|
| Status |
Public on May 08, 2024 |
| Title |
RNAseq GSK343 rep1 |
| Sample type |
SRA |
| |
|
| Source name |
K562 cells with GSK343
|
| Organism |
Homo sapiens |
| Characteristics |
treatment: treated with 5µmGSK343 for 72hours
cell line: K562
|
| Treatment protocol |
EV, S1KO, S2KO and DKO cells undergoes the CRISPR excision. The drug treated K562 cells were either treated with DMSO-72h, 5uM GSK343-72h, 100uM X5050-72h or combinational treatment (5uM GSK343+100uM X5050-72h). The time course GR treatment was done for either 8h or 24h treatment of the same concentration of GR as 72h.
|
| Growth protocol |
All the cells were cultured in RPMI-1640 supplemented with 10% Fetal Bovine Serum (FBS) and 1% penicillin-streptomycin. All cultures were maintained at 37 °C, 5% CO2 in a humidified incubator.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA were isolated from the cells using RNeasy Mini Kit (Qiagen) together with performing on-column DNase digestion (Qiagen). RNA concentrations were determined by Nanodrop ND1000 (Thermo Scientific).
The quality of the RNA extracted was analyzed using Agilent RNA 6000 Nano Kit (Agilent), and quantitated using Nanodrop ND1000 (Thermo Scientific). Library construction was performed using TruSeq Stranded Total RNA LT (w/ Ribo-Zero Gold) Set A (Illumina) as per protocol.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Data processing |
For the RNA-seq reads of K562 EV, S1KO, S2KO and DKO, we first trimmed off adaptors by trimmomatic (0.38) using option ILLUMINACLIP:TruSeq3-PE.fa:2:30:10 LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:36. Only those paired-end reads that remained properly paired would be then mapped to the hg19 reference GTF file using kallisto(0.44.0) with option '-b 100'. Differentially expressed genes were called using sleuth(0.29.0) with gene-level aggregation and wald test.
For the RNA-seq reads of K562 DMSO, X5050, GSK343, combined 8H, combined 24H and combined 72H, we first trimmed off adaptors by trimmomatic (0.38) using option ILLUMINACLIP:TruSeq3-PE.fa:2:30:10 LEADING:3 TRAILING:3 SLIDINGWINDOW:4:15 MINLEN:36. Only those paired-end reads that remained properly paired would be then mapped to the hg19 reference GTF file using STAR (2.7.3a) with parameters ”--runThreadN 16 --outSAMtype BAM SortedByCoordinate --outReadsUnmapped Fastx --quantMode GeneCounts --readFilesCommand zcat --twopassMode Basic --outFilterType BySJout --outFilterMultimapNmax 20 --outFilterMismatchNmax 999 --outFilterMismatchNoverReadLmax 0.04 --alignSJoverhangMin 8 --alignSJDBoverhangMin 1 --chimSegmentMin 20 --chimJunctionOverhangMin 20 --chimOutJunctionFormat 1 --alignIntronMin 20 --alignIntronMax 1000000 --alignMatesGapMax 1000000 --chimSegmentReadGapMax 0 --alignSJstitchMismatchNmax 0 -1 0 0 ”. Differentially expressed genes were called using Deseq2(0.29.0) .
Genome_build: hg19
Supplementary_files_format_and_content: *.tsv : *.tsv is a plaintext file of the abundance estimates generated by Kallisto. Fields contained: 1. target_id. Transcript ID; 2. length. Transcript length; 3. eff_length. Effective length of the transcript; 4. est_counts. Estimated read counts given by the bootstrapping process of Kallisto; 5. tpm. Esmated transcript per million (TPM).
Supplementary_files_format_and_content: *.tab: *.tab is a plaintext file of the abundance estimates generated by STAR with --quantMode GeneCounts option. Fields contained: 1. gene id ; 2. counts for unstranded RNA-seq; 3. counts for the 1st read strand aligned with RNA; 4. counts for the 2nd read strand aligned with RNA.
|
| |
|
| Submission date |
Jan 11, 2022 |
| Last update date |
May 08, 2024 |
| Contact name |
KAIJING CHEN |
| E-mail(s) |
KAIJING001@e.ntu.edu.sg
|
| Phone |
98667053
|
| Organization name |
Nanyang Technological University
|
| Department |
school of biological science
|
| Lab |
Melissa Fullwood
|
| Street address |
60 Nanyang Drive
|
| City |
Singapore |
| ZIP/Postal code |
637551 |
| Country |
Singapore |
| |
|
| Platform ID |
GPL20301 |
| Series (2) |
| GSE193486 |
Super-silencer perturbation by EZH2 and REST inhibition leads to large loss of chromatin interactions and reduction in cancer growth [RNA-seq] |
| GSE193489 |
Super-silencer perturbation by EZH2 and REST inhibition leads to large loss of chromatin interactions and reduction in cancer growth |
|
| Relations |
| BioSample |
SAMN24849083 |
| SRA |
SRX13731854 |
