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| Status |
Public on Dec 24, 2016 |
| Title |
WT1minusKTS cells replicate 2 Streptavidin |
| Sample type |
SRA |
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| Source name |
K562 cell line
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| Organism |
Homo sapiens |
| Characteristics |
cell line: K562
transfections apart from pef1abirav5-neo: pEF1a-Flagbiotin-(WT1+17AA/-KTS)-puro
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| Treatment protocol |
Stable transfectants w pEF1aBirAv5-neo plasmid
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| Growth protocol |
Cultured in RPMI 1640 medium w 10% FBS w 1 mg/ml G418
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
After crosslinking with 1% formaldehyde for 10 minutes, chromatin shearing was done by sonication in a Bioruptor UCD-200 (Diagenode, Liège, Belgium). The resulting lysate was divided into one aliquot for streptavidin capture (the equivalent of 9-10 million cells), another for Histone 3 K4 tri-methylation (H3K4me3; ab 8580, Abcam, Cambridge, UK) immunoprecipitation (the equivalent of 1-2 million cells) and a third aliquot for non-immunoprecipitated lysate to use as input control. The H3K4me3 immunoprecipitation and the input control preparation were done using the MagnaChip kit, according to the manufacturer’s instructions, increasing the reaction volumes to accommodate the input cell number. For the streptavidin capture, Dynabeads M-280 Streptavidin (Invitrogen) were used, according to the manufacturer’s recommendations with minor modifications. Briefly, beads were washed with PBS three times prior to use, 3 mg of beads were used for each sample, incubation was done at room temperature, and washing after incubation was done five times with PBS with 0.1% BSA. After washing, the streptavidin beads were resuspended in Chip Elution Buffer from the MagnaChip kit and all samples proceeded to Proteinase K treatment and DNA purification according to the MagnaChip protocol.
ThruPlex DNA-seq kit (Rubicon Genomics, Ann Arbor, Michigan, USA) according to manufacturer’s recommendations.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
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| Data processing |
Illumina RTA 1.17.21.3 was used for base-calling
ChIP-seq reads were aligned to hg19 using Bowtie2-2.2.7 with parameters –local –no-unal
Peaks where called using MACS2-2.1.1.20160226 with default settings
Genome_build: hg19
Supplementary_files_format_and_content: BED files with the found peaks
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| Submission date |
May 02, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Linnea Järvstråt |
| E-mail(s) |
[email protected]
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| Organization name |
Lund University
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| Department |
Department of Hematology and Transfusion Medicine
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| Lab |
Björn Nilsson
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| Street address |
Klinikgatan 26
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| City |
Lund |
| State/province |
Scania |
| ZIP/Postal code |
22184 |
| Country |
Sweden |
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| Platform ID |
GPL18573 |
| Series (1) |
| GSE81009 |
Global binding of isoforms of Wilms' Tumor gene 1 and H3K4me3 in K562 cells |
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| Relations |
| BioSample |
SAMN04931607 |
| SRA |
SRX1738501 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2140607_Streptavidin_pooled_exp2_peaks.bed.gz |
46.0 Kb |
(ftp)(http) |
BED |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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