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Status |
Public on Mar 31, 2010 |
Title |
Patient ID 53 Gleason grade 6 |
Sample type |
RNA |
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|
Channel 1 |
Source name |
patient tissue
|
Organism |
Homo sapiens |
Characteristics |
disease state: Gleason grade 6
|
Biomaterial provider |
Prof.Klocker, University of Innsbruck
|
Extracted molecule |
total RNA |
Extraction protocol |
RNA isolated from laser-capture microdissected epithelial prostate tumor cells of tissues from patients who had undergone radical prostatectomy were subjected to a two-round amplification using the MessageAmpTM II aRNA Amplification Kit (Applied Biosystems/Ambion, Austin, USA).
|
Label |
Cy5
|
Label protocol |
direct fluorescent labelling of 2 µg amplified RNA, anneling of 0.5µg Random Primer and reverse-transcribed into cDNA with Superscript III reverse transcriptase for 1h at 42°C in the presence of 0.4 mM dGTP, 0.4 mM dATP, 0.4 mM dTTP, 0.24 mM dCTP, 0.125 mM Cy3-dCTP. After hydrolysis of RNA in 0.2 M NaOH, Cy3-labeled probes were purified with Microcon YM-30 column.
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|
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Channel 2 |
Source name |
Universal human reference RNA (Stratagene), two rounds amplified
|
Organism |
Homo sapiens |
Characteristics |
sample: common reference
|
Biomaterial provider |
Stratagene
|
Extracted molecule |
total RNA |
Extraction protocol |
2 µg total RNA was amplified two-rounds using the MessageAmpTM II aRNA Amplification Kit (Applied Biosystems/Ambion, Austin, USA)
|
Label |
Cy3
|
Label protocol |
direct fluorescent labelling of 2 µg amplified RNA, anneling of 500ng Random Primer and reverse-transcribed into cDNA with Superscript III reverse transcriptase for 1h at 42°C in the presence of 0.4 mM dGTP, 0.4 mM dTTP, 0.4 mM dATP, 0.24 mM dCTP, 0.125mM Cy-5 dCTP . After hydrolysis of RNA in 0.2 M NaOH, Cy5-labeled probes were purified with Microcon YM-30 column.
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|
|
|
Hybridization protocol |
ch1 and ch2 together are solved in 50 µl 1x DIG-Easy hybridization buffer (Roche Diagnostics, Mannheim, Germany), containing 10x Denhardt’s solution and 2 ng/µl Cot1-DNA (Invitrogen); sample was denaturated at 65°C for 2min, hybridzation of arrays were performed in Corning chambers 14h at 37°C; slides were washed twice in 1xSSC+0.1SDS and once in 0.1xSSC+ 0.1SDS before air pressure drying and scanning.
|
Scan protocol |
arrays were scanned with the GenePix 4000B microarray scanner and analyzed using GenePix Pro 4.1 software (Axon Instruments)
|
Description |
patient.ID 53
|
Data processing |
raw data processing was performed using ArrayMagic (Buness et al., 2005) and VSN normalization method; Low quality measurements were excluded from further analysis. generalized log2 ratios (test/reference) from the remaining cDNA clones were given in the data table
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|
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Submission date |
Mar 31, 2009 |
Last update date |
Apr 02, 2009 |
Contact name |
Nicole Chui Pressinotti |
E-mail(s) |
n.pressinotti@dkfz-heidelberg.de
|
Organization name |
DKFZ
|
Street address |
Im Neuenheimer Feld 580
|
City |
Heidelberg |
ZIP/Postal code |
69120 |
Country |
Germany |
|
|
Platform ID |
GPL3050 |
Series (1) |
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