|
| Status |
Public on Sep 27, 2023 |
| Title |
SEM_biol_rep_2_KMT2A |
| Sample type |
SRA |
| |
|
| Source name |
cell line
|
| Organism |
Homo sapiens |
| Characteristics |
tissue: cell line
cell line: Acute Lymphoblastic Leukemia cell line SEM
cell type: Paediatric pro B-cell line derived from ALL with t(4;11)(q21;q23) translocation
genotype: t(4;11)(q21;q23) translocation
treatment: None
chip antibody: KMT2A ( Bethyl labs A300-086-A lot. 6)
|
| Treatment protocol |
SEM and SEMPINORES cells were cultured in the presence of 50 µM of the DOT1L inhibitor pinometostat (EPZ5676, Selleckchem) for 1 week, with passage of the cells after 3 days with fresh inhibitor
|
| Growth protocol |
SEM and SEMPINORES cells were continuously grown in in RPMI-1640 medium containing GlutaMAXTM supplemented with 10% fetal calf serum, 100 IU/ml Penicillin and Streptomycin, and 0.125µg/ml Amphotericin B (Life Technologies), at 37°C under a 5% CO2 containing atmosphere. Cell lines passed every 3-4 days and routinely tested for the absence of mycoplasma and DNA fingerprinted to assure cell line authenticity.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Up to 20 million leukemic cells were crosslinked and lysed using the SimpleChIP kit (Cell Signaling Technology®, #9003) according to the manufacturer’s recommendations and subsequently sonicated using a Bioruptor sonicator (Diagenode) to generate 150-300 bp fragment size. Next, immunoprecipitation and antibody-protein-DNA precipitation were performed according to the guidelines of the manufacturer. Antibodies against KMT2A (Bethyl labs; cat.nr. A300-086-A), AFF1 (Abcam; cat.nr. ab31812), H3K4Me3, H3K27Ac and H3K79Me2 (Diagenode; cat. nrs. pAB-003-050, C15410196 and C15410051) were used. ChIP-seq DNA libraries were generated using the NEB Next Ultra DNA library preparation kit for Illumina (New England Biolabs) according to the manufacturer’s recommendations
ChIP-seq DNA libraries were generated using the NEB Next Ultra DNA library preparation kit for Illumina (New England Biolabs, E7645) according to the manufacturer’s recommendations
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
ChIPseq
|
| Data processing |
QC was performed with fastQC (https://www.bioinformatics.babraham.ac.uk/projects/fastqc/)
reads were trimmed using trim_galore (https://www.bioinformatics.babraham.ac.uk/projects/trim_galore/)
Reads were then aligned to the human genome, hg19, with bowtie2
Duplicate reads were removed using DeepTools alignmentSieve, with the flag –ignore duplicates
BigWigs were generated using the DeepTools bamCoverage command, with the flags –extendReads –normalize using RPKM
Assembly: hg19
Supplementary files format and content: bigWig
|
| |
|
| Submission date |
Apr 27, 2023 |
| Last update date |
Sep 27, 2023 |
| Contact name |
Ronald W. Stam |
| E-mail(s) |
r.w.stam@prinsesmaximacentrum.nl
|
| Organization name |
Prinses Maxima Centrum
|
| Street address |
Heidelberglaan 25
|
| City |
Utrecht |
| ZIP/Postal code |
3702 VK |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL18573 |
| Series (2) |
| GSE230806 |
Modelling acquired resistance to DOT1L inhibition exhibits the adaptive potential of KMT2A-rearranged Acute Lymphoblastic Leukemia’ [ChIP-seq] |
| GSE230807 |
Modelling acquired resistance to DOT1L inhibition exhibits the adaptive potential of KMT2A-rearranged Acute Lymphoblastic Leukemia’ |
|
| Relations |
| BioSample |
SAMN34411432 |
| SRA |
SRX20121530 |
