|
| Status |
Public on May 13, 2016 |
| Title |
BJEL_RNA_PolII_ChIP-seq |
| Sample type |
SRA |
| |
|
| Source name |
Immortalized BJEL fibroblast cells (pretransformed)
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: BJEL
cell type: foreskin fibroblast cells
chip antibody: polyclonal anti-RNA-PolII (H-224)
chip antibody supplier: Santa Cruz Biotechnology
chip antibody catalog#: sc-9001
chip antibody lot #: I0513
|
| Growth protocol |
BJEL cells were cultured in monolayer conditions Dulbecco’s modified Eagle’s medium (DMEM)/M199 (4:1) (with 4.5 g/l glucose) supplemented with 10% of heat inactivated fetal calf serum (FCS), G-418 (400 µg/µl) and Hygromycin (400 µg/µl).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
cells were fixed with 1% para-formaldehyde (Electron Microscopy Sciences) for 30 minutes at room temperature. ChIP assays were performed following previously described conditions: chromatin sonication and immunoprecipitation in lysis buffer (50mM Tris–HCl pH=8, 140mM NaCl, 1mM EDTA, 1% Triton, 0.1% Na-deoxycholate) complemented with protease inhibitor cocktail (Roche 11873580001); 2x washes with lysis buffer; 2x washes with lysis buffer containing 500mM NaCl; 2x washes with washing buffer (10mM Tris–HCl pH=8, 250mM LiCl, 0.5% NP-40, 1mM EDTA, 0.5% Na-deoxycholate); 2x washes with 1x TE buffer; elution at 65°1C; 15 min at 65°C in elution buffer (50mM Tris–HCl pH=8, 10mM EDTA, 1% SDS); overnight decrosslinking at 65C followed by PCI purification. qPCR-qualified ChIP assays were quantified (Qubit dsDNA HS assay kit; Invitrogen); 10 ng of the ChIPed material was used for preparing Illumina sequencing libraries.
Libraries were prepared following a multiplexing approach (NEXTflexTM ChIP-seq Bioo Scientific kit; ref: 5143-02).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
Immortalized BJEL fibroblast cells (pretransformed)
ChIP against RNA PolII
|
| Data processing |
Prepared sequencing libraries were sequenced on the Illumina instrument HiSeq2000. Regular Illumina pipelines were used for image processing and base calling.
bed files obtained from the aligned profiles using peak calling tools MeDiChi-seq (for RNA PolII and H3K9ac, H3K4me3, H3K27ac marks) ann SICER tool (for H3K27me3) with the input datasets as a negative control
Genome_build: hg19
Supplementary_files_format_and_content: wig and bed (peak annotation information)
|
| |
|
| Submission date |
Sep 04, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Valeriya Malysheva |
| E-mail(s) |
Valeriya.MALYSHEVA@igbmc.fr
|
| Organization name |
IGBMC
|
| Street address |
1 rue Laurent Fries
|
| City |
Illkirch; Strasbourg |
| ZIP/Postal code |
67404 |
| Country |
France |
| |
|
| Platform ID |
GPL11154 |
| Series (2) |
| GSE72532 |
Reconstructing gene regulatory networks of tumorigenesis [ChIP-Seq] |
| GSE72533 |
Reconstructing gene regulatory networks of tumorigenesis |
|
| Relations |
| BioSample |
SAMN04032797 |
| SRA |
SRX1184131 |
