|
| Status |
Public on Jan 14, 2014 |
| Title |
ChIP-seq_JMJD6-re1-HEK293T |
| Sample type |
SRA |
| |
|
| Source name |
HEK293T cell
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: Human Embryonic Kidney 293 cells
chip antibody: JMJD6
chip antibody details: ab10526 (Abcam)
|
| Treatment protocol |
For experiments not involving siRNA transfection, cells grown to ~90% confluent were fixed directly; For experiments involving siRNA transfection, cells were subjected to two rounds of siRNAs transfection in 3 days, followed by fixation.
|
| Growth protocol |
HEK293T and HeLa cells were cultured in DMEM and MEM alpha medium supplemented with 10% FBS, respectively.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
For ChIP-seq, fixation was stopped by adding Glycine (0.125M) and incubating for 5 min at RT, followed by washing with PBS twice. Chromatin DNA was sheared to 300500 bp average in size through sonication. Resultant was immunoprecipitated with control IgG or specific antibodies overnight at 4°C, followed by incubation with protein G magnetic beads (Invitrogen) for an additional 2 h. After washing and elution, the proteinDNA complex was reversed by heating at 65°C overnight. For samples fixed with glutaraldehyde, proteinase K (0.3mg/ml) (Ambion) was added during de-crosslinking. Immunoprecipitated DNA was purified by using QIAquick spin columns (Qiagen) ; 7SK snRNA ChIRP was performed with an single anti-sense DNA oligonucleotide (5-accttgagagcttgtttggagg-3) following the protocol exactly as described previously (Chu et al., 2012). This DNA oligonucleotide sequence has been shown to specifically target to and efficiently pull down 7SK snRNA (Yang et al., 2001).
the libraries were constructed following Illumina’s Chip-Seq Sample prep kit. Briefly, Chip DNA was end-blunted and added with an ‘A’ base so the adaptors from Illumina with a ‘T’ can ligate on the ends. Then 200–400 bp fragments are gel-isolated and purified. The library was amplified by 18 cycles of PCR.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
JMJD6 ChIP-seq in HEK293T cells
|
| Data processing |
Basecalls performed using CASAVA version 1.4
reads were aligned to the hg18 genome assembly using Bowtie version 2.0.1.
The bigwig files were generated by using Homer v3.9, which the total tags are normalized to 1.00e+07.
Genome_build: hg18
Supplementary_files_format_and_content: bigwig files report normalized total tags
|
| |
|
| Submission date |
Oct 24, 2013 |
| Last update date |
May 15, 2019 |
| Contact name |
Qi Ma |
| E-mail(s) |
q1ma@ucsd.edu
|
| Organization name |
UCSD
|
| Street address |
9500 Gilman Dr,
|
| City |
San Diego |
| ZIP/Postal code |
92093 |
| Country |
USA |
| |
|
| Platform ID |
GPL11154 |
| Series (1) |
| GSE51633 |
Brd4 and JMJD6-associated Anti-pause Enhancers in Regulation of Transcriptional Pause Release |
|
| Relations |
| BioSample |
SAMN02383465 |
| SRA |
SRX367310 |
| Named Annotation |
GSM1249879_JMJD6-re1-HEK293T.bigWig |
