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. 2020 Jun 4;78(5):975–985.e7. doi: 10.1016/j.molcel.2020.03.027
REAGENT or RESOURCE SOURCE IDENTIFIER
Antibodies
Mouse monoclonal anti-yeast Rad53 Susan Gasser; Hauer et al., 2017 N/A
Rat monoclonal anti-tubulin, clone YL1/2 Sigma-Aldrich Cat# 92092402-1VL, RRID: CVCL_J781
Rabbit polyclonal anti-DNA Ligase 1 Elabscience Cat# E-AB-31210
Mouse monoclonal anti-Chk1, clone 2G1D5 Cell Signaling Technology Cat# 2360, RRID: AB_2080320
Rabbit polyclonal anti-phospho-Chk1 (Ser345) Cell Signaling Technology Cat# 2341, RRID: AB_330023
Goat anti-mouse immunoglobulins, HRP Dako Cat# P0447, RRID: AB_2617137
Goat anti-rabbit immunoglobulins, HRP Dako Cat# P0448, RRID: AB_2617138
Goat anti-rat immunoglobulins, HRP Dako Cat# P0450, RRID: AB_2630354
Goat anti-rat IgG Secondary Antibody, IRDye® 800CW LI-COR Cat# 926-32219, RRID: AB_1850025
Chemicals, Peptides, and Recombinant Proteins
Zymolyase 20T AMS Biotechnology Cat# 120491-1
T4 DNA ligase, 20,000,000 U/mL New England Biolabs Cat# B0202
Q5® High-Fidelity DNA polymerase New England Biolabs Cat# M0491
RNase A from bovine pancreas Sigma-Aldrich Cat# 10109169001
Proteinase K Roche Cat# 3115801001
Phenylmethanesulfonyl fluoride Sigma-Aldrich Cat# P7626
β-Agarase I, 1000 U/mL New England Biolabs Cat# M0392
Alpha-Factor peptide (WHWLQLKPGQPMY), > 95% ProteoGenix SAS Cat# GM-PT001
BsrDI New England Biolabs Cat# R0574
Nb.BsrDI New England Biolabs Cat# R0648
NotI New England Biolabs Cat# R0189
Nb.BtsI New England Biolabs Cat# R0707
Antarctic Phosphatase New England Biolabs Cat# M0289
T4 Endonuclease V (T4 PDG) New England Biolabs Cat# M0308
APE1 New England Biolabs Cat# M0282
hAAG New England Biolabs Cat# M0313
Lipofectamine RNAiMAX Transfection Reagent Thermo Fisher Scientific Cat# 13778150
Benzonase Sigma-Aldrich Cat# E1014-25KU
PhosSTOP Phosphatase Inhibitor Sigma-Aldrich Cat# 04906837001
Opti-MEM® Reduced Serum Medium Thermo Fisher Scientific Cat# 11058021
Critical Commercial Assays
AMPure XP beads Beckman Coulter Cat# A63881
NEBNext® Ultra II DNA Library Prep Kit for Illumina® New England Biolabs Cat# E7645
Phusion Flash high-fidelity PCR master mix Thermo Fisher Scientific Cat# F-548
Dynabeads MyOne Streptavidin C1 Life Technologies Cat# 65001
High Sensitivity D1000 ScreenTape Agilent Technologies Cat# 5067-5584
High Sensitivity D1000 ScreenTape reagents Agilent Technologies Cat# 5067-5585
RNA ScreenTape Agilent Technologies Cat# 5067-5576
RNA ScreenTape sample buffer Agilent Technologies Cat# 5067-5577
RNA ScreenTape ladder Agilent Technologies Cat# 5067-5578
Qubit dsDNA HS Assay Kit Invitrogen Cat# Q32854
Agilent Bioanalyzer High Sensitivity DNA Kit Agilent Technologies Cat# 5067-4646
NextSeq 550 System High-Output Kit Illumina Cat# 20024906
NextSeq 550 System Mid-Output Kit Illumina Cat# 20024904
NuSieve GTG Agarose Lonza Cat# 859081
Yeast Genomic DNA Extraction Kit QIAGEN Cat# 10243
Bio-Rad Protein Assay Dye Reagent Bio-Rad Cat# 500-0006
NuPAGE LDS sample buffer Thermo Fisher Scientific Cat# NP0007
NuPAGE 4-12% Bis-Tris Protein Gels Thermo Fisher Scientific Cat# NP0322
Deposited Data
Raw and analyzed data This paper GEO: GSE134225
Human reference genome UCSC GRCh37/hg19 UCSC Genome Browser ftp://hgdownload.soe.ucsc.edu/goldenPath/hg19/chromosomes/
Yeast reference genome UCSC sacCer3 UCSC Genome Browser ftp://hgdownload.soe.ucsc.edu/goldenPath/sacCer3/chromosomes/
3′ end maps of Nb.BtsI-digested DNA, WT and pol1-L868M, Fastq (Figure 2A, reanalyzed with GLOE-Pipe) Reijns et al., 2015 GEO: GSM1573437, GSM1573438
Motifs for Abf1, Reb1, Rap1 in yeast, GFF (Figure 4B) MacIsaac et al., 2006 https://downloads.yeastgenome.org/published_datasets/MacIsaac_2006_PMID_16522208/MacIsaac_high_confidence_with_sequence.gff
