| Antibodies |
| H3K27ac |
Abcam |
cat# ab4729, RRID:AB_2118291 |
| H3K4me1 |
Abcam |
cat# ab8895, RRID:AB_306847 |
| H3K4me3 |
Abcam |
cat# ab8580, RRID:AB_306649 |
| Gatal |
Abcam |
cat# ab11852, RRID:AB_298635 |
| Rad21 |
Abcam |
cat# ab992, RRID:AB_2176601 |
| Polli |
Santa Cruz Biotechnology |
cat# sc-899, RRID:AB_632359 |
| Nf-e2 |
Santa Cruz Biotechnology |
cat# sc-22827, RRID:AB_2152924 |
| Medi |
Bethyl |
cat# IHC-00149, RRID:AB_2144026, Discontinued |
| Brd4 |
Bethyl |
cat# IHC-00396, RRID:AB_1604188, |
| CTCF |
Cambridge Bioscience, supplier: Active Motif |
cat# 61311, RRID:AB_2614975 |
| CD71-FITC |
eBioscience |
cat# 11–0711-85, RRID:AB_465125 |
| ter119-PE |
BD Pharmingen |
cat # 553673, RRID:AB_394986 |
| Anti-FITC magnetic microbeads |
Miltenyi |
cat# 130–048-701, RRID:AB_244371 |
|
| Biological Samples |
|
| mouse embryonic blood |
this paper |
N/A |
| mouse adult blood |
this paper |
N/A |
| mouse adult brain |
this paper |
N/A |
| mouse fetal liver cells |
this paper |
N/A |
|
| Chemicals, peptides, and recombinant proteins |
|
| PFHM II |
Gibco |
cat# 12040077 |
| L-Ascorbic Acid |
Sigma |
cat# A4544–25G |
| Transferrin |
Merck |
cat# 10652202001 |
| LIF |
Cell Bioscience Cell Guidance Systems (supplier) |
cat# GFM200–100 |
| FCS used in mESC cultures |
Gibco |
cat# 10270–106 |
| Cesium chloride powder |
Merck |
cat# C4036–100g |
| UltraPure Ethidium bromide |
Life |
cat# 15585011 |
| OptiMEM medium |
Thermofisher Scientific |
cat# 31985062 |
| DpnII enzyme for Tiled-C library prep |
New England Biolabs |
cat# R0543M |
| T4 DNA Ligase, HC (30 U/uL)-5,000 units |
Life Technologies Ltd |
cat# EL0013 |
| Immolase DNA Polymerase |
Bioline |
cat# BIO-21046 |
|
| Critical commercial assays |
|
| Tapestation High-Sensitivity D1000 reagents |
Agilent |
cat# 5067–5585 |
| Tapestation High-Sensitivity D1000 screentapes |
Agilent |
cat# 5067–5584 |
| Tapestation D1000 ScreeTape |
Agilent |
cat# 5067–5582 |
| Tapestation D1000 reagents |
Agilent |
cat# 5067–5583 |
| RNA Screentape |
Agilent |
cat# 5067–5576 |
| Qubit dsDNA BR Assay Kit |
Invitrogen |
cat# Q32850
|
| Qubit dsDNA HS Assay Kit |
Invitrogen |
cat# Q32851
|
| Qubit BR RNA Assay Kit |
Invitrogen |
cat# Q10211
|
| Qubit HS RNA Assay Kit |
Invitrogen |
cat# Q32855
|
| KAPA Quantification kit |
KAPA |
cat# KK4824 |
| Illumina Tagment DNA Enzyme and Buffer Large Kit |
Illumina |
cat# 20034198 |
| NEBNext High-Fidelity 2x PCR Master Mix |
NEB |
cat# M0541 |
| NextSeq® 500/550 High Output Kit v2.5(75 cycles) |
illumina |
cat# 20024906 |
| NEBNext Ultra II Directional RNA Library Prep Kit |
NEB |
cat# E7760 |
| NEBNext rRNA Depletion Kit (Human/ Mouse/Rat) |
NEB |
cat# E6310 |
| NEBNext® Ultra™ II DNA Library Prep with Sample Purification Beads |
NEB |
cat# E7103S |
| Applied BiosystemsTaqMan Universal PCR Master Mix - 5mL |
Thermofisher Scientific |
cat# 4304437 |
| NEBNext Multiplex Oligos for Illumina |
NEB |
cat# E7335/E7500 |
| SuperScript™ III First-Strand Synthesis SuperMix for qRT-PCR |
Thermofisher Scientific |
cat# 11752050 |
| Lipofectamine™ LTX Reagent with PLUS™ Reagent |
Thermofisher Scientific |
cat# 15338100 |
| Hba-a1/2 (FAM-MGB) |
Thermofisher Scientific |
Assay ID Mm02580841_g1 |
| Hbb-b (FAM-MGB) |
Thermofisher Scientific |
Assay ID Mm01611268_g1 |
| Hbb-y (FAM-MGB) |
Thermofisher Scientific |
Assay ID Mm00433936_g1 |
