|
| Status |
Public on May 24, 2016 |
| Title |
HCT116 MYC 3' TBE1 (WT) - 2 |
| Sample type |
SRA |
| |
|
| Source name |
colorectal carcinoma cell line
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: HCT116
genotype: wild type
|
| Growth protocol |
HCT116 MYC 3' TBE1 (WT) and KO cells were maintained in Dulbecco's Modified Eagle's Medium (DMEM) supplemented with 10% FBS, 50 units/ml penicillin, 2 mM Glutamax, and 0.1 mg/ml streptomycin at 37°C in 5% CO2
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
RNA was extracted from two biological replicates of 7.5 x 105 MYC 3' TBE1 (WT) or KO cells using the mirVana kit (Ambion, AM1560). The extracted RNAs were quantified using a NanoDrop apparatus (Thermo Scientific) which also ensured that the A260:A280 ratio was 1.9 or above. The RNA integration number (RIN) for each sample was measured using an RNA 6000 Nano Kit (Agilent, 5067-1511), and a RIN value of above 7 was required to proceed.
The cDNA libraries were prepared using the NEXTflex™ Rapid RNA Sequencing Kit and NEXTflex™ RNA-Seq Barcodes – 24 Kit (BioO Scientific) as per the manufacturer’s instructions. The final product was assessed for its size distribution using Bioanalyzer DNA High Sensitivity Kit (Agilent Technologies) and for its concentration using Kapa library quantification kit (Kapa Biosystems). The samples were sequenced using a HiSeq 2500 that was set to run for 50 cycles using a single-read recipe (TruSeq SBS Kit v3, FC-401-3002) according to the manufacturer’s instructions.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Data processing |
Bcl converion and demultiplexing using CASAVA 1.8.2. Adapter masked fastq files were generated.
Quality filter by FASTX Toolkit 0.0.13 (fastq_quality_filter: -Q33 -q 20 -p 80 was used as a parameter, fastq_quality_trimmer: -Q33 -t 20 -l 10 was used as a parameter). Fastq files were generated.
Read alignment using Tophat 2.0.9 using hg19 reference genome. BAM files were generated.
FPKM calculation using Cufflinks 2.2.1. Plain text (can be read by excel) files were generated. Both Tophat and Cufflinks used Gencode v19 gtf file as guided annotation file.
Genome_build: hg19
Supplementary_files_format_and_content: xls files for FPKM values for each sample and result of differential gene expression analysis
|
| |
|
| Submission date |
Jul 13, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Yuka Imamura Kawasawa |
| E-mail(s) |
yui102@psu.edu
|
| Organization name |
Penn State University
|
| Department |
College of Medicine, Pharmacology
|
| Street address |
500 University Dr.
|
| City |
Hershey |
| State/province |
PA |
| ZIP/Postal code |
17033 |
| Country |
USA |
| |
|
| Platform ID |
GPL16791 |
| Series (1) |
| GSE70833 |
HCT116 MYC 3' TBE1 (WT) and KO RNA-Seq |
|
| Relations |
| BioSample |
SAMN03856561 |
| SRA |
SRX1092623 |
