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| Status |
Public on Mar 06, 2022 |
| Title |
3D7_AP2-G5-GFP_T_input |
| Sample type |
SRA |
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| Source name |
trophozoite (25-30 hpi)
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| Organism |
Plasmodium falciparum |
| Characteristics |
tissue: whole body
developmental stage: trophozoite (25-30 hpi)
strain: Pf 3D7
genotype: AP2-G5-GFP
host: Homo sapiens
chip antibody: None
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| Treatment protocol |
Ring stage cultures were regularly synchronized by treatments of 5% sorbitol.
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| Growth protocol |
Parasites were cultured in fresh O-type human erythrocytes in complete RPMI 1640 medium (Gibco) with 0.5% Albumax I (Invitrogen) and a gas phase maintained under 5% CO2, 5% O2 and 90% N2 at 37°C.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Synchronized parasites were harvested at the ring (10-15 hpi), trophozoite (25-30 hpi), and schizont (40-45 hpi) stages respectively and crosslinked immediately with 1% paraformaldehyde (Sigma) by rotating for 10 minutes at 37°C, which was then quenched with 0.125 M glycine for 5 minutes on ice. Parasite nuclei released from infected red blood cells were sheared for 20-30 minutes using an M220 sonicator (Covaris) at 5% duty cycle, 200 cycles per burst, and 75 W of peak incident power to generate 100-500 bp fragments in length. 20 µL of the sheared chromatin was reserved as the input control. Chromatin was subsequently immunoprecipitated overnight at 4°C using 0.5 µg of antibodies against GFP (Abcam, ab290) or HP1 and protein A/G magnetic beads (ThermoFisher Scientific, 26162). After extensive washes, the immunoprecipitated materials were eluted with elution buffer. Then crosslinking was reversed by overnight incubation at 45°C, followed by treatments of RNase A at 37°C for 30 minutes and Proteinase K at 45°C for two hours. The ChIPped DNA was finally purified according to the MinElute PCR purification kit (Qiagen, 28006) instructions.
1.5 ng of ChIP-DNA was subjected to end-repair (Epicentre, ER81050), 3' adenylation (NEB, M0212L), and adapter ligation (NEB, M2200L). Then after purification with Agencourt AMPure XP beads (Beckman Coulter), libraries were amplified using the KAPA HiFi PCR Kit (KAPA Biosystems, KB2500) under the following conditions: 1 minute of initial denaturation at 98°C, 12 cycles of 10 seconds at 98°C and 1 minute at 65°C, and 5 minutes of final extension at 65°C.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
HiSeq X Ten |
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| Data processing |
To remove residual adapters and low-quality bases, sliding window trimming was performed on reads with Trimmomatic v0.39 using a window size of 4 and average quality of the window above 15. A minimum read length of 50 bp and average read quality above 20 were required after read clipping.
Trimmed ChIP-seq reads were mapped to the P. falciparum 3D7 reference genome (release 47) using Bowtie2 v2.4.2 and default parameters.
Peaks were identified using the callpeak function of MACS2 v2.2.7.1 and a q-value cutoff of 0.05. The options --call-summits and --broad were applied to the detection of PfApiAP2 and PfHP1 binding peaks, respectively. Peaks with fold enrichment below 1.5 were removed from downstream analysis. Furthermore, a transfectant with episomal GFP expression was used as a negative control to exclude ChIP background signals of the anti-GFP antibody. PfApiAP2 binding peaks with their fold enrichment/background GFP fold enrichment lower than 1.5 were also removed from further studies.
Normalized signal tracks of log2-transformed ChIP/input fold enrichment were generated with MACS2 bdgcmp function.
Genome_build: Pf 3D7 v47
Supplementary_files_format_and_content: bigWig files of log2-transformed ChIP/input fold enrichment
Supplementary_files_format_and_content: BED files of peak summits for PfApiAP2 ChIP-seqs
Supplementary_files_format_and_content: BED files of peaks for PfHP1 ChIP-seqs
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| Submission date |
Sep 23, 2021 |
| Last update date |
Mar 06, 2022 |
| Contact name |
Mei Jiang |
| E-mail(s) |
gingerplum@hotmail.com
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| Phone |
862165985138
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| Organization name |
Tongji University
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| Street address |
1239 Siping Road
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| City |
Shanghai |
| State/province |
Shanghai |
| ZIP/Postal code |
200092 |
| Country |
China |
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| Platform ID |
GPL26835 |
| Series (2) |
| GSE184658 |
A coordinated regulatory network of ApiAP2 transcription factors involved in heterochromatic gene expression during Plasmodium falciparum blood-stage development [ChIP-Seq] |
| GSE184659 |
A coordinated regulatory network of ApiAP2 transcription factors involved in heterochromatic gene expression during Plasmodium falciparum blood-stage development |
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| Relations |
| BioSample |
SAMN21572561 |
| SRA |
SRX12307826 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data not applicable for this record |
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