|
| Status |
Public on Jul 29, 2023 |
| Title |
sgBuGZ-1_SETD1A |
| Sample type |
SRA |
| |
|
| Source name |
MOLM-13;iCas9
|
| Organism |
Homo sapiens |
| Characteristics |
vector: iCas9-puro;sgBuGZ-1-GFP
chip antibody: SETD1A (#61702 Cell Signaling)
stimulation: Doxycycline
time: 4d
|
| Treatment protocol |
HA-FKBP-hSETD1A expression vector were introduced with lentivirus and stable expressing cell was established. Doxycycline-inducible Cas9 expression vector was introduced with lentivirus and stable expressing single cell clone was established. sgRNA-GFP vector was introduced with lentivirus and GFP-positive cells were sorted. GFP sorted cells were treated with doxycycline for 4 days.
|
| Growth protocol |
MOLM-13 cells were cultured in RPMI-1640 containing penicillin-streptomycin, supplemented with 10% fetal bovine serum.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
For ChIP-seq, cells were fixed with 2mM DSG and 1% formaldehyde, and samples from sonicated nuclear fraction were incubated with each antibody and histone-DNA complexes were isolated. For CUT&Tag, human MOLM-13 cells were mixed with 1/10 number of mouse MLL-AF9 transduced cells for spike-in control, and fixed with 0.1% formaldehyde. Cells were mixed with conA beads, and then incubated with SETD1A antibody.
ChIP-seq libraries were prepared using NEBNext ChIP-Seq Library Preparation Set for Illumina (New England Biolabs) following the manufacturer’s protocol. For CUT&Tag, tagmentation was performed using pA-Tn5 with adaptor complex.
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Description |
CUT&Tag
|
| Data processing |
library strategy: CUT&Tag
CUT&Tag library were sequenced with paired-end, but analyzed from read R1 fastq files.
BCL2FASTQ version1.8.4 software was used for basecalling.
ChIP-seq reads were aligned to hg19 reference human genome, using bowtie. CUT&Tag reads were aligned to hg19 and mm10.
The mapped sequence reads were extended to 500 bp and converted to the continuous signal data in wig format or converted to bigwig format using HOMER software. Wig files were converted to bigwig files using UCSC wigToBigWig program.
Assembly: hg19, mm10
|
| |
|
| Submission date |
Jun 15, 2023 |
| Last update date |
Jul 29, 2023 |
| Contact name |
Masaki Fukuyo |
| E-mail(s) |
fukuyo@chiba-u.jp
|
| Organization name |
Chiba University
|
| Department |
Department of Molecular Oncology
|
| Street address |
1-8-1 Inohana, Chuo-ku
|
| City |
Chiba |
| ZIP/Postal code |
260-8670 |
| Country |
Japan |
| |
|
| Platform ID |
GPL24676 |
| Series (2) |
| GSE159144 |
SETD1A function is mediated through interaction with mitotic regulators BuGZ/BUB3 in leukemia [ChIP-seq] |
| GSE159146 |
SETD1A function is mediated through interaction with mitotic regulators BuGZ/BUB3 in leukemia. |
|
| Relations |
| BioSample |
SAMN35743227 |
| SRA |
SRX20686964 |
