|
| Status |
Public on Aug 30, 2016 |
| Title |
HCT116_J2_Combo |
| Sample type |
SRA |
| |
|
| Source name |
HCT116
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: human colon carcinoma cell line HCT116
treatment: Combination
antibody: J2 antibody
|
| Treatment protocol |
HCT116 (3 x 10^5) cells were treated with 0.3 μM of 5-Aza-CdR (Sigma-Aldrich). After 24 hours, medium was replaced and maintained at Day 5.
|
| Growth protocol |
HCT116 were maintained in McCoy’s 5A medium and cultured according to standard mammalian tissue culture protocols and sterile technique.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The 5-Aza-CdR treated cells were lysed with lysis buffer (30 mM HEPES-KOH at pH 7.4, 20mM MgOAc, 5mM DTT, 150mM KOAc, 0.1% NP-40, protease inhibitors). RNase T1(single-strand-specific RNA endonuclease) was added into the lysis buffer. The cell lysates were used for dsRNA pull-down with 3μg of J2 antibody (Scicons) on Dynabeads Protein G (Invitrogen) for 2 hours at 4°C. The beads were washed with wash buffer (30 mM HEPES-KOH at pH 7.4, 20mM MgOAc, 5mM DTT, 150mM KOAc, 0.1% NP-40, protease inhibitors). Digestions of single-stranded RNA were carried out by RNase T1 (Roche). Digestions of DNA were carried out by TURBO DNase (Thermo Fisher). dsRNA was purified and fragmented into ∼200 bp by shearing (Covaris). cDNA library preparation. RNA-seq libraries were prepared by using the NEBNext Multiplex Small RNA Library Prep Set for Illumina.
|
| |
|
| Library strategy |
OTHER |
| Library source |
transcriptomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
double stranded RNA
20160114_JonesLabVAI_J2RNAseq_ERVcnt.xlsx
|
| Data processing |
Count reads aligned to annotation from RepeatMasker
Genome_build: hg19
Supplementary_files_format_and_content: dsRNA measurement from J2 antibody pull down experiment in HCT116 cell line
|
| |
|
| Submission date |
Jan 20, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Peter A Jones |
| E-mail(s) |
Peter.Jones@vai.org
|
| Organization name |
Van Andel Research Institute
|
| Department |
Center for Epigenetics
|
| Lab |
Jones Lab
|
| Street address |
333 Bostwick Ave
|
| City |
Grand Rapids |
| State/province |
MI |
| ZIP/Postal code |
49503 |
| Country |
USA |
| |
|
| Platform ID |
GPL18573 |
| Series (1) |
| GSE77031 |
Double-stranded RNA sequencing of HCT116 cells with 5-Aza-CdR and combination treatment compared to untreated cells |
|
| Relations |
| BioSample |
SAMN04430469 |
| SRA |
SRX1538732 |
