title: Heart:Atria, Male (MoHe.MaAt)

description:
Strain: C57BL6
Heart:Atria, Male total RNA (MoHe.MaAt) 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 MoHe.MaAt.sig21
Cell type Heart:Atria, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoHe.MaAt_sig21.5212F.f-20	12/10/2004	391535	9649	QC Passed
MoHe.MaAt_sig21.5212W.c-20	11/20/2004	333899	6280	QC Passed
MoHe.MaAt_sig21.5212W.b-20	11/15/2004	233558	6062	QC Passed
MoHe.MaAt_sig21.5212F.c-20	11/12/2004	184449	5122	QC Passed
MoHe.MaAt_sig21.5212F.b-20	11/12/2004	248225	7372	QC Passed
MoHe.MaAt_sig21.5212F.a-20	10/5/2004	267703	7905	QC Passed
MoHe.MaAt_sig21.5212W.a-20	10/5/2004	347060	8815	QC Passed
Run group:
Total Beads successfully sequenced - 2006429
Processed Signatures - 14751

source_name: Heart:Atria, Male total RNA


title: CEBPe ChIP from Retinoic Acid Stimulated HL60 Cells, 2 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 CEBPe 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, CEBPe, HL60 Cells, 2 hours Retinoic Acid

molecule: genomic DNA

organism: Homo sapiens

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


title: CEBPe 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 CEBPe 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, CEBPe, HL60 Cells, 8 hours Retinoic Acid

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


title: H3K27T 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 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, 0 hours Retinoic Acid

source_name: H3K27TMe ChIP from Retinoic Acid Stimulated HL60 Cells, 0 hours

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


title: Embryo E18, Male/Female (MoEm.MF18)

status: Public on Jan 28 2005

type: MPSS

city: Hayward

first: Daixing

last: Zhou

address: 25861 Industrial Blvd

department: Bioinformatics

institute: Solexa

description:
Strain: C57BL6
Embryo E18, Male/Female total RNA (MoEm.MF18) 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 MoEm.MF18.sig21
Cell type Embryo E18, Male/Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoEm.MF18_sig21.5323W.a-20	1/17/2005	588926	13280	QC Passed
MoEm.MF18_sig21.5323W.b-20	1/17/2005	541669	13473	QC Passed
MoEm.MF18_sig21.5323F.a-20	1/15/2005	503235	12810	QC Passed
MoEm.MF18_sig21.5210F.a-20	10/7/2004	436012	9384	QC Passed
Run group:
Total Beads successfully sequenced - 2069842
Processed Signatures - 18592

molecule: total RNA

organism: Mus musculus

source_name: Embryo E18, Male/Female total RNA


title: Brain:OlfactoryTubercle,PrefrontalCortex,Septum, Male (MoBr.MTCS)

description:
Strain: C57BL6
Brain:OlfactoryTubercle,PrefrontalCortex,Septum, Male total RNA (MoBr.MTCS) 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.MTCS.sig22
Cell type Brain:OlfactoryTubercle,PrefrontalCortex,Septum, 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.MTCS_sig22.5290F.d-20	1/6/2005	604654	13817	QC Passed
MoBr.MTCS_sig22.5290W.c-20	1/1/2005	406645	10549	QC Passed
MoBr.MTCS_sig22.5290W.a-20	12/15/2004	597159	17663	QC Passed
MoBr.MTCS_sig22.5290F.a-20	12/10/2004	467714	10457	QC Passed
Run group:
Total Beads successfully sequenced - 2076172
Processed Signatures - 17743


title: Heart: atria, Female (MoHe.FeAt)

description:
Strain: C57BL6
Heart: atria, Female total RNA (MoHe.FeAt) 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 MoHe.FeAt.sig21
Cell type Heart: atria, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoHe.FeAt_sig21.5311W.b-20	1/18/2005	405582	2242	QC Passed
MoHe.FeAt_sig21.5311F.b-20	1/18/2005	229484	3445	QC Passed
MoHe.FeAt_sig21.5311W.d-20	1/17/2005	458621	3982	QC Passed
MoHe.FeAt_sig21.5311F.c-20	1/15/2005	367322	4596	QC Passed
MoHe.FeAt_sig21.5311F.a-20	12/17/2004	297373	3404	QC Passed
MoHe.FeAt_sig21.5141F.c-20	10/18/2004	293934	3339	QC Passed
MoHe.FeAt_sig21.5141W.b-20	8/18/2004	492639	3144	QC Passed
Run group:
Total Beads successfully sequenced - 2544955
Processed Signatures - 6970