| Hba-x (FAM-MGB) |
Thermofisher Scientific |
Assay ID Mm00439255_m1 |
| Snrnp25 (FAM-MGB) |
Thermofisher Scientific |
Assay ID Mm00547218_m1 |
| Hbb-bh1 (FAM-MGB) |
Thermofisher Scientific |
Assay ID Mm00433932_g1 |
| Mpg (FAM-MGB) |
Thermofisher Scientific |
Assay ID Mm00447872_m1 |
| Nprl3 (FAM-MGB) |
Thermofisher Scientific |
Assay ID Mm01193449_m1 |
| Rhbdf1 (FAM-MGB) |
Thermofisher Scientific |
Assay ID Mm00711711_m1 |
| RPS18 (FAM-MGB) |
Thermofisher Scientific |
Assay ID Mm02601777_g1 |
| LS selection columns |
Miltenyi 130–042-401 |
cat# 130–042-401 |
| Direct-zol MicroPrep Kit |
Zymo Research |
cat# R2060 |
| AMPure XP Beads |
Beckman Coulter |
cat# A63881 |
| Nextera XT Library Preparation kit |
illumina |
cat# FC-131–1024 |
| Dynabeads M-280 Streptavidin |
ThermoFisher |
cat# 11205D |
| NEBNext Poly(A) mRNA Magnetic Isolation Module |
NEB |
cat# E7490S |
|
| Deposited data |
|
| ChIP-seq, ATAC-seq, RNA-seq and NG Capture-C data (sequence reads and processed files) |
This paper, Gene Expression Omnibus |
GEO: GSE220463
|
|
| Experimental models: cell lines |
|
| 1- mDist mouse ESC line (RMGR ready) |
modified in Prof. Doug Higgs Lab |
DOI: https://doi.org/10.1016/j.cell.2006.11.044
|
| 2- R2-only mouse ESC line (RMGR ready) |
This paper, derived from mDist mouse ESC line, heterozygote for the R2-only BAC inserted allele. Used to generate the R2- only mouse model |
N/A |
| 3- Hemizygous mDist mouse ESC line (RMGR-ready on the undeleted allele) |
This paper, used as Wildtype and base line for other genetically modified clones |
N/A |
| 4- Hemizygous mDist mouse ESC line (RMGR-ready and and R2-only on the undeleted allele) |
This paper, used as R2-only mESC clone and base line for other genetically modified clones |
N/A |
| 5- R1-only mESC |
This paper, derived from mESC #4 in this list |
N/A |
| 6- R1R2-only mESC |
This paper, derived from mESC #4 in this list |
N/A |
| 7- R1R2R3-only mESC |
This paper, derived from mESC #3 in this list |
N/A |
| 8- R1R2R3Rm-only mESC |
This paper, derived from mESC #3 in this list |
N/A |
| 9- R1R2R3R4-only mESC |
This paper, derived from mESC #3 in this list |
N/A |
| 10- R1R3RmR4-only mESC |
This paper, derived from mESC #3 in this list |
N/A |
| 11- R2R3RmR4-only mESC |
This paper, derived from mESC #3 in this list |
N/A |
| 12- RmR3R4-only mESC |
This paper, derived from mESC #3 in this list |
N/A |
| 13- R1R2Rm-only mESC |
This paper, derived from mESC #3 in this list |
N/A |
| 14- R1R2R4-only mESC |
This paper, derived from mESC #3 in this list |
N/A |
| 15- R1R2R4-only mESC |
This paper, derived from mESC #3 in this list |
N/A |
| 16- R2R4[R1] mESC |
This paper, derived from mESC #4 in this list |
N/A |
| 17- R2[R4] mESC |
This paper, derived from mESC #4 in this list |
N/A |
| 18- R1R2R3[R4] mESC |
This paper, derived from mESC #4 in this list |
N/A |
| 19- R1R2HS1[R4] mESC |
This paper, derived from mESC #4 in this list |
N/A |
| 20- Δα-SE mESC |
This paper, derived from mESC #4 in this list |
N/A |
|
| Experimental models: organisms/strains |
|
| mouse R2-only model WT, Heterozygotes, Homozygotes analyzed from the same colony |
This paper, derived by microinjecting mDist-R2-only mouse ESCs into blastocysts from C57BL/6 mice and implanted into pseudopregnant females. |
N/A |
|
| Oligonucleotides |
| Guide RNA sequences |
