Comment[ArrayExpressAccession] E-GEOD-5650 Investigation Title Genomewide demarcation of RNA PolII transcription units by physical fractionation of chromatin- ORF-INT enrichment Comment[Submitted Name] Genomewide demarcation of RNA PolII transcription units by physical fractionation of chromatin- ORF-INT enrichment Publication Title Genomewide demarcation of RNA polymerase II transcription units revealed by physical fractionation of chromatin. Publication Author List Nagy PL, Cleary ML, Brown PO, Lieb JD Publication DOI PubMed ID 12750471 Experimental Design other Experimental Design Term Source REF Public Release Date 2006-08-30 Experiment Description Epigenetic modifications of chromatin serve an important role in regulating the expression and accessibility of genomic DNA. We report here a genomewide approach for fractionating yeast chromatin into two functionally distinct parts, one containing RNA polymerase II transcribed sequences, and the other comprising noncoding sequences and genes transcribed by RNA polymerases I and III. Noncoding regions could be further fractionated into promoters and segments lacking promoters. The observed separations were apparently based on differential crosslinking efficiency of chromatin in different genomic regions. The results reveal a genomewide molecular mechanism for marking promoters and genomic regions that have a license to be transcribed by RNA polymerase II, a previously unrecognized level of genomic complexity that may exist in all eukaryotes. Our approach has broad potential use as a tool for genome annotation and for the characterization of global changes in chromatin structure that accompany different genetic, environmental, and disease states. Keywords: Genomewide mapping of regulatory elements through differential fractionation of crosslinked chromatin based on nucleosome occupancy. ORF-INT enrichment- Whole cells were fixed by addition of 37% formaldehyde/11% methanol to the growth medium to a final concentration of 1% formaldehyde at 30°C for 30 min. Glycine was added to 125mM from a 2.5 M stock and incubated for 5 min. The cells were centrifuged in a Sorvall RT7 at 3,000 rpm for 5 min at 4°C and washed twice with PBS and once with sterile water. Without reversing crosslinks, extracts were prepared by glass-bead disruption, sonication (fragment size 200-2,000 bp, peak at 900 bp), and standard phenol-chloroform extraction. Date of Experiment Term Source Name EFO Term Source Version Term Source File http://efo.svn.sourceforge.net/viewvc/efo/trunk/src/efoinowl/efo.owl Person Last Name Lieb Person First Name Jason Person Mid Initials D Person Email jlieb@bio.unc.edu Person Affiliation University of North Carolina at Chapel Hill Person Phone 919-843-3229 Person Fax Person Address Biology, University of North Carolina at Chapel Hill, 203 Fordham Hall, Chapel Hill, NC, USA Person Roles submitter Person Roles Term Source REF Person Roles Term Accession Number Normalization Type Normalization Term Accession Number Normalization Term Source REF Replicate Type Replicate Term Accession Number Replicate Term Source REF Quality Control Type Quality Control Term Accession Number Quality Control Term Source REF Protocol Name P-GSE5650-1 P-GSE5650-2 Protocol Description ID_REF = ID_REF
CH1I_MEAN = Mean feature pixel intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale
CH2I_MEAN = Mean feature pixel intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale
CH1B_MEDIAN = The median feature background intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale; Channel: Cy3 Channel; Background
CH2B_MEDIAN = The median feature background intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale; Channel: Cy5 channel; Background
CH1D_MEAN = The mean feature pixel intensity at wavelength 532 nm with the median background subtracted.; Type: integer; Scale: linear_scale; Channel: Cy3 Channel