source_name: Heart: atria, Female total RNA


title: Testis, Male (MoTe.Male)

description:
Strain: C57BL6
Testis, Male total RNA (MoTe.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 MoTe.Male.sig22
Cell type Testis, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoTe.Male_sig22.5293W.e-20	1/25/2005	444518	10931	QC Passed
MoTe.Male_sig22.5293W.c-20	12/18/2004	427540	10508	QC Passed
MoTe.Male_sig22.5293F.c-20	12/15/2004	258674	6863	QC Passed
MoTe.Male_sig22.5293F.b-20	12/8/2004	473105	10697	QC Passed
MoTe.Male_sig22.5168W.b-20	9/20/2004	352636	10339	QC Passed
MoTe.Male_sig22.5168F.b-20	9/17/2004	375232	10394	QC Passed
Run group:
Total Beads successfully sequenced - 2331705
Processed Signatures - 16297

source_name: Testis, Male total RNA


title: Cartilage:Xiphoid, Female (MoCg.FeXi)

description:
Strain: C57BL6
Cartilage:Xiphoid, Female total RNA (MoCg.FeXi) 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 MoCg.FeXi.sig21
Cell type Cartilage:Xiphoid, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoCg.FeXi_sig21.5282W.c-20	12/13/2004	343777	6848	QC Passed
MoCg.FeXi_sig21.5282F.c-20	12/10/2004	527334	11493	QC Passed
MoCg.FeXi_sig21.5282F.b-20	12/6/2004	533161	15835	QC Passed
MoCg.FeXi_sig21.5179W.a-20	9/21/2004	414715	12450	QC Passed
MoCg.FeXi_sig21.5179W.b-20	9/21/2004	341833	7220	QC Passed
MoCg.FeXi_sig21.5179F.b-20	9/20/2004	363341	9799	QC Passed
Run group:
Total Beads successfully sequenced - 2524161
Processed Signatures - 17897

source_name: Cartilage:Xiphoid, Female total RNA


title: Pituitary, Male (MoPi.Male)

status: Public on Jan 05 2005

description:
Strain: C57BL6
Pituitary, Male total RNA (MoPi.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 MoPi.Male.sig22
Cell type Pituitary, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoPi.Male_sig22.5253W.g-20	12/21/2004	458280	10125	QC Passed
MoPi.Male_sig22.5253W.e-20	12/5/2004	316417	6378	QC Passed
MoPi.Male_sig22.5253F.d-20	12/3/2004	396477	3789	QC Passed
MoPi.Male_sig22.5253F.c-20	11/30/2004	367465	3747	QC Passed
MoPi.Male_sig22.5253W.d-20	11/30/2004	251051	6468	QC Passed
MoPi.Male_sig22.5253F.b-20	11/15/2004	270215	2094	QC Passed
Run group:
Total Beads successfully sequenced - 2059905
Processed Signatures - 10905

source_name: Pituitary, Male total RNA


title: Large intestine, Female (MoLI.Feml)

description:
Strain: C57BL6
Large intestine, Female total RNA (MoLI.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 MoLI.Feml.sig21
Cell type Large 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
MoLI.Feml_sig21.5329W.a-20	12/19/2004	426977	9024	QC Passed
MoLI.Feml_sig21.5249F.c-20	11/20/2004	409731	6087	QC Passed
MoLI.Feml_sig21.5249W.a-20	11/9/2004	765978	12837	QC Passed
MoLI.Feml_sig21.5249F.b-20	11/8/2004	446781	7452	QC Passed
Run group:
Total Beads successfully sequenced - 2049467
Processed Signatures - 13829

source_name: Large intestine, Female total RNA


title: Brg1 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 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, 8 hours Retinoic Acid


title: Cartilage:Xiphoid, Male (MoCg.MaXi)

description:
Strain: C57BL6
Cartilage:Xiphoid, Male total RNA (MoCg.MaXi) 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 MoCg.MaXi.sig21
Cell type Cartilage:Xiphoid, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoCg.MaXi_sig21.5281F.c-20	12/28/2004	363535	8872	QC Passed
MoCg.MaXi_sig21.5281W.c-20	12/18/2004	611892	15465	QC Passed
MoCg.MaXi_sig21.5281F.b-20	12/9/2004	315224	5725	QC Passed
MoCg.MaXi_sig21.5281F.a-20	12/2/2004	451731	8284	QC Passed
MoCg.MaXi_sig21.5281W.b-20	12/1/2004	472341	10175	QC Passed
Run group:
Total Beads successfully sequenced - 2214723
Processed Signatures - 16766

