Virus:Article Title: Long-range regulation of transcription scales with genomic distance in a gene-specific manner.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Cas9- mouse monoclonal Active Motif Cat# 61757; RRID: AB_2793760 H3K27ac- rabbit polyclonal Active Motif Cat# 39133 RNA Pol II-mouse monoclonal Biolegend Cat# 664906; RRID: AB_2565554 H3K27me3- mouse monoclonal Active Motif Cat# 61017; RRID: AB_2614987 Bacterial and virus strains 10-beta Competent E. coli (high efficiency) NEB C3019H Stable Competent E. coli (high efficiency) NEB C3040H Chemicals, peptides, and recombinant proteins Accutase Sigma-Aldrich Cat# A6964-100ML DMEM/F12 Gibco Cat# 11320082 GSK3b CHIR99021 Selleck Chemicals Cat# S2924 MEK inhibitor PD0325901 Selleck Chemicals Cat# S1036 Gem21 NeuroPlex, without vitamin A Gemini Bio Cat# 400-161-010 N2 Neuroplex Gemini Bio Cat# 400-163-005 Sodium pyruvate (100mM) Gibco Cat# 11360070 2-mercaptoethanol Gibco Cat# 21985023 Anti-Anti (100x) Sigma-Aldrich Cat# A5955-100ML MEM NEAA (100x) ThemoFisher Cat# 11140050 LIF homemade N/A Laminin Gibco Cat# 23017015 Poly-l-ornithine Sigma-Aldrich Cat# P4638 BSA, Serum replacement Gemini Bio Cat# 700-104-100 Doxycycline hyclate EMD Millipore Cat# 32438501GM EED226 Selleck Chemicals Cat# S8496 Trizol reagent Invitrogen Cat# 10296010 Blasticidin, (10mg/mL) Invivogen Cat# ant-bl-1 G418, (100mg/mL) Invivogen Cat# ant-gn-1 Puromycin (10mg/mL) Invivogen Cat# ant-pr-1 Fibronectin ThermoFisher Cat# FC01010MG cOmplete, EDTA-free Protease inhibitor cocktail Millipore Sigma Cat# 11873580001 PMSF Sigma-Aldrich Cat# 10837091001 RNaseOUT Invitrogen Cat# 10777019 oligo dT (20mer) IDT N/A BsrGI NEB Cat# R3575 10xTango Buffer ThermoFisher Cat# BY5 10x T4 Ligase Buffer NEB Cat# B0202S T7 DNA Ligase NEB Cat# M0318S T4 DNA Ligase (30U/uL) ThermoFisher Cat# EL0013 BpiI ThermoFisher Cat# ER1011 Herculease II Dusion DNA Polymerase Agilent Technologies Cat# 600675 Critical commercial assays Lipofectamine 2000 ThermoFisher Cat# 11668019 Dynabeads Protein G Immunoprecipitation Invitrogen Cat# 10004D (Continued on next page) e1 Molecular Cell 85, 1–15.e1–e7, January 16, 2025 .. REAGENT or RESOURCE SOURCE IDENTIFIER ChIP Clean & Concentrator-5 Zymo Cat# D5205 Direct-zol RNA miniprep Zymo Cat# R2052 DNA Clean and Concentrator-5 Zymo Cat# D4014 High-Capacity cDNA Reverse Transcripase kit ThermoFisher Cat# 4368814 Sensifast SYBR No-Rox 2x mix Meridian Bioline Cat# 98020 Plasmid Safe ATP-dependent DNase Lucigen Cat# 3110K Gentra Puregene kit Qiagen Cat# 158745 Deposited data CRISPRa screen This paper Geo: GSE278772 ORCA processed datasets This paper 4DN data portal: 4DNESPB3DGGV 4DNESSKEUB9C 4DNESWTEQEBQ Experimental models: Cell lines R1 mESC WT Nagy et al.49 N/A R1 mESC, Fgf5-t2A-eGFP/tetO-dCas9-VPR This paper N/A R1 mESC, tetO-dCas9-VPR, clone 1 This paper N/A R1 mESC, tetO-dCas9-VPR, clone 2 This paper N/A