|
| Status |
Public on Jan 28, 2009 |
| Title |
T Cells Replicate 2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Tonsilar T cells
|
| Organism |
Homo sapiens |
| Characteristics |
Extracted from reactive tonsils
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using mirVana™ miRNA Isolation Kit (Ambion) following manufacturer's instructions
|
| Label |
Cy3
|
| Label protocol |
The assay started with 5 µg of total RNA sample. The RNA was size-fractionated using a YM-100 Microcon centrifugal filter (GE Millipore), and the 3′ ends of the small RNAs (<300 nt) were extended with a poly(A) tail using poly(A) polymerase. An oligonucleotide tag was then ligated to the poly(A) tail for later fluorescent dye staining; two different tags were used for the two RNA samples in the dual-sample experiments
|
| |
|
| Channel 2 |
| Source name |
FirstChoice® Human Skeletal Muscle Total RNA (Ambion)
|
| Organism |
Homo sapiens |
| Characteristics |
Total RNA from healthy human skeletal muscle
Tissue: skeletal muscle
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extracted using mirVana™ miRNA Isolation Kit (Ambion) following manufacturer's instructions
|
| Label |
Cy5
|
| Label protocol |
The assay started with 5 µg of total RNA sample. The RNA was size-fractionated using a YM-100 Microcon centrifugal filter (GE Millipore), and the 3′ ends of the small RNAs (<300 nt) were extended with a poly(A) tail using poly(A) polymerase. An oligonucleotide tag was then ligated to the poly(A) tail for later fluorescent dye staining; two different tags were used for the two RNA samples in the dual-sample experiments
|
| |
|
| |
| Hybridization protocol |
Hybridization was performed overnight on a µParaflo microfluidic chip using a microcirculation pump. Hybridization solution consisted of 100 µL of 6× SSPE buffer (0.90 M NaCl, 60 mM Na2HPO4, 6 mM EDTA, pH 6.8) containing 25% formamide at 34°C. After hybridization, the miRNAs were detected by fluorescence labeling using tag-specific Cy3 and Cy5 dyes
|
| Scan protocol |
Scanned on a GenePix 4000B (Molecular Device) laser scanner
Images were quantified using Array-Pro image analysis software (Media Cybernetics)
|
| Description |
Biological replicate 2 of 3. T cell, untreated, purified from reactive tonsils
|
| Data processing |
Median normalized, background subtracted data was used to calculate the ratio of processed Green signal/processed Red signal, and ratios were then log2 transformed. Probes hsa-miR-377 and hsa-miR-542-5p were excluded due to systematic dye bias, following manufacturer instructions.
|
| |
|
| Submission date |
Sep 25, 2008 |
| Last update date |
Jan 28, 2009 |
| Contact name |
Raquel Malumbres |
| E-mail(s) |
rmalumbres@yahoo.com
|
| Phone |
+34 948194700
|
| Organization name |
University of Navarra/ Clínica Universidad de Navarra
|
| Department |
Hemato-Oncology
|
| Lab |
Multiple Myeloma
|
| Street address |
Avenida Pio XII 55
|
| City |
Pamplona |
| State/province |
Navarra |
| ZIP/Postal code |
31008 |
| Country |
Spain |
| |
|
| Platform ID |
GPL7372 |
| Series (1) |
| GSE12934 |
miRNA profiles of tonsilar B and T lymphocytes |
|
