Isolation:Article Title: Rewiring of the epigenome and chromatin architecture by exogenously induced retinoic acid signaling during zebrafish embryonic development
Article Snippet: .. Isolated cells were lysed (lysis buffer: 10 mM Tris–HCl pH 8, 10 mM NaCl, 0.3% Igepal CA-630 [Sigma-Aldrich, I8896] and 1x protease inhibitor cocktail [Complete, Roche, 11697498001]), and the DNA was digested with DpnII (New England BioLabs, R0543M) and Csp6I (Fermentas, Thermo Scientific, FD0214) as primary and secondary enzymes, respectively. .. T4 DNA ligase (Promega, M1804) was used for both ligation steps.
Article Title: Nucleosome placement and polymer mechanics explain genomic contacts on 100kbp scales
Article Snippet: .. Chromatin was fragmented by subjecting isolated nuclei to primary restriction enzyme digestion using the 4-bp cutter Csp6I (ThermoFisher, Cat. ER0211) to achieve optimal contact-mapping resolution, followed by in situ ligation in a reaction mix containing ligase buffer and ligase, incubated overnight at 16 o C. Post-ligation, cross-links were reversed by overnight incubation with Proteinase K at 65 o C. DNA was purified using SPRI beads and subjected to a second restriction enzyme digestion using NlaIII (NEB, Cat. R0125L) to generate sticky ends for circularization. .. The digested DNA was then ligated again, and the resulting 3C templates were purified using AMPure XP beads (Beckman Coulter, Cat. A63881), and quantified using the Qubit dsDNA BR Assay Kit.
Article Title: Long-range enhancer-controlled genes are hypersensitive to regulatory factor perturbations.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Mouse monoclonal anti-RAD21 Merck Cat#05–908; RRID:AB_417383 Mouse monoclonal anti-GAPDH Santa Cruz Cat#sc-32233; RRID:AB_627679 Mouse monoclonal anti-FLAG Merck Cat#F1804; RRID:AB_262044 Mouse monoclonal anti-SF3B1 Santa Cruz Cat#sc-514655; RRID:AB_2941964 Goat monoclonal anti-Mouse IgG (H + L), HRP Conjugate Promega Cat#W4021; RRID:AB_430834 Mouse monoclonal anti-BrdU BD Pharmingen Cat#555627; RRID:AB_395993 Bacterial and virus strains ElectroMAX Stbl4 Competent Cells Invitrogen Cat#11635018 One Shot TOP10 Chemically Competent E. coli Invitrogen Cat#C404010 Chemicals, peptides, and recombinant proteins RPMI 1640 Gibco Cat#72400021 FBS Sigma-Aldrich Cat#F7524 Pen-strep Gibco Cat#15070063 Tryple Gibco Cat#12605010 Puromycin Merck Cat#P9620 Blasticidin Sigma-Aldrich Cat#15205 PEI Polysciences Cat#23966 Polybrene Sigma-Aldrich Cat#H9268-5G 5-Ph-IAA MedChemExpress Cat#HY-134653 dTAG7 TOCRIS Cat#6912 5-Bromouridine Sigma-Aldrich Cat#850187 BlpI ThermoFisher Cat#ER0091 BstXI ThermoFisher Cat#ER1021 Csp6I ThermoFisher CatR0125 NlaIII NEB Cat#ER0211 T4 DNA Ligase Roche Cat#10799009001 Proteinase K Roche Cat#3115801001 TRIzol Invitrogen Cat#15596026 RNAseOUT Invitrogen Cat#10777019 Critical commercial assays NEBNext Ultra II Directional RNA Library Prep Kit NEB Cat#E7760S NEBNext Poly(A) mRNA Magnetic Isolation Module NEB Cat#E7490S Q5 High-Fidelity DNA Polymerase NEB Cat# M0491S P-beads Macherey Nagel Cat#744100.34 Expand Long Template PCR system Roche Cat#11759060001 AMPure-XP beads Beckman Coulter Cat#A63881 Protein G Dynabeads ThermoFisher Cat#10004D SuperSignal West Pico PLUS Chemiluminescent Substrate ThermoFisher Cat#34580 (Continued on next page) Cell Genomics 5, 100778, March 12, 2025 e1 .. REAGENT or RESOURCE SOURCE IDENTIFIER 4–15% Mini-PROTEAN TGX