R1 mESC, tetO-dCas9-VPR, P2FF2P-promoter swap This paper N/A R1 mESC, tetO-dCas9-VPR, delLTR, clone 12 This paper N/A Oligonucleotides Oligonucleotides used in this paper, Table S1 This paper N/A Guides in CRISPRa screen, Table S2 This paper N/A CARGO oligos and guides, Table S3 This paper N/A Recombinant DNA Super piggyBac Transposase expression vector System Biosciences (SBI) Cat# PB210PA-1 PhiC31 Integrase expression vector System Biosciences (SBI) Cat# FC200PA-1 pAAV-Ef1a-mCherry-IRES-Cre Fenno et al.50 Addgene Cat# 55632 pCAG-Flpe Matsuda et al.51 Addgene Cat#1 3787 pk335-BFP This paper N/A pX458-mCherry Gu et al.35 Addgene Cat# 161974 pCLJ-Fgf5-t2A-eGFP This paper N/A pCLJ-P2F-Frt-PGK-Puro-Frt This paper N/A pCLJ-F2P-LoxP-PGK-NeoR-LoxP This paper N/A pCLJ-F2P-LoxP-PGK-PuroR-LoxP This paper N/A pk335-CARGO-6mer-26kb-USP This paper N/A pk335-CARGO-6mer-24kb-USP This paper N/A pk335-CARGO-6mer-20kb-USP This paper N/A pk335-CARGO-6mer-11kb-USP This paper N/A pk335-CARGO-6mer-7.5kb-USP This paper N/A pk335-CARGO-6mer-4.5kb-USP This paper N/A pk335-CARGO-6mer-3.3kb-USP This paper N/A pk335-CARGO-6mer-2.1kb-USP This paper N/A pk335-CARGO-6mer-700bp-USP This paper N/A pk335-CARGO-6mer-PrPro This paper N/A pk335-CARGO-6mer-58kb-USF This paper N/A pk335-CARGO-6mer-55kb-USF This paper N/A pk335-CARGO-6mer-42kb-USF This paper N/A (Continued on next page) Molecular Cell 85, 1–15.e1–e7, January 16, 2025 e2
Recombinant:Article Title: Long-range regulation of transcription scales with genomic distance in a gene-specific manner.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Cas9- mouse monoclonal Active Motif Cat# 61757; RRID: AB_2793760 H3K27ac- rabbit polyclonal Active Motif Cat# 39133 RNA Pol II-mouse monoclonal Biolegend Cat# 664906; RRID: AB_2565554 H3K27me3- mouse monoclonal Active Motif Cat# 61017; RRID: AB_2614987 Bacterial and virus strains 10-beta Competent E. coli (high efficiency) NEB C3019H Stable Competent E. coli (high efficiency) NEB C3040H Chemicals, peptides, and recombinant proteins Accutase Sigma-Aldrich Cat# A6964-100ML DMEM/F12 Gibco Cat# 11320082 GSK3b CHIR99021 Selleck Chemicals Cat# S2924 MEK inhibitor PD0325901 Selleck Chemicals Cat# S1036 Gem21 NeuroPlex, without vitamin A Gemini Bio Cat# 400-161-010 N2 Neuroplex Gemini Bio Cat# 400-163-005 Sodium pyruvate (100mM) Gibco Cat# 11360070 2-mercaptoethanol Gibco Cat# 21985023 Anti-Anti (100x) Sigma-Aldrich Cat# A5955-100ML MEM NEAA (100x) ThemoFisher Cat# 11140050 LIF homemade N/A Laminin Gibco Cat# 23017015 Poly-l-ornithine Sigma-Aldrich Cat# P4638 BSA, Serum replacement Gemini Bio Cat# 700-104-100 Doxycycline hyclate EMD Millipore Cat# 32438501GM EED226 Selleck Chemicals Cat# S8496 Trizol reagent Invitrogen Cat# 10296010 Blasticidin, (10mg/mL) Invivogen Cat# ant-bl-1 G418, (100mg/mL) Invivogen Cat# ant-gn-1 Puromycin (10mg/mL) Invivogen Cat# ant-pr-1 Fibronectin