|
| Status |
Public on Feb 26, 2020 |
| Title |
p53_MUT8-COMB |
| Sample type |
SRA |
| |
|
| Source name |
Cell Line
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: Calu-1
genotype/variation: p53 R158G mutant
treatment: Belinostat
chip antibody: p53 (#9282, Cell Signaling)
|
| Treatment protocol |
Cell lines were cultured in petri dishes and treated with either 0.01% DMSO or 0.1uM belinostat with 10uM cisplatin for 48 hours.
|
| Growth protocol |
Cell lines were obtained directly from the American Type Culture Collection (ATCC), and were maintained in a humidified 37°C incubator, in culture medium (RPMI for Calu-1) supplemented with 10% fetal bovine serum, 2mM L-glutamine, 100 μg/mL streptomycin and 100 U/mL penicillin.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
In brief, cells were crosslinked with 1% formaldehyde, glycine quenched, lysed with cell lysis buffer (Sigma, #2978) supplemented with 1 mM PMSF, and subjected to pulse sonication (20s with 30s internal, 5 cycles). The supernatant was then immunoprecipitated with the p53 and rabbit IgG antibodies (Cell Signaling, #9282 and #2729) at 4°C overnight and incubated with Magna ChIP Protein G magnetic beads (Millipore, #16-662).
Libraries were prepared according to Illumina's instructions accompanying the DNA Sample Kit (Part# 0801-0303). Briefly, DNA was end-repaired using a combination of T4 DNA polymerase, E. coli DNA Pol I large fragment (Klenow polymerase) and T4 polynucleotide kinase. The blunt, phosphorylated ends were treated with Klenow fragment (32 to 52 exo minus) and dATP to yield a protruding 3- 'A' base for ligation of Illumina's adapters which have a single 'T' base overhang at the 3’ end. After adapter ligation DNA was PCR amplified with Illumina primers for 15 cycles and library fragments of ~250 bp (insert plus adaptor and PCR primer sequences) were band isolated from an agarose gel. The purified DNA was captured on an Illumina flow cell for cluster generation. Libraries were sequenced on the Illumina HiSeq following the manufacturer's protocols.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
S4
p53 R158G mutant cell line treated with Belinostat.
|
| Data processing |
bcl2fastq 2.17.1.14 were used for basecalling.
Sequenced reads were mapped to hg19 using bowtie v2.1.053 with parameters -N 23 1 –sensitive -p 2 –no-unal
Peaks were called using MACS2 v2.0.936.
Genome_build: hg19 (GRCh37)
Supplementary_files_format_and_content: ChIP-seq signal in bigwig for each sample.
|
| |
|
| Submission date |
Mar 21, 2019 |
| Last update date |
Feb 29, 2020 |
| Contact name |
Tuan Zea Tan |
| E-mail(s) |
csittz@nus.edu.sg
|
| Organization name |
National University of Singapore
|
| Department |
Cancer Science Institute Singapore
|
| Street address |
14 Medical Drive, #11-01, Singapore
|
| City |
Singapore |
| ZIP/Postal code |
117599 |
| Country |
Singapore |
| |
|
| Platform ID |
GPL11154 |
| Series (2) |
| GSE128673 |
Targeting codon 158 p53-mutant cancers via the induction of p53 acetylation [ChIP-seq] |
| GSE129027 |
Targeting codon 158 p53-mutant cancers via the induction of p53 acetylation |
|
| Relations |
| BioSample |
SAMN11191534 |
| SRA |
SRX5552347 |
