|
Status |
Public on Jan 28, 2009 |
Title |
Centroblasts Replicate 3 |
Sample type |
RNA |
|
|
Channel 1 |
Source name |
Tonsilar centroblasts
|
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 3 of 4. Centroblast, 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 |
|