source_name: Cartilage:Xiphoid, Male total RNA


title: Lymph nodes: mesenteric, Male (MoLN.MaMs)

description:
Strain: C57BL6
Lymph nodes: mesenteric, Male total RNA (MoLN.MaMs) 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.MaMs.sig22
Cell type Lymph nodes: mesenteric, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoLN.MaMs_sig22.5296F.c-20	12/28/2004	482515	10656	QC Passed
MoLN.MaMs_sig22.5296W.c-20	12/24/2004	427898	10123	QC Passed
MoLN.MaMs_sig22.5296F.d-20	12/23/2004	418331	8371	QC Passed
MoLN.MaMs_sig22.5296F.a-20	12/11/2004	355335	7891	QC Passed
MoLN.MaMs_sig22.5296W.b-20	12/9/2004	343705	8572	QC Passed
MoLN.MaMs_sig22.5296W.a-20	12/8/2004	322824	9027	QC Passed
Run group:
Total Beads successfully sequenced - 2350608
Processed Signatures - 16339

source_name: Lymph nodes: mesenteric, Male total RNA


title: Thymus, Female (MoTh.Feml)

description:
Strain: C57BL6
Thymus, Female total RNA (MoTh.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 MoTh.Feml.sig21
Cell type Thymus, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoTh.Feml_sig21.5280W.b-20	12/13/2004	654547	17677	QC Passed
MoTh.Feml_sig21.5280F.a-20	12/2/2004	343847	10228	QC Passed
MoTh.Feml_sig21.5280F.b-20	12/1/2004	449061	11663	QC Passed
MoTh.Feml_sig21.5151F.a-20	9/22/2004	339696	9987	QC Passed
MoTh.Feml_sig21.5151W.a-20	9/2/2004	561755	15139	QC Passed
Run group:
Total Beads successfully sequenced - 2348906
Processed Signatures - 21390

source_name: Thymus, Female total RNA


title: Thyroid/parathyroid, Male (MoTp.Male)

description:
Strain: C57BL6
Thyroid/parathyroid, Male total RNA (MoTp.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 MoTp.Male.sig21
Cell type Thyroid/parathyroid, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoTp.Male_sig21.5220W.e-20	12/24/2004	467804	10523	QC Passed
MoTp.Male_sig21.5220F.e-20	12/5/2004	383049	8938	QC Passed
MoTp.Male_sig21.5220F.c-20	11/26/2004	586207	11782	QC Passed
MoTp.Male_sig21.5220W.c-20	11/23/2004	596661	10124	QC Passed
Run group:
Total Beads successfully sequenced - 2033721
Processed Signatures - 15436

source_name: Thyroid/parathyroid, Male total RNA


title: CEBPe 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 CEBPe 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, CEBPe, HL60 Cells, 32 hours Retinoic Acid

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


title: Pituitary, Female (MoPi.Feml)

description:
Strain: C57BL6
Pituitary, Female total RNA (MoPi.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 MoPi.Feml.sig21
Cell type Pituitary, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoPi.Feml_sig21.5247W.f-20	12/15/2004	308219	6823	QC Passed
MoPi.Feml_sig21.5247W.d-20	11/19/2004	227558	7363	QC Passed
MoPi.Feml_sig21.5247F.b-20	11/3/2004	337952	5405	QC Passed
MoPi.Feml_sig21.5247F.a-20	11/3/2004	365615	5426	QC Passed
MoPi.Feml_sig21.5247W.b-20	11/2/2004	341752	6655	QC Passed
MoPi.Feml_sig21.5203F.b-20	10/4/2004	425412	7658	QC Passed
Run group:
Total Beads successfully sequenced - 2006508
Processed Signatures - 11777

source_name: Pituitary, Female total RNA


title: CTCF 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 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, 8 hours Retinoic Acid


title: CEBPe 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 CEBPe 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, CEBPe, HL60 Cells, 0 hours Retinoic Acid


title: Large intestine, Male (MoLI.Male)

status: Public on Dec 02 2004

description:
Strain: C57BL6
Large intestine, Male total RNA (MoLI.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 MoLI.Male.sig21
Cell type Large 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
MoLI.Male_sig21.5193F.d-20	11/4/2004	429787	4715	QC Passed
MoLI.Male_sig21.5193F.c-20	10/27/2004	322659	3331	QC Passed
MoLI.Male_sig21.5193W.b-20	9/28/2004	386294	4631	QC Passed
MoLI.Male_sig21.5193W.a-20	9/28/2004	405911	4330	QC Passed
MoLI.Male_sig21.5193F.b-20	9/27/2004	390518	5880	QC Passed
Run group:
Total Beads successfully sequenced - 1935169
Processed Signatures - 7488

