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| Status |
Public on Apr 07, 2021 |
| Title |
NMYC_RB_1803_2_Input |
| Sample type |
SRA |
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| Source name |
NMYC_RB_Input
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| Organism |
Mus musculus |
| Characteristics |
genotype: Pb-Cre+; Pten f/f; Rb1 f/f; MYCN+
tissue: Mouse tumor
chip antibody: none
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| Treatment protocol |
Tumors were harvested and frozen in OCT. Frozen tumor blocks were sliced into 30 um thick cryosections.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Fifteen cryosections were collected and crosslinked with 1% methanol-free formaldehyde for 10 minutes and fixation was quenched using 2.5 M glycine for 5 minutes. The cell pellets were centrifuged and washed twice in cold PBS. Each pellet was resuspended in 1 ml of lysis buffer (50 mM Tris HCl pH 8, 0.5% SDS, 10 mM EDTA with protease and phosphatase inhibitors (Thermo Scientific)) and lysed for 20 minutes at 4°C. The nuclei were collected by centrifugation and resuspended in a second lysis buffer (10 mM Tris HCl pH 7.5, 150 mM NaCl, 0.1% SDS, 1 mM EDTA, 1% NP-40, 1% sodium deoxycholate, with protease and phosphatase inhibitors (Thermo Scientific)). The protein-bound chromatin was sheared by sonication (Diagenode, Bioruptor Pico). Input sheared chromatin was reserved as a control for downstream ChIP-seq analysis. Equal volumes of sheared chromatin were immunoprecipitated with specified antibodies.
Libraries were generated using the Hyper Prep Kit (Kapa Biosystems) following the manufacturer's recommended protocol.
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 4000 |
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| Data processing |
Quality control of raw sequencing reads was performed using FastQC
Low-quality reads were removed using Trimmomatic with a sliding window of 4bp and quality threshold of 20
Reads passing QC were aligned to mm10 using Bowtie2
BAM file creation, sorting, removal of PCR duplicates and indexing were performed using samtools
ChIP-seq peaks were called using MACS2 with input chromatin as the control for each set of data.
Genome_build: mm10
Supplementary_files_format_and_content: ChIP-seq signal (bigWig) called by MACS2 using input chromatin as control
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| Submission date |
May 29, 2020 |
| Last update date |
Apr 07, 2021 |
| Contact name |
Nicholas Brady |
| E-mail(s) |
njb2003@med.cornell.edu
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| Organization name |
Weill Cornell Medicine
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| Department |
Pathology and Laboratory Medicine
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| Street address |
1188 York Ave
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| City |
New York |
| State/province |
NY |
| ZIP/Postal code |
10065 |
| Country |
USA |
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| Platform ID |
GPL21103 |
| Series (2) |
| GSE151424 |
Temporal evolution of cellular heterogeneity during the progression to advanced, AR-negative prostate cancer [ChIP-seq] |
| GSE151426 |
Temporal evolution of cellular heterogeneity during the progression to advanced, AR-negative prostate cancer |
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| Relations |
| BioSample |
SAMN15053674 |
| SRA |
SRX8423451 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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