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| Status |
Public on Sep 15, 2015 |
| Title |
DACOR1 |
| Sample type |
SRA |
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| Source name |
patient-derived colon cancer cell line induced with DACOR1
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| Organism |
Homo sapiens |
| Characteristics |
cancer type: colorectal cancer
Sex: Male
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| Treatment protocol |
The ChIRP-seq protocol was carried out as previously described by Chu et al. Briefly, 5 x 108 V852 cells with DACOR1 lentivirus were first crosslinked using 1% formaldehyde for 10 minutes. The cells were spun down, suspended in Buffer A (Hepes 20mM, KCl 10mM, MgCl2 1.5 mM, DTT 0.5mM, 1% Empigen), and dounced before collecting the nuclei by centrifugation. The nuclei were sonicated in nuclei lysis buffer (20mM Tris-HCl pH 7.5, 10mM EDTA , 1% SDS, 1mM DTT, protease inhibitor cocktail, RNaseOut 80 U/ml) to produce 100-500 bp DNA fragments. LiCl2 was added at 0.5M to nuclear lysates. Equal amounts of nuclear lysates were incubated with either DACOR1 specific or non-specific DNA probes modified with a TEG linker and Biotin at their 5’ ends, and incubated for 24 hour at 37°C with rotation. The next day, RiboMinus™ streptavidin-coated magnetic beads (Life Technologies) were blocked with 800 ug/ml yeast tRNA and 800 ug/ml BSA for 1 hour at 37°C in hybridization buffer (5mM Tris-HCl pH 7.5, 10mM EDTA, 500mM LiCl2) before washing and adding to nuclear lysates for 30 minutes. The beads were then washed three times with nuclear lysis buffer, wash buffer (5mM Tris-HCl pH 7.5, 0.5mM EDTA, 1M NaCl), and PBS. The beads were suspended in 200 ul of PBS and incubated at 75°C for 5 minutes, the supernatant was collected from the beads.
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| Growth protocol |
Grown in MEM media with 2% FBS, 2.4 mM L-glutamine, 10 ug/ml insulin, 2 ug/ml human transferrin, 2 ug/ml selenium, 1 ug/ml hydrocortisone, 50 ug/ml gentamicin
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
The supernatant was incubated at 65°C overnight to reverse crosslinking before extracting DNA using DNeasy Blood & Tissue Kit (Qiagen).
Illumina ChIP-seq library preparation kit
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| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina HiSeq 2500 |
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| Data processing |
100 bp paired-end DNA sequencing was performed on a HiSeq 2500 at Otogenetics Corporation.
DNA reads were mapped against human genome (hg19) using Bowtie 2.
Peak calling was performed by using MACS2.
Peak annotation was completed using ChIPpeakAnno.
Genomic hits were filtered based on a 15 or greater fold enrichment.
Genome_build: hg19
Supplementary_files_format_and_content: Tab-delinated text are provides DACOR1 ChIRP hits with genomic location, nearest gene, fold enrichment, p-value, and q-value
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| Submission date |
Aug 14, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Ahmad M Khalil |
| E-mail(s) |
amk175@case.edu
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| Organization name |
Case Western Reserve University
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| Department |
Genetics and Genome Sciences
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| Lab |
BRB719
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| Street address |
2109 Adelbert Rd
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| City |
Cleveland |
| State/province |
OH |
| ZIP/Postal code |
44106 |
| Country |
USA |
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| Platform ID |
GPL16791 |
| Series (1) |
| GSE58989 |
DNMT1-associated long non-coding RNA regulate global gene expression and DNA methylation in colon cancer |
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| Relations |
| BioSample |
SAMN03994230 |
| SRA |
SRX1150170 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1854427_DACOR1_ChIRP_analysis.txt.gz |
33.9 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Processed data are available on Series record |
| Raw data are available in SRA |
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