Antigen

Antigen Class

Unclassified

Antigen

Unclassified

Cell type

Cell type Class

Bone

Cell type

U2OS

Tissue

bone

Lineage

mesoderm

Description

osteosarcoma from the tibia of a 15 year old, J. Ponten and E. Saksela derived this line (originally 2T) in 1964 from a moderately differentiated sarcoma, viruses were not detected during co-cultivation with WI-38 cells or in CF tests against SV40, RSV or adenoviruses, mycoplasma contamination was detected and eliminated in 1972, (PMID: 6081590)

Sample

ENA checklist

ERC000011

INSDC center alias

University of Manchester

INSDC center name

University of Manchester

INSDC first public

2016-06-27T11:01:18Z

INSDC last update

2018-03-08T18:50:38Z

INSDC status

public

SRA accession

ERS496029

Sample Name

ERS496029

alias

E-MTAB-2701:FOXO3_R1

broker name

ArrayExpress

cell line

U2OS

cell type

osteosarcoma

organism

Homo sapiens

sex

female

title

FOXO3_R1

Sequenced DNA Library

library_name

FOXO3_R1

library_strategy

ChIP-Seq

library_source

GENOMIC

library_selection

ChIP

library_construction_protocol

After reverse cross-link and protein digestion with Proteinase K (Roche), DNA was purified using Qiaquick, PCR purification kit (Qiagen) following the manufacturer's instructions. The stable cell lines were grown in DMEM supplement with 10% foetal bovine serum. Media contained 100 micro-g/ml of Hygromycin. Expression of ZMYM2(WT) or ZMYM2(SIM2mut) was induced by treating the cells for 48 hours with 100 ng/ml [for ZMYM2(WT)] or 50 ng/ml [for ZMYM2(SIM2mut)] doxycycline (Sigma). Expression of FOXO3 was induced by treating the cells for 48 hours with 1 ng/ml doxycycline (Sigma). Cells were double cross-linked with dimethyl adipimidate (DMA) for 20 minutes and subsequently with Formaldehyde for 10 minutes. ChIP-seq was performed as described in Schmidt et al., 2009, using 60 x 106 U2OS-ZMYM2(WT) or U2OS-ZMYM2(SIM2mut). Immuno-precipitation was performed overnight at 4 degrees C by incubating the shared DNA-protein complex with 6 ug of anti-Flag (Sigma) or control IgG (Millipore) antibodies previously coupled to Dynabeads protein G (Invitrogen). Schmidt D, Wilson MD, Spyrou C, Brown GD, Hadfield J, Odom DT. (2009) ChIP-seq: using high-throughput sequencing to discover protein-DNA interactions. Methods 48: 240-248. Note: paired-end sequencing libraires were created in this experiment but only the forward (R1) reads are available in this submission. The 'LIBRARY_LAYOUT' attribute is therefore set to be 'SINGLE' rather than 'PAIRED'.

Sequencing Platform

instrument_model

Illumina HiSeq 2000

hg38

Number of total reads

32806296

Reads aligned (%)

98.9

Duplicates removed (%)

79.2

Number of peaks

1844 (qval < 1E-05)

hg19

Number of total reads

32806296

Reads aligned (%)

98.0

Duplicates removed (%)

80.9

Number of peaks

1560 (qval < 1E-05)