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Sample GSM426758 Query DataSets for GSM426758
Status Public on May 27, 2010
Title Embryonic Stem Cell LaminB1 1
Sample type genomic
 
Channel 1
Source name Dam-LaminB1 DamID DNA from Embryonic Stemm Cells
Organism Mus musculus
Characteristics strain: 129/ola
Extracted molecule genomic DNA
Extraction protocol 1-2x10e5 cells were infected with lenti-virus expressing either Dam-LaminB1 or only Dam. After 48 hours gDNA was isolated with the Qiagen Blood and Tissue kit per the manufacturers protocol. 2.5 μg of gDNA was digested with DpnI (New England Biolabs)), adapters were ligated with T4 ligase ((New England Biolabs)), and the ligation reaction was subsequently digested with DpnII ((New England Biolabs)). 500 ng of the DpnII digested material was used as input for a PCR amplification of methylated fragments (MePCR), see Vogel et al., 2008, Nature Protocols for details. MePCR fragments were purified by Qiaquick Spin columns (Qiagen).
Label Cy5
Label protocol 1.5 µg MePCR DNA was directly labeled by Klenow (New England Biolabs) random priming with Cy3/Cy5 nonamers per manufacturer's protocol (http://www.nimblegen.com/products/lit/lit.html). Labeling time was extended to 12 hours to increase yield.
 
Channel 2
Source name Dam DamID DNA from Embryonic Stemm Cells
Organism Mus musculus
Characteristics strain: 129/ola
Extracted molecule genomic DNA
Extraction protocol 1-2x10e5 cells were infected with lenti-virus expressing either Dam-LaminB1 or only Dam. After 48 hours gDNA was isolated with the Qiagen Blood and Tissue kit per the manufacturers protocol. 2.5 μg of gDNA was digested with DpnI (New England Biolabs)), adapters were ligated with T4 ligase ((New England Biolabs)), and the ligation reaction was subsequently digested with DpnII ((New England Biolabs)). 500 ng of the DpnII digested material was used as input for a PCR amplification of methylated fragments (MePCR), see Vogel et al., 2008, Nature Protocols for details. MePCR fragments were purified by Qiaquick Spin columns (Qiagen).
Label Cy3
Label protocol 1.5 µg MePCR DNA was directly labeled by Klenow (New England Biolabs) random priming with Cy3/Cy5 nonamers per manufacturer's protocol (http://www.nimblegen.com/products/lit/lit.html). Labeling time was extended to 12 hours to increase yield.
 
 
Hybridization protocol The labeled MePCR DNA was precipitated with 0.1 volume 5M NaCl and 1 volume isopropanol, 80 μg per sample was combined into 33.6 μl water, and mixed with 2.4 μl Alignment oligo, 24 μl hybridization component A, and 60 μl 2x Hybridization Buffer. The entire 120 μl was used for hybridization in Tecan hybridization stations for 16-18 h at 42C. Arrays were then washed in 42 °C Nimblegen washbuffer1, and room temperature Nimblegen washbuffer 2 and 3. All buffers and washes were completed using manufacturer's protocols (http://www.nimblegen.com/products/lit/lit.html)
Scan protocol Microarrays were scanned in a Agilent Scanner (Model G2505C, Serial number US22502518), which uses Scan Control software (Version A.8.1.3). Slides were first placed in a slide holder before putting it in the Scanner Carousel, which was installed in the scanner. Slides were double-pass scanned at maximum PMT for each channel, at a 2 um per pixel resolution.
Description none
Data processing Arrays were quantified with NimbleScan 2.5. Raw .pair files were read with the Ringo package for R and the loess normalized. Subsequently, single channel data was obtained and subjected to quantile-quantile normalisation over all available cell types and replicates.
 
Submission date Jul 10, 2009
Last update date Dec 20, 2018
Contact name Bas van Steensel
E-mail(s) b.v.steensel@nki.nl
Phone + 31 20 512 2040
Fax +31 20 669 1383
URL http://www.nki.nl/nkidep/vansteensel
Organization name Netherlands Cancer Institute
Department division of Molecular Biology
Lab van Steensel group
Street address Plesmanlaan 121
City Amsterdam
ZIP/Postal code 1066 CX
Country Netherlands
 
Platform ID GPL8840
Series (2)
GSE17051 Reorganization of nuclear lamina – genome interactions upon differentiation of embryonic stem cells.
GSE51334 DNA replication-timing boundaries separate stable chromosome domains with cell-type-specific functions
Relations
Reanalyzed by GSE97092

Data table header descriptions
ID_REF
VALUE The score was calculated as the log2 Dam-LaminB1: Dam ratio of the loess-quantile normalized data.

Data table
ID_REF VALUE
MUS00000000000000000000000859724 1.16511162394244
MUS00000000000000000000000859725 -0.00141956618871575
MUS00000000000000000000000859726 0.106166210462757
MUS00000000000000000000000859727 -0.423837039223675
MUS00000000000000000000000859728 -0.487724925425664
MUS00000000000000000000000859729 1.10723524074106
MUS00000000000000000000000859730 1.28440558903419
MUS00000000000000000000000859731 0.740342439068172
MUS00000000000000000000000859732 0.98231346495709
MUS00000000000000000000000859733 1.01459612473673
MUS00000000000000000000000859734 -0.454521276025874
MUS00000000000000000000000859735 0.426736331956484
MUS00000000000000000000000859736 -0.299550934863008
MUS00000000000000000000000859737 0.869105461009191
MUS00000000000000000000000859738 0.885282980029524
MUS00000000000000000000000859739 2.50807213909649
MUS00000000000000000000000859740 0.66036976443395
MUS00000000000000000000000859741 1.01013823317380
MUS00000000000000000000000859742 0.00481369502269935
MUS00000000000000000000000859743 0.0414827143633563

Total number of rows: 2121749

Table truncated, full table size 105936 Kbytes.




Supplementary file Size Download File type/resource
GSM426758_CMF_HD285098_Cy3_ESC1.pair.gz 43.2 Mb (ftp)(http) PAIR
GSM426758_CMF_HD285098_Cy5_ESC1.pair.gz 43.2 Mb (ftp)(http) PAIR
Processed data included within Sample table
Processed data are available on Series record

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