NCBI Logo
GEO Logo
   NCBI > GEO > Accession DisplayHelp Not logged in | LoginHelp
GEO help: Mouse over screen elements for information.
          Go
Sample GSM1591774 Query DataSets for GSM1591774
Status Public on Sep 10, 2015
Title BrdUIP_rif1∆
Sample type SRA
 
Source name KYP1328
Organism Schizosaccharomyces pombe
Characteristics genotype/variation: rif1delta
epitope-tag: adh1-Thymidine kinase
antibody: Anti-bromodeoxyuridine (MBL), Subclass; mouse IgG1, Clone; 2B1, Cat No.; MI-11-3,
strain: KYP1328
Treatment protocol Cells during G1 phase were fixed with 1% Formaldehyde at 20min after release.
Cells were arrested at early S-phase in 25mM HU at 60min after release from M-phase arrest and then the early replicated regions were labeled with 200µg/ml BrdU.
Growth protocol The stains were grown until mid log phase in YES medium at 30ºC. Cells containing nda3-KM311 mutation were arrested at M-phase at 20ºC for 5 hr. Cells were released into sequential cell cycle at 30ºC.
Extracted molecule genomic DNA
Extraction protocol The cell wall was disrupted by zirconia beads and multi-beads shocker (Yasui Kikai Co.). Lysates were prepared by sonication and clarified by centrifugation. Protein and DNA complexes were immuno-precipitated by antibody conjugated with Protein G Dynabeads.
BrdU-incorporated genomic DNA was purified by Qiagen Genomic-tip and genomic buffer set. Purified genomic DNA was sheared by sonication. The fragmented DNA was heat-denatured and BrdU-substituted DNA was immuno-precipitated by Anti-BrdU antibody conjugated with anti-IgG1a-dynabeads.
ChIP, BrdUIP, and Input DNA was sheared to around 150bp by S220 Focused-ultrasonicators (COVARIS, INC.). The fragmented DNA were end-repaired, ligated to sequencing adapters and amplified according to the protocol of NEBNext® ChIP-Seq Library Prep Master Mix Set and NEBNext Multiplex Oligos for Illumina (New England Biolabs) to prepare library. The size and concentration of libraries were checked by Bioanalyzer 2000 and real-time PCR using KAPA Library Quantification Kits, respectively.
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina MiSeq
 
Description rif1∆
BrdUIP_HU_rif1delta.bedgraph
Data processing All fastq files were aligned to S.pombe reference genomic ASM294v2 from Pombase by Bowtie1.0.0 with defalt setting.
The generated SAM files by Bowtie were converted to BAM and sorted by Samtools.
Peaks were called with Model-based analysis of ChIP-seq (MACS2.0.10) using following parameters; macs2 callpeak -t ChIP.sam -c Input.sam -f SAM -g 1.4e107 -n result_file -B -q 0.01 --nomodel
The generated treat_pileup.bedgraph files were loaded on Integrated Genome Browser.
Genome_build: ASM294v2
Supplementary_files_format_and_content: bedgraph
 
Submission date Jan 26, 2015
Last update date May 15, 2019
Contact name Yutaka Kanoh
E-mail(s) kanou-yt@igakuken.or.jp
Organization name Tokyo Metropolitan Institute of Medical Science
Department Genome Medicine
Lab Genome Dynamics Projec
Street address 2-1-6 Kamikitazawa, Setagaya-ku
City Tokyo
ZIP/Postal code 156-8506
Country Japan
 
Platform ID GPL16192
Series (1)
GSE65293 Rif1 binds to G-quadruplexes and suppresses replication over a long distances
Relations
BioSample SAMN03295237
SRA SRX853384

Supplementary data files not provided
SRA Run SelectorHelp
Processed data are available on Series record
Raw data are available in SRA

| NLM | NIH | GEO Help | Disclaimer | Accessibility |
NCBI Home NCBI Search NCBI SiteMap