title: HisH4 ChIP from Retinoic Acid Stimulated HL60 Cells, 2 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against H4-TetAc overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, H4-TetAc, HL60 Cells, 2 hours Retinoic Acid

source_name: H4-TetAc ChIP from Retinoic Acid Stimulated HL60 Cells, 2 hours


title: HisH4 ChIP from Retinoic Acid Stimulated HL60 Cells, 8 hours (NCBIv34)

status: Public on Jun 17 2005

type: genomic

city: Santa Clara

first: Thomas

middle: R

last: Gingeras

state: CA

address: 3380 Central Expressway

country: USA

institute: Affymetrix, Inc.

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against H4-TetAc overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, H4-TetAc, HL60 Cells, 8 hours Retinoic Acid

molecule: genomic DNA

organism: Homo sapiens

source_name: H4-TetAc ChIP from Retinoic Acid Stimulated HL60 Cells, 8 hours

source_name: Whole Cell Extract (WCE) DNA from Retinoic Acid Stimulated HL60 Cells, 8 hours


title: RARecA ChIP from Retinoic Acid Stimulated HL60 Cells, 2 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against RARecA overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, RARecA, HL60 Cells, 2 hours Retinoic Acid

source_name: Whole Cell Extract (WCE) DNA from Retinoic Acid Stimulated HL60 Cells, 2 hours


title: SIRT1 ChIP from Retinoic Acid Stimulated HL60 Cells, 8 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against SIRT1 overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, SIRT1, HL60 Cells, 8 hours Retinoic Acid


title: SIRT1 ChIP from Retinoic Acid Stimulated HL60 Cells, 32 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against SIRT1 overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, SIRT1, HL60 Cells, 32 hours Retinoic Acid

source_name: Whole Cell Extract (WCE) DNA from Retinoic Acid Stimulated HL60 Cells, 32 hours


title: Spleen, Male (MoSp.Male)

status: Public on Apr 04 2005

type: MPSS

city: Hayward

first: Daixing

last: Zhou

address: 25861 Industrial Blvd

department: Bioinformatics

institute: Solexa

description:
Strain: C57BL6
Spleen, Male total RNA (MoSp.Male) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoSp.Male.sig22
Cell type Spleen, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoSp.Male_sig22.5126F.a-20	10/11/2004	321176	6283	QC Passed
MoSp.Male_sig22.5126F.c-20	10/26/2004	331164	5770	QC Passed
MoSp.Male_sig22.5126W.b-20	8/14/2004	560691	10110	QC Passed
MoSp.Male_sig22.5126W.d-20	10/24/2004	496253	8016	QC Passed
MoSp.Male_sig22.5292F.d-20	2/18/2005	430141	6601	QC Passed
Run group:
Total Beads successfully sequenced - 2139425
Processed Signatures - 12146

molecule: total RNA

organism: Mus musculus

source_name: Spleen, Male total RNA


title: SkeletalMuscle:Thigh, Male (MoSM.MaTh)

description:
Strain: C57BL6
SkeletalMuscle:Thigh, Male total RNA (MoSM.MaTh) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoSM.MaTh.sig21
Cell type SkeletalMuscle:Thigh, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoSM.MaTh_sig21.5312F.a-20	12/21/2004	524744	5770	QC Passed
MoSM.MaTh_sig21.5312F.e-20	1/24/2005	518029	5223	QC Passed
MoSM.MaTh_sig21.5312W.c-20	1/10/2005	527843	7104	QC Passed
MoSM.MaTh_sig21.5417W.a-20	2/28/2005	545568	6582	QC Passed
Run group:
Total Beads successfully sequenced - 2116184
Processed Signatures - 9299

source_name: SkeletalMuscle:Thigh, Male total RNA


title: Pol2 ChIP from Retinoic Acid Stimulated HL60 Cells, 32 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against Pol2 overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, Pol2, HL60 Cells, 32 hours Retinoic Acid


title: PU1 ChIP from Retinoic Acid Stimulated HL60 Cells, 32 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against PU1 overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, PU1, HL60 Cells, 32 hours Retinoic Acid


title: P300 ChIP from Retinoic Acid Stimulated HL60 Cells, 0 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against P300 overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, P300, HL60 Cells, 0 hours Retinoic Acid

source_name: Whole Cell Extract (WCE) DNA from Retinoic Acid Stimulated HL60 Cells, 0 hours


