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Sample GSM450280 Query DataSets for GSM450280
Status Public on Nov 25, 2009
Title Replication timing of EpiSC5, clone O
Sample type genomic
 
Channel 1
Source name Early-replicating DNA of EpiSC5, clone O
Organism Mus musculus
Characteristics cell line: EpiSC5, clone O
developmental stage: Epiblast-derived stem cells
Extracted molecule genomic DNA
Extraction protocol Replication timing data were obtained by hybridizing early and late replication intermediates to NimbleGen oligonucleotide arrays, as described in Hiratani et al [PLoS Biology (2008) 6: e245]. Briefly, replication intermediates are prepared from cells that are first pulse-labeled with BrdU and then sorted into early (1st half of S) and late (2nd half of S) stages of S-phase by flow cytometry, followed by anti-BrdU immunoprecipitation of the BrdU-substituted (nascent) replication intermediates that were synthesized either early or late during S-phase. Samples were labeled after unbiased amplification of recovered DNA by whole-genome amplification (WGA; Sigma, GenomePlex). Further details can be found in Hiratani et al [PLoS Biology (2008) 6: e245].
Label Cy3,Cy5
Label protocol Standard NimbleGen protocol (www.nimblegen.com/products/lit/index.html). Briefly, randomly-amplified BrdU-immunoprecipitated DNA samples (1 ug each) were Cy3- or Cy5-labeled by Klenow reaction using end-labeled Cy3- or Cy5-random 9-mers.
 
Channel 2
Source name Late-replicating DNA of EpiSC5, clone O
Organism Mus musculus
Characteristics cell line: EpiSC5, clone O
developmental stage: Epiblast-derived stem cells
Extracted molecule genomic DNA
Extraction protocol Replication timing data were obtained by hybridizing early and late replication intermediates to NimbleGen oligonucleotide arrays, as described in Hiratani et al [PLoS Biology (2008) 6: e245]. Briefly, replication intermediates are prepared from cells that are first pulse-labeled with BrdU and then sorted into early (1st half of S) and late (2nd half of S) stages of S-phase by flow cytometry, followed by anti-BrdU immunoprecipitation of the BrdU-substituted (nascent) replication intermediates that were synthesized either early or late during S-phase. Samples were labeled after unbiased amplification of recovered DNA by whole-genome amplification (WGA; Sigma, GenomePlex). Further details can be found in Hiratani et al [PLoS Biology (2008) 6: e245].
Label Cy5,Cy3
Label protocol Standard NimbleGen protocol (www.nimblegen.com/products/lit/index.html). Briefly, randomly-amplified BrdU-immunoprecipitated DNA samples (1 ug each) were Cy3- or Cy5-labeled by Klenow reaction using end-labeled Cy3- or Cy5-random 9-mers.
 
 
Hybridization protocol Standard NimbleGen protocol (www.nimblegen.com/products/lit/index.html). Briefly, Cy3 and Cy5 labeled DNA samples (6 ug each) were co-hybridized to Nimblegen CGH arrays containing one oligonucleotide probe every 5.8 kb across the mouse genome (Nimblegen, 2006-07-26_MM8_WG_CGH).
Scan protocol GenePix 4000B scanner (Molecular Devices) and GenePix software were used per NimbleGen's standard protocol (www.nimblegen.com/products/lit/index.html).
Description merge replicates 1 and 2
Data processing NimbleScan software was used to obtain .pair raw data per manufacturer's instructions. Raw early/late data (i.e. from .pair files) from two independent biological replicates in which early and late replicating DNA were labeled reciprocally were loess-normalized to remove signal intensity-dependent bias, scaled to a reference data set to have the same median absolute deviation and averaged (limma package, R/Bioconductor).The mean early/late ratios were used to generate a smoothed profile (i.e. processed data) using local polynomial smoothing (loess, 300-kb span) for each chromosome using basic functions in the statistical language R. Processed data sets can be graphically displayed (and are also downloadable) on our web site (http://www.replicationdomain.org). Further details can be found in Hiratani et al [PLoS Biology (2008) 6: e245].
 
Submission date Sep 04, 2009
Last update date Apr 16, 2013
Contact name David M. Gilbert
E-mail(s) gilbert@bio.fsu.edu
Phone 8506457583
Organization name Florida State University
Street address 319 Stadium Drive
City Tallahassee
State/province Florida
ZIP/Postal code 32306-4295
Country USA
 
Platform ID GPL9156
Series (4)
GSE17983 Replication Timing Reveal An Epigenetic Commitment To Differentiation Prior To Germ Layer Specification (WG_CGH, RT)
GSE18019 Genome-Wide Dynamics of Replication Timing Revealed by In Vitro Models of Mouse Embryogenesis
GSE49847 A comparative encyclopedia of DNA elements in the mouse genome

Data table header descriptions
ID_REF
VALUE Log2 transformed early/late replication timing ratio

Data table
ID_REF VALUE
CHR01FS003001832 -0.584110281
CHR01FS003018759 -0.466276154
CHR01FS003036253 -0.360754596
CHR01FS003041992 -0.329737336
CHR01FS003053606 -0.272407878
CHR01FS003065156 -0.22261519
CHR01FS003076536 -0.180596974
CHR01FS003087994 -0.145351067
CHR01FS003093673 -0.130552506
CHR01FS003105423 -0.105758884
CHR01FS003110919 -0.096866353
CHR01FS003116559 -0.089538616
CHR01FS003122452 -0.083832122
CHR01FS003134224 -0.07841813
CHR01FS003156928 -0.090630333
CHR01FS003162762 -0.098723201
CHR01FS003174487 -0.121070071
CHR01FS003180251 -0.134648608
CHR01FS003185980 -0.149587787
CHR01FS003191525 -0.165242791

Total number of rows: 384849

Table truncated, full table size 11047 Kbytes.




Supplementary file Size Download File type/resource
GSM450280_15483802_532.pair.gz 6.4 Mb (ftp)(http) PAIR
GSM450280_15483802_635.pair.gz 6.4 Mb (ftp)(http) PAIR
GSM450280_15513002_532.pair.gz 6.4 Mb (ftp)(http) PAIR
GSM450280_15513002_635.pair.gz 6.4 Mb (ftp)(http) PAIR
Processed data included within Sample table

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