ThermoFisher Cat# FC01010MG cOmplete, EDTA-free Protease inhibitor cocktail Millipore Sigma Cat# 11873580001 PMSF Sigma-Aldrich Cat# 10837091001 RNaseOUT Invitrogen Cat# 10777019 oligo dT (20mer) IDT N/A BsrGI NEB Cat# R3575 10xTango Buffer ThermoFisher Cat# BY5 10x T4 Ligase Buffer NEB Cat# B0202S T7 DNA Ligase NEB Cat# M0318S T4 DNA Ligase (30U/uL) ThermoFisher Cat# EL0013 BpiI ThermoFisher Cat# ER1011 Herculease II Dusion DNA Polymerase Agilent Technologies Cat# 600675 Critical commercial assays Lipofectamine 2000 ThermoFisher Cat# 11668019 Dynabeads Protein G Immunoprecipitation Invitrogen Cat# 10004D (Continued on next page) e1 Molecular Cell 85, 1–15.e1–e7, January 16, 2025 .. REAGENT or RESOURCE SOURCE IDENTIFIER ChIP Clean & Concentrator-5 Zymo Cat# D5205 Direct-zol RNA miniprep Zymo Cat# R2052 DNA Clean and Concentrator-5 Zymo Cat# D4014 High-Capacity cDNA Reverse Transcripase kit ThermoFisher Cat# 4368814 Sensifast SYBR No-Rox 2x mix Meridian Bioline Cat# 98020 Plasmid Safe ATP-dependent DNase Lucigen Cat# 3110K Gentra Puregene kit Qiagen Cat# 158745 Deposited data CRISPRa screen This paper Geo: GSE278772 ORCA processed datasets This paper 4DN data portal: 4DNESPB3DGGV 4DNESSKEUB9C 4DNESWTEQEBQ Experimental models: Cell lines R1 mESC WT Nagy et al.49 N/A R1 mESC, Fgf5-t2A-eGFP/tetO-dCas9-VPR This paper N/A R1 mESC, tetO-dCas9-VPR, clone 1 This paper N/A R1 mESC, tetO-dCas9-VPR, clone 2 This paper N/A R1 mESC, tetO-dCas9-VPR, P2FF2P-promoter swap This paper N/A R1 mESC, tetO-dCas9-VPR, delLTR, clone 12 This paper N/A Oligonucleotides Oligonucleotides used in this paper, Table S1 This paper N/A Guides in CRISPRa screen, Table S2 This paper N/A CARGO oligos and guides, Table S3 This paper N/A Recombinant DNA Super piggyBac Transposase expression vector System Biosciences (SBI) Cat# PB210PA-1 PhiC31 Integrase expression vector System Biosciences (SBI) Cat# FC200PA-1 pAAV-Ef1a-mCherry-IRES-Cre Fenno et al.50 Addgene Cat# 55632 pCAG-Flpe Matsuda et al.51 Addgene Cat#1 3787 pk335-BFP This paper N/A pX458-mCherry Gu et al.35 Addgene Cat# 161974 pCLJ-Fgf5-t2A-eGFP This paper N/A pCLJ-P2F-Frt-PGK-Puro-Frt This paper N/A pCLJ-F2P-LoxP-PGK-NeoR-LoxP This paper N/A pCLJ-F2P-LoxP-PGK-PuroR-LoxP This paper N/A pk335-CARGO-6mer-26kb-USP This paper N/A pk335-CARGO-6mer-24kb-USP This paper N/A pk335-CARGO-6mer-20kb-USP This paper N/A pk335-CARGO-6mer-11kb-USP This paper N/A pk335-CARGO-6mer-7.5kb-USP This paper N/A pk335-CARGO-6mer-4.5kb-USP This paper N/A pk335-CARGO-6mer-3.3kb-USP This paper N/A pk335-CARGO-6mer-2.1kb-USP This paper N/A pk335-CARGO-6mer-700bp-USP This paper N/A pk335-CARGO-6mer-PrPro This paper N/A pk335-CARGO-6mer-58kb-USF This paper N/A pk335-CARGO-6mer-55kb-USF This paper N/A pk335-CARGO-6mer-42kb-USF This paper N/A (Continued on next page) Molecular Cell 85, 1–15.e1–e7, January 16, 2025 e2