title: Small intestine, Male (MoSI.Male)

description:
Strain: C57BL6
Small intestine, Male total RNA (MoSI.Male) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoSI.Male.sig22
Cell type Small intestine, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoSI.Male_sig22.5413W.a-20	2/28/2005	456974	9137	QC Passed
MoSI.Male_sig22.5244W.a-20	11/8/2004	602854	13552	QC Passed
MoSI.Male_sig22.5244F.a-20	11/3/2004	500990	10812	QC Passed
MoSI.Male_sig22.5184F.a-20	9/24/2004	308144	7577	QC Passed
MoSI.Male_sig22.5184F.b-20	9/21/2004	357465	9065	QC Passed
Run group:
Total Beads successfully sequenced - 2226427
Processed Signatures - 17866

source_name: Small intestine, Male total RNA


title: P300 ChIP from Retinoic Acid Stimulated HL60 Cells, 8 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against P300 overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, P300, HL60 Cells, 8 hours Retinoic Acid


title: Liver:right lobe, Female (MoLi.FeRL)

description:
Strain: C57BL6
Liver:right lobe, Female total RNA (MoLi.FeRL) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoLi.FeRL.sig23
Cell type Liver:right lobe, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoLi.FeRL_sig23.5411W.c-20	3/22/2005	598092	6428	QC Passed
MoLi.FeRL_sig23.5411F.b-20	2/23/2005	580104	6631	QC Passed
MoLi.FeRL_sig23.5411F.a-20	2/23/2005	589213	5793	QC Passed
MoLi.FeRL_sig23.5297W.a-20	12/10/2004	645483	5304	QC Passed
Run group:
Total Beads successfully sequenced - 2412892
Processed Signatures - 8605

source_name: Liver:right lobe, Female total RNA


title: Prostate, Male (MoPr.Male)

description:
Strain: C57BL6
Prostate, Male total RNA (MoPr.Male) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoPr.Male.sig21
Cell type Prostate, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoPr.Male_sig21.5314F.c-20	1/24/2005	551963	7282	QC Passed
MoPr.Male_sig21.5314F.d-20	1/28/2005	406873	5625	QC Passed
MoPr.Male_sig21.5314W.c-20	1/31/2005	476782	3453	QC Passed
MoPr.Male_sig21.5314W.d-20	1/31/2005	374776	3452	QC Passed
MoPr.Male_sig21.5314W.h-20	3/14/2005	538288	4593	QC Passed
Run group:
Total Beads successfully sequenced - 2348682
Processed Signatures - 8206

source_name: Prostate, Male total RNA


title: Brain:Cortical mantle, Male (MoBr.MaCM)

status: Public on Mar 02 2005

description:
Strain: C57BL6
Brain:Cortical mantle, Male total RNA (MoBr.MaCM) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoBr.MaCM.sig22
Cell type Brain:Cortical mantle, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoBr.MaCM_sig22.5324W.d-20	2/6/2005	423872	13250	QC Passed
MoBr.MaCM_sig22.5324F.b-20	1/19/2005	463112	14890	QC Passed
MoBr.MaCM_sig22.5324F.a-20	1/19/2005	548363	19111	QC Passed
MoBr.MaCM_sig22.5206W.a-20	10/12/2004	479773	18097	QC Passed
MoBr.MaCM_sig22.5206F.a-20	10/8/2004	258711	8641	QC Passed
Run group:
Total Beads successfully sequenced - 2173831
Processed Signatures - 25563

source_name: Brain:Cortical mantle, Male total RNA


title: Brain:Cerebellum, Female (MoBr.FeCb)

description:
Strain: C57BL6
Brain:Cerebellum, Female total RNA (MoBr.FeCb) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoBr.FeCb.sig21
Cell type Brain:Cerebellum, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoBr.FeCb_sig21.5400W.b-20	2/28/2005	602326	21733	QC Passed
MoBr.FeCb_sig21.5254F.g-20	1/6/2005	534044	19578	QC Passed
MoBr.FeCb_sig21.5254F.b-20	11/15/2004	425563	16937	QC Passed
MoBr.FeCb_sig21.5254W.a-20	11/9/2004	637236	20102	QC Passed
Run group:
Total Beads successfully sequenced - 2199169
Processed Signatures - 31523

source_name: Brain:Cerebellum, Female total RNA


title: Brain:Hippocampus, Female (MoBr.FeHi)

