KEY RESOURCES TABLE
| REAGENT or RESOURCE | SOURCE | IDENTIFIER |
|---|---|---|
| Antibodies | ||
| Anti-ACTB (WB) | Sigma-Aldrich # A2228 | RRID:AB_476697 |
| Anti-CTCF (IP, ChIP) | Abcam # ab128873 | RRID:AB_11144295 |
| Anti-CTCF (WB) | Millipore # 07–729 | RRID:AB_441965 |
| Anti-FLAG (IP) | Sigma-Aldrich # F7425 | RRID:AB_439687 |
| Anti-FLAG (WB, IP) | Sigma-Aldrich # F3165 | RRID:AB_259529 |
| Anti-H3 (WB) | Abcam # ab1791 | RRID:AB_302613 |
| Anti-HA tag (WB) | Abcam # ab9110 | RRID:AB_307019 |
| Anti-HaloTag (WB) | Promega # G9211 | RRID:AB_2688011 |
| Anti-Smc1a (ChIP) | Bethyl # A300–055A | RRID:AB_2192467 |
| Anti-V5 (IP) | Abcam # ab9166 | RRID:AB_307024 |
| Anti-V5 (WB) | Thermo Fisher Scientific # R960–25 | RRID:AB_2556564 |
| Mouse IgG (IP, ChIP) | Jackson ImmunoResearch Labs # 015–000-003 |
RRID:AB_2337188 |
| Rabbit IgG (IP, ChIP) | Jackson ImmunoResearch Labs # 011–000-003 |
RRID:AB_2337118 |
| ANTI-FLAG M2 Affinity Gel (in vitro RNA binding assay) | Sigma-Aldrich #A2220 | RRID:AB_10063035 |
| Chemicals, Peptides, and Recombinant Proteins | ||
| DAPI 4′,6-Diamidine-2′-phenylindole dihydrochloride | Sigma-Aldrich | Cat. # 10236276001 |
| HaloTag TMR ligand | Promega | Cat. # G8251 |
| HaloTag PA-JF549 ligand | Grimm et al., 2016 | N/A |
| Benzonase | Millipore | Cat. # 71205 |
| RNase A | Thermo Scientific | Cat. # EN0531 |
| DNase I | Ambion | Cat. # AM2222 |
| Formaldehyde | Polysciences | Cat. # 1881420 |
| 10 mM dNTPs | KAPA Biosystems | Cat. # KK1017 |
| 10 mM ATP | New England Biolabs | Cat. # P0756S |
| 5 U/μl T4 DNA polymerase | Invitrogen | Cat. # 18005025 |
| 5 U/μl Taq DNA polymerase | ThermoFisher Scientific | Cat. # EP0401 |
| KAPA HS HIFI polymerase | KAPA Biosystems | Cat. # KK2502 |
| Rapid DNA ligase | Enzymatics | Cat. # L6030-HC-L |
| AMPure XP beads | Agencourt | Cat. # A63880 |
| DSG (disuccinimidyl glutarate) | ThermoFisher Scientific | Cat. # 20593 |
| Micrococcal Nuclease | Worthington Biochem | Cat. # LS004798 |
| 100mM ATP | ThermoFisher Scientific | Cat. # R1441 |
| DNA Polymerase I, Large (Klenow) Fragment | New England Biolabs | Cat. # M0210 |
| T4 Polynucleotide Kinase | New England Biolabs | Cat. # M0201 |
| T4 DNA Ligase | New England Biolabs | Cat. # M0202 |
| Exonuclease III (E. coli) | New England Biolabs | Cat. # M0206 |
| Biotin-14-dATP | Jena Bioscience | Cat. # NU-835-BIO14 |
| Biotin-11-dCTP | Jena Bioscience | Cat. # NU-809-BIOX |
| 20X Proteinase K solution | Sigma Aldrich | Cat. # 3115879001 |
| Dynabeads MyOne Streptavidin C1 | ThermoFisher Scientific | Cat. # 65001 |
| SuperScript III Reverse Transcriptase | Invitrogen | Cat. # 18080044 |
| SYBR Gold Nucleic Acid Gel Stain | Invitrogen | Cat. # S11494 |
| PageBlue Protein Staining Solution | ThermoFisher Scientific | Cat. # 24620 |
| TRIzol Reagent | ThermoFisher Scientific | Cat. # 15596026 |
| Ni-NTA agarose | QIAGEN | Cat. # 30210 |
| 3X FLAG Peptide | Sigma Aldrich | Cat. # F4799 |
| recombinant 3xFLAG-Halo-wt-CTCF-6xHis protein | This manuscript | r-wt-CTCF |
| recombinant 3xFLAG-Halo-ΔRBRi-CTCF-6xHis protein | This manuscript | r-ΔRBRi-CTCF |
| EMPIGEN BB detergent | Sigma-Aldrich | Cat. # 30326 |
| Critical Commercial Assays | ||
| Click-iT EdU Alexa Fluor 488 Flow Cytometry Assay Kit | ThermoFisher Scientific | Cat. # C10425 |
| NEBNext Ultra II | New England Biolabs | Cat. # E7645 |
| End-It DNA End-Repair | Lucigen | Cat. # ER81050 |
| RNeasy Mini Kit | QIAGEN | Cat. # 74104 |
| DNA-free DNA Removal Kit | Invitrogen | Cat. # AM1906 |
| Ribo-Zero rRNA Removal Kit | Illumina | Cat. # MRZH116 |
| TruSeq RNA Sample Preparation v2 Kit | Illumina | Cat. # RS-122–2001 |
| HiScribe T7 Quick High Yield RNA Synthesis Kit | New England Biolabs | Cat. # E2050S |
| Bac-to-Bac Baculovirus Expression System | ThermoFisher Scientific | Cat. # 10359016 |
| Deposited Data | ||
