|
| Status |
Public on May 24, 2016 |
| Title |
HT10801b_CenpA_X_(20151015_2) |
| Sample type |
SRA |
| |
|
| Source name |
Immunoprecipitated DNA
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: fibrosarcoma
cell line: HT1080-1b
chip antibody: Anti-FLAG M2 magnetic beads (Signa Cat # M8823)
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
The HT1080-1b fibrosarcoma cell line was grown in RPMI medium using standard protocols. Cross-linking ChIP was performed as described previously (PMID: 24737864) with few modifications. Briefly, for each IP 200 million cells were fixed at a density equal to 2 million/ml using formaldehyde to a final concentration of 1%. Cross-linking was quenched with 125mM glycine. Chromatin was released from cells by incubating in lysis buffer (1% SDS, 10mM EDTA and 50 mM Tris-HCl (pH 8.1)) on ice for 10 min. The resulting chromatin solution was diluted 10 times with IP dilution buffer (1% Triton X-100, 2mM EDTA, 150 mM NaCl and 20mM Tris-HCl (pH8.1)) and digested with Micrococcal nuclease (MNase, Sigma, Cat # N3755) at 37°C for 45 min. MNase digested chromatin was sonicated as described before ( PMID: 24737864)and solubilized chromatin was precleared using protein A sepharose beads for 30 minutes at 4 °C. The precleared chromatin was subjected to crosslinking immunoprecipitation using Anti-FLAG-M2 magnetic beads (Sigma Cat # M8823).
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
MNase |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
HT1080-1b cross-linked ChIP CENP-A
|
| Data processing |
250bp single end reads were trimmed with trim-galore version 0.3.7 with parameters --quality 20 --adapter AGATCGGAAGAGC --stringency 3 --phred33 --length 25 and then aligned with various human alpha-satellite sequences using BWA 0.7.10 aln and sampe functions saving up to 10 alignments per pair, otherwise with default options. Trimmed reads mapped to two consensus sequences representing 20 human centromere sequences are included as supplementary bed files (Cen1-like and Cen13-like).
|
| |
|
| Submission date |
Dec 23, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Jorja Henikoff |
| E-mail(s) |
jorja@fhcrc.org
|
| Phone |
206-667-4850
|
| Organization name |
Fred Hutchinson Cancer Research Center
|
| Department |
Basic Sciences
|
| Lab |
Henikoff
|
| Street address |
1100 Fairview AV N, A1-162
|
| City |
Seattle |
| State/province |
WA |
| ZIP/Postal code |
98109-1024 |
| Country |
USA |
| |
|
| Platform ID |
GPL16791 |
| Series (1) |
| GSE69839 |
CENP-T and CENP-C bridge adjacent CENP-A nucleosomes on young alpha-satellite dimers |
|
| Relations |
| BioSample |
SAMN04364782 |
| SRA |
SRX1500120 |