description:
Strain: C57BL6
Brain:Hippocampus, Female total RNA (MoBr.FeHi) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoBr.FeHi.sig21
Cell type Brain:Hippocampus, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoBr.FeHi_sig21.5420F.d-20	3/18/2005	489602	16681	QC Passed
MoBr.FeHi_sig21.5420F.f-20	3/31/2005	399267	12775	QC Passed
MoBr.FeHi_sig21.5420F.g-20	3/31/2005	381118	12185	QC Passed
MoBr.FeHi_sig21.5420W.b-20	2/28/2005	440070	12758	QC Passed
MoBr.FeHi_sig21.5420W.c-20	3/22/2005	495684	18076	QC Passed
MoBr.FeHi_sig21.5420W.d-20	3/28/2005	456966	12301	QC Passed
Run group:
Total Beads successfully sequenced - 2662707
Processed Signatures - 25456

source_name: Brain:Hippocampus, Female total RNA


title: Brain:Amygdala, Female (MoBr.FeAm)

description:
Strain: C57BL6
Brain:Amygdala, Female total RNA (MoBr.FeAm) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoBr.FeAm.sig23
Cell type Brain:Amygdala, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoBr.FeAm_sig23.5291F.a-20	12/21/2004	420704	10097	QC Passed
MoBr.FeAm_sig23.5291F.d-20	1/10/2005	406312	9513	QC Passed
MoBr.FeAm_sig23.5291W.a-20	12/20/2004	349156	12290	QC Passed
MoBr.FeAm_sig23.5291W.b-20	12/22/2004	304943	9255	QC Passed
MoBr.FeAm_sig23.5412F.a-20	3/22/2005	466176	12690	QC Passed
MoBr.FeAm_sig23.5412W.a-20	2/28/2005	609044	14326	QC Passed
Run group:
Total Beads successfully sequenced - 2556335
Processed Signatures - 19342

source_name: Brain:Amygdala, Female total RNA


title: Placenta - E18, Female (MoPl.FE18)

description:
Strain: C57BL6
Placenta - E18, Female total RNA (MoPl.FE18) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoPl.FE18.sig21
Cell type Placenta - E18, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoPl.FE18_sig21.5208F.a-20	10/7/2004	593548	11333	QC Passed
MoPl.FE18_sig21.5208F.b-20	10/5/2004	600437	12576	QC Passed
MoPl.FE18_sig21.5208W.c-20	11/19/2004	672002	14842	QC Passed
MoPl.FE18_sig21.5322W.b-20	1/31/2005	484199	9584	QC Passed
MoPl.FE18_sig21.5322W.d-20	2/18/2005	556369	11548	QC Passed
Run group:
Total Beads successfully sequenced - 2906555
Processed Signatures - 20318

source_name: Placenta - E18, Female total RNA


title: Brain:Olfactory Bulb, Male (MoBr.MaOB)

description:
Strain: C57BL6
Brain:Olfactory Bulb, Male total RNA (MoBr.MaOB) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoBr.MaOB.sig22
Cell type Brain:Olfactory Bulb, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoBr.MaOB_sig22.5213F.b-20	11/19/2004	373480	15817	QC Passed
MoBr.MaOB_sig22.5213W.a-20	10/13/2004	348766	12963	QC Passed
MoBr.MaOB_sig22.5367F.a-20	2/22/2005	604542	25078	QC Passed
MoBr.MaOB_sig22.5367W.a-20	1/27/2005	384602	16752	QC Passed
MoBr.MaOB_sig22.5367W.d-20	2/23/2005	455157	25667	QC Passed
Run group:
Total Beads successfully sequenced - 2166547
Processed Signatures - 34415

source_name: Brain:Olfactory Bulb, Male total RNA


title: HisH4 ChIP from Retinoic Acid Stimulated HL60 Cells, 32 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against H4-TetAc overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, H4-TetAc, HL60 Cells, 32 hours Retinoic Acid

source_name: H4-TetAc ChIP from Retinoic Acid Stimulated HL60 Cells, 32 hours


title: SkeletalMuscle:Thigh, Female (MoSM.FeTh)

