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Sample GSM1468968 Query DataSets for GSM1468968
Status Public on Aug 12, 2015
Title no-tag FLAG ChIP-seq
Sample type SRA
 
Source name fission yeast cells, WT, no-tag FLAG, FLAG ChIP
Organism Schizosaccharomyces pombe
Characteristics strain/background: SPO573
genotype/variation: h- ade6-M26 ura4-D18 ade6::ade6+-Padh15-skp1-AtTIR1-2NLS-9myc::hphMX, nmt81-Cnd2-IAA17::kanMX + pRep4
tagged protein: no-tag FLAG
culture condition: asynchronous cultures
chip antibody: mouse monoclonal anti-Flag M2 (Sigma, Cat. F3165, Lot. SLBH1191V)
Growth protocol Fission yeast cells (5 x 10^8 cells at 1 x 10^7 cells/ml) were grown at 30°C in YEA medium for Myc ChIP samples. For FLAG ChIP samples, cells were grown at 30°C in EMM medium and further cultured in EMM containing 15 mM thiamine and 0.5 mM auxin (1-naphthaleneacetic acid) for 2 hours (5 x 10^8 cells at 1 x 10^7 cells/ml), followed by pFA fixation. Using an auxin-based degron system (Kanke et al., 2011, BMC Cell Biol vol.12: 8), endogenous Cnd2 proteins were depleted by adding auxin and thiamine into culture medium. Wild-type and mutant Cnd2 proteins tagged with 3FLAG at their C-termini were continuously expressed from plasmids carrying the endogenous cnd2 promoter.
Extracted molecule genomic DNA
Extraction protocol Fission yeast cells grown at 30°C in YEA were shifted to 18°C for 5 min before 30 min fixation in 3% paraformaldehyde. Soluble chromatin fractions prepared from fixed cells were sonicated for 30 min by a Bioruptor (Diagenode). Tagged proteins were purified by each antibody and protein G-coupled Dynabeads (Life Technologies).
Libraries were prepared from 5 ng of DNA using the TruSeq DNA sample prep kit (Illumina) for Myc samples or using the NEBNext® ChIP-Seq Library Prep Master Mix Set for Illumina® with the NEBNext® Multiplex Oligos for Illumina® (index Primers Set 1) for FLAG samples, following manufacturer's protocol. Sequencing was performed by an Illumina Genome Analyzer II (Myc) or an Illumina HiSeq 2000 (FLAG).
 
Library strategy ChIP-Seq
Library source genomic
Library selection ChIP
Instrument model Illumina HiSeq 2000
 
Data processing Basecalls performed using Illumina CASAVA version 8.
ChIP-seq reads were aligned to the S. pombe genome (20090706; download from http://ebi.edu.au/ftp/databases/pombase/pombe/Chromosome_contigs/OLD/20090706/) using Maq version 0.7.1 for MyC samples or aligned to the S. pombe genome (ASM294v1.18; download from ftp://ftp.ensemblgenomes.org/pub/fungi/release-18/fasta/schizosaccharomyces_pombe/dna/) using BWA version 0.6.2 with the option -n 100 for FLAG samples.
Fragment size was estimated by using HOMER version 4.4.
The scores of the peaks were determined by the subtraction of control read density from treatment read density and calculate 200bp window average.
Genome_build: ASM294v1
Supplementary_files_format_and_content: wig files. Scores represent subtraction of control read density from treatment read density.
 
Submission date Aug 08, 2014
Last update date May 15, 2019
Contact name Hideki Tanizawa
E-mail(s) hidekit@uoregon.edu
Organization name University of Oregon
Department Institute of Molecular Biology
Street address 1370 Franklin Blvd
City Eugene
State/province OR
ZIP/Postal code 97403
Country USA
 
Platform ID GPL13988
Series (1)
GSE60273 Interaction between TBP and condensing drives the organization and faithful segregation of mitotic chromosomes
Relations
BioSample SAMN02980038
SRA SRX673293

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data not applicable for this record

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