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| Status |
Public on Apr 14, 2021 |
| Title |
ChIp-seq_Pfhmgb1_KO_h3k9me3_rep1 |
| Sample type |
SRA |
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| Source name |
ChIp-seq_Pfhmgb1_KO_h3k9me3_rep1
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| Organism |
Plasmodium falciparum |
| Characteristics |
development stage: Ring
strain: Pf 3D7
antibody: Rabbit anti-H3K9me3 antibody (Abcam, ab8898)
genotype: Pfhmgb1 Knock Down
host: Homo sapiens
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| Treatment protocol |
synchronized parasites at ring stage were cross-linked with 1% paraformaldehyde for 10 min, stopped with 0.125 M glycine for 5min and released with 0.15% saponin.
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| Growth protocol |
P. falciparum 3D7strain with 2% hematocrit was cultured in medium containing 10.44g/L RPMI-1640, 25mM HEPES, 10% Albumax I, 20μg/ml gentamicin, 0.1 mM hypoxanthine, a gas phase with 5% O2, 3% CO2 at 37 °C. Parasites were synchronized by 5% sorbitol and a 40%/70% Percoll gradient centrifugation.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Nuclei were isolated by Lysis Buffer and Dounce homogenization, followed by sonication to generate 200–600 bp fragments in length on the highest power setting for 10-20 min at 30s intervals. Protein-DNA complexes are selectively immunoprecipitated using 3 µg of Rabbit anti-H3K9me3 antibody (Abcam, ab8898), 3 μg of Rabbit anti-H3K9Ac antibody (Millipore, 07-352), or rabbit IgG (Sigma) as control. Formaldehyde induced cross-links are reversed with NaCl and DNA extracted using Proteinase K and MinElute PCR purification kit (Qiagen, 28006).
The ChIP DNAs were used to prepare sequencing libraries, including end-repair (Invitrogen No.16123), adding protruding 3’ A base (NEB No.M0212L), adapter ligation(NEB No.M2200L), size selection with Ampure beads (for 250 bp inserts) and amplification of libraries(KAPA Biosystems KB2500) (PCR program: 1 min at 98 ℃ followed by 12 cycles of 10 s at 98 ℃, 1 min at 65 ℃; finally extended 5 min at 65 ℃). Amplified libraries were cleand with Ampure beads
The library were sequenced on an Illumina HiSeq Xten systerm with PE150 strategy.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
HiSeq X Ten |
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| Description |
ChIp-seq_Pfhmgb1_KO_h3k9me3_rep1
Pfhmgb1_KO_h3k9me3_rep1.bed
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| Data processing |
Low-quality and the adaptor sequences were trimmed from the reads using cutadapt (v1.18) with parameters: -a AGATCGGAAGAGC -A AGATCGGAAGAGC --trim-n -m 50 -q 20,20. Then, the reads were mapped to the Plasmodium falciparum 3D7 genome (Pf 3D7 v32, obtained from PlasmoDB) using Bowtie2 (v2.3.4.3) with parameters: -N 0 --no-discordant --no-mixed --no-unal. The Samtools (v1.9) was used to transfer the mapping results from sam format to position sorted bam format. Next, the duplicated reads were removed by markdup from sambamba (v0.6.8). Then, the bam files were converted to bigwig files using bamCoverage from the deeptools suite (v3.1.3) with parameters: --normalizeUsing RPKM --binSize 25. The bigwig files from ChIP sample were normalized to input sample by bigwigCompare from the deeptools suite (v3.1.3) with parameters: --pseudocount 1 for further analysis. The Integrative Genomics Viewer (IGV) was used to show the signal of histone modification in certain genomic region in a track view.
Genome_build: Pf 3D7 v32
Supplementary_files_format_and_content: *.bed: HM bindings sites.
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| Submission date |
Dec 10, 2019 |
| Last update date |
Apr 14, 2021 |
| Contact name |
meng liu |
| E-mail(s) |
1710949@tongji.edu.cn
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| Organization name |
TongJi University
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| Street address |
1239,SiPing Road
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| City |
shanghai |
| ZIP/Postal code |
20082 |
| Country |
China |
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| Platform ID |
GPL26835 |
| Series (2) |
| GSE141760 |
HMGB1 is required for virulence gene expression via configuration of genome architecture in Plasmodium falciparum [HM-ChIP] |
| GSE141762 |
HMGB1 is required for virulence gene expression via configuration of genome architecture in Plasmodium falciparum |
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| Relations |
| BioSample |
SAMN13523972 |
| SRA |
SRX7299413 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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