Protease Inhibitor:Article Title: Long-range regulation of transcription scales with genomic distance in a gene-specific manner.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Cas9- mouse monoclonal Active Motif Cat# 61757; RRID: AB_2793760 H3K27ac- rabbit polyclonal Active Motif Cat# 39133 RNA Pol II-mouse monoclonal Biolegend Cat# 664906; RRID: AB_2565554 H3K27me3- mouse monoclonal Active Motif Cat# 61017; RRID: AB_2614987 Bacterial and virus strains 10-beta Competent E. coli (high efficiency) NEB C3019H Stable Competent E. coli (high efficiency) NEB C3040H Chemicals, peptides, and recombinant proteins Accutase Sigma-Aldrich Cat# A6964-100ML DMEM/F12 Gibco Cat# 11320082 GSK3b CHIR99021 Selleck Chemicals Cat# S2924 MEK inhibitor PD0325901 Selleck Chemicals Cat# S1036 Gem21 NeuroPlex, without vitamin A Gemini Bio Cat# 400-161-010 N2 Neuroplex Gemini Bio Cat# 400-163-005 Sodium pyruvate (100mM) Gibco Cat# 11360070 2-mercaptoethanol Gibco Cat# 21985023 Anti-Anti (100x) Sigma-Aldrich Cat# A5955-100ML MEM NEAA (100x) ThemoFisher Cat# 11140050 LIF homemade N/A Laminin Gibco Cat# 23017015 Poly-l-ornithine Sigma-Aldrich Cat# P4638 BSA, Serum replacement Gemini Bio Cat# 700-104-100 Doxycycline hyclate EMD Millipore Cat# 32438501GM EED226 Selleck Chemicals Cat# S8496 Trizol reagent Invitrogen Cat# 10296010 Blasticidin, (10mg/mL) Invivogen Cat# ant-bl-1 G418, (100mg/mL) Invivogen Cat# ant-gn-1 Puromycin (10mg/mL) Invivogen Cat# ant-pr-1 Fibronectin ThermoFisher Cat# FC01010MG cOmplete, EDTA-free Protease inhibitor cocktail Millipore Sigma Cat# 11873580001 PMSF Sigma-Aldrich Cat# 10837091001 RNaseOUT Invitrogen Cat# 10777019 oligo dT (20mer) IDT N/A BsrGI NEB Cat# R3575 10xTango Buffer ThermoFisher Cat# BY5 10x T4 Ligase Buffer NEB Cat# B0202S T7 DNA Ligase NEB Cat# M0318S T4 DNA Ligase (30U/uL) ThermoFisher Cat# EL0013 BpiI ThermoFisher Cat# ER1011 Herculease II Dusion DNA Polymerase Agilent Technologies Cat# 600675 Critical commercial assays Lipofectamine 2000 ThermoFisher Cat# 11668019 Dynabeads Protein G Immunoprecipitation Invitrogen Cat# 10004D (Continued on next page) e1 Molecular Cell 85, 1–15.e1–e7, January 16, 2025 .. REAGENT or RESOURCE SOURCE IDENTIFIER ChIP Clean & Concentrator-5 Zymo Cat# D5205 Direct-zol RNA miniprep Zymo Cat# R2052 DNA Clean and Concentrator-5 Zymo Cat# D4014 High-Capacity cDNA Reverse Transcripase kit ThermoFisher Cat# 4368814 Sensifast SYBR No-Rox 2x mix Meridian Bioline Cat# 98020 Plasmid Safe ATP-dependent DNase Lucigen Cat# 3110K Gentra Puregene kit Qiagen Cat# 158745 Deposited data CRISPRa screen This paper Geo: GSE278772 ORCA processed datasets This paper 4DN data portal: 4DNESPB3DGGV 4DNESSKEUB9C 4DNESWTEQEBQ Experimental models: Cell lines R1 mESC WT Nagy et al.49 N/A R1 mESC, Fgf5-t2A-eGFP/tetO-dCas9-VPR This paper N/A R1 mESC, tetO-dCas9-VPR, clone 1 This paper N/A R1 mESC, tetO-dCas9-VPR, clone 2 This paper N/A R1 mESC, tetO-dCas9-VPR, P2FF2P-promoter swap This paper N/A R1 mESC, tetO-dCas9-VPR, delLTR, clone 12 This paper N/A Oligonucleotides Oligonucleotides used in this paper, Table S1 This paper N/A Guides in CRISPRa screen, Table S2 This paper N/A CARGO oligos and guides, Table S3 This paper N/A Recombinant DNA Super piggyBac Transposase expression vector System Biosciences (SBI) Cat# PB210PA-1 PhiC31 Integrase expression vector System Biosciences (SBI) Cat# FC200PA-1 pAAV-Ef1a-mCherry-IRES-Cre Fenno et al.50 Addgene Cat# 55632 pCAG-Flpe Matsuda et al.51 Addgene Cat#1 3787 pk335-BFP This paper N/A pX458-mCherry Gu et al.35 Addgene Cat# 161974 pCLJ-Fgf5-t2A-eGFP This paper N/A pCLJ-P2F-Frt-PGK-Puro-Frt This paper N/A pCLJ-F2P-LoxP-PGK-NeoR-LoxP This paper N/A pCLJ-F2P-LoxP-PGK-PuroR-LoxP This paper N/A pk335-CARGO-6mer-26kb-USP This paper N/A pk335-CARGO-6mer-24kb-USP This paper N/A pk335-CARGO-6mer-20kb-USP This paper N/A pk335-CARGO-6mer-11kb-USP This paper N/A pk335-CARGO-6mer-7.5kb-USP This paper N/A pk335-CARGO-6mer-4.5kb-USP This paper N/A pk335-CARGO-6mer-3.3kb-USP This paper N/A pk335-CARGO-6mer-2.1kb-USP This paper N/A pk335-CARGO-6mer-700bp-USP This paper N/A pk335-CARGO-6mer-PrPro This paper N/A pk335-CARGO-6mer-58kb-USF This paper N/A pk335-CARGO-6mer-55kb-USF This paper N/A pk335-CARGO-6mer-42kb-USF This paper N/A (Continued on next page) Molecular Cell 85, 1–15.e1–e7, January 16, 2025 e2
Immunoprecipitation:Article Title: Long-range regulation of transcription scales with genomic distance in a gene-specific manner.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Cas9- mouse monoclonal Active Motif Cat# 61757; RRID: AB_2793760 H3K27ac- rabbit polyclonal Active Motif Cat# 39133 RNA Pol II-mouse monoclonal Biolegend Cat# 664906; RRID: AB_2565554 H3K27me3- mouse monoclonal Active Motif Cat# 61017; RRID: AB_2614987 Bacterial and virus strains 10-beta Competent E. coli (high efficiency) NEB C3019H Stable Competent E. coli (high efficiency) NEB C3040H Chemicals, peptides, and recombinant proteins Accutase Sigma-Aldrich Cat# A6964-100ML DMEM/F12 Gibco Cat# 11320082 GSK3b CHIR99021 Selleck Chemicals Cat# S2924 MEK inhibitor PD0325901 Selleck Chemicals Cat# S1036 Gem21 NeuroPlex, without vitamin A Gemini Bio Cat# 400-161-010 N2 Neuroplex Gemini Bio Cat# 400-163-005 Sodium pyruvate (100mM) Gibco Cat# 11360070 2-mercaptoethanol Gibco Cat# 21985023 Anti-Anti (100x) Sigma-Aldrich Cat# A5955-100ML MEM NEAA (100x) ThemoFisher Cat# 11140050 LIF homemade N/A Laminin Gibco Cat# 23017015 Poly-l-ornithine Sigma-Aldrich Cat# P4638 BSA, Serum replacement Gemini Bio Cat# 700-104-100 Doxycycline hyclate EMD Millipore Cat# 32438501GM EED226 Selleck Chemicals Cat# S8496 Trizol reagent Invitrogen Cat# 10296010 Blasticidin, (10mg/mL) Invivogen Cat# ant-bl-1 G418, (100mg/mL) Invivogen Cat# ant-gn-1 Puromycin (10mg/mL) Invivogen Cat# ant-pr-1 Fibronectin ThermoFisher Cat# FC01010MG cOmplete, EDTA-free Protease inhibitor cocktail Millipore Sigma Cat# 11873580001 PMSF Sigma-Aldrich Cat# 10837091001 RNaseOUT Invitrogen Cat# 10777019 oligo dT (20mer) IDT N/A BsrGI NEB Cat# R3575 10xTango Buffer ThermoFisher Cat# BY5 10x T4 Ligase Buffer NEB Cat# B0202S T7 DNA Ligase NEB Cat# M0318S T4 DNA Ligase (30U/uL) ThermoFisher Cat# EL0013 BpiI ThermoFisher Cat# ER1011 Herculease II Dusion DNA Polymerase Agilent Technologies Cat# 600675 Critical commercial assays Lipofectamine 2000 ThermoFisher Cat# 11668019 Dynabeads Protein G Immunoprecipitation Invitrogen Cat# 10004D (Continued on next page) e1 Molecular Cell 85, 1–15.e1–e7, January 16, 2025 .. REAGENT or RESOURCE SOURCE IDENTIFIER ChIP Clean & Concentrator-5 Zymo Cat# D5205 Direct-zol RNA miniprep Zymo Cat# R2052 DNA Clean and Concentrator-5 Zymo Cat# D4014 High-Capacity cDNA Reverse Transcripase kit ThermoFisher Cat# 4368814 Sensifast SYBR No-Rox 2x mix Meridian Bioline Cat# 98020 Plasmid Safe ATP-dependent DNase Lucigen Cat# 3110K Gentra Puregene kit Qiagen Cat# 158745 Deposited data CRISPRa screen This paper Geo: GSE278772 ORCA processed datasets This paper 4DN data portal: 4DNESPB3DGGV 4DNESSKEUB9C 4DNESWTEQEBQ Experimental models: Cell lines R1 mESC WT Nagy et al.49 N/A R1 mESC, Fgf5-t2A-eGFP/tetO-dCas9-VPR This paper N/A R1 mESC, tetO-dCas9-VPR, clone 1 This paper N/A R1 mESC, tetO-dCas9-VPR, clone 2 This paper N/A R1 mESC, tetO-dCas9-VPR, P2FF2P-promoter swap This paper N/A R1 mESC, tetO-dCas9-VPR, delLTR, clone 12 This paper N/A Oligonucleotides Oligonucleotides used in this paper, Table S1 This paper N/A Guides in CRISPRa screen, Table S2 This paper N/A CARGO oligos and guides, Table S3 This paper N/A Recombinant DNA Super piggyBac Transposase expression vector System Biosciences (SBI) Cat# PB210PA-1 PhiC31 Integrase expression vector System Biosciences (SBI) Cat# FC200PA-1 pAAV-Ef1a-mCherry-IRES-Cre Fenno et al.50 Addgene Cat# 55632 pCAG-Flpe Matsuda et al.51 Addgene Cat#1 3787 pk335-BFP This paper N/A pX458-mCherry Gu et al.35 Addgene Cat# 161974 pCLJ-Fgf5-t2A-eGFP This paper N/A pCLJ-P2F-Frt-PGK-Puro-Frt This paper N/A pCLJ-F2P-LoxP-PGK-NeoR-LoxP This paper N/A pCLJ-F2P-LoxP-PGK-PuroR-LoxP This paper N/A pk335-CARGO-6mer-26kb-USP This paper N/A pk335-CARGO-6mer-24kb-USP This paper N/A pk335-CARGO-6mer-20kb-USP This paper N/A pk335-CARGO-6mer-11kb-USP This paper N/A pk335-CARGO-6mer-7.5kb-USP This paper N/A pk335-CARGO-6mer-4.5kb-USP This paper N/A pk335-CARGO-6mer-3.3kb-USP This paper N/A pk335-CARGO-6mer-2.1kb-USP This paper N/A pk335-CARGO-6mer-700bp-USP This paper N/A pk335-CARGO-6mer-PrPro This paper N/A pk335-CARGO-6mer-58kb-USF This paper N/A pk335-CARGO-6mer-55kb-USF This paper N/A pk335-CARGO-6mer-42kb-USF This paper N/A (Continued on next page) Molecular Cell 85, 1–15.e1–e7, January 16, 2025 e2
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