Precast Protein Gel Biorad Cat#4561083 Deposited data Raw western blot images This study Mendeley Data; https://doi.org/10.17632/ r9878ks8nm.1 Reporter CRISPRi screen This study GEO; GSE281575 RAD21-AID_BrU-seq This study GEO; GSE281753 MED23-FKBP_BrU-seq This study GEO; GSE281753 LDB1-FKBP_BrU-seq This study GEO; GSE281753 CRISPRi_KD_RNAseq_highthroughput This study GEO; GSE281752 CRISPRi_KD_RNAseq_total This study GEO; GSE280672 4C-seq This study GEO; GSE281419; GSE281420; GSE281576 ENCODE-rE2G predictions, K562 Gschwind et al.10; ENCODE ENCFF202FID MED14-AID_SLAMseq Bell et al.48 GEO; GSE198944 JQ1_SLAMseq Muhar et al.53 GEO; GSE100708 LDB1_ChIPseq Guo et al.49 GEO; GSE142228 GENCODE Human reference genome GRCh38/hg38 GENCODE https://ftp.ebi.ac.uk/pub/databases/ gencode/Gencode_human/release_44/ GRCh38.primary_assembly.genome.fa.gz GENCODE Comprehensive gene annotation V44 GENCODE https://ftp.ebi.ac.uk/pub/databases/ gencode/Gencode_human/release_44/ gencode.v44.annotation.gtf.gz UCSC Human reference genome GRCh38/hg38 UCSC http://hgdownload.cse.ucsc.edu/ goldenpath/hg38 UCSC Human reference genome GRCh37/hg19 UCSC http://hgdownload.cse.ucsc.edu/ goldenpath/hg19 Experimental models: Cell lines K562_E0-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_E-50-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_E100-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_EC100-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_EC100-GFP_RAD21-AID; LV-Tir1 This study N/A K562_EC100-GFP_MED23-FKBP This study N/A K562_EC100-GFP_LDB1-FKBP This study N/A Oligonucleotides sgRNA library for CRISPRi screen Table S1 N/A Other oligonucleotides Table S3 N/A Recombinant DNA pU6-sgRNA EF1Alpha-puro-T2A-BFP Gilbert et al.96 Addgene #60955 pHR-SFFV-dCas9-BFP-KRAB Gilbert et al.96 Addgene #46911 pMK393 (mAID-mCherry-BSD) Yesbolatova et al.43 Addgene #121194 pMK381 (AAVS1 CMV-OsTIR1F74G) Yesbolatova et al.43 Addgene #140536 pSpCas9(BB)-2A-BFP This study N/A RAD21-AID targeting construct This study N/A pLV_Tir1 construct This study N/A MED23-FKBP targeting construct This study N/A LDB1-FKBP targeting construct This study N/A (Continued on next page) e2 Cell Genomics 5, 100778, March 12, 2025 OPEN ACCESS
Lysis:Article Title: Rewiring of the epigenome and chromatin architecture by exogenously induced retinoic acid signaling during zebrafish embryonic development
Article Snippet: .. Isolated cells were lysed (lysis buffer: 10 mM Tris–HCl pH 8, 10 mM NaCl, 0.3% Igepal CA-630 [Sigma-Aldrich, I8896] and 1x protease inhibitor cocktail [Complete, Roche, 11697498001]), and the DNA was digested with DpnII (New England BioLabs, R0543M) and Csp6I (Fermentas, Thermo Scientific, FD0214) as primary and secondary enzymes, respectively. .. T4 DNA ligase (Promega, M1804) was used for both ligation steps.
Protease Inhibitor:Article Title: Rewiring of the epigenome and chromatin architecture by exogenously induced retinoic acid signaling during zebrafish embryonic development
Article Snippet: .. Isolated cells were lysed (lysis buffer: 10 mM Tris–HCl pH 8, 10 mM NaCl, 0.3% Igepal CA-630 [Sigma-Aldrich, I8896] and 1x protease inhibitor cocktail [Complete, Roche, 11697498001]), and the DNA was digested with DpnII (New England BioLabs, R0543M) and Csp6I (Fermentas, Thermo Scientific, FD0214) as primary and secondary enzymes, respectively. .. T4 DNA ligase (Promega, M1804) was used for both ligation steps.
