|
| Status |
Public on Dec 20, 2016 |
| Title |
TRAF7/KLF4_ChIP-Seq [MN-191] |
| Sample type |
SRA |
| |
|
| Source name |
meningioma
|
| Organism |
Homo sapiens |
| Characteristics |
tissue type: meningioma
mutation subtype: TRAF7/KLF4
grade: benign
chip antibody: H3k27ac (Abcam, ab4729)
|
| Treatment protocol |
All samples are untreated, primary specimens from brain tumor patients.
|
| Growth protocol |
Samples were excised during neurosugical procedures and frozen in OCT for later use. H&E slides were prepared for each sample to confirm tumor histology and assess tissue density. On the day of each experiment, sections were cut using a cryostat (20uM) and immediately prepared for immunoprecipitation.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
10 to 15 frozen sections of each tumor block or dura sample were collected for each experiment. Tissue was crosslinked with 1% formaldehyde, quenched with glycine, and washed with PBS. Nuclei were extracted by dounce homogenization and resuspended in nuclear lysis buffer containing 0.3% SDS. Chromatin was sheared by sonication with a Q800R2 Sonicator by QSonica (60 minutes total, amplitude 30, 10 second pulses, 10 second rest). Soluble chromatin was incubated with magnetic beads coated with either H3K27ac antibody (ab4729) overnight at 4°C. Chromatin was precipitated using a magnet, washed extensively, and eluted with TE+1% SDS. Crosslinks were reversed, and samples were purified.
Prior to sequencing, the enrichment of each sample was calculated using qPCR and well-established control primers for H3K27ac.. Samples were subjected to standard Illumina paired-end multiplexed library construction. H3K27ac ChIP and input samples were sequenced for each tumor (1x75bp, HiSeq 2000).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
MN-191_ChIP-Seq
processed data file:
mn_201_h3k27ac_peaks_Gateway_SuperEnhancers_forDiffBind.bed
|
| Data processing |
H3K27ac reads were aligned uniquely with bowtie (0.12.7) 77 to the human genome (hg19). Regions of enrichment were identified with MACS (v1.4) 78.
Super-enhancers were separated from typical enhancers using ROSE pipeline with parameters -s (stitching) 12500, -t (promoter exclusion zone) 2000. We used DiffBind to estimate significance of super-enhancer read change (adjusted p-value threshold = 0.05) between meningioma subtypes.
Genome_build: hg19
Supplementary_files_format_and_content: bed
|
| |
|
| Submission date |
Dec 09, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Akdes Serin Harmanci |
| Organization name |
Yale University
|
| Department |
Departments of Neurosurgery and Genetics
|
| Lab |
Murat Gunel
|
| Street address |
300 Cedar Street TAC S330
|
| City |
New Haven |
| State/province |
Connecticut |
| ZIP/Postal code |
06510 |
| Country |
USA |
| |
|
| Platform ID |
GPL11154 |
| Series (2) |
| GSE91372 |
Integrated genomic analyses of de novo pathways underlying atypical meningiomas [ChIP-Seq] |
| GSE91376 |
Integrated genomic analyses of de novo pathways underlying atypical meningiomas |
|
| Relations |
| BioSample |
SAMN06128229 |
| SRA |
SRX2409198 |
