|
| Status |
Public on Oct 15, 2021 |
| Title |
mir-4423 overexpression, 24 h post-transfection, replicate 2 |
| Sample type |
RNA |
| |
|
| Source name |
HeLa cells + mir-4423 plasmid
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: HeLa
plasmid: mir-4423 expression plasmid
time point: 24 h
|
| Treatment protocol |
The cells were transiently transfected with the mir-199a-1 expression plasmid (HmiR0296-MR04, GeneCopoeia), the mir-4423 expression plasmid (HmiR1083-MR04, GeneCopoeia) or the negative control plasmid (CmiR0001-MR04, GeneCopoeia) using the FuGENE HD transfection reagent (Promega).
|
| Growth protocol |
The HeLa cells were cultured in DMEM (Thermo Fisher Scientific) supplemented with 10% FBS (Sigma-Aldrich) and 50 U/ml penicillin-streptomycin (Thermo Fisher Scientific) in T25 flasks (Greiner Bio-One).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated 24 h and 48 h after transfection with the miRNeasy Mini Kit (Qiagen).
|
| Label |
Biotin
|
| Label protocol |
According to the GeneChip Whole Transcript (WT) manual, cRNA was prepared from 210 ng total RNA. The cRNA was then used to generate single-stranded DNA, which was fragmented and biotinylated.
|
| |
|
| Hybridization protocol |
Labeled single-stranded DNA in the sense orientation was hybridized for 16 hours at 45 °C on Clariom D microarrays. The microarrays were washed and stained with a streptavidin-phycoerythrin conjugate in a GeneChip Fluidics Station 450. Signal amplification with antibodies was applied following the instructions provided by Thermo Fisher Scientific.
|
| Scan protocol |
The microarrays were scanned with a GeneChip Scanner 3000 7G (Affymetrix).
|
| Description |
Transcriptome data for HeLa cell culture after transfection of mir-4423 plasmid
|
| Data processing |
Data preprocessing of the raw microarray scans was performed using the Affymetrix GeneChip Command Console software version 4.0. The signal intensities of the >6 million oligonucleotide probes were further processed using the Transcriptome Analysis Console (TAC) version 4.0.2 (Applied Biosystems) in the default configuration. The signal space transformation robust multi-array average (SST-RMA) algorithm was applied for background adjustment, quantile normalization, gene-level probe set summarization and log2 transformation. The TAC software was also utilized for differential gene expression analysis.
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| |
|
| Submission date |
Oct 13, 2021 |
| Last update date |
Oct 16, 2021 |
| Contact name |
Michael Hecker |
| E-mail(s) |
michael.hecker@rocketmail.com
|
| Organization name |
University of Rostock
|
| Department |
Department of Neurology
|
| Lab |
Division of Neuroimmunology
|
| Street address |
Gehlsheimer Str. 20
|
| City |
Rostock |
| ZIP/Postal code |
18147 |
| Country |
Germany |
| |
|
| Platform ID |
GPL23126 |
| Series (1) |
| GSE185859 |
Screening for potential target genes of hsa-mir-199a-1 and hsa-mir-4423 using Clariom D arrays |
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