Concentration Assay:
In Situ:
Article Title: Nucleosome placement and polymer mechanics explain genomic contacts on 100kbp scales
Article Snippet: .. Chromatin was fragmented by subjecting isolated nuclei to primary restriction enzyme digestion using the 4-bp cutter Csp6I (ThermoFisher, Cat. ER0211) to achieve optimal contact-mapping resolution, followed by in situ ligation in a reaction mix containing ligase buffer and ligase, incubated overnight at 16 o C. Post-ligation, cross-links were reversed by overnight incubation with Proteinase K at 65 o C. DNA was purified using SPRI beads and subjected to a second restriction enzyme digestion using NlaIII (NEB, Cat. R0125L) to generate sticky ends for circularization. .. The digested DNA was then ligated again, and the resulting 3C templates were purified using AMPure XP beads (Beckman Coulter, Cat. A63881), and quantified using the Qubit dsDNA BR Assay Kit.
Ligation:Article Title: Nucleosome placement and polymer mechanics explain genomic contacts on 100kbp scales
Article Snippet: .. Chromatin was fragmented by subjecting isolated nuclei to primary restriction enzyme digestion using the 4-bp cutter Csp6I (ThermoFisher, Cat. ER0211) to achieve optimal contact-mapping resolution, followed by in situ ligation in a reaction mix containing ligase buffer and ligase, incubated overnight at 16 o C. Post-ligation, cross-links were reversed by overnight incubation with Proteinase K at 65 o C. DNA was purified using SPRI beads and subjected to a second restriction enzyme digestion using NlaIII (NEB, Cat. R0125L) to generate sticky ends for circularization. .. The digested DNA was then ligated again, and the resulting 3C templates were purified using AMPure XP beads (Beckman Coulter, Cat. A63881), and quantified using the Qubit dsDNA BR Assay Kit.
Incubation:Article Title: Nucleosome placement and polymer mechanics explain genomic contacts on 100kbp scales
Article Snippet: .. Chromatin was fragmented by subjecting isolated nuclei to primary restriction enzyme digestion using the 4-bp cutter Csp6I (ThermoFisher, Cat. ER0211) to achieve optimal contact-mapping resolution, followed by in situ ligation in a reaction mix containing ligase buffer and ligase, incubated overnight at 16 o C. Post-ligation, cross-links were reversed by overnight incubation with Proteinase K at 65 o C. DNA was purified using SPRI beads and subjected to a second restriction enzyme digestion using NlaIII (NEB, Cat. R0125L) to generate sticky ends for circularization. .. The digested DNA was then ligated again, and the resulting 3C templates were purified using AMPure XP beads (Beckman Coulter, Cat. A63881), and quantified using the Qubit dsDNA BR Assay Kit.
Purification:Article Title: Nucleosome placement and polymer mechanics explain genomic contacts on 100kbp scales
Article Snippet: .. Chromatin was fragmented by subjecting isolated nuclei to primary restriction enzyme digestion using the 4-bp cutter Csp6I (ThermoFisher, Cat. ER0211) to achieve optimal contact-mapping resolution, followed by in situ ligation in a reaction mix containing ligase buffer and ligase, incubated overnight at 16 o C. Post-ligation, cross-links were reversed by overnight incubation with Proteinase K at 65 o C. DNA was purified using SPRI beads and subjected to a second restriction enzyme digestion using NlaIII (NEB, Cat. R0125L) to generate sticky ends for circularization. .. The digested DNA was then ligated again, and the resulting 3C templates were purified using AMPure XP beads (Beckman Coulter, Cat. A63881), and quantified using the Qubit dsDNA BR Assay Kit.
