|
| Status |
Public on Nov 27, 2023 |
| Title |
RAC1_rep2 |
| Sample type |
SRA |
| |
|
| Source name |
Brain
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: Brain
cell line: IDH-wt-hGICs-MGT#4-dEGFP dCas9-VPR
cell type: Transformed neural progenitor cells from the subventricular zone
genotype: RAC1 overexpressing
|
| Growth protocol |
The IDH-wt-hGICs-MGT#4-dEGFP dCas9-VPR were propagated in RHB-A medium (Takara), supplemented with EGF (20 ng/ml; R&D), bFGF (20 ng/ml; R&D), heparin (1 μg/ml; Sigma), PDGF-aa (20 ng/ml; R&D), 1% penicillin and streptomycin. Cells were cultured at 37°C in a 5% CO2, 3% O2 and 95% humidity incubator.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA extraction from IDH-wt-hGICs-MGT#4-dEGFP dCas9-VPR cells was performed using the TRIzol Reagent (Invitrogen, 15596026), followed by isopropanol precipitation and subsequent AMPure XP beads (Beckman Coulter, #A63881) purification.
Multiplexed 3'-cDNA libraries were constructed starting from 60 ng of total RNA per sample as input, utilizing barcoded oligo-dT primers (Datlinger et al., 2021) in an adapted version of Bulk RNA barcoding and sequencing (BRB-seq) protocol (Alpern et al., 2019).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
RAC1 overexpressing cells, sorted for high levels of MGT#4 activation
|
| Data processing |
Illumina index demultiplexing was performed using the bcl2fastq conversion software (v2.20.0).
The oligo-dT barcode sequences from Read 1 were extracted using cutadapt (v2.1) for subsequent internal demultiplexing of reads using BRBseqTools-1.6.jar (http://github.com/DeplanckeLab/BRB-seqTools).
The BRBseqTools demultiplexed data was aligned to a custom genome (MGT#4-containing GRCh38) using STAR (v2.7.8a).
HTSeq (v2.0.2) was used to generate the count matrices.
Assembly: GRCh38.107 + MGT#4
Supplementary files format and content: Tab-delimited text file including raw counts for each sample.
|
| |
|
| Submission date |
Jun 29, 2023 |
| Last update date |
Nov 27, 2023 |
| Contact name |
Gaetano Gargiulo |
| E-mail(s) |
Gaetano.Gargiulo@mdc-berlin.de
|
| Phone |
+49 30 9406-3861
|
| Organization name |
Max Delbrück Center for Molecular Medicine
|
| Department |
Molecular Oncology
|
| Street address |
Robert-Rössle-Straße 10
|
| City |
Berlin |
| ZIP/Postal code |
13125 |
| Country |
Germany |
| |
|
| Platform ID |
GPL24676 |
| Series (1) |
| GSE236153 |
Transcriptomic profiling of IDH-wt-hGICs-MGT#4-dEGFP cells upon overexpression of CRISPRa-screen predicted pro-mesenchymal phenotypic drivers. |
|
| Relations |
| BioSample |
SAMN36072587 |
| SRA |
SRX20827516 |