source_name: Large intestine, Male total RNA


title: Eye, Female (MoEy.Feml)

description:
Strain: C57BL6
Eye, Female total RNA (MoEy.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 MoEy.Feml.sig21
Cell type Eye, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoEy.Feml_sig21.5222W.f-20	12/7/2004	513231	21516	QC Passed
MoEy.Feml_sig21.5222F.f-20	12/6/2004	395823	10839	QC Passed
MoEy.Feml_sig21.5222F.c-20	11/24/2004	338104	8177	QC Passed
MoEy.Feml_sig21.5222W.c-20	11/24/2004	537293	15712	QC Passed
MoEy.Feml_sig21.5222F.b-20	10/18/2004	269919	7823	QC Passed
Run group:
Total Beads successfully sequenced - 2054370
Processed Signatures - 24332

source_name: Eye, Female total RNA


title: Brg1 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 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, 0 hours Retinoic Acid


title: Brain:Thalamus, Male (MoBr.MaTh)

description:
Strain: C57BL6
Brain:Thalamus, Male total RNA (MoBr.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 MoBr.MaTh.sig21
Cell type Brain:Thalamus, 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.MaTh_sig21.5330W.b-20	12/20/2004	397571	10946	QC Passed
MoBr.MaTh_sig21.5330W.a-20	12/16/2004	517628	14352	QC Passed
MoBr.MaTh_sig21.5191F.c-20	10/28/2004	384348	9313	QC Passed
MoBr.MaTh_sig21.5191W.a-20	10/4/2004	393714	8867	QC Passed
MoBr.MaTh_sig21.5191F.a-20	9/29/2004	421958	11215	QC Passed
MoBr.MaTh_sig21.5191F.b-20	9/27/2004	382185	10400	QC Passed
Run group:
Total Beads successfully sequenced - 2497404
Processed Signatures - 21857

source_name: Brain:Thalamus, Male total RNA


title: Brain:Hypothalamus/preoptic area, Female (MoBr.FHyP)

description:
Strain: C57BL6
Brain:Hypothalamus/preoptic area, Female total RNA (MoBr.FHyP) 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.FHyP.sig23
Cell type Brain:Hypothalamus/preoptic area, 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.FHyP_sig23.5295F.b-20	12/11/2004	361615	8832	QC Passed
MoBr.FHyP_sig23.5295F.a-20	12/10/2004	440241	11313	QC Passed
MoBr.FHyP_sig23.5295W.c-20	12/24/2004	394049	10126	QC Passed
MoBr.FHyP_sig23.5295W.e-20	12/24/2004	536480	13832	QC Passed
MoBr.FHyP_sig23.5295F.c-20	12/23/2004	477675	10035	QC Passed
Run group:
Total Beads successfully sequenced - 2210060
Processed Signatures - 17652


title: Brain:Olfactory Bulb, Female (MoBr.FeOB)

status: Public on Nov 30 2004

description:
Strain: C57BL6
Brain:Olfactory Bulb, Female total RNA (MoBr.FeOB) 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.FeOB.sig22
Cell type Brain:Olfactory Bulb, 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.FeOB_sig22.5245W.b-20	11/8/2004	549511	19166	QC Passed
MoBr.FeOB_sig22.5245F.b-20	11/1/2004	489600	14969	QC Passed
MoBr.FeOB_sig22.5245F.a-20	11/1/2004	319150	10371	QC Passed
MoBr.FeOB_sig22.5245W.a-20	11/1/2004	418673	17439	QC Passed
MoBr.FeOB_sig22.5177F.b-20	9/22/2004	352443	12264	QC Passed
Run group:
Total Beads successfully sequenced - 2129377
Processed Signatures - 25616

source_name: Brain:Olfactory Bulb, Female total RNA


title: Heart:Aorta, Male (MoAo.Male)

description:
Strain: C57BL6
Heart:Aorta, Male total RNA (MoAo.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 MoAo.Male.sig21
Cell type Heart:Aorta, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoAo.Male_sig21.5300W.a-20	12/5/2004	500357	10569	QC Passed
MoAo.Male_sig21.5300F.b-20	12/3/2004	366114	6908	QC Passed
MoAo.Male_sig21.5153F.c-20	10/19/2004	464519	9504	QC Passed
MoAo.Male_sig21.5153F.a-20	9/15/2004	353865	9213	QC Passed
MoAo.Male_sig21.5153W.b-20	9/8/2004	419765	8216	QC Passed
Run group:
Total Beads successfully sequenced - 2104620
Processed Signatures - 14322