Virus:Article Title: Long-range enhancer-controlled genes are hypersensitive to regulatory factor perturbations.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Mouse monoclonal anti-RAD21 Merck Cat#05–908; RRID:AB_417383 Mouse monoclonal anti-GAPDH Santa Cruz Cat#sc-32233; RRID:AB_627679 Mouse monoclonal anti-FLAG Merck Cat#F1804; RRID:AB_262044 Mouse monoclonal anti-SF3B1 Santa Cruz Cat#sc-514655; RRID:AB_2941964 Goat monoclonal anti-Mouse IgG (H + L), HRP Conjugate Promega Cat#W4021; RRID:AB_430834 Mouse monoclonal anti-BrdU BD Pharmingen Cat#555627; RRID:AB_395993 Bacterial and virus strains ElectroMAX Stbl4 Competent Cells Invitrogen Cat#11635018 One Shot TOP10 Chemically Competent E. coli Invitrogen Cat#C404010 Chemicals, peptides, and recombinant proteins RPMI 1640 Gibco Cat#72400021 FBS Sigma-Aldrich Cat#F7524 Pen-strep Gibco Cat#15070063 Tryple Gibco Cat#12605010 Puromycin Merck Cat#P9620 Blasticidin Sigma-Aldrich Cat#15205 PEI Polysciences Cat#23966 Polybrene Sigma-Aldrich Cat#H9268-5G 5-Ph-IAA MedChemExpress Cat#HY-134653 dTAG7 TOCRIS Cat#6912 5-Bromouridine Sigma-Aldrich Cat#850187 BlpI ThermoFisher Cat#ER0091 BstXI ThermoFisher Cat#ER1021 Csp6I ThermoFisher CatR0125 NlaIII NEB Cat#ER0211 T4 DNA Ligase Roche Cat#10799009001 Proteinase K Roche Cat#3115801001 TRIzol Invitrogen Cat#15596026 RNAseOUT Invitrogen Cat#10777019 Critical commercial assays NEBNext Ultra II Directional RNA Library Prep Kit NEB Cat#E7760S NEBNext Poly(A) mRNA Magnetic Isolation Module NEB Cat#E7490S Q5 High-Fidelity DNA Polymerase NEB Cat# M0491S P-beads Macherey Nagel Cat#744100.34 Expand Long Template PCR system Roche Cat#11759060001 AMPure-XP beads Beckman Coulter Cat#A63881 Protein G Dynabeads ThermoFisher Cat#10004D SuperSignal West Pico PLUS Chemiluminescent Substrate ThermoFisher Cat#34580 (Continued on next page) Cell Genomics 5, 100778, March 12, 2025 e1 .. REAGENT or RESOURCE SOURCE IDENTIFIER 4–15% Mini-PROTEAN TGX Precast Protein Gel Biorad Cat#4561083 Deposited data Raw western blot images This study Mendeley Data; https://doi.org/10.17632/ r9878ks8nm.1 Reporter CRISPRi screen This study GEO; GSE281575 RAD21-AID_BrU-seq This study GEO; GSE281753 MED23-FKBP_BrU-seq This study GEO; GSE281753 LDB1-FKBP_BrU-seq This study GEO; GSE281753 CRISPRi_KD_RNAseq_highthroughput This study GEO; GSE281752 CRISPRi_KD_RNAseq_total This study GEO; GSE280672 4C-seq This study GEO; GSE281419; GSE281420; GSE281576 ENCODE-rE2G predictions, K562 Gschwind et al.10; ENCODE ENCFF202FID MED14-AID_SLAMseq Bell et al.48 GEO; GSE198944 JQ1_SLAMseq Muhar et al.53 GEO; GSE100708 LDB1_ChIPseq Guo et al.49 GEO; GSE142228 GENCODE Human reference genome GRCh38/hg38 GENCODE https://ftp.ebi.ac.uk/pub/databases/ gencode/Gencode_human/release_44/ GRCh38.primary_assembly.genome.fa.gz GENCODE Comprehensive gene annotation V44 GENCODE https://ftp.ebi.ac.uk/pub/databases/ gencode/Gencode_human/release_44/ gencode.v44.annotation.gtf.gz UCSC Human reference genome GRCh38/hg38 UCSC http://hgdownload.cse.ucsc.edu/ goldenpath/hg38 UCSC Human reference genome GRCh37/hg19 UCSC http://hgdownload.cse.ucsc.edu/ goldenpath/hg19 Experimental models: Cell