|
| Status |
Public on Apr 21, 2021 |
| Title |
S1_H3K9me3 |
| Sample type |
SRA |
| |
|
| Source name |
Streaming cells (AX4) replicate 1
|
| Organism |
Dictyostelium discoideum |
| Characteristics |
developmental stage: Streaming
strain: AX4
|
| Growth protocol |
We cultured te strain at 22 °C shaken in the incubator at 180 rpm in HL5 medium with 300 μg/ml streptomycin
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
10 million D. discoideum cells were cross-linked with 1x linking buffer (11 mM HEPES-NaOH, 110 mM NaCl, 1.1 mM EDTA and 1.1 mM EGTA) with 1% formaldehyde for 10 minutes at RT and quenched by 2M glycine. The samples were fragmented by applying a biorupter for 25 cycles in the cold room with 30 seconds on and 30 seconds off per cycle. The precleared chromatin was incubated with antibody-coupled bead slurries and rotated at 4°C overnight. DNA was eluted from the beads by 200 µl TE with 1% SDS.
Histone ChIP-Seq libraries were generated using the Ovation Ultralow V2 kit (Nugen, 0344-32) according to the manufacturer’s instructions. Libraries were PCR amplified for 13-16 cycles depending on the antibody, and library quality was assessed using the Agilent 2100 Bioanalyzer (Agilent Technologies). Seventy-five bp reads were generated on the Illumina Nextseq 500 (Illumina). ChIP-seq were sequenced to a depth of at least 8 million total reads/sample.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
The raw fastq files were trimmed for the first and last 8 nucleotides, the multiqc was use to summarize the fastQC results, and the trimmed files were mapped to the Dictyostelium discoideum genome using bowtie2
The duplicated reads were removed by picard and sorted bam files were subject to peak calling with MACS2.
Differential binding analyses were performed by R package Diffbind using log2 normalized ChIP read counts with FDR < 0.05
Genome_build: AX4
Supplementary_files_format_and_content: Peak files generated by macs2
|
| |
|
| Submission date |
Sep 17, 2019 |
| Last update date |
Apr 21, 2021 |
| Contact name |
Eric Greer |
| E-mail(s) |
Eric.Greer@childrens.harvard.edu
|
| Organization name |
Boston Children's Hospital/Harvard Medical School
|
| Department |
Newborn Medicine/Department of Pediatrics
|
| Lab |
Greer Lab
|
| Street address |
320 Longwood Avenue
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02115 |
| Country |
USA |
| |
|
| Platform ID |
GPL26360 |
| Series (2) |
| GSE137599 |
Determine the genome-wide localization of histone and histone marks in different stages of D. discoideum |
| GSE137604 |
Role of Epigenetics in Unicellular to Multicellular Transition in Dictyostelium |
|
| Relations |
| BioSample |
SAMN12776345 |
| SRA |
SRX6862307 |
