|
| Status |
Public on Feb 28, 2013 |
| Title |
WT_B |
| Sample type |
SRA |
| |
|
| Source name |
WT [yIW160]
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
strain: WT [yIW160]
cell cycle stage: asynchronous culture
|
| Growth protocol |
Cells were grown at 30 degrees in YEP + 2% glucose to O.D. 0.1, and transferred to fresh medium containing either 2% glucose or 2% galactose for 4 hours. Doxycycline was subsequently added to 40ug/ml for 2.5 hours.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
DNA extraction was per the medium resolution double-strand break protocol in Murakami et al., 2009 (PMID 19799180). Adaptor ligation followed the protocol described in Smith and Whitehouse, 2012 (PMID 22419157). Genomic DNA was purified and heat-denatured: DNA was bound to Source 15Q resin in batch at 300mM NaCl, pH12, and eluted in 50mM steps. Fractions from 800-900 mM were collected, neutralized and concentrated by precipitation. Double-stranded sequencing adaptors with N6 random overhangs ligated to each end; after second-strand synthesis using Taq, material from 200-1000bp was gel-purified and amplified by PCR. Products from 200-500 bp were gel-purified from two successive 2.5% agarose gels.
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
Replicate wild type.
Okazaki fragments.
Columns in sgr files are as follows:
column 1: chromosome
column 2: base pair
column 3: sum of counts of Okazaki fragments overlapping this position
Columns in the average.tab file are as follows:
column 1: chromosome
column 2: base pair
column 3: base pair
column 4: Weighted Average Time
column 5: Sum of Probabilities in Weighted Average
Columns in replicationProfile.tab files:
column 1: chromosome
column 2: line segment's first base pair
column 3: line segment's first time
column 4: line segment's last base pair
column 5: line segment's last time in minutes
column 6: line segment's probability (0-1)
column 7: direction of replication fork progression
column 8: red value (red for forward blue for reverse)
column 9: green value (red for forward blue for reverse)
column 10: blue value (red for forward blue for reverse)
Columns in replicationProfile250bp.tab represent 250 bp interpolated sampling of replicationProfile.tab line segment values:
column 1: chromosome
column 2: base pair sampled by interpolation
column 3: time in minutes
column 4: probability (0-1)
column 5: direction of replication fork progression
column 6: red value (red for forward blue for reverse)
column 7: green value (red for forward blue for reverse)
column 8: blue value (red for forward blue for reverse)
The scripts in the SCRIPTS folder can be run inside of R when they are placed in the same folder as the replication profile files in order to plot the replication profiles per chromosome. Instructions for using the scripts are written in comments inside the scripts.
|
| Data processing |
Data for WT_A and WT_B were aligned to the May 2008 build of the S. cerevisiae genome using in-house software: paired-end reads each beginning with a 20bp perfect match to a unique genomic locus and within 1kb of one another were considered. The count of Okazaki fragments overlapping each base per DNA strand is reported.
Data for GAL_E, GAL_G, and GLU_F were aligned to the May 2008 build of the S. cerevisiae genome using tmap software using the command "tmap mapall -r $(cat fastqName.tab) -f $(cat yeastName.tab).fasta -a 0 -Q 0 -S 0 -n 10 stage1 -v map1 map2 map3 > TMAP/out.sam": single-end reads with a score greater than or equal to 20 matching a unique genomic locus were considered. The count of Okazaki fragments overlapping each base per DNA strand is reported.
For all data sets, Okazaki fragment counts per base per strand were processed using in-house software as described in the coinciding literature in order to produce replication profiles. Scripts are provided for viewing the data in R.
Genome_build: May 2008, supplemented by strain-appropriate recombinant DNA.
Supplementary_files_format_and_content: Strand-specific sgr file scores represent counts of Okazaki fragments that overlap each base. Tables (tab files) contain timing data calculated for forks moving in a specified direction at a specified probability or proportion of cell population most likely represented at such time. Table files named average.tab contain the average time of replication per base. Table files named replicationProfile.tab contain line segments representing fork motion in a specified direction from an origin to the most likely point of merger from a folded probability method described in the coinciding literature at the specified probability or proportion of cell population most likely represented at such times. Tables files named replicationProfile250bp.tab contain the same values as those in replicationProfile.tab but at intervals matching previously reported data. R script files are included with commented instructions for how to plot these values in R.
|
| |
|
| Submission date |
Sep 07, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Iestyn Whitehouse |
| E-mail(s) |
whitehoi@mskcc.org
|
| Organization name |
Sloan-Kettering Institute
|
| Department |
Molecular Biology
|
| Street address |
1275 York Avenue
|
| City |
New York |
| State/province |
New York |
| ZIP/Postal code |
10065 |
| Country |
USA |
| |
|
| Platform ID |
GPL13821 |
| Series (1) |
| GSE40696 |
Quantitative, genome-wide analysis of eukaryotic replication initiation and termination |
|
| Relations |
| Reanalysis of |
GSM835651 |
| SRA |
SRX185766 |
| BioSample |
SAMN01163780 |