description:
Strain: C57BL6
SkeletalMuscle:Thigh, Female total RNA (MoSM.FeTh) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoSM.FeTh.sig21
Cell type SkeletalMuscle:Thigh, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoSM.FeTh_sig21.5418F.a-20	2/28/2005	465810	4777	QC Passed
MoSM.FeTh_sig21.5418F.d-20	3/22/2005	449998	5537	QC Passed
MoSM.FeTh_sig21.5418F.e-20	3/23/2005	346950	3295	QC Passed
MoSM.FeTh_sig21.5418W.a-20	2/28/2005	510428	6544	QC Passed
MoSM.FeTh_sig21.5418W.c-20	3/14/2005	492804	6236	QC Passed
Run group:
Total Beads successfully sequenced - 2265990
Processed Signatures - 8867

source_name: SkeletalMuscle:Thigh, Female total RNA


title: Brain:Midbrain, Female (MoBr.FeMb)

description:
Strain: C57BL6
Brain:Midbrain, Female total RNA (MoBr.FeMb) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoBr.FeMb.sig21
Cell type Brain:Midbrain, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoBr.FeMb_sig21.5287F.b-20	12/8/2004	383799	9951	QC Passed
MoBr.FeMb_sig21.5366F.a-20	1/27/2005	395313	13065	QC Passed
MoBr.FeMb_sig21.5366F.b-20	1/27/2005	464797	13860	QC Passed
MoBr.FeMb_sig21.5366W.b-20	1/28/2005	520335	16443	QC Passed
MoBr.FeMb_sig21.5366W.c-20	2/23/2005	653725	17394	QC Passed
Run group:
Total Beads successfully sequenced - 2417969
Processed Signatures - 23595

source_name: Brain:Midbrain, Female total RNA


title: Lymph nodes: mesenteric, Female (MoLN.FeMs)

description:
Strain: C57BL6
Lymph nodes: mesenteric, Female total RNA (MoLN.FeMs) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoLN.FeMs.sig21
Cell type Lymph nodes: mesenteric, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoLN.FeMs_sig21.5325F.b-20	1/24/2005	605810	10326	QC Passed
MoLN.FeMs_sig21.5325F.a-20	1/22/2005	576408	9848	QC Passed
MoLN.FeMs_sig21.5325W.a-20	1/18/2005	525495	9285	QC Passed
MoLN.FeMs_sig21.5129W.b-20	8/12/2004	474185	9246	QC Passed
Run group:
Total Beads successfully sequenced - 2181898
Processed Signatures - 14235


title: Brain:Cortical mantle, Female (MoBr.FeCM)

description:
Strain: C57BL6
Brain:Cortical mantle, Female total RNA (MoBr.FeCM) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoBr.FeCM.sig22
Cell type Brain:Cortical mantle, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoBr.FeCM_sig22.5357W.b-20	1/28/2005	666803	20715	QC Passed
MoBr.FeCM_sig22.5357W.a-20	1/27/2005	471595	15196	QC Passed
MoBr.FeCM_sig22.5357F.a-20	1/26/2005	349794	20166	QC Passed
MoBr.FeCM_sig22.5246F.b-20	11/9/2004	524890	16133	QC Passed
MoBr.FeCM_sig22.5246F.a-20	11/9/2004	379323	14409	QC Passed
Run group:
Total Beads successfully sequenced - 2392405
Processed Signatures - 28120

source_name: Brain:Cortical mantle, Female total RNA


title: HisH4 ChIP from Retinoic Acid Stimulated HL60 Cells, 0 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against H4-TetAc overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, H4-TetAc, HL60 Cells, 0 hours Retinoic Acid

source_name: H4-TetAc ChIP from Retinoic Acid Stimulated HL60 Cells, 0 hours


title: H3K27T ChIP from Retinoic Acid Stimulated HL60 Cells, 8 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against H3K27TMe overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, H3K27TMe, HL60 Cells, 8 hours Retinoic Acid

source_name: H3K27TMe ChIP from Retinoic Acid Stimulated HL60 Cells, 8 hours

source_name: Whole Cell Extract (WCE) DNA from Retinoic Acid Stimulated HL60 Cells, 8 hours


title: H3K27T ChIP from Retinoic Acid Stimulated HL60 Cells, 2 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against H3K27TMe overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, H3K27TMe, HL60 Cells, 2 hours Retinoic Acid

source_name: H3K27TMe ChIP from Retinoic Acid Stimulated HL60 Cells, 2 hours

source_name: Whole Cell Extract (WCE) DNA from Retinoic Acid Stimulated HL60 Cells, 2 hours


