|
| Status |
Public on Jun 29, 2024 |
| Title |
HepG2 cell, shEPDR1 |
| Sample type |
SRA |
| |
|
| Source name |
HepG2
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: HepG2
cell type: hepatocarcinoma cell
genotype: shEPDR1
|
| Treatment protocol |
HepG2 cells were were infected with lentivirus expressing shEPDR1 and selected with 0.5 µg.ml–1 puromycin to establish stable cells, with a NTC as control
|
| Growth protocol |
HepG2 cells were cultured in DMEM supplemented with 10% FBS and 1% penicillin‒streptomycin. Cells were grown at 37°C and 5% CO2.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from cell lines using TRIzol Reagent (Life Technologies). RNA integrity was assessed by RNA integrity number and determined using an Agilent 2100 Bioanalyzer.
Total RNA was used as input material for the RNA sample preparations. Sequencing libraries were generated using NEBNext® UltraTM RNA Library Prep Kit for Illumina® (NEB, USA) following manufacturer’s recommendations and index codes were added to attribute sequences to each sample. Briefly, mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. Fragmentation was carried out using divalent cations under elevated temperature in NEBNext First Strand Synthesis Reaction Buffer(5X). First strand cDNA was synthesized using random hexamer primer and M-MuLV Reverse Transcriptase(RNase H). Second strand cDNA synthesis was subsequently performed using DNA Polymerase I and RNase H. Remaining overhangs were converted into blunt ends via exonuclease/polymerase activities. After adenylation of 3’ ends of DNA fragments, NEBNext Adaptor with hairpin loop structure were ligated to prepare for hybridization. In order to select cDNA fragments of preferentially 250~300 bp in length, the library fragments were purified with AMPure XP system (Beckman Coulter, Beverly, USA). Then 3 µl USER Enzyme (NEB, USA) was used with sizeselected, adaptor-ligated cDNA at 37°C for 15 min followed by 5 min at 95 °C before PCR. Then PCR was performed with Phusion High-Fidelity DNA polymerase, Universal PCR primers and Index (X) Primer. At last, PCR products were purified (AMPure XP system) and library quality was assessed on the Agilent Bioanalyzer 2100 system.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Reads were aligned to the human genome hg38. TopHat2 v.2.1.0 and cufflinks v.2.2.1 were used to analyze RNA-seq data. Gene differential expression analysis was carried out with the DEGSeq R package (1.26.0).
Assembly: hg38
Supplementary files format and content: fpkm_tracking
|
| |
|
| Submission date |
Dec 14, 2023 |
| Last update date |
Jun 29, 2024 |
| Contact name |
Xiaoyu Qian |
| E-mail(s) |
xiaoyu8274687@outlook.com
|
| Organization name |
South China University of Technology
|
| Street address |
Guangzhou Panyu University City
|
| City |
Guangzhou |
| State/province |
Guangzhou |
| ZIP/Postal code |
510641 |
| Country |
China |
| |
|
| Platform ID |
GPL24676 |
| Series (1) |
| GSE250169 |
EPDR1 promotes PD-L1 expression and antitumor immune evasion by inhibiting TRIM21-dependent ubiquitylation if IKBKB. |
|
| Relations |
| BioSample |
SAMN38836397 |
| SRA |
SRX22885345 |
