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Status |
Public on Aug 29, 2008 |
Title |
UCI101 pre-mir-424 2 |
Sample type |
RNA |
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Source name |
Ovary
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Organism |
Homo sapiens |
Characteristics |
Papillary ovarian carcinoma Gender: female
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Treatment protocol |
Transfected with precursor miRNA hsa-miR-424 (Ambion) using siPORT NeoFX (Ambion)
|
Extracted molecule |
total RNA |
Extraction protocol |
RNeasy Mini Kit (Qiagen)
|
Label |
Cy3
|
Label protocol |
Streptavidin-Cy3 bound to cRNA amplified with biotin-16-UTP using the Illumina TotalPrep RNA Amplification Kit (Ambion; Austin, TX, cat # IL1791). A 0.5ug aliquot of total RNA from each sample was labeled using the Illumina TotalPrep RNA Amplification Kit (Ambion; Austin, TX, cat # IL1791). RNA was first converted into single-stranded cDNA using reverse transcription and an oligo-dT primer containing the T7 RNA polymerase promoter then the single-stranded cDNA was copied to produce double-stranded cDNA molecules. The double-stranded cDNA was used in an overnight in vitro transcription reaction where single-stranded RNA (cRNA) was generated and labeled by incorporating biotin-16-UTP.
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Hybridization protocol |
A total of 0.85ug of biotin-labeled cRNA was hybridized for 16 hours to Illumina's Sentrix MouseRef-8 Expression BeadChips (Illumina, San Diego, CA). Each BeadChip has 24,000 well-annotated RefSeq transcripts per sample with approximately 30-fold redundancy. The arrays were washed at both 55 degrees C and room temperature and then blocked. The biotinylated cRNA was detected with streptavidin-Cy3, washed a final time and the arrays were dried by spinning at 500 RPM for 4 minutes.
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Scan protocol |
Slides were scanned using Illumina's BeadStation 500X Genetic Analysis Systems scanner. The image data was extracted using BeadStudio software, version 3. (Illumina)
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Description |
Microarray expression data generated from Illumina Human_Ref-8_V2 Sentrix BeadChips hybridized and scanned using a single channel of Cy3 fluorescent label.
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Data processing |
Raw microarray data was analyzed using DIANE 6.0, a spreadsheet-based microarray analysis program. Microarray intensities were subjected to Z normalization. i.e. [(intensity of sample #1 from array A - avarage of all intensities from array A)/ standard diviation of all of intensities on array A]. Differently expressed cRNA species were identified between the different sample groups by the ratio of these normalized intensities.
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Submission date |
Aug 29, 2008 |
Last update date |
Jun 22, 2020 |
Contact name |
Supriyo De |
Organization name |
NIA-IRP, NIH
|
Department |
Laboratory of Genetics and Genomics
|
Lab |
Computational Biology & Genomics Core
|
Street address |
251 Bayview Blvd
|
City |
Baltimore |
State/province |
Maryland |
ZIP/Postal code |
21224 |
Country |
USA |
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Platform ID |
GPL6104 |
Series (1) |
GSE12615 |
Ovarian Cancer Cell Lines pre-microRNA Transfection |
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