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Sample GSM346696 Query DataSets for GSM346696
Status Public on Feb 27, 2009
Title Nuclear scaffold and matrix attachment of HeLa chromatin (NaCl_2)
Sample type genomic
 
Channel 1
Source name Scaffold attachment profile of HeLa DNA: NaCl Loop Replicate 2
Organism Homo sapiens
Characteristics gender: female
cell type: cultured HeLa S3 cells
Extracted molecule genomic DNA
Extraction protocol Nuclear halos were prepared in solution from isolated HeLa nuclei with exposure to 2 M NaCl using approximately 1 x 107 cells. After extraction, the halos were pelleted at 1000 x g for 5 minutes at 4°C then washed gently in REact® 3 restriction buffer (Invitrogen, Carlsbad, CA) on a rocker platform for 20 minutes at room temperature then centrifuged at 1000 x g for 5 minutes at 4°C then the supernatant discarded. This washing procedure was repeated an additional two times. After the third wash, the halos were resuspended in restriction buffer and the loops separated from the nuclear matrix associated DNA by digestion with 400 U of EcoRI (Invitrogen, Carlsbad, CA) at 37°C for 3 hours. Subsequent to restriction digestion, the matrix fraction was pelleted by centrifugation at 16,000 x g for 5 minutes at 4°C and the loop containing supernatant was removed and placed in a separate tube. The matrix fraction was resuspended, and then washed in restriction buffer, immediately pelleted at 16,000 x g for 5 minutes at 4°C and supernatant discarded. The nuclear matrix containing pellet fraction was washed an additional two times. Both loop and matrix restriction fragments were then freed from any nuclear proteins by overnight digestion at 55°C with 50 µg/mL of proteinase K buffered with 50 mM Tris-HCl buffer, pH 8.0, containing 50 mM NaCl, 25 mM EDTA and 0.5% SDS. DNA was recovered and purified using a Quantum-prep matrix (BioRad, Hercules, CA) then resuspended in deionized water.
Label Cy3
Label protocol Labeled at NimbleGen facilities in accordance with manufacturer's protocol
 
Channel 2
Source name Scaffold attachment profile of HeLa DNA: NaCl Matrix Replicate 2
Organism Homo sapiens
Characteristics gender: female
cell type: cultured HeLa S3 cells
Extracted molecule genomic DNA
Extraction protocol Nuclear halos were prepared in solution from isolated HeLa nuclei with exposure to 2 M NaCl using approximately 1 x 107 cells. After extraction, the halos were pelleted at 1000 x g for 5 minutes at 4°C then washed gently in REact® 3 restriction buffer (Invitrogen, Carlsbad, CA) on a rocker platform for 20 minutes at room temperature then centrifuged at 1000 x g for 5 minutes at 4°C then the supernatant discarded. This washing procedure was repeated an additional two times. After the third wash, the halos were resuspended in restriction buffer and the loops separated from the nuclear matrix associated DNA by digestion with 400 U of EcoRI (Invitrogen, Carlsbad, CA) at 37°C for 3 hours. Subsequent to restriction digestion, the matrix fraction was pelleted by centrifugation at 16,000 x g for 5 minutes at 4°C and the loop containing supernatant was removed and placed in a separate tube. The matrix fraction was resuspended, and then washed in restriction buffer, immediately pelleted at 16,000 x g for 5 minutes at 4°C and supernatant discarded. The nuclear matrix containing pellet fraction was washed an additional two times. Both loop and matrix restriction fragments were then freed from any nuclear proteins by overnight digestion at 55°C with 50 µg/mL of proteinase K buffered with 50 mM Tris-HCl buffer, pH 8.0, containing 50 mM NaCl, 25 mM EDTA and 0.5% SDS. DNA was recovered and purified using a Quantum-prep matrix (BioRad, Hercules, CA) then resuspended in deionized water.
Label Cy5
Label protocol Labeled at NimbleGen facilities in accordance with manufacturer's protocol
 
 
Hybridization protocol Hybridized at NimbleGen facilities in accordance with NimbleGen protocol
Scan protocol Scanned at NimbleGen facilities in accordance with NimbleGen protocol
Description The nuclear scaffold/matrix provides an anchor for higher order genome structure that has both structural and functional implications. Different extraction protocols, i.e., utilizing either 25 mM LIS or 2 M NaCl, isolate somewhat different protein constituents of either the nuclear scaffold or nuclear matrix respectively. We have mapped, by array CGH, the locations of attachment to each of these residual protein bodies relative to non-attached DNA along the entire length of human chromosomes 14, 15, 16, 17 and 18 in HeLa cells.
Data processing Dual channel array data was q spline normalized using the NimbleGen data analysis suite. Normalized signal ratios were analyzed for enrichment in either the loop or matrix fraction.
 
Submission date Dec 01, 2008
Last update date Dec 31, 2009
Contact name Stephen A Krawetz
E-mail(s) steve@compbio.med.wayne.edu
Organization name Wayne State University
Street address 275 East Hancock, CS Mott Center
City Detroit
State/province MI
ZIP/Postal code 48207
Country USA
 
Platform ID GPL5820
Series (1)
GSE13774 Nuclear scaffold and matrix attachment of HeLa chromatin

Data table header descriptions
ID_REF
VALUE Normalized Loop/Normalized Matrix signal

Data table
ID_REF VALUE
CHR14P024717751 -0.195
CHR14P024718401 0.003
CHR14P024719156 0.051
CHR14P024719901 0.104
CHR14P024720554 0.032
CHR14P024721260 0.198
CHR14P024721998 -0.491
CHR14P024728902 -0.24
CHR14P024729612 -0.43
CHR14P024730375 -0.388
CHR14P024731054 -0.272
CHR14P024731714 0.027
CHR14P024732434 -0.067
CHR14P024733134 -0.556
CHR14P024733814 -0.268
CHR14P024734534 -0.615
CHR14P024735294 -0.589
CHR14P024735959 -0.32
CHR14P024736694 -0.148
CHR14P024737349 0.025

Total number of rows: 385573

Table truncated, full table size 8420 Kbytes.




Supplementary file Size Download File type/resource
GSM346696_12118102_01_532.pair.gz 6.2 Mb (ftp)(http) PAIR
GSM346696_12118102_01_635.pair.gz 6.2 Mb (ftp)(http) PAIR
Processed data included within Sample table

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