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Sample GSM303604 Query DataSets for GSM303604
Status Public on Dec 01, 2008
Title C666-1 Nasopharyngeal carcinoma cell line, EBV-positive
Sample type RNA
Source name MiRNAs from C666-1 cell line, grown to log phase
Organism human gammaherpesvirus 4
Characteristics C666-1 is a nasopharyngeal carcinoma cell line that maintains EBV without drug selection. Cells were grown to confluency in log phase and then harvested for RNA isolation.
Extracted molecule total RNA
Extraction protocol RNA was extracted using TRIzol reagent. MicroRNAs were precipitated with linear acrylamide.
Label Cy3
Label protocol For detection of miRNAs by microarray, miRNAs were amplified from total RNA isolate as previously described (47). Briefly, miRNAs were ligated to 3’ linker (5’ AppCTG TAG GCA CCA TCA ATddC 3’, Integrated DNA Technologies) and a 5’ linker (5’ ATC GTrA rGrGrC rArCrC rUrGrA 3’, ribonucleic acids noted with an r). The RNAs were reverse transcribed with a reverse transcription primer (5’ ATT GAT GGT GCC TAC 3’) using Superscript II (Invitrogen) and PCR amplified with forward (5’ GGC CAG TGA ATT GTA ATA CGA CTC ACT ATA GGG TTC TCG TGT TCC GTT TGT ACT CTA AGG TGG AAT CGT AGG CAC CTG AAA 3’) and reverse (5’ ATT GAT GGT GCC TAC AG 3’) primers. The resulting amplicons were transcribed in vitro overnight for 12 to 16 hours using T7 Megascript (Ambion).
Hybridization protocol RNAs were mixed with 1x SDS-based hybridization buffer (Genisphere, Inc.), and warmed to 70 degrees Celsius for 10 minutes. RNAs were allowed to cool to room temperature, and then hybridized to microarray slides at 55 degrees Celsius overnight. Slides were washed for 15 minutes in each of the following solutions pre-warmed to 55 degrees Celsius: 0.2% SDS, 2x SSC; 2x SSC; 0.2x SSC. Slides were centrifuged to dry slides. 3DNA dendrimer (Genisphere, Inc.) was warmed to 70 degrees Celsius with 1x SDS-based hybridization buffer for 10 minutes and then allowed to cool to room temperature. Microarray slides were hybridized with the 3DNA dendrimer, which binds the capture sequence incorporated into the RNAs during the in vitro transcription. Microarray slides were hybridized at 42 degrees Celsius for 4 hours. Slides were washed as after the first hybridization, but wash buffers were pre-warmed to 42 degrees Celsius. Slides were centrifuged to dry.
Scan protocol Slides were scanned on GenePix 4000B microarray scanner at 500 PMT, 33% laser power, 532 nm wavelength, 5 micromolar resolution. Data image was collected in GenePix Pro 6.0.
Description The modification (label protocol) of the RNAs allows for selective amplification of RNAs with 3' hydroxyl and 5' phosphate groups.
Data processing Background-corrected, raw-scale intensity values were obtained using GenePix Pro 6.0 (Molecular Devices) after minor manipulation to align and identify microarray probe spots. The averages of the median intensities of quadruplicate features were normalized to the average median intensities of the small nucleolar RNAs (snoRNAs), U38B, U43, U44, and U49.
Submission date Jul 06, 2008
Last update date Jul 09, 2008
Contact name Zachary Pratt
Organization name University of Wisconsin-Madison
Department Oncology
Lab Sugden
Street address 1400 University Avenue
City Madison
State/province WI
ZIP/Postal code 53706
Country USA
Platform ID GPL7022
Series (1)
GSE12014 Expression of EBV miRNAs in numerous EBV-positive cell lines

Data table header descriptions
VALUE normalized signal intensity

Data table
10 0.0045
1 0.0012
6 0.0003
36 0.2999
16 0.1908
14 0.0353
18 3.2033
26 0.1943
23 0.6312
21 0.2338
30 0.4545
27 0.0036
20 2.0093
7 0.3196
2 1.1042
11 0.4249
33 0.0102
13 0.2827
24 0.0571
17 0.0021

Total number of rows: 39

Table truncated, full table size <1 Kbytes.

Supplementary file Size Download File type/resource
GSM303604.txt.gz 5.8 Kb (ftp)(http) TXT
Processed data included within Sample table

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