REAGENT or RESOURCE	SOURCE	IDENTIFIER
Antibodies
Rabbit-polyclonal anti-phospho-ATR-T1989p	GenTex	#GTX128145; RRID: AB_2687562
Mouse-monoclonal anti-α-tubulin (Clone B512)	Sigma-Aldrich	#T5168; RRID: AB_477579
Mouse-monoclonal anti-AIM-1 (AuroraB)	BD Biosciences	#611082; RRID: AB_2227708
Rabbit-polyclonal anti-CHMP4B	ProteinTech	#13683-1-AP; RRID: AB_2877971
Rabbit-polyclonal anti-ATRIP	Bethyl	#A300-095A; RRID: AB_242514
Rabbit-polyclonal anti-phospho-RPA32-S33p	Bethyl	#A300-246A; RRID: AB_2180847
Mouse-monoclonal anti-ATR (C-1)	SantaCruz Biotechnology	#sc-515173; RRID: AB_2893291
Mouse