|
| Status |
Public on Jul 14, 2015 |
| Title |
PBMCs sample_TR023 |
| Sample type |
RNA |
| |
|
| Source name |
Peripheral blood mononuclear cells (PBMCs)
|
| Organism |
Homo sapiens |
| Characteristics |
time: MTH4
site: Cali
individual: 319N
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Whole blood mRNA was extracted using TempusTM Blood RNA Tube isolation kits provided by the manufacturer Applied Biosystems, and the quality was determined based on the Agilent Bioanalyzer 2100 RNA Integrity score (RIN).
|
| Label |
EvaGreen
|
| Label protocol |
The qRT-PCR was completed in three steps. (1) Total whole blood RNA was converted to single stranded cDNA using polyT priming of reverse transcription, (2) the 96 targeted genes were pre-amplified in a single 13-cycle PCR reaction for each sample following conditions outlined in the manufacturer’s protocol, by combining 100 ng cDNA with the pooled primers and EvaGreen Pre-Amp Mastermix (Fluidigm BioMark™), (3) exonuclease step was perform to clean any unincorporated primers and (4) qPCR reactions were performed for each sample and individual gene on each sample on a 96×96 array with 30 amplification cycles. Average Ct value was calculated at a point in which every reaction is in the exponential phase to ensure accuracy and precision of amplification.
|
| |
|
| Hybridization protocol |
n/a
|
| Scan protocol |
n/a
|
| Description |
Test
|
| Data processing |
We subtracted each Ct from 30, setting missing values to 0. Since small Ct values correspond to high transcript abundance, this subtraction yields values ranging from 0 (no expression) to 30 (very high abundance). And Then mean normalization was perform using JMP genomics software. The non-normalized data, normalized data and fold-change data are available on the series record. Fold-changes were calculated by dividing the non-normalized test values by the non-normalized control value. Normalization was not performed for the control sample data.
|
| |
|
| Submission date |
Mar 31, 2015 |
| Last update date |
Jul 14, 2015 |
| Contact name |
Monica L. Rojas-Peña |
| E-mail(s) |
monica.rojas@gatech.edu
|
| Organization name |
Georgia Institute of Technology
|
| Department |
School of Biology
|
| Lab |
Center for Integrative Genomics (CIG)
|
| Street address |
310 Ferst Dr.
|
| City |
Atlanta |
| State/province |
GA |
| ZIP/Postal code |
30332 |
| Country |
USA |
| |
|
| Platform ID |
GPL19957 |
| Series (2) |
| GSE67469 |
Transcription Profiling of Malaria-Naïve and Semi-Immune Colombian Volunteers in a Plasmodium vivax Sporozoite Challenge [RT-PCR] |
| GSE67470 |
Transcription Profiling of Malaria-Naïve and Semi-Immune Colombian Volunteers in a Plasmodium vivax Sporozoite Challenge |
|
