|
| Status |
Public on Jul 28, 2016 |
| Title |
Input-shControl |
| Sample type |
SRA |
| |
|
| Source name |
DU145 cells, shLuciferase, input
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: DU145
cell type: prostate cancer cell line
tissue derivation: prostate; derived from brain metastatic site
treatment: shLuciferase
chip antibody: None
|
| Treatment protocol |
Control shLuciferase DU145 cells.
|
| Growth protocol |
DU145 were cultured in High Glucose Dulbecco's Modified Eagle's Medium supplemented with L-glutamine, sodium bicarbonate, pyridoxine.HCI, sodium pyruvate and 10% fetal bovine serum.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
ChIP with reference exogenous genome (ChIP-Rx) was performed to compare H3K4me1 and H3K4me3 levels in shLuciferase, shJARID1D or shZMYND8 DU145 human prostate cancer cells. Drosophila melanogaster chromatin and Drosophila-specific H2Av antibody were spiked-in to each ChIP reaction as a minor fraction. Cells were cross-linked with 1% formaldehyde at room temperature for 10 min and then lysed and sonicated to fragment genomic DNA sizes to 200-500 bp. Chromatin samples were incubated with specific antibodies overnight at 4℃. The protein-DNA complexes were immobilized on protein A/G beads. The bound fractions were washed with low salt, high salt, LiCI wash buffer and TE buffer. Reverse-crosslinking of the eluted DNA were carried out at 67℃ for 8 hours. After RNase A and protease K digestion, DNA fraction was purified using PCR purification kit.
ChIP DNA libraries were prepared for sequencing using standard Illumina protocols.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
Processed data file: Input-Control.uniq.wig
|
| Data processing |
Illumina Casava1.7 software used for basecalling.
Total read numbers of ChIP-Seq samples without spike-in control are normalized to be 25 million, and ChIP-Seq samples with spiked-in Control are normalized according to the number of spiked-in Drosophila melanogaster chromatin.
ChIP-seq reads were aligned to the hg19 genome assembly using bowtie v1.1.2 with parameters -p 8 -m 1 --chunkmbs 512 --best
ChIP-Seq peaks were called using DANPOS2 v2.2.2 with the following parameters: --smooth_width 0 -e 1 --frsz 200 --extend 200 --pheight 1e-50 --extend_pheight 1e-20
RNA-Seq reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to hg19 genome assembly using tophat v2.0.12 with parameters --mate-std-dev 200 -p 8 -r 203
The number of fragments that originated from each gene in each sample is estimated using cuffdiff with default parameters. Then, the differentially expressed genes based on read counts are inferred using edgeR R package v3.12.0.
Genome_build: hg19 (GRCh37)
Supplementary_files_format_and_content: For RNA-Seq data, tab-delimited text file (genes.count_tracking.txt) includes raw counts of reads, RPKM values for each gene of each sample.
Supplementary_files_format_and_content: For ChIP-Seq data, wig files showing the average signal density across genome are generated using DANPOS2 v2.2.2.
|
| |
|
| Submission date |
Jun 03, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
JIE Lv |
| E-mail(s) |
lvjcmb@gmail.com
|
| Organization name |
St Jude Children's Research Hospital
|
| Department |
Department of Computational Biology
|
| Street address |
262 Danny Thomas Place
|
| City |
Memphis |
| State/province |
TN |
| ZIP/Postal code |
38105 |
| Country |
USA |
| |
|
| Platform ID |
GPL16791 |
| Series (1) |
| GSE82260 |
ZMYND8 reads the dual histone mark H3K4me1-H3K14ac to antagonize the expression of metastasis-linked genes |
|
| Relations |
| BioSample |
SAMN05204115 |
| SRA |
SRX1819618 |
