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Sample GSM177476 Query DataSets for GSM177476
Status Public on Oct 23, 2007
Title HV3-20-59CGH
Sample type genomic
 
Channel 1
Source name male DNA
Organism Homo sapiens
Characteristics male DNA
Extracted molecule genomic DNA
Extraction protocol DNA was isolated using the QiAmp DNA isolation kit (Qiagen, Hilden, Germany) and subsequently quantified by a fluorometric assay (Quant-iT Pico Green dsDNA Kit, Invitrogen, Karlsruhe, Germany). Fluorescence was measured using a TECAN (Salzburg, Austria) SpectrafluorPLUS microplate fluorescence reader with an excitation wavelength at 485 nm and an emission wavelength at 535 nm. DNA concentrations were calculated from a standard curve of double-stranded control DNA provided with the kit that was measured in triplicate at the concentrations 30, 3, 0.3, and 0.03 ng/µl (measured by fluorometry).A volume of 2.5 µl out of 4 ng/µl dilutions from cell lines and control DNA was used as starting material for the amplification. The phi29-amplification was carried out according to the Repli-G kit manufacturer's instructions (Qiagen) using an incubation time of 16 h. Repli-G reactions were checked by a real time based intra Alu-PCR assay. Additionally DNA concentration was fluorimetrically measured with Quant-iT Pico Green dsDNA Kit (Invitrogen) and ranged between 10-30 Ž¼g.
Label cy3
Label protocol 10 µg of amplified and purified DNA were labeled using the BioPrime Array CGH Genomic Labeling kit (Invitrogen) according to the manufacturer¢â¬â„¢s instructions in a volume of 50 µl with a modified dNTP mix containing 120 µM each of dATP, dGTP, and dCTP; 60 µM dTTP; and 60 µM Cy5-dUTP or Cy3-dUTP. Labeled DNA was purified using QIAquick PCR purification columns (Qiagen). Tumor and reference DNA were pooled, mixed with 50 µg Cot-1 DNA (Invitrogen) and concentrated to a volume of 20 µl in a vacuum centrifuge. DNA was then mixed with an equal amount of 2x formamide buffer, denaturated at 95°C for 5 min and preincubated at 37°C for 30 min in a waterbath.
 
Channel 2
Source name DU145 HOM
Organism Homo sapiens
Characteristics male DNA
Extracted molecule genomic DNA
Extraction protocol DNA was isolated using the QiAmp DNA isolation kit (Qiagen, Hilden, Germany) and subsequently quantified by a fluorometric assay (Quant-iT Pico Green dsDNA Kit, Invitrogen, Karlsruhe, Germany). Fluorescence was measured using a TECAN (Salzburg, Austria) SpectrafluorPLUS microplate fluorescence reader with an excitation wavelength at 485 nm and an emission wavelength at 535 nm. DNA concentrations were calculated from a standard curve of double-stranded control DNA provided with the kit that was measured in triplicate at the concentrations 30, 3, 0.3, and 0.03 ng/µl (measured by fluorometry).A volume of 2.5 µl out of 4 ng/µl dilutions from cell lines and control DNA was used as starting material for the amplification. The phi29-amplification was carried out according to the Repli-G kit manufacturer's instructions (Qiagen) using an incubation time of 16 h. Repli-G reactions were checked by a real time based intra Alu-PCR assay. Additionally DNA concentration was fluorimetrically measured with Quant-iT Pico Green dsDNA Kit (Invitrogen) and ranged between 10-30 Ž¼g.
Label cy5
Label protocol 10 µg of amplified and purified DNA were labeled using the BioPrime Array CGH Genomic Labeling kit (Invitrogen) according to the manufacturer¢â¬â„¢s instructions in a volume of 50 µl with a modified dNTP mix containing 120 µM each of dATP, dGTP, and dCTP; 60 µM dTTP; and 60 µM Cy5-dUTP or Cy3-dUTP. Labeled DNA was purified using QIAquick PCR purification columns (Qiagen). Tumor and reference DNA were pooled, mixed with 50 µg Cot-1 DNA (Invitrogen) and concentrated to a volume of 20 µl in a vacuum centrifuge. DNA was then mixed with an equal amount of 2x formamide buffer, denaturated at 95°C for 5 min and preincubated at 37°C for 30 min in a waterbath.
 
 
Hybridization protocol DNA samples were hybridized onto the array for 16-18 hours at 42°C utilizing the MAUI 4-bay hybridization system and MAUI Mixer A0 (BioMicro System, Salt Lake City, UT, USA). After hybridization arrays and MAUI Mixer were disassembled in 42°C 1xSSC and 0.05% SDS (wash solution 1) and subsequently washed two times in wash solution 1 for 5 minutes followed by two washes in 0.1xSSC (wash solution 2) for 5 minutes.
Scan protocol Dried array slides were scanned using a ScanLite Express microarray scanner (Perkin Elmer, Wellesley, MA, USA). Raw image files of the arrays were processed using DeArray software (Chen et al, 1997) and resulting image data were imported into R environment using bioconductor packages (Gentleman RC et al., 2004) for further analysis.
Description male DNA
Data processing Loess normalization
 
Submission date Mar 27, 2007
Last update date Mar 13, 2008
Contact name Sean Davis
E-mail(s) sdavis2@mail.nih.gov
Phone 301-435-2652
Organization name National Cancer Institute
Lab Genetics Branch
Street address 37 Convent Drive, Room 6138
City Bethesda
State/province MD
ZIP/Postal code 20892
Country USA
 
Platform ID GPL5046
Series (1)
GSE7376 Detection of novel amplification units in prostate cancer

Data table header descriptions
ID_REF
VALUE log2 ratio (sample/reference)

Data table
ID_REF VALUE
24 -0.0311480211151054
12 -0.0280856409140917
120 -0.225916138912059
108 -0.0455562046104101
216 0.443811510323216
204 0.387746907507329
312 0.227458099445471
300 0.255656358852142
408 0.27369435454057
396 0.161551438103794
504 0.273397029123345
492 -0.196206875202934
600 0.625815220463027
588 1.05320194188654
696 -0.115016039527309
684 -0.0877447465728689
792 0.847435314621546
780 -0.276786406973779
888 0.142623629297737
876 NULL

Total number of rows: 36288

Table truncated, full table size 827 Kbytes.




Supplementary file Size Download File type/resource
GSM177476_Result.txt.gz 2.8 Mb (ftp)(http) TXT

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