title: Brown Fat, Male (MoBF.Male)

description:
Strain: C57BL6
Brown Fat, Male total RNA (MoBF.Male) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoBF.Male.sig21
Cell type Brown Fat, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoBF.Male_sig21.5209F.a-20	10/19/2004	364704	6310	QC Passed
MoBF.Male_sig21.5209W.a-20	10/18/2004	544420	6890	QC Passed
MoBF.Male_sig21.5209W.c-20	11/15/2004	621278	8371	QC Passed
MoBF.Male_sig21.5364F.a-20	1/26/2005	509470	8766	QC Passed
MoBF.Male_sig21.5364F.b-20	2/18/2005	402025	6076	QC Passed
Run group:
Total Beads successfully sequenced - 2441897
Processed Signatures - 12633

source_name: Brown Fat, Male total RNA


title: Brain:Caudate,Putamen,Accumbens,BasalForebrain, Female (MoBr.FCPA)

description:
Strain: C57BL6
Brain:Caudate,Putamen,Accumbens,BasalForebrain, Female total RNA (MoBr.FCPA) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoBr.FCPA.sig22
Cell type Brain:Caudate,Putamen,Accumbens,BasalForebrain, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoBr.FCPA_sig22.5326W.d-20	2/15/2005	389456	12374	QC Passed
MoBr.FCPA_sig22.5326W.a-20	1/14/2005	552695	18211	QC Passed
MoBr.FCPA_sig22.5326F.b-20	1/13/2005	473852	15214	QC Passed
MoBr.FCPA_sig22.5326F.a-20	1/13/2005	438895	15493	QC Passed
MoBr.FCPA_sig22.5211F.a-20	10/18/2004	434148	17293	QC Passed
Run group:
Total Beads successfully sequenced - 2289046
Processed Signatures - 25736


title: Adrenal, Male (MoAd.Male)

description:
Strain: C57BL6
Adrenal, Male total RNA (MoAd.Male) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoAd.Male.sig21
Cell type Adrenal, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoAd.Male_sig21.5365W.a-20	1/31/2005	595782	15050	QC Passed
MoAd.Male_sig21.5202W.d-20	11/2/2004	482653	11219	QC Passed
MoAd.Male_sig21.5202F.c-20	11/1/2004	525010	10632	QC Passed
MoAd.Male_sig21.5202F.b-20	10/18/2004	639412	17318	QC Passed
Run group:
Total Beads successfully sequenced - 2242857
Processed Signatures - 20609

source_name: Adrenal, Male total RNA


title: CTCF ChIP from Retinoic Acid Stimulated HL60 Cells, 2 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against CTCF overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, CTCF, HL60 Cells, 2 hours Retinoic Acid


title: Heart:Aorta, Female (MoAo.Feml)

description:
Strain: C57BL6
Heart:Aorta, Female total RNA (MoAo.Feml) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoAo.Feml.sig21
Cell type Heart:Aorta, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoAo.Feml_sig21.5327W.b-20	1/31/2005	580600	13050	QC Passed
MoAo.Feml_sig21.5178F.c-20	11/3/2004	302649	7005	QC Passed
MoAo.Feml_sig21.5178F.a-20	9/27/2004	300296	6798	QC Passed
MoAo.Feml_sig21.5178F.b-20	9/22/2004	351163	7283	QC Passed
MoAo.Feml_sig21.5178W.a-20	9/22/2004	371143	9091	QC Passed
MoAo.Feml_sig21.5178W.b-20	9/22/2004	357657	10197	QC Passed
Run group:
Total Beads successfully sequenced - 2263508
Processed Signatures - 16234

source_name: Heart:Aorta, Female total RNA


title: Brown Fat, Female (MoBF.Feml)

description:
Strain: C57BL6
Brown Fat, Female total RNA (MoBF.Feml) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoBF.Feml.sig21
Cell type Brown Fat, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoBF.Feml_sig21.5328F.c-20	2/2/2005	401368	9214	QC Passed
MoBF.Feml_sig21.5328W.a-20	1/20/2005	634258	11470	QC Passed
MoBF.Feml_sig21.5328F.b-20	1/19/2005	443075	9754	QC Passed
MoBF.Feml_sig21.5219F.d-20	11/21/2004	211953	4106	QC Passed
MoBF.Feml_sig21.5219W.b-20	10/12/2004	366808	7483	QC Passed
Run group:
Total Beads successfully sequenced - 2057462
Processed Signatures - 14859

