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Sample GSM1847207 Query DataSets for GSM1847207
Status Public on Aug 17, 2015
Title Skud_2
Sample type SRA
 
Source name meiotic culture
Organism Saccharomyces kudriavzevii
Characteristics strain: ZP591
epitope tag: epitope tag: Spo11-Flag
meiotic time-point: 9 hr in meiosis
Growth protocol Cells were grown in YPA (1% yeast extract, 2% Bacto Peptone, 1% potassium acetate) for 14 hr at 23°C, harvested, resuspended to OD600=6.0 in 1% potassium acetate, 0.2 × supplements (2 μg/mL adenine, 2 μg/mL histidine, 6 μg/mL leucine, 2 μg/mL tryptophan, 2 μg/mL uracil) and sporulated at 23°C.
Extracted molecule genomic DNA
Extraction protocol Based on Pan et al. 2011 Cell 144:719-731. Whole cell extracts were prepared by grinding frozen cells in a bead mill and resuspending powder in 10%TCA. Pellets were then extracted with SDS and soluble protein was subjected to affinity purification of Spo11-Flag fusion with Protein-G Dynabeads. Protein was digested with proteinase K and Spo11-associated oligonucleotides were recovered by ethanol precipitation.
Based on Pan et al. 2011 Cell 144:719-731. Oligonucleotides were tailed with ribo-G using rGTP and terminal transferase, then ligated to duplex DNA adaptors with a dC overhang using T4 RNA ligase 2. Complementary strands were synthesized with Klenow, then purified by denaturing polyacrylamide electrophoresis. Purified complementary strands were subjected to 3' tailing with ribo-G using rGTP and terminal transferase, then another duplex adaptor was ligated using T4 RNA ligase 2. Ligated material was amplified with 16 cycles of PCR to add Illumina HiSeq adaptors
 
Library strategy OTHER
Library source genomic
Library selection other
Instrument model Illumina HiSeq 2500
 
Description Spo11-associated DNA oligonucleotides
Data processing Base calls perfomed using HiSeq Control Software v.2.0.10.0 for samples sequenced on Illumina HiSeq 2500
Adapters (AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC) were clipped from reads using fastx_clipper, part of the FASTX Toolkit 0.0.13.2, with parameters -l 15 -n -v -Q33 -i
First five bases (diffusion primer added in library prep) in each sequence were trimmed and trimmed sequence was recorded in the FASTQ ID tag for downstream tracking
Clipped reads were mapped using SHRiMP 2.1.1b with parameters -N 15 -U -g -1000 -q -1000 -m 10 -i -20 -h 100 -r 50% -n 1 -s 1111111111,11110111101111,1111011100100001111,1111000011001101111 -o 1001 -Q -E --sam-unaligned --strata
Sequences clipped from a read during mapping should be polyC on the 3' end or polyG on the 5' end. If clipped bases were not polyC or polyG the alignment score was corrected by deducting the mismatch score for each non-C,G base in the clipped sequence.
Maps of 5' ends of sequence reads were compiled and analyzed in R (RStudio version 0.98.1091, R version 3.0.1)
Genome_build: For wild-derived S. cerevisiae strains YPS128 and UWOPS03-461.4, reads were mapped to the SGD assembly released in June 2008, called "sacCer2" by UCSC: http://genome.ucsc.edu/cgi-bin/hgGateway?hgsid=340067645; for S. paradoxus YPS138, reads were mapped to the strain's genome assembly from SGRP (Saccharomyces Genome Resequencing Project, Wellcome Trust Sanger Institute); for S. mikatae IFO1815 and S. kudriavzevii ZP591, reads were mapped to the strains' genome assemblies from Scannell et al. 2011 G3 1: 11-25, available on http://www.saccharomycessensustricto.org.
Supplementary_files_format_and_content: wig files include RPM (reads per million mapped) for each sample at each non-zero base position in the genome
 
Submission date Aug 10, 2015
Last update date May 15, 2019
Contact name Isabel Lam
Organization name Memorial Sloan Kettering Cancer Center
Department Molecular Biology
Lab Keeney Lab
Street address 1275 York Avenue
City New York
State/province New York
ZIP/Postal code 10065
Country USA
 
Platform ID GPL20789
Series (2)
GSE71887 Spo11-oligo mapping in Saccharomyces species (S. paradoxus, S. mikatae, S. kudriavzevii) and wild-derived S. cerevisiae strains (YPS128, UWOPS03-461.4)
GSE71930 Non-paradoxical evolutionary stability of the recombination initiation landscape in Saccharomycetes
Relations
BioSample SAMN03979999
SRA SRX1141315

Supplementary file Size Download File type/resource
GSM1847207_20150526_Skud3_normalized_hitmap.wig.gz 3.1 Mb (ftp)(http) WIG
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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