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Status |
Public on Jul 15, 2020 |
Title |
GORD 6 |
Sample type |
RNA |
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Source name |
peripheral blood serum exosomes
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Organism |
Homo sapiens |
Characteristics |
tissue: peripheral blood serum exosomes patient diagnosis: Gastro Oesophageal Reflux (GORD)
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Treatment protocol |
Peripheral blood was collected into serum clot activator tubes
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Growth protocol |
n/a
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Extracted molecule |
total RNA |
Extraction protocol |
Exosomes were isolated from the serum samples using ExoquickTm (System Biosciences, EXOQ20A-1). Extraction of miRNA from exosomes was performed using a miRNeasy Serum/Plasma kit (Qiagen, #217184) with a final elution volume of 24 microlitres ultra pure water
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Label |
FAM
|
Label protocol |
PCR assays were performed by following the protocol for TaqMan@OpenArray Human MicroRNA Panel (4461104, Life technologies). For each sample, 3μl of RNA was reverse transcribed using pre-defined RT-primers (MegaplexTM Primer Human Pool A and Pool B) and the TaqMan® microRNA Reverse Transcription Kit (Life technologies, #4366596). Pre-amplifications were carried out with MegaplexTM PreAmp Pools and TaqMan PreAmp Master Mix on 7.5μl cDNA/ sample for each pool. The pre-amplified products (4μl per sample) were diluted at the recommended 1:40 dilution with 156μl of RNase-free ultra pure water before loading onto the 384-well TaqMan OpenArray loading plate.
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Hybridization protocol |
n/a
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Scan protocol |
n/a
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Description |
GORD serum from patient with Gastro Oesophageal Reflux disease, without cancer microRNA GORD 6 -- SAB 11473
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Data processing |
Expression data were processed using Quant Studio Flex software qPCR Analysis Software (Applied Biosystems™, version 1.2.2). Fluorescence data was exported to CSV files by the software for use in R statistical software. Data were analysed in R version 3.4.2. The relative expression of each miRNA was calculated as 2(40-Ct). All possible permutations of miRNA ratios were generated from the relative expression values using R version 3.4.2. Nested cross validation and Lasso regression were used to select a panel of micrRNA ratios to build a logisitc regression model that could differentiate healthy controls and GORDs from OPSCCs. Matrix normalized worksheet reports the miRNA ratio expression values generated by R version 3.0.2 Fold Change worksheet reports the fold difference of expression of the microRNA ratios between the indicated study groups
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Submission date |
Sep 09, 2019 |
Last update date |
Jul 15, 2020 |
Contact name |
Damian James Hussey |
E-mail(s) |
damian.hussey@flinders.edu.au
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Organization name |
Flinders University
|
Street address |
Flinders Drive
|
City |
Bedford Park |
ZIP/Postal code |
5042 |
Country |
Australia |
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|
Platform ID |
GPL17837 |
Series (1) |
GSE137109 |
Cross validated serum exosomal microRNAs for the detection of oropharyngeal squamous cell carcinoma |
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