Nucleosome positions in yeast, BED (Figure 4B) Whitehouse et al., 2007 https://static-content.springer.com/esm/art%3A10.1038%2Fnature06391/MediaObjects/41586_2007_BFnature06391_MOESM319_ESM.xls
Okazaki fragment maps in Cdc9-depleted yeast, Fastq, rep 1 & rep2 (Figure 4C, reanalyzed with GLOE-Pipe) Smith and Whitehouse, 2012 GEO: GSM835650, GSM835651
END-seq data in HCT116, Fastq (Figure 5, reanalyzed with GLOE-Pipe) Tubbs et al., 2018 GEO: GSM3227952
DNase I hypersensitivity data in HCT116, BigWig, rep1 & rep2 (Figure 6B) ENCODE Project Consortium, 2012 GEO: GSM736493, GSM736600
OK-Seq and S50 data in HeLa and GM06990, rep1 & rep2 (BedGraph, received from Chunlong Chen, chunlong.chen@curie.fr) (Figure 6B) Petryk et al., 2016 GEO SRA: SRP065949 (SRX1427549, SRX1427548, SRX1424659, SRX1424656)
Experimental Models: Cell Lines
HCT116 Cancer Research UK London Research Institute Cell Services N/A
HCT116 LIG3−/−:mL3 Oh et al., 2014 N/A
Experimental Models: Organisms/Strains
S. cerevisiae: strain W303 ATCC ATCC: 201238
S. cerevisiae: strain JKM179 Lee et al., 1998 N/A
S. cerevisiae: strain DF5 ATCC ATCC: 200912
S. cerevisiae: strain DF5 apn1::KanMX apn2::KanMX This paper N/A
S. cerevisiae: strain cdc9AID Kubota et al., 2013 N/A
Oligonucleotides
Primer #3898: CTACACGACGCTCTTCCGATCTNNN
NN∗N-NH2 (∗phosphorothioate bond, IDT code ∗; NH2: 3′-amino modification, IDT code /3AmMO/)		 Integrated DNA Technologies N/A
Primer #3899: PO4-AGATCGGAAGAGCGTCGTGTAG
GGAAAGAGTG TAGATCTCGTTTT-Bio (PO4: 5′-phosphorylation, IDT code /5Phos/; T-Bio: 3′-biotin-dT, IDT code /3BiodT/)		 Integrated DNA Technologies N/A
Primer #3790: CGAGATCTACACTCTTTCCCTACA
CGACGCTCTTCCGATCT		 Integrated DNA Technologies N/A
Primer #3791: GACTGGAGTTCAGACGTGTGCTC
TTCCGATCT		 Integrated DNA Technologies N/A
Primer #3792: GATCGGAAGAGCACACGTCTG
AACTCCAGTC		 Integrated DNA Technologies N/A
Primer P5: AATGATACGGCGACCACCGAGATCT
ACACTCTTTCCCTACACGACGCTCTTCCGATCT		 Integrated DNA Technologies N/A
Primer P7: CAAGCAGAAGACGGCATACGAGAT
(X)6GTGACTGGAGTTCAGACGTGTGCT
CTTCCGATCT		 Integrated DNA Technologies N/A
siRNA targeting sequence: Silencer Select Negative Control No. 1 Thermo Fisher Scientific Cat# 4390843
siRNA targeting sequence: Ligase 1: Silencer Select s8174 Thermo Fisher Scientific Cat# 4390824
Software and Algorithms
bcl2fastq, version 2.19 Illumina https://support.illumina.com/sequencing/sequencing_software/bcl2fastq-conversion-software.html
Bpipe, version 0.9.9.3 Sadedin et al., 2012 http://docs.bpipe.org/
NGSpipe2go Institute of Molecular Biology gGmbH https://github.com/imbforge/NGSpipe2go
FastQC, version 0.11.5 Andrews, 2019 https://www.bioinformatics.babraham.ac.uk/projects/fastqc/
Trimmomatic, version 0.36 Bolger et al., 2014 http://www.usadellab.org/cms/?page=trimmomatic
Bowtie 2, version 2.3.4 Langmead and Salzberg, 2012 http://bowtie-bio.sourceforge.net/bowtie2/index.shtml
Samtools, version 1.5 Li et al., 2009 http://samtools.sourceforge.net/
BEDTools, version 2.25.0 Quinlan and Hall, 2010 https://bedtools.readthedocs.io/en/latest/
bedGraphToBigWig, version 365 Kent et al., 2010 https://github.com/ENCODE-DCC/kentUtils
MACS2 callpeak, version 2.1.1 Zhang et al., 2008 https://github.com/taoliu/MACS
ChIPseeker package, version 1.14.1 Yu et al., 2015 https://bioconductor.org/packages/release/bioc/html/ChIPseeker.html
deepTools, version 3.1.0 Ramírez et al., 2014 https://deeptools.readthedocs.io/en/develop/
Replicon Gindin et al., 2014 https://github.com/RepliconBioinfo/Replicon [downloaded 20.12.2019]
FlowJo, version 10.6.1 FlowJo, LLC https://www.flowjo.com/
GLOE-Pipe This paper https://github.com/helle-ulrich-lab/ngs-gloepipe
Other
Detailed protocol for the preparation of GLOE-Seq libraries This paper See Supplemental Information
3D model of a custom-made mold for agarose plugs This paper See Data S1
3D model of a tool for extrusion of agarose plugs from custom-made mold This paper See Data S1