source_name: Brown Fat, Female total RNA


title: CTCF ChIP from Retinoic Acid Stimulated HL60 Cells, 32 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against CTCF overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, CTCF, HL60 Cells, 32 hours Retinoic Acid

source_name: Whole Cell Extract (WCE) DNA from Retinoic Acid Stimulated HL60 Cells, 32 hours


title: Lung, Male (MoLu.Male)

status: Public on Jan 28 2005

description:
Strain: C57BL6
Lung, Male total RNA (MoLu.Male) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoLu.Male.sig21
Cell type Lung, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoLu.Male_sig21.5214W.e-20	12/7/2004	641693	15972	QC Passed
MoLu.Male_sig21.5214W.d-20	12/6/2004	444752	9421	QC Passed
MoLu.Male_sig21.5214W.b-20	11/24/2004	431247	8805	QC Passed
MoLu.Male_sig21.5214F.c-20	11/20/2004	356336	6807	QC Passed
MoLu.Male_sig21.5214F.a-20	10/12/2004	369931	7566	QC Passed
Run group:
Total Beads successfully sequenced - 2243959
Processed Signatures - 16526

source_name: Lung, Male total RNA


title: Small intestine, Female (MoSI.Feml)

description:
Strain: C57BL6
Small intestine, Female total RNA (MoSI.Feml) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoSI.Feml.sig21
Cell type Small intestine, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoSI.Feml_sig21.5350F.a-20	1/22/2005	555757	11273	QC Passed
MoSI.Feml_sig21.5350W.a-20	1/18/2005	494687	9053	QC Passed
MoSI.Feml_sig21.5180W.d-20	11/23/2004	473824	11813	QC Passed
MoSI.Feml_sig21.5180F.d-20	11/19/2004	484120	9090	QC Passed
MoSI.Feml_sig21.5180F.b-20	9/21/2004	411334	8832	QC Passed
Run group:
Total Beads successfully sequenced - 2419722
Processed Signatures - 16808

source_name: Small intestine, Female total RNA


title: H3K27T ChIP from Retinoic Acid Stimulated HL60 Cells, 32 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against H3K27TMe overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, H3K27TMe, HL60 Cells, 32 hours Retinoic Acid

source_name: H3K27TMe ChIP from Retinoic Acid Stimulated HL60 Cells, 32 hours


title: Cervix and vagina, Female (MoCV.Feml)

description:
Strain: C57BL6
Cervix and vagina, Female total RNA (MoCV.Feml) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoCV.Feml.sig23
Cell type Cervix and vagina, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoCV.Feml_sig23.5313F.f-20	1/24/2005	614465	7898	QC Passed
MoCV.Feml_sig23.5313F.a-20	1/3/2005	481835	6324	QC Passed
MoCV.Feml_sig23.5313W.b-20	12/24/2004	513024	9308	QC Passed
MoCV.Feml_sig23.5313W.a-20	12/24/2004	479190	8670	QC Passed
Run group:
Total Beads successfully sequenced - 2088514
Processed Signatures - 11569

source_name: Cervix and vagina, Female total RNA


title: Lung, Female (MoLu.Feml)

description:
Strain: C57BL6
Lung, Female total RNA (MoLu.Feml) received from National Human Genome Research Institute was analyzed through Bioanalyzer and passed our quality control test.  The mRNA was processed according to the MPSS protocol as outlined in the previous publications (Brenner, S., et al. (2000) Proc Natl Acad Sci U S A 97(4): 1665-1670 and Brenner, S. et al., Nat. Biotechnol 18(6): 630-634).  Briefly, the mRNA was reverse transcribed and the cDNA was digested with Dpn II. The 20 bases adjacent to the 3? most Dpn II site was cloned into a Megaclone vector.  The resulting library was amplified and loaded onto microbeads. About 1.6 million microbeads were loaded into each flow cell and the signature sequences were determined by a series of enzymatic reactions as outlined in the above publications. The abundance for each signature was converted to transcripts per million (tpm) for the purpose of comparisons between samples.
Cells/tissue:
Library MoLu.Feml.sig21
Cell type Lung, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoLu.Feml_sig21.5286W.d-20	1/11/2005	289684	9496	QC Passed
MoLu.Feml_sig21.5286W.b-20	12/18/2004	268693	6630	QC Passed
MoLu.Feml_sig21.5286F.a-20	12/6/2004	430604	10702	QC Passed
MoLu.Feml_sig21.5286F.b-20	12/6/2004	491316	10063	QC Passed
MoLu.Feml_sig21.5149F.a-20	10/12/2004	394448	10130	QC Passed
MoLu.Feml_sig21.5149W.a-20	8/30/2004	581725	13890	QC Passed
Run group:
Total Beads successfully sequenced - 2456470
Processed Signatures - 19159