This paper, designed and provided by the Genome Engineering Facility at the Weatherall Institute of Molecular Medicine by Dr Philip Hublitz; see Table S2
|
N/A |
| PCR primers for genome engineering screening |
This paper, designed and provided by the Genome Engineering Facility at the Weatherall Institute of Molecular Medicine by Dr Philip Hublitz; see Table S3
|
N/A |
| Tiled-C Capture oligonucleotides |
STAR Methods
|
DOI: https://doi.org/10.1038/s41467-020-16598-7
|
|
| Recombinant DNA |
|
| pSpCas9(BB)-2A-GFP (pX458) vector |
Addgene plasmid |
http://n2t.net/addgene:48138;RRID:Addgene_48138
|
| pSpCas9(BB)-2A-Ruby (pX458) |
This paper, modified version of the pX458-GFP vector, Provided by the Genome Engineering Facility at the Weatherall Institute of Molecular Medicine by Dr Philip Hublitz |
N/A |
| HDR donor vectors |
this paper, GeneArt Gene Synthesis custom design and in-house cloning |
N/A |
| pCAGGS-Cre-IRESpuro plasmid |
This paper, provided by the Genome Engineering Facility at the Weatherall Institute of Molecular Medicine |
DOI: https://doi.org/10.1038/sj.onc.1205530
|
| Flippase (Flp)-expressing vector |
This paper, provided by the Genome Engineering Facility at the Weatherall Institute of Molecular Medicine |
https://doi.org/10.1002/gene.1076
|
|
| Software and algorithms |
|
| CRISPOR |
https://doi.org/10.1093/nar/gky354
|
http://crispor.tefor.net/
|
| BreakingCas |
https://doi.org/10.1093/nar/gkw407
|
https://bioinfogp.cnb.csic.es/tools/breakingcas/
|
| ggplot2 |
https://doi.org/10.1007/978-3-319-24277-4
|
https://ggplot2.tidyverse.org/
|
| bowtie2 |
https://doi.org/10.1038/nmeth.1923
|
https://github.com/BenLangmead/bowtie2
|
| SAMtools |
https://doi.org/10.1093/bioinformatics/btp352
|
http://samtools.sourceforge.net
|
| deepTools (bamcoverage, FilteredRNAstrand) |
deeptools.ie-freiburg.mpg.de
|
https://doi.org/10.1093/nar/gkw257
|
| MACS2 |
https://doi.org/10.1186/GB-2008-9-9-R137
|
https://hbctraining.github.io/Intro-to-ChIPseq/lessons/05_peak_calling_macs.html
|
| DESeq2 |
https://genomebiology.biomedcentral.com/articles/10.1186/s13059-014-0550-8
|
http://www.bioconductor.org/packages/release/bioc/html/DESeq2.html
|
| MEME suite |
https://doi.org/10.1093/NAR/GKV416
|
http://meme-suite.org
|
| Diffbind, rgl, magick for Principal Component Analysis (PCA) |
RStudio |
http://bioconductor.org/packages/release/bioc/vignettes/DiffBind/inst/doc/DiffBind.pdf
|
| HiC-Pro |
https://doi.org/10.1186/S13059-015-0831-X
|
http://github.com/nservant/HiC-Pro
|
| star alignment tool |
https://doi.org/10.1093/BIOINFORMATICS/BTS635
|
http://code.google.com/p/rna-star/
|
| Rsubread featurecounts |
https://doi.org/10.1093/nar/gkz114
|
http://www.bioconductor.org
|
| edgeR expression differential analysis |
RStudio |
https://www.bioconductor.org/packages/devel/bioc/vignettes/edgeR/inst/doc/edgeRUsersGuide.pdf
|
| bedtools |
https://doi.org/10.1093/BIOINFORMATICS/BTQ033
|
http://code.google.com/p/bedtools
|
| JASPAR |
https://doi.org/10.1093/nar/gkab1113
|
https://jaspar.genereg.net/
|
| SASQUATCH |
https://doi.org/10.1101/gr.220202.117
|
https://github.com/Hughes-Genome-Group/sasquatch
|
| Plotgardner |
https://doi.org/10.1093/bioinformatics/btac057
|
https://github.com/PhanstielLab/plotgardener
|
|
| Other |
|
| Beckman Coulter OptiSealpolypropylene Centrifuge tube 56 tubes and plugs (13 × 48mm) |
Beckman Coulter |
cat# 361621 |