CH2D_MEAN = .The mean feature pixel intensity at wavelength 635 nm with the median background subtracted.; Type: integer; Scale: linear_scale; Channel: Cy5 channel
CH1I_MEDIAN = Median feature pixel intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale
CH2I_MEDIAN = Median feature pixel intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale
CH1B_MEAN = The mean feature background intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale; Background
CH2B_MEAN = The mean feature background intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale; Background
CH1D_MEDIAN = The median feature pixel intensity at wavelength 532 nm with the median background subtracted.; Type: integer; Scale: linear_scale
CH2D_MEDIAN = The median feature pixel intensity at wavelength 635 nm with the median background subtracted.; Type: integer; Scale: linear_scale
CH1_PER_SAT = The percentage of feature pixels at wavelength 532 nm that are saturated.; Type: integer; Scale: linear_scale
CH2_PER_SAT = The percentage of feature pixels at wavelength 635 nm that are saturated.; Type: integer; Scale: linear_scale
CH1I_SD = The standard deviation of the feature intensity at wavelength 532 nm.; Type: integer; Scale: linear_scale; Channel: Cy3 Channel
CH2I_SD = The standard deviation of the feature pixel intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale; Channel: Cy5 channel
CH1B_SD = The standard deviation of the feature background intensity at wavelength 532 nm.; Type: float; Scale: linear_scale; Channel: Cy3 Channel; Background
CH2B_SD = The standard deviation of the feature background intensity at wavelength 635 nm.; Type: integer; Scale: linear_scale; Channel: Cy5 channel; Background
PERGTBCH1I_1SD = The percentage of feature pixels with intensities more than one standard deviation above the background pixel intensity, at wavelength 532 nm.; Type: integer; Scale: linear_scale
PERGTBCH2I_1SD = The percentage of feature pixels with intensities more than one standard deviation above the background pixel intensity, at wavelength 635 nm.; Type: integer; Scale: linear_scale
PERGTBCH1I_2SD = The percentage of feature pixels with intensities more than two standard deviations above the background pixel intensity, at wavelength 532 nm.; Type: integer; Scale: linear_scale
PERGTBCH2I_2SD = The percentage of feature pixels with intensities more than two standard deviations above the background pixel intensity, at wavelength 532 nm.; Type: integer; Scale: linear_scale
SUM_MEAN = The sum of the arithmetic mean intensities for each wavelength, with the median background subtracted.; Type: integer; Scale: linear_scale
SUM_MEDIAN = The sum of the median intensities for each wavelength, with the median background subtracted.; Type: integer; Scale: linear_scale
RAT1_MEAN = Ratio of the arithmetic mean intensities of each spot for each wavelength, with the median background subtracted. Channel 1/Channel 2 ratio, (CH1I_MEAN - CH1B_MEDIAN)/(CH2I_MEAN - CH2B_MEDIAN) or Green/Red ratio.; Type: float; Scale: linear_scale
RAT2_MEAN = The ratio of the arithmetic mean intensities of each feature for each wavelength, with the median background subtracted.; Type: float; Scale: linear_scale
RAT2_MEDIAN = The ratio of the median intensities of each feature for each wavelength, with the median background subtracted.; Type: float; Scale: linear_scale
PIX_RAT2_MEAN = The geometric mean of the pixel-by-pixel ratios of pixel intensities, with the median background subtracted.; Type: float; Scale: linear_scale
PIX_RAT2_MEDIAN = The median of pixel-by-pixel ratios of pixel intensities, with the median background subtracted.; Type: float; Scale: linear_scale
RAT2_SD = The geometric standard deviation of the pixel intensity ratios.; Type: float; Scale: linear_scale
TOT_SPIX = The total number of feature pixels.; Type: integer; Scale: linear_scale