source_name: Lung, Female total RNA


title: CTCF ChIP from Retinoic Acid Stimulated HL60 Cells, 0 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against CTCF overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, CTCF, HL60 Cells, 0 hours Retinoic Acid

source_name: Whole Cell Extract (WCE) DNA from Retinoic Acid Stimulated HL60 Cells, 0 hours


title: Brg1 ChIP from Retinoic Acid Stimulated HL60 Cells, 32 hours (NCBIv34)

description:
HL60 suspension cells were grown in Iscove's Modified Dulbecco's Medium (IMDM), GlutMAX; supplemented with 20% FBS, and 1% Penicillin-Streptomycin, at 37oC in 5% CO2 to a density of 0.75 x 10^6 cells/ml (medium and supplemental components were purchased from Invitrogen).  Cells were then induced with 1 uM all-trans-retinoic acid (ATRA, Sigma) for the period of time specified.
At the appropriate time, suspension cells are fixed with 1% formaldehyde (J.T. Baker) at room temperature for 10 minutes.  Subsequently, 1/20 volume of 2.5 M glycine (American Bioanalytical) was added and incubation proceeded for 5 minutes.  Crosslinked cells were extensively washed resulting in nuclear pellet formation.  A nuclear pellet, equivalent to 2 x 10^8 cells, was treated with 20U  of MNase (USB) for 10 min at 37C to generate fragmented DNA of 500-1000 bp average size, followed by sonication.
Cellular debris was separated from solubilized chromatin by high-speed centrifugation.  Isolated chromatin, equivalent to 2.5 x 10^7 cells, was pre-cleared for 30 min with Protein A/G sepharose beads (Amersham Biosciences) at 4C.  This pre-cleared chromatin was separated from the beads and immunoreacted with a monoclonal antibody against Brg1 overnight at 4C.  Protein A/G resin were subsequently added and allowed to bind for 3 hours at 4C.  Recovered, immunoprecipitated chromatin was extensively washed, crosslinks reversed, and purified immunoprecipitated (IP) DNA recovered using a QIAquick PCR purification kit (Qiagen).  A mock IP was preformed in parallel to isolate total DNA referred to as INPUT.
Two rounds of linear amplification were separately performed on 30% of IP and INPUT DNA using Sequenase (USB, per manufacturer's protocol) and a random primer (GTTTCCCAGTCACGATCNNNNNNNNN).  DNA was recovered using a QIAquick PCR purification kit (Qiagen).  PCR was performed using Invitrogen products, per manufacturer's protocol, on 75% of linearly amplified DNA using a primer (GTTTCCCAGTCACGATC) for 30 cycles.  DNA was purified via a QIAquick PCR purification kit (Qiagen), and 90% of PCR amplified DNA was subjected to an additional 25 rounds of PCR, with subsequent DNA recovered as stated above.  Amplified DNA was verified on 1.5% agarose gel to be an average size of 300 bp.
PCR amplified DNA, 5-10 ug, was subjected to further fragmentation to 50-100 bp by DNAse I (1U/ul; Epicentre) treatment in 1X One-Phor-All buffer (Pharmacia). Following DNAse I inactivation at 99C for 10 min, the size ditsribution of fragmented DNA was verified on a 1% agarose gel.
The PCR amplified, fragmented DNA from DNAse I treatment was end-labeled with 70nM of bio-ddATP (Perkin Elmer) using 6-10U of terminal transferase (TdT, Roche) per 1ug of fragmented DNA in 1X TdT buffer (Roche) and 5mM CoCl2 (Roche) for 2 hours at 37C. The labeled DNA material was subsequently hybridized to the chips  (previously prehybridized for 1 hour in 1X MES-triton at 45C) for 18 hours at 45C in a 3 M TMAC/1X MES-based solution.
Keywords = Homo sapiens, Brg1, HL60 Cells, 32 hours Retinoic Acid