source_name: Heart:Aorta, Male total RNA


title: Eye, Male (MoEy.Male)

description:
Strain: C57BL6
Eye, Male total RNA (MoEy.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 MoEy.Male.sig21
Cell type Eye, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoEy.Male_sig21.5201W.c-20	11/8/2004	384776	13126	QC Passed
MoEy.Male_sig21.5201W.a-20	10/12/2004	520366	16376	QC Passed
MoEy.Male_sig21.5201W.b-20	10/11/2004	449908	15753	QC Passed
MoEy.Male_sig21.5201F.b-20	10/11/2004	303836	9150	QC Passed
MoEy.Male_sig21.5201F.a-20	10/11/2004	362297	8970	QC Passed
Run group:
Total Beads successfully sequenced - 2021183
Processed Signatures - 21536

source_name: Eye, Male total RNA


title: Bone:Femur, Female (MoBn.FeFm)

description:
Strain: C57BL6
Bone:Femur, Female total RNA (MoBn.FeFm) 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 MoBn.FeFm.sig21
Cell type Bone:Femur, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoBn.FeFm_sig21.5248W.c-20	11/23/2004	693088	8770	QC Passed
MoBn.FeFm_sig21.5248F.c-20	11/20/2004	333519	5293	QC Passed
MoBn.FeFm_sig21.5248F.b-20	11/8/2004	369141	8714	QC Passed
MoBn.FeFm_sig21.5248W.a-20	11/8/2004	600356	6274	QC Passed
MoBn.FeFm_sig21.5248F.a-20	11/8/2004	337614	5858	QC Passed
Run group:
Total Beads successfully sequenced - 2333718
Processed Signatures - 11115

source_name: Bone:Femur, Female total RNA


title: Brain:Caudate,Putamen,Accumbens,BasalForebrain, Male (MoBr.MCPA)

description:
Strain: C57BL6
Brain:Caudate,Putamen,Accumbens,BasalForebrain, Male total RNA (MoBr.MCPA) 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.MCPA.sig21
Cell type Brain:Caudate,Putamen,Accumbens,BasalForebrain, 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.MCPA_sig21.5198F.d-20	11/16/2004	489621	12880	QC Passed
MoBr.MCPA_sig21.5198W.a-20	10/1/2004	672816	21425	QC Passed
MoBr.MCPA_sig21.5198W.b-20	9/28/2004	432775	11543	QC Passed
MoBr.MCPA_sig21.5198F.a-20	9/23/2004	553805	16543	QC Passed
Run group:
Total Beads successfully sequenced - 2149017
Processed Signatures - 23334


title: Heart: ventricles and septum, Male (MoHe.MaVS)

description:
Strain: C57BL6
Heart: ventricles and septum, Male total RNA (MoHe.MaVS) 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 MoHe.MaVS.sig21
Cell type Heart: ventricles and septum, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoHe.MaVS_sig21.5278W.b-20	11/30/2004	507782	7242	QC Passed
MoHe.MaVS_sig21.5278F.c-20	11/29/2004	536764	8990	QC Passed
MoHe.MaVS_sig21.5278F.b-20	11/26/2004	423821	8002	QC Passed
MoHe.MaVS_sig21.5278W.a-20	11/19/2004	499533	7307	QC Passed
Run group:
Total Beads successfully sequenced - 1967900
Processed Signatures - 12215


title: Thymus, Male (MoTh.Male)

description:
Strain: C57BL6
Thymus, Male total RNA (MoTh.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 MoTh.Male.sig21
Cell type Thymus, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoTh.Male_sig21.5189F.c-20	11/15/2004	170248	3968	QC Passed
MoTh.Male_sig21.5189F.b-20	11/12/2004	295741	7718	QC Passed
MoTh.Male_sig21.5189W.b-20	11/10/2004	310187	6707	QC Passed
MoTh.Male_sig21.5189F.a-20	9/27/2004	320327	8382	QC Passed
MoTh.Male_sig21.5189W.a-20	9/24/2004	335613	8617	QC Passed
MoTh.Male_sig21.5131W.b-20	8/9/2004	305324	11556	QC Passed
MoTh.Male_sig21.5131W.a-20	8/9/2004	330656	14413	QC Passed
Run group:
Total Beads successfully sequenced - 2068096
Processed Signatures - 16562

source_name: Thymus, Male total RNA


title: Brg1 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 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, 2 hours Retinoic Acid

