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| Status |
Public on May 13, 2014 |
| Title |
KYSE70 shTP63(2) Day 0 |
| Sample type |
SRA |
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| Source name |
KYSE70_shTP63_Day 0
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| Organism |
Homo sapiens |
| Characteristics |
cell line: KYSE70
cell type: human esophageal squamous carcinoma cells
genotype/variation: stably expressing pLKO-Tet-On-shTP63
treated with: none
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| Treatment protocol |
Doxycycline 50ng/ml for 4 days
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| Growth protocol |
RPMI1640 +10% FBS
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
For each sample, polyA+ RNA was isolated from 5 ug of total RNA using Dynabeads mRNA DIRECT Kit (Ambion). RNA was fragmented by incubation at 98 °C for 41 min in 1× RNA restoration buffer (Ambion). cDNA was synthesized with 20-40 ng of polyA+ RNA with a peak length around 600-700 bases with the use of SuperScript III Reverse Transcriptase, SuperScript Double-Stranded cDNA Synthesis kit, and random primers (Invitrogen). Primers were annealed at room temperature for 10 min followed by first strand synthesis for 1 h at 50°C and second strand synthesis for 2.5 h at 16°C. Following cDNA size selection with 1.8× AMPure XP beads (Beckman Coulter), end repair, and A-tailing, adapter ligation was performed using the TruSeq DNA Sample Preparation kit (Illumina) according to manufacturer’s instruction. Adaptor-ligated cDNAs were cleaned up with 0.8× AMPure XP beads followed by 8 cycles of PCR with primers complementary to the adaptor sequences with indexes. The PCR products were purified with 0.8× AMPure XP beads and then subject to HiSeq 2000 sequencing.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
Sample 4
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| Data processing |
Aligned to human reference genome (hg19) and exon-exon junctions (ensembl v64) with PRADA pipeline.
Transcriptome was collapsed to gene level using the GENCODE v12 transcriptome model.
RPKM values were generated by RNA-seQC for each experiment, log2 transformed (log2 (RPKM+1)), and quantile-normalized.
Gene expressions for each condition (shSOX2, and shTP63) were represented by the mean of duplicates, and were used to calculate the log2 fold change between two experimental conditions.
Genome_build: hg19 for human genome, ensembl v64 for exon-exon junctions.
Supplementary_files_format_and_content: Matrix text file, quantile-normalized log2 (RPKM+1)) values for each sample
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| Submission date |
May 17, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Shouyong Peng |
| E-mail(s) |
shouyongpeng@gmail.com
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| Organization name |
Dana-Farber Cancer Institute
|
| Department |
Medical Oncology
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| Lab |
Bass
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| Street address |
450 Brookline Avenue
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| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02215 |
| Country |
USA |
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| Platform ID |
GPL11154 |
| Series (1) |
| GSE47058 |
Differential gene expression by suppression of either SOX2 or TP63 in KYSE70 human esophageal squamous carcinoma cell line. |
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| Relations |
| BioSample |
SAMN02146676 |
| SRA |
SRX282062 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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