|
| Status |
Public on Jul 14, 2023 |
| Title |
MLLAF9 screen control |
| Sample type |
SRA |
| |
|
| Source name |
AML
|
| Organism |
Mus musculus |
| Characteristics |
cell type: MLL-AF9
time point: day 9
|
| Treatment protocol |
K562 and MLLAF9 Cas9 cells (100x10^6 per cell line) were infected with the pooled lentiviral sgRNA libraries at a multiplicity of infection of 0.3. Infected cells were selected with 2ug/ml puromycin for 72hrs, commencing 48hrs after transduction. ~ 100x10^6 cells were fixed/sonicated and ChIP performed at day 6 (H3K4me3) and day 9 (H3K79me2).
|
| Growth protocol |
Cells were cultured in RPMI-1640 supplemented with 2mM Glutamax, 100IU/ml Penicillin, 100 ug/ml Streptomycin and 10%HI-FCS in 5% CO2 37degrees.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genomic DNA was extracted using Blood and tissue genomic DNA kit, Qiagen
One-step PCR was performed, containing adaptors for Illumina sequencing.
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
Custom targeted epigenetic library
|
| Data processing |
Library strategy: CRISPR-seq
Bcl2fastq (Illumina) was used to convert BCL files to Fastq files
Reads were trimmed to remove adapter with fastx clipper or cutadapt
Trimmed reads were aligned to the reference sgRNA library with bowtie2
After filtering to remove multi-aligning reads, the read counts were computed for each sgRNA.
The RSA algorithm was used to rank the genes for which targeting sgRNA were significantly enriched in the sorted populations compared to the control unsorted populations grown in parallel.
Supplementary_files_format_and_content: Guide Counts file
|
| |
|
| Submission date |
Dec 23, 2021 |
| Last update date |
Jul 14, 2023 |
| Contact name |
Mark Dawson |
| E-mail(s) |
mark.dawson@petermac.org
|
| Organization name |
Peter MacCallum Cancer Centre
|
| Department |
Cancer Research
|
| Street address |
305 Grattan Street
|
| City |
Melbourne |
| ZIP/Postal code |
3000 |
| Country |
Australia |
| |
|
| Platform ID |
GPL19057 |
| Series (2) |
| GSE192544 |
CRISPR-ChIP to identify mechanisms of chromatin regulation [CRISPR-ChIP] |
| GSE192562 |
CRISPR-ChIP to identify mechanisms of chromatin regulation |
|
| Relations |
| BioSample |
SAMN24367728 |
| SRA |
SRX13496587 |