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


title: Brain:OlfactoryTubercle,PrefrontalCortex,Septum, Female (MoBr.FTCS)

description:
Strain: C57BL6
Brain:OlfactoryTubercle,PrefrontalCortex,Septum, Female total RNA (MoBr.FTCS) 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.FTCS.sig22
Cell type Brain:OlfactoryTubercle,PrefrontalCortex,Septum, 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.FTCS_sig22.5194W.f-20	11/29/2004	286475	7456	QC Passed
MoBr.FTCS_sig22.5194W.d-20	11/12/2004	343606	11472	QC Passed
MoBr.FTCS_sig22.5194F.c-20	11/4/2004	377077	13756	QC Passed
MoBr.FTCS_sig22.5194F.d-20	11/3/2004	307087	9119	QC Passed
MoBr.FTCS_sig22.5194W.a-20	10/1/2004	452939	15531	QC Passed
MoBr.FTCS_sig22.5194F.a-20	9/24/2004	365495	11926	QC Passed
Run group:
Total Beads successfully sequenced - 2132679
Processed Signatures - 22198


title: Esophagus, Female (MoEs.Feml)

status: Public on Nov 15 2004

description:
Strain: C57BL6
Esophagus, Female total RNA (MoEs.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 MoEs.Feml.sig22
Cell type Esophagus, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoEs.Feml_sig22.5252W.b-20	11/3/2004	541556	9777	QC Passed
MoEs.Feml_sig22.5252F.a-20	11/3/2004	558014	9207	QC Passed
MoEs.Feml_sig22.5252F.b-20	11/1/2004	449047	6239	QC Passed
MoEs.Feml_sig22.5252W.a-20	11/1/2004	514616	8253	QC Passed
MoEs.Feml_sig22.5125W.d-20	10/12/2004	546976	8551	QC Passed
Run group:
Total Beads successfully sequenced - 2610209
Processed Signatures - 14353

source_name: Esophagus, Female total RNA


title: Spleen, Female (MoSp.Feml)

description:
Strain: C57BL6
Spleen, Female total RNA (MoSp.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 MoSp.Feml.sig22
Cell type Spleen, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoSp.Feml_sig22.5243F.b-20	11/12/2004	483843	9030	QC Passed
MoSp.Feml_sig22.5243W.a-20	11/9/2004	576958	9809	QC Passed
MoSp.Feml_sig22.5243F.a-20	11/8/2004	433860	10177	QC Passed
MoSp.Feml_sig22.5130W.e-20	8/26/2004	599543	13887	QC Passed
Run group:
Total Beads successfully sequenced - 2094204
Processed Signatures - 16105

source_name: Spleen, Female total RNA


title: RNAP ChIP from Retinoic Acid Stimulated HL-60 Cells, 0 hours (NCBIv34)

status: Public on Jan 31 2005

description:
HL-60 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 RNA Polymerase II, RNAP (Covance Research Products) 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, RNAP, HL-60 Cells, 0 hours Retinoic Acid

source_name: Whole Cell Extract (WCE) DNA from Retinoic Acid Stimulated HL-60 Cells, 0 hours


title: Spinal cord:entire, Male (MoSC.MaEn)

description:
Strain: C57BL6
Spinal cord:entire, Male total RNA (MoSC.MaEn) 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 MoSC.MaEn.sig21
Cell type Spinal cord:entire, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoSC.MaEn_sig21.5255F.g-20	12/13/2004	343077	9118	QC Passed
MoSC.MaEn_sig21.5255F.f-20	12/1/2004	350659	8488	QC Passed
MoSC.MaEn_sig21.5255W.a-20	11/11/2004	710993	17315	QC Passed
MoSC.MaEn_sig21.5134F.b-20	9/16/2004	281300	10689	QC Passed
MoSC.MaEn_sig21.5134W.c-20	8/14/2004	418243	12924	QC Passed
MoSC.MaEn_sig21.5134W.b-20	8/7/2004	413845	14758	QC Passed
Run group:
Total Beads successfully sequenced - 2518117
Processed Signatures - 22592

source_name: Spinal cord:entire, Male total RNA


title: Thyroid/parathyroid, Female (MoTp.Feml)