| Raw imaging data (all raw data for This manuscript) | This manuscript and Hansen et al., 2018b | https://zenodo.org/record/2208323 |
| Micro-C, ChIP-Seq and RNA-Seq data | This manuscript | https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE123636 |
| Raw images and uncropped gels and blots deposited in Mendeley Data | This manuscript | DOI: https://doi.org/10.17632/5zdrpcsbt9.2 |
| CTCF ChIP-Seq in mESCs | Chen et al., 2008 | GSE11431 |
| Smc1a ChIP-Seq in mESCs | Kagey et al., 2010 | GSE22562 |
| RNA-seq in mESCs after CTCF degradation | Nora et al., 2017 | GSE98671 |
| Experimental Models: Cell Lines | ||
| Mouse: JM8.N4 mouse embryonic stem cells | Pettitt et al., 2009 and UC Davis KOMP Repository | https://www.komp.org/pdf.php?cloneID=8669 |
| mESC C59 FLAG-Halo-CTCF; Rad21- SNAPf-V5 (knock-in) |
Hansen et al., 2017 | C59 |
| mESC C59D2 ΔRBRi-FLAG-Halo-CTCF (e10::3xHA); Rad21-SNAPf-V5 (knock-in) | This manuscript | C59D2 or ΔRBRi |
| mESC C62 3xFLAG-Halo-CTCF (allele 1); V5-SNAPf-Halo-CTCF (allele 2) (both knock-in) | This manuscript | C62 |
| Oligonucleotides | ||
| See Table S1 | N/A | |
| Recombinant DNA | ||
| pBlueScript SK II (+) | Addgene # 212205 | GenBank: X52328.1 |
| pBSII HR mCtcf. delRBRi(link-3XHA) Repair Vector | This manuscript | N/A |
| pUC57 V5 Snap(f) mCTCF Repair Vector | This manuscript | N/A |
| pUC57 3xFLAG Halo TEV mCTCF Repair Vector | This manuscript | N/A |
| pFastBac Dual Expression Vector | ThermoFisher Scientific | Cat. # 10712024 |
| Software and Algorithms | ||
| MATLAB 2014b | The Mathworks | 2014b |
| PALM analysis pipeline (MATLAB) | This manuscript | https://gitlab.com/anders.sejr.hansen/palm_pipeline |
| FCSREAD (MATLAB) | Mathworks File Exchange | https://www.mathworks.com/matlabcentral/fileexchange/8430-flow-cytometry-data-reader-and-visualization |
| MTT-Algorithm (MATLAB implementation) | Sergé et al., 2008 | https://gitlab.com/tjian-darzacq-lab/SPT_LocAndTrack |
| SimpleTracker (MATLAB) | Jean-Yves Tinevez | https://www.mathworks.com/matlabcentral/fileexchange/34040-simple-tracker |
| Spatial Point Patterns Analysis (ads package on CRAN) | Pélissier and Goreaud, 2015 | https://cran.r-project.org/web/packages/ads/index.html |
| RStudio | RStudio | https://www.rstudio.com |
| BaSDI (MATLAB) | Elmokadem and Yu, 2015 | https://github.com/jiyuuchc/BaSDI |
| ImageJ (https://imagej.net/) | Schindelin et al., 2012 | RRID:SCR_003070 |
| Samtools | Li et al., 2009 | RRID:SCR_002105 |
| Integrative Genomics Viewer |
Robinson et al., 2011; Thorvaldsdóttir et al., 2013 |
RRID:SCR_011793 |
| Bowtie | Langmead et al., 2009 | RRID:SCR_005476 |
| deepTools | Ramírez et al., 2016 | RRID:SCR_016366 |
| FastQC | http://www.bioinformatics.babraham.ac.uk/projects/fastqc | RRID:SCR_014583 |
| MACS2 | Zhang et al., 2008 | https://github.com/taoliu/MACS/ |
| Python 3.7 | Python | https://www.python.org/ |
| Anaconda 3.7 | Anaconda | https://www.anaconda.com/ |
| HiC-Pro | Servant et al., 2015 | https://github.com/nservant/HiC-Pro |
| Cooler | Abdennur and Mirny, 2019 | https://github.com/mirnylab/cooler |
| Juicer tools | Durand et al., 2016 | https://github.com/aidenlab/juicer |
| Higlass | Kerpedjiev et al., 2018 | http://higlass.io/ |
| STAR | Dobin et al., 2013 | https://github.com/alexdobin/STAR |
| HTSeq | Anders et al., 2015 | https://pypi.org/project/HTSeq/ |
| edgeR | Robinson et al., 2010 | https://bioconductor.org/packages/release/bioc/html/edgeR.html |
| Liu et al., 2015 | ||
| Galaxy |
Blankenberg et al., 2010; Giardine et al., 2005; Goecks et al., 2010 |
https://usegalaxy.org/ |
| DESeq2 | Love et al., 2014 | https://bioconductor.org/packages/release/bioc/html/DESeq2.html |
| DAVID Bioinformatics Resources 6.8 | Huang et al., 2009a, 2009b | https://david.ncifcrf.gov/home.jsp |
| ViSP | El Beheiry and Dahan, 2013 | https://www.nature.com/articles/nmeth.2566 |