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Sample GSM8636782

Query DataSets for GSM8636782

Status

Public on Nov 22, 2024

Title

Input

Sample type

SRA

Source name

Bladder

Organism

Homo sapiens

Characteristics

tissue: Bladder
cell line: UMUC3
cell type: Bladder cancer

Treatment protocol

Cells were treated with cigaretter smoke media (10%) for 3 days

Growth protocol

UMUC3 cells were grown in EMEM supplemented with 10% fetal bovine serum (FBS) and antibiotics in humidified atmosphere with 5% CO2 at 37 degree C

Extracted molecule

genomic DNA

Extraction protocol

The cells were harvested, lysed and ysates were immunoprecipitation with acK130-H2a antibodies (2 ug) for 3 hours. The magnetic beads were used to capture immunoprecipitates, washed and bound chromatin was eluted, followed by protease treatment.
Ten nanograms of immunoprecipitated DNA was fragmented to 300 base pairs using a Covaris M220 Focused-ultrasonicator (Covaris, Inc., Woburn, MA) and then used to generate sequencing libraries using the Kapa Hyper Prep Kit (Roche Sequencing Solutions Inc., Pleasanton, CA).

Library strategy

ChIP-Seq

Library source

genomic

Library selection

ChIP

Instrument model

Illumina NovaSeq 6000

Data processing

The size and quality of the library was evaluated using the Agilent BioAnalyzer (Agilent Technologies, Inc., Santa Clara, CA), and the library was quantitated with the Kapa Library Quantification Kit.
Each enriched DNA library was then sequenced on an Illumina NextSeq 500 sequencer to generate 40-50 million 75-base paired-end reads (Illumina, Inc., San Diego, CA).
The raw sequence data were aligned using BowTie 2 [1], and the binding sites were identified using the MACS peak-finding software
Raw fastq files were aligned against human genome hg38 assembly using Burrows-Wheeler Alignment tool and then processed using methylQA to generate bed and bigwig files.
The MACS2 peak caller was used to compare the ChIP-Seq signal to a corresponding input control to identify narrow regions of enrichment (peaks) using default parameters.
Assembly: ChIP-Seq signals and peak locations were further visualized using UCSC Genome Browser.
Supplementary files format and content: The top 500-1000 ChIP-Seq peaks were assigned to the nearest genes using the annotatePeaks function from HOMER2, motif binding sites were determined using HOMer2, and GO and pathway enrichment analyses were performed by EnrichR.
Supplementary files format and content: Assembly: hg38, mm10

Submission date

Nov 18, 2024

Last update date

Nov 22, 2024

Contact name

Tiandao Li

Organization name

Washington University

Street address

4444 Forest Park Ave

City

St Louis