description:
Strain: C57BL6
Thyroid/parathyroid, Female total RNA (MoTp.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 MoTp.Feml.sig21
Cell type Thyroid/parathyroid, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoTp.Feml_sig21.5221W.c-20	11/23/2004	614111	10758	QC Passed
MoTp.Feml_sig21.5221F.c-20	11/21/2004	399777	8563	QC Passed
MoTp.Feml_sig21.5221F.a-20	10/14/2004	343637	8004	QC Passed
MoTp.Feml_sig21.5221W.a-20	10/14/2004	512678	9151	QC Passed
MoTp.Feml_sig21.5221W.b-20	10/14/2004	416909	7915	QC Passed
MoTp.Feml_sig21.5221F.b-20	10/12/2004	327600	7789	QC Passed
Run group:
Total Beads successfully sequenced - 2614712
Processed Signatures - 15140

source_name: Thyroid/parathyroid, Female total RNA


title: Skin:hairy, from back, Male (MoSk.MaHB)

description:
Strain: C57BL6
Skin:hairy, from back, Male total RNA (MoSk.MaHB) 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 MoSk.MaHB.sig21
Cell type Skin:hairy, from back, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoSk.MaHB_sig21.5279W.b-20	11/22/2004	648252	18338	QC Passed
MoSk.MaHB_sig21.5279F.c-20	11/19/2004	389315	9252	QC Passed
MoSk.MaHB_sig21.5279F.a-20	11/15/2004	329988	8613	QC Passed
MoSk.MaHB_sig21.5195W.a-20	10/1/2004	386214	12161	QC Passed
MoSk.MaHB_sig21.5195W.b-20	9/28/2004	356446	9148	QC Passed
MoSk.MaHB_sig21.5195F.b-20	9/24/2004	329975	10975	QC Passed
Run group:
Total Beads successfully sequenced - 2440190
Processed Signatures - 21058

source_name: Skin:hairy, from back, Male total RNA


title: Mammary gland, Female (MoMG.Feml)

description:
Strain: C57BL6
Mammary gland, Female total RNA (MoMG.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 MoMG.Feml.sig23
Cell type Mammary gland, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoMG.Feml_sig23.5181F.c-20	11/8/2004	479212	11380	QC Passed
MoMG.Feml_sig23.5181W.b-20	9/21/2004	409501	10777	QC Passed
MoMG.Feml_sig23.5181W.a-20	9/20/2004	407911	10095	QC Passed
MoMG.Feml_sig23.5181F.b-20	9/20/2004	374774	10618	QC Passed
MoMG.Feml_sig23.5181F.a-20	9/17/2004	429686	10751	QC Passed
Run group:
Total Beads successfully sequenced - 2101084
Processed Signatures - 17648

source_name: Mammary gland, Female total RNA


title: Ovary, Female (MoOv.Feml)

status: Public on Nov 01 2004

description:
Strain: C57BL6
Ovary, Female total RNA (MoOv.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 MoOv.Feml.sig21
Cell type Ovary, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoOv.Feml_sig21.5139F.e-20	10/29/2004	516520	13346	QC Passed
MoOv.Feml_sig21.5139F.a-20	10/1/2004	415347	12780	QC Passed
MoOv.Feml_sig21.5139F.b-20	9/30/2004	378384	13337	QC Passed
MoOv.Feml_sig21.5139W.b-20	8/16/2004	553347	19533	QC Passed
MoOv.Feml_sig21.5139W.a-20	8/16/2004	448425	15811	QC Passed
Run group:
Total Beads successfully sequenced - 2312023
Processed Signatures - 24639

source_name: Ovary, Female total RNA


title: Bone:Femur, Male (MoBn.MaFm)

description:
Strain: C57BL6
Bone:Femur, Male total RNA (MoBn.MaFm) 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 MoBn.MaFm.sig21
Cell type Bone:Femur, Male
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoBn.MaFm_sig21.5158F.d-20	10/26/2004	412620	7689	QC Passed
MoBn.MaFm_sig21.5158F.a-20	10/1/2004	384296	7075	QC Passed
MoBn.MaFm_sig21.5158F.b-20	10/1/2004	462739	6799	QC Passed
MoBn.MaFm_sig21.5158W.b-20	9/7/2004	590876	7785	QC Passed
MoBn.MaFm_sig21.5158W.a-20	9/3/2004	575732	7219	QC Passed
Run group:
Total Beads successfully sequenced - 2426263
Processed Signatures - 11765

source_name: Bone:Femur, Male total RNA


title: White fat, Male (MoWF.MaAb)