TOT_BPIX = The total number of background pixels.; Type: integer; Scale: linear_scale
REGR = The regression ratio of every pixel in a 2-feature-diameter circle around the center of the feature.; Type: float; Scale: linear_scale
CORR = The correlation between channel1 (Cy3) & Channel 2 (Cy5) pixels within the spot, and is a useful quality control parameter. Generally, high LOG_RAT2N_MEDIANs imply better fit & good spot quality.; Type: float; Scale: linear_scale
DIAMETER = The diameter in um of the feature-indicator.; Type: integer; Scale: linear_scale
X_COORD = X-coordinate of the center of the spot-indicator associated with the spot, where (0,0) is the top left of the image.; Type: integer; Scale: linear_scale
Y_COORD = Y-coordinate of the center of the spot-indicator associated with the spot, where (0,0) is the top left of the image.; Type: integer; Scale: linear_scale
TOP = Box top: int(((centerX - radius) - Xoffset) / pixelSize).; Type: integer; Scale: linear_scale
BOT = Box bottom: int(((centerX + radius) - Xoffset) / pixelSize).; Type: integer; Scale: linear_scale
LEFT = Box left: int(((centerY - radius) - yoffset) / pixelSize).; Type: integer; Scale: linear_scale
RIGHT = Box right: int(((centerY + radius) - yoffset) / pixelSize); Type: integer; Scale: linear_scale
FLAG = The type of flag associated with a feature: -100 = user-flagged null spot; -50 = software-flagged null spot; 0 = spot valid.; Type: integer; Scale: linear_scale
CH2IN_MEAN = Normalized LOG_RAT2N_MEDIAN of mean Channel 2 (usually 635 nm) intensity (CH2I_MEAN/Normalization factor).; Type: integer; Scale: linear_scale; Channel: Cy5 channel
CH2BN_MEDIAN = Normalized LOG_RAT2N_MEDIAN of median Channel 2 (usually 635 nm) background (CH2B_MEDIAN/Normalization factor).; Type: integer; Scale: linear_scale; Channel: Cy5 channel; Background
CH2DN_MEAN = Normalized LOG_RAT2N_MEDIAN of mean Channel 2 (usually 635 nm) intensity with normalized background subtracted (CH2IN_MEAN - CH2BN_MEDIAN).; Type: integer; Scale: linear_scale; Channel: Cy5 channel
RAT2N_MEAN = Type: float; Scale: linear_scale
CH2IN_MEDIAN = Normalized LOG_RAT2N_MEDIAN of median Channel 2 (usually 635 nm) intensity (CH2I_MEDIAN/Normalization factor).; Type: integer; Scale: linear_scale
CH2DN_MEDIAN = Normalized LOG_RAT2N_MEDIAN of median Channel 2 (usually 635 nm) intensity with normalized background subtracted (CH2IN_MEDIAN - CH2BN_MEDIAN).; Type: integer; Scale: linear_scale
RAT1N_MEAN = Ratio of the means of Channel 1 (usually 532 nm) intensity to normalized Channel 2 (usually 635 nm) intensity with median background subtracted (CH1D_MEAN/CH2DN_MEAN). Channel 1/Channel 2 ratio normalized or Green/Red ratio normalized.; Type: float; Scale: linear_scale
RAT2N_MEDIAN = Channel 2/Channel 1 ratio normalized, RAT2_MEDIAN/Normalization factor or Red/Green median ratio normalized.; Type: float; Scale: linear_scale
VALUE = Log (base 2) of the ratio of the mean of Channel 2 (usually 635 nm) to Channel 1 (usually 532 nm) [log (base 2) (RAT2N_MEAN)].; Type: float; Scale: log_base_2
LOG_RAT2N_MEDIAN = Log (base 2) of the ratio of the median of Channel 2 (usually 635 nm) to Channel 1 (usually 532 nm) [log (base 2) (RAT2N_MEDIAN)].; Type: float; Scale: log_base_2 Log (base 2) of the ratio of the mean of Channel 2 (usually 635 nm) to Channel 1 (usually 532 nm) Protocol Software Protocol Hardware Protocol Contact Protocol Type bioassay_data_transformation feature_extraction Protocol Term Source REF Protocol Term Accession Number Experimental Factor Name DNA PREPARED FROM IP INPUT, AMPLIFIED NOTE Experimental Factor Type DNA prepared from IP Input, Amplified NOTE Experimental Factor Term Source REF Experimental Factor Term Accession Number Comment[SecondaryAccession] GSE5650 Comment[GEOLastUpdateDate] 2009-03-15 Comment[GEOReleaseDate] 2006-08-29 Comment[ArrayExpressSubmissionDate] 2006-08-27 SDRF File E-GEOD-5650.sdrf.txt