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Sample GSM5708380 Query DataSets for GSM5708380
Status Public on Dec 20, 2021
Title rpb1-T,FSP1, MNase-seq
Sample type SRA
 
Source name Whole yeast cells
Organism Saccharomyces cerevisiae
Characteristics growth protocol: Rich medium, 30°, log phase
genotype: rpb1-T,FSP1
sample: 9930-1-3 #1
Growth protocol Cultures were grown to logarithmic phase in rich medium at 30° C, treated with 1% formaldehyde for 20 minutes, then harvested by centrigugation.
Extracted molecule genomic DNA
Extraction protocol Glycine (final 0.2 M) was added to quench crosslinking and cells were washed with 1X Tris-buffered saline. Cell walls were removed with 500 U lyticase, 37° 20 minutes, or until >90% of cells were spheroplasted and collected by centrifugation. Cells were lysed in MNase digestion buffer and 10 U MNase, then incubated at 37° for 15 minutes, and the reaction stopped, diluted, and crosslinks reversed by incubating 5 hours at 65°. DNA was precipitated with ethanol, then treated with RNase A, followed by Proteinase K digestion and purification on Qiagen Qiaquick columns.
Sequencing libraries were constructed using NEBNext® Multiplex Oligos (Index Primers Set 1 and Set 2) and NEBNext® ChIP-seq Library Prep Reagent Set for Illumina. Ligated samples were subjected to 50 bp paired-end sequencing in an Illumina NovaSeq 6000 sequencer.
 
Library strategy MNase-Seq
Library source genomic
Library selection MNase
Instrument model Illumina NovaSeq 6000
 
Description MNase resistant fraction
rpb1-T-FSP.normal.bw
rpb1-T-FSP.skinny.bw
rpb1-T-FSP.subnuc.bw
Data processing Reads were aligned to the yeast genome version SacCer3 using Novocraft novoalign version 3.7.1, with adapter trimming, allowing one random multi-mapping alignment.
Duplicate alignments were randomly sub-sampled to a level of 40% relative to non-duplicate reads using bam_partial_dedup, version 1.7, https://github.com/tjparnell/HCI-Scripts/tree/master/BamFile.
Fragment coverage was generated using bam2wig version 1.52 (https://github.com/tjparnell/biotoolbox) with an extension of 150 bp, skipping chrMito and high-copy number features including telomeric and rDNA loci, depth normalized to Reads Per Million, averaging between replicates, and converting to bigWig format.
Genome_build: SacCer3
Supplementary_files_format_and_content: bigWig files representing the nucleosomal fragment coverage (140-170 bp) or subnucleosomal fragments (90-120 bp), normalized to Reads Per Million, and averaged between replicates were generated.
 
Submission date Nov 29, 2021
Last update date Dec 20, 2021
Contact name Tim Formosa
E-mail(s) [email protected]
Phone 8015815435
Organization name University of Utah School of Medicine
Department Biochemistry
Street address 15 N Medical Dr East RM 4100
City Salt Lake City
State/province UT
ZIP/Postal code 84112-5650
Country USA
 
Platform ID GPL27812
Series (2)
GSE184955 Spt6-tSH2 domain and Rpb1
GSE189831 The interaction between the Spt6-tSH2 domain and Rpb1 affects multiple functions of RNA Polymerase II [MNase-seq]
Relations
BioSample SAMN23496230
SRA SRX13257139

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

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