status: Public on Oct 26 2004

description:
Strain: C57BL6
White fat, Male total RNA (MoWF.MaAb) 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 MoWF.MaAb.sig22
Cell type White 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
MoWF.MaAb_sig22.5128F.b-20	9/30/2004	512059	11247	QC Passed
MoWF.MaAb_sig22.5128F.a-20	9/10/2004	608656	13982	QC Passed
MoWF.MaAb_sig22.5128W.b-20	8/12/2004	569426	11747	QC Passed
MoWF.MaAb_sig22.5128W.a-20	8/11/2004	492276	12108	QC Passed
Run group:
Total Beads successfully sequenced - 2182417
Processed Signatures - 17942

source_name: White fat, Male total RNA


title: Uterus:Pregnant E18, Female (MoUt.FP18)

description:
Strain: C57BL6
Uterus:Pregnant E18, Female total RNA (MoUt.FP18) 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 MoUt.FP18.sig21
Cell type Uterus:Pregnant 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
MoUt.FP18_sig21.5223F.c-20	11/3/2004	596625	10187	QC Passed
MoUt.FP18_sig21.5223W.b-20	10/24/2004	676917	14646	QC Passed
MoUt.FP18_sig21.5223W.a-20	10/21/2004	593076	12000	QC Passed
MoUt.FP18_sig21.5223F.a-20	10/19/2004	445622	9573	QC Passed
Run group:
Total Beads successfully sequenced - 2312240
Processed Signatures - 17084

source_name: Uterus:Pregnant E18, Female total RNA


title: Brain:Hypothalamus/preoptic area, Male (MoBr.MHyP)

description:
Strain: C57BL6
Brain:Hypothalamus/preoptic area, Male total RNA (MoBr.MHyP) 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.MHyP.sig21
Cell type Brain:Hypothalamus/preoptic area, 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.MHyP_sig21.5140F.f-20	10/20/2004	287116	8708	QC Passed
MoBr.MHyP_sig21.5140F.d-20	9/30/2004	367118	10986	QC Passed
MoBr.MHyP_sig21.5140F.b-20	9/24/2004	326060	9242	QC Passed
MoBr.MHyP_sig21.5140W.b-20	8/17/2004	529051	16354	QC Passed
MoBr.MHyP_sig21.5140W.a-20	8/16/2004	588504	18328	QC Passed
Run group:
Total Beads successfully sequenced - 2097849
Processed Signatures - 21719


title: RNAP ChIP from Retinoic Acid Stimulated HL-60 Cells, 8 hours (NCBIv34)

description:
HL-60 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 RNA Polymerase II, RNAP (Covance Research Products) 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, RNAP, HL-60 Cells, 8 hours Retinoic Acid

source_name: Whole Cell Extract (WCE) DNA from Retinoic Acid Stimulated HL-60 Cells, 8 hours


title: Stomach,  Female (MoSt.Feml)

description:
Strain: C57BL6
Stomach,  Female total RNA (MoSt.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 MoSt.Feml.sig23
Cell type Stomach,  Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoSt.Feml_sig23.5192W.d-20	11/8/2004	587290	9329	QC Passed
MoSt.Feml_sig23.5192F.c-20	10/8/2004	487261	9279	QC Passed
MoSt.Feml_sig23.5192W.a-20	9/27/2004	523276	8269	QC Passed
MoSt.Feml_sig23.5192F.b-20	9/27/2004	558246	9097	QC Passed
Run group:
Total Beads successfully sequenced - 2156073
Processed Signatures - 13378

source_name: Stomach,  Female total RNA


title: Skin:hariy, from back, Female (MoSk.FeHB)

description:
Strain: C57BL6
Skin:hariy, from back, Female total RNA (MoSk.FeHB) 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 MoSk.FeHB.sig22
Cell type Skin:hariy, from back, Female
Source NHGRI
RNA isolation LYNX mRNA QC passed
cDNA library:
Library 	DpnII restriction - (signature cloning using MmeI)
Sequence length	20 bp
MPSS:
runs
MoSk.FeHB_sig22.5170F.c-20	10/4/2004	337741	8158	QC Passed
MoSk.FeHB_sig22.5170W.a-20	9/17/2004	412511	10638	QC Passed
MoSk.FeHB_sig22.5170W.b-20	9/16/2004	409310	9671	QC Passed
MoSk.FeHB_sig22.5170F.b-20	9/16/2004	412695	10127	QC Passed
MoSk.FeHB_sig22.5170F.a-20	9/15/2004	390908	10615	QC Passed
Run group:
Total Beads successfully sequenced - 1963165
Processed Signatures - 15628

source_name: Skin:hariy, from back, Female total RNA