lines K562_E0-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_E-50-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_E100-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_EC100-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_EC100-GFP_RAD21-AID; LV-Tir1 This study N/A K562_EC100-GFP_MED23-FKBP This study N/A K562_EC100-GFP_LDB1-FKBP This study N/A Oligonucleotides sgRNA library for CRISPRi screen Table S1 N/A Other oligonucleotides Table S3 N/A Recombinant DNA pU6-sgRNA EF1Alpha-puro-T2A-BFP Gilbert et al.96 Addgene #60955 pHR-SFFV-dCas9-BFP-KRAB Gilbert et al.96 Addgene #46911 pMK393 (mAID-mCherry-BSD) Yesbolatova et al.43 Addgene #121194 pMK381 (AAVS1 CMV-OsTIR1F74G) Yesbolatova et al.43 Addgene #140536 pSpCas9(BB)-2A-BFP This study N/A RAD21-AID targeting construct This study N/A pLV_Tir1 construct This study N/A MED23-FKBP targeting construct This study N/A LDB1-FKBP targeting construct This study N/A (Continued on next page) e2 Cell Genomics 5, 100778, March 12, 2025 OPEN ACCESS
Recombinant:Article Title: Long-range enhancer-controlled genes are hypersensitive to regulatory factor perturbations.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Mouse monoclonal anti-RAD21 Merck Cat#05–908; RRID:AB_417383 Mouse monoclonal anti-GAPDH Santa Cruz Cat#sc-32233; RRID:AB_627679 Mouse monoclonal anti-FLAG Merck Cat#F1804; RRID:AB_262044 Mouse monoclonal anti-SF3B1 Santa Cruz Cat#sc-514655; RRID:AB_2941964 Goat monoclonal anti-Mouse IgG (H + L), HRP Conjugate Promega Cat#W4021; RRID:AB_430834 Mouse monoclonal anti-BrdU BD Pharmingen Cat#555627; RRID:AB_395993 Bacterial and virus strains ElectroMAX Stbl4 Competent Cells Invitrogen Cat#11635018 One Shot TOP10 Chemically Competent E. coli Invitrogen Cat#C404010 Chemicals, peptides, and recombinant proteins RPMI 1640 Gibco Cat#72400021 FBS Sigma-Aldrich Cat#F7524 Pen-strep Gibco Cat#15070063 Tryple Gibco Cat#12605010 Puromycin Merck Cat#P9620 Blasticidin Sigma-Aldrich Cat#15205 PEI Polysciences Cat#23966 Polybrene Sigma-Aldrich Cat#H9268-5G 5-Ph-IAA MedChemExpress Cat#HY-134653 dTAG7 TOCRIS Cat#6912 5-Bromouridine Sigma-Aldrich Cat#850187 BlpI ThermoFisher Cat#ER0091 BstXI ThermoFisher Cat#ER1021 Csp6I ThermoFisher CatR0125 NlaIII NEB Cat#ER0211 T4 DNA Ligase Roche Cat#10799009001 Proteinase K Roche Cat#3115801001 TRIzol Invitrogen Cat#15596026 RNAseOUT Invitrogen Cat#10777019 Critical commercial assays NEBNext Ultra II Directional RNA Library Prep Kit NEB Cat#E7760S NEBNext Poly(A) mRNA Magnetic Isolation Module NEB Cat#E7490S Q5 High-Fidelity DNA Polymerase NEB Cat# M0491S P-beads Macherey Nagel Cat#744100.34 Expand Long Template PCR system Roche Cat#11759060001 AMPure-XP beads Beckman Coulter Cat#A63881 Protein G Dynabeads ThermoFisher Cat#10004D SuperSignal West Pico PLUS Chemiluminescent Substrate ThermoFisher Cat#34580 (Continued on next page) Cell Genomics 5, 100778, March 12, 2025 e1 .. REAGENT or RESOURCE SOURCE IDENTIFIER 4–15% Mini-PROTEAN TGX Precast Protein Gel Biorad Cat#4561083 Deposited data Raw western blot images This study Mendeley Data; https://doi.org/10.17632/ r9878ks8nm.1 Reporter CRISPRi screen This study GEO; GSE281575 RAD21-AID_BrU-seq This study GEO; GSE281753 MED23-FKBP_BrU-seq This study GEO; GSE281753 LDB1-FKBP_BrU-seq This study GEO; GSE281753 CRISPRi_KD_RNAseq_highthroughput This study GEO; GSE281752 CRISPRi_KD_RNAseq_total This study GEO; GSE280672 4C-seq This study GEO; GSE281419; GSE281420; GSE281576 ENCODE-rE2G predictions, K562 Gschwind et al.10; ENCODE ENCFF202FID MED14-AID_SLAMseq Bell et al.48 GEO; GSE198944 JQ1_SLAMseq Muhar et al.53 GEO; GSE100708 LDB1_ChIPseq Guo et al.49 GEO; GSE142228 GENCODE Human reference genome GRCh38/hg38 GENCODE https://ftp.ebi.ac.uk/pub/databases/ gencode/Gencode_human/release_44/ GRCh38.primary_assembly.genome.fa.gz GENCODE Comprehensive gene annotation V44 GENCODE https://ftp.ebi.ac.uk/pub/databases/ gencode/Gencode_human/release_44/ gencode.v44.annotation.gtf.gz UCSC Human reference genome GRCh38/hg38 UCSC http://hgdownload.cse.ucsc.edu/ goldenpath/hg38 UCSC Human reference genome GRCh37/hg19 UCSC http://hgdownload.cse.ucsc.edu/ goldenpath/hg19 Experimental models: Cell lines K562_E0-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_E-50-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_E100-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_EC100-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_EC100-GFP_RAD21-AID; LV-Tir1 This study N/A K562_EC100-GFP_MED23-FKBP This study N/A K562_EC100-GFP_LDB1-FKBP This study N/A Oligonucleotides sgRNA library for CRISPRi screen Table S1 N/A Other oligonucleotides Table S3 N/A Recombinant DNA pU6-sgRNA EF1Alpha-puro-T2A-BFP Gilbert et al.96 Addgene #60955 pHR-SFFV-dCas9-BFP-KRAB Gilbert et al.96 Addgene #46911 pMK393 (mAID-mCherry-BSD) Yesbolatova et al.43 Addgene #121194 pMK381 (AAVS1 CMV-OsTIR1F74G) Yesbolatova et al.43 Addgene #140536 pSpCas9(BB)-2A-BFP This study N/A RAD21-AID targeting construct This study N/A pLV_Tir1 construct This study N/A MED23-FKBP targeting construct This study N/A LDB1-FKBP targeting construct This study N/A (Continued on next page) e2 Cell Genomics 5, 100778, March 12, 2025 OPEN ACCESS
Polymerase Chain Reaction:Article Title: Long-range enhancer-controlled genes are hypersensitive to regulatory factor perturbations.
Article Snippet: .. REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Mouse monoclonal anti-RAD21 Merck Cat#05–908; RRID:AB_417383 Mouse monoclonal anti-GAPDH Santa Cruz Cat#sc-32233; RRID:AB_627679 Mouse monoclonal anti-FLAG Merck Cat#F1804; RRID:AB_262044 Mouse monoclonal anti-SF3B1 Santa Cruz Cat#sc-514655; RRID:AB_2941964 Goat monoclonal anti-Mouse IgG (H + L), HRP Conjugate Promega Cat#W4021; RRID:AB_430834 Mouse monoclonal anti-BrdU BD Pharmingen Cat#555627; RRID:AB_395993 Bacterial and virus strains ElectroMAX Stbl4 Competent Cells Invitrogen Cat#11635018 One Shot TOP10 Chemically Competent E. coli Invitrogen Cat#C404010 Chemicals, peptides, and recombinant proteins RPMI 1640 Gibco Cat#72400021 FBS Sigma-Aldrich Cat#F7524 Pen-strep Gibco Cat#15070063 Tryple Gibco Cat#12605010 Puromycin Merck Cat#P9620 Blasticidin Sigma-Aldrich Cat#15205 PEI Polysciences Cat#23966 Polybrene Sigma-Aldrich Cat#H9268-5G 5-Ph-IAA MedChemExpress Cat#HY-134653 dTAG7 TOCRIS Cat#6912 5-Bromouridine Sigma-Aldrich Cat#850187 BlpI ThermoFisher Cat#ER0091 BstXI ThermoFisher Cat#ER1021 Csp6I ThermoFisher CatR0125 NlaIII NEB Cat#ER0211 T4 DNA Ligase Roche Cat#10799009001 Proteinase K Roche Cat#3115801001 TRIzol Invitrogen Cat#15596026 RNAseOUT Invitrogen Cat#10777019 Critical commercial assays NEBNext Ultra II Directional RNA Library Prep Kit NEB Cat#E7760S NEBNext Poly(A) mRNA Magnetic Isolation Module NEB Cat#E7490S Q5 High-Fidelity DNA Polymerase NEB Cat# M0491S P-beads Macherey Nagel Cat#744100.34 Expand Long Template PCR system Roche Cat#11759060001 AMPure-XP beads Beckman Coulter Cat#A63881 Protein G Dynabeads ThermoFisher Cat#10004D SuperSignal West Pico PLUS Chemiluminescent Substrate ThermoFisher Cat#34580 (Continued on next page) Cell Genomics 5, 100778, March 12, 2025 e1 .. REAGENT or RESOURCE SOURCE IDENTIFIER 4–15% Mini-PROTEAN TGX Precast Protein Gel Biorad Cat#4561083 Deposited data Raw western blot images This study Mendeley Data; https://doi.org/10.17632/ r9878ks8nm.1 Reporter CRISPRi screen This study GEO; GSE281575 RAD21-AID_BrU-seq This study GEO; GSE281753 MED23-FKBP_BrU-seq This study GEO; GSE281753 LDB1-FKBP_BrU-seq This study GEO; GSE281753 CRISPRi_KD_RNAseq_highthroughput This study GEO; GSE281752 CRISPRi_KD_RNAseq_total This study GEO; GSE280672 4C-seq This study GEO; GSE281419; GSE281420; GSE281576 ENCODE-rE2G predictions, K562 Gschwind et al.10; ENCODE ENCFF202FID MED14-AID_SLAMseq Bell et al.48 GEO; GSE198944 JQ1_SLAMseq Muhar et al.53 GEO; GSE100708 LDB1_ChIPseq Guo et al.49 GEO; GSE142228 GENCODE Human reference genome GRCh38/hg38 GENCODE https://ftp.ebi.ac.uk/pub/databases/ gencode/Gencode_human/release_44/ GRCh38.primary_assembly.genome.fa.gz GENCODE Comprehensive gene annotation V44 GENCODE https://ftp.ebi.ac.uk/pub/databases/ gencode/Gencode_human/release_44/ gencode.v44.annotation.gtf.gz UCSC Human reference genome GRCh38/hg38 UCSC http://hgdownload.cse.ucsc.edu/ goldenpath/hg38 UCSC Human reference genome GRCh37/hg19 UCSC http://hgdownload.cse.ucsc.edu/ goldenpath/hg19 Experimental models: Cell lines K562_E0-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_E-50-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_E100-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_EC100-GFP_dCas9-KRAB-BFP Rinzema et al.32 N/A K562_EC100-GFP_RAD21-AID; LV-Tir1 This study N/A K562_EC100-GFP_MED23-FKBP This study N/A K562_EC100-GFP_LDB1-FKBP This study N/A Oligonucleotides sgRNA library for CRISPRi screen Table S1 N/A Other oligonucleotides Table S3 N/A Recombinant DNA pU6-sgRNA EF1Alpha-puro-T2A-BFP Gilbert et al.96 Addgene #60955 pHR-SFFV-dCas9-BFP-KRAB Gilbert et al.96 Addgene #46911 pMK393 (mAID-mCherry-BSD) Yesbolatova et al.43 Addgene #121194 pMK381 (AAVS1 CMV-OsTIR1F74G) Yesbolatova et al.43 Addgene #140536 pSpCas9(BB)-2A-BFP This study N/A RAD21-AID targeting construct This study N/A pLV_Tir1 construct This study N/A MED23-FKBP targeting construct This study N/A LDB1-FKBP targeting construct This study N/A (Continued on next page) e2 Cell Genomics 5, 100778, March 12, 2025 OPEN ACCESS
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