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Sample GSM702294 Query DataSets for GSM702294
Status Public on Dec 07, 2011
Title Cse4_5min_(20110115_6_Cse45)
Sample type SRA
 
Source name DNA from Cse4-FLAG chromatin immunoprecipitation
Organism Saccharomyces cerevisiae
Characteristics strain: Sby5146
medium: YEPD grown:OD=0.8
antibody: Sigma A2220 Anti Flag M2 Affinity gel; lot 020M6014
sample type: MNase-protected DNA fragments from Cse4-FLAG immunoprecipitation of native chromatin, 5 minute digestion
Extracted molecule genomic DNA
Extraction protocol Yeast nuclei were prepared as described [Furuyama, S., and Biggins, S. (2007), Centromere identity is specified by a single centromeric nucleosome in budding yeast, Proc Natl Acad Sci USA 104, 14706-14711], flash-frozen in liquid nitrogen and stored at -80oC. Nuclei were thawed at room temperature and digested with MNase followed by chromatin preparation as described [Furuyama and Biggins, (2007)], except that after MNase digestion, the slurry was passed four times through a 20 guage needle, then four times through a 26 guage needle [Jin, C., and Felsenfeld, G. (2007), Nucleosome stability mediated by histone variants H3.3 and H2A.Z, Genes Dev 21, 1519-1529], and the combined S1+S2 supernatants were clarified by centrifugation in a fixed-angle Sorvall SS34 rotor at 12,000 rpm at 4oC for 10 min at least twice or until no visible pellet remained. ChIP was performed as previously described [Furuyama and Biggins, (2007)], and DNA was extracted using a standard protocol [Mito, Y., Henikoff, J., and Henikoff, S. (2005), Genome-scale profiling of histone H3.3 replacement patterns, Nat Genet 37, 1090-1097]. A modified Illumina Solexa library protocol was used as described in supplementary file Solexa_libarary_protocol_GEO.pdf.
For Flag IP, we used Sigma A2220 Anti Flag M2 Affinity gel; lot 020M6014.
 
Library strategy MNase-Seq
Library source genomic
Library selection MNase
Instrument model Illumina HiSeq 2000
 
Description MNase-protected DNA fragments from Cse4-FLAG immunoprecipitation of native chromatin, 5 minute digestion
Illumina HiSeq 2000
Data processing 1. We used Novoalign to map paired-end reads to release 61 (UCSC sacCer2) of the S.cervisiae genomic sequence obtained from downloads.yeastgenome.org. 50bp reads were trimmed to 20bp. If a read was mapped to multiple locations, one location was picked at random. (Supplementary file Cse45.Novoalign.sam) 2. For each base pair in the genome, we counted the number of paired-end fragments aligned over it. 3. We normalized base pair counts by dividing by the total number of counts for all base pairs and then multiplying by the total number of base pairs in the genome. (Supplementary file Cse45.wig) 4. We broke down aligned paired-end fragments into sub-groups by insert size length and repeated steps 2. and 3. for the paired-end fragments in each sub-group.
 
Submission date Apr 06, 2011
Last update date Jun 11, 2013
Contact name Jorja Henikoff
E-mail(s) jorja@fhcrc.org
Phone 206-667-4850
Organization name Fred Hutchinson Cancer Research Center
Department Basic Sciences
Lab Henikoff
Street address 1100 Fairview AV N, A1-162
City Seattle
State/province WA
ZIP/Postal code 98109-1024
Country USA
 
Platform ID GPL13821
Series (1)
GSE28298 Tripartite organization of centromeric chromatin in budding yeast
Relations
BioSample SAMN02198539

Supplementary file Size Download File type/resource
GSM702294.wig.gz 44.9 Mb (ftp)(http) WIG
GSM702294_henikoff_Cse45.Novoalign.sam.gz 830.2 Mb (ftp)(http) SAM
Processed data provided as supplementary file
Raw data not provided for this record

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