1% considered significant. These results were reduced to unique genes by eliminating all but the highest scoring clones for each gene. A Pearson correlation coefficient was calculated in Excel to assess the strength of the linear relationship between NPF cyclin D1/NPF and CAF/NPF average log2 ratios.
|
| |
|
| Submission date |
Feb 01, 2007 |
| Last update date |
Feb 05, 2007 |
| Contact name |
Denise Mauldin |
| E-mail(s) |
[email protected]
|
| Phone |
2066673480
|
| Fax |
2066672917
|
| URL |
http://www.pedb.org
|
| Organization name |
Fred Hutchinson Cancer Research Center
|
| Department |
Human Biology
|
| Lab |
Peter Nelson
|
| Street address |
1100 Fairview Ave N D4-100
|
| City |
Seattle |
| State/province |
WA |
| ZIP/Postal code |
98109 |
| Country |
USA |
| |
|
| Platform ID |
GPL4768 |
| Series (1) |
| GSE6936 |
Comparison of Normal Prostate Fibroblasts Overexpressing Cyclin D1 and Carcinoma Associated Fibroblasts |
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