|
| Status |
Public on Aug 10, 2024 |
| Title |
786-O_TFE3KD_dox_rep3 |
| Sample type |
SRA |
| |
|
| Source name |
cell line
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: cell line
cell line: 786-O
cell type: Cancer cell
genotype: TFE3 knockdown
treatment: dox
|
| Treatment protocol |
ccRCC cell line (786-O) and tRCC cell line (FU-UR-1) were transduced with lentivirus expressing doxycycline shRNA against wild type TFE3 and ASPSCR1-TFE3 and selected with 500 mg/mL of G418. Subsequently, the cells were treated with doxycycline at a concentration of 1mg/mL for 5 days. s-TFE tRCC cells were transduced with control sgRNA or sgRNA targeting ASPSCR1-TFE3 for 5 days
|
| Growth protocol |
DMEM, 10% fetal bovine serum, 1x penicillin/streptomyicin. Cells were cultured at 37C at 5% CO2.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using RNeasy Plus Mini Kit (QIAGEN, #74136)
After the QC procedures, mRNA is enriched using oligo(dT) beads. Then, the mRNA is fragmented randomly by adding fragmentation buffer, then the cDNA is synthesized by using mRNA template and random hexamers primer, after which a custom second-strand synthesis buffer (Illumina), dNTPs, RNase H and DNA polymerase I are added to initiate the second-strand synthesis. After a series of terminal repair, A ligation and sequencing adaptor ligation, the double-stranded cDNA library is completed through size selection and PCR enrichment.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
Paired-end RNA-seq reads were aligned to a Bowtie2 indexed human genome (hg38 sourced from UCSC) using STAR with default settings. For better alignment, the first aligned splicing junctions detected by STAR (SJ.out.tab) were used for STAR alignment again with parameters “--sjdbFileChrStartEnd SJ.out.tab --readFilesIn $R1 $R2 --quantMode TranscriptoomeSAM GeneCounts --sjdbGTFfile $GTF_file --outSAMtype BAM Unsorted SortedByCoordinate”. Then the aligned to transcriptome bam files (AlignedtoTranscriptome.out.bam) were used to quantify gene expression levels by rsem-calculate-expression function from RSEM. Tag per million reads (TPM) from gene expression results files (.genes.results) were extracted and assembled for subsequent differential gene expression analysis by DESeq2.
Assembly: hg38
Supplementary files format and content: genes.results.gz
|
| |
|
| Submission date |
May 03, 2024 |
| Last update date |
Aug 10, 2024 |
| Contact name |
Kaimeng Huang |
| E-mail(s) |
Kaimeng_huang@dfci.harvard.edu
|
| Organization name |
Dana-Farber Cancer Institute
|
| Street address |
4 Blackfan Circle
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02215 |
| Country |
USA |
| |
|
| Platform ID |
GPL24676 |
| Series (1) |
| GSE266517 |
Metabolic reprogramming driven by TFE3 fusions unveils novel vulnerabilities in translocation renal cell carcinoma [RNA-Seq] |
|
| Relations |
| BioSample |
SAMN41200437 |
| SRA |
SRX24468949 |
