Antigen

Antigen Class

Input control

Antigen

Input control

Cell type

Cell type Class

Others

Cell type

Mesothelioma

MeSH Description

A tumor derived from mesothelial tissue (peritoneum, pleura, pericardium). It appears as broad sheets of cells, with some regions containing spindle-shaped, sarcoma-like cells and other regions showing adenomatous patterns. Pleural mesotheliomas have been linked to exposure to asbestos. (Dorland, 27th ed)

Sample

source_name

Mesothelioma tumor cells

tissue

Mesothelioma tumor cells

genotype

Nf2-/-;Cdkn2ab-/-

antibody

input

Sequenced DNA Library

library_strategy

ChIP-Seq

library_source

GENOMIC

library_selection

ChIP

library_construction_protocol

for RNA se: RNA was isolated from tumor cell lines and mouse tumors with the Qiagen All Prep DNA/RNA kit. Quantification and quality assessment for RNA were performed with a Bioanalyzer (RIN >6.5) (Agilent, Santa Clara, CA, USA). For DNAseq: Genomic DNA were isolated from frozen tumor sample using the Allprep DNA/RNA/Protein mini kit (Qiagen) according to the manufacturer's instructions. The amount of double stranded DNA in genomic DNA samples was quantified using the Qubit® dsDNA HS Assay Kit (Invitrogen). Subsequently, 250 ng of double stranded genomic DNA was fragmented by Covaris shearing and samples were purified with the Agencourt AMPure XP PCR Purification beads according to manufacturer's instructions (Beckman Coulter, cat no A63881). for ChIp seq: ChIP-Seq were performed for H3K27me3 and H2A119Ub1 with the protocol provided by Diagenode (iDeal ChIp-seq kit for histones Cat #C01010051). In brief, samples were crosslinked with formaldehyde for 10 minutes at room temperature and subsequently quenched with glycine. Samples lysed and sonicated for at least 20 cycles of 30 s On and 30 s Off using Diagenode Bioruptor Pico. For ChIP, 5 μg of antibody was conjugated with 50 μl of protein A magnetic beads. Immunoprecipitated DNA were processed for library preparation. for RNAseq: Sequencing libraries were constructed with a TruSeq mRNA Library Preparation Kit using poly-A-enriched RNA (Illumina, San Diego, CA, USA). The Samples were run on a HiSeq 2500 Illumina sequencer generating 65 base-pair single end reads. , for DNAseq: DNA library preparation for Illumina sequencing was done with the TruSeq® DNA LT Sample Preparation kit (Illumina). Up to ten uniquely indexed samples were pooled equimolarly, and each pool was then sequenced as single end 65 bp run using an Illumina HiSeq2500 machine according to manufacturer's instructions., for ChIpseq: DNA were processed for library preparation. Libraries were sequenced using Illumina HiSeq2500 genome analyzed (65bp, single end) and aligned to mm10 using TopHat with the default setting. Reads with mapping quality >20 were selected, peak calling over input control was performed using MACS2 .

Sequencing Platform

instrument_model

Illumina HiSeq 2500

mm10

Number of total reads

18171228

Reads aligned (%)

95.9

Duplicates removed (%)

23.8

Number of peaks

554 (qval < 1E-05)

mm9

Number of total reads

18171228

Reads aligned (%)

95.5

Duplicates removed (%)

23.8

Number of peaks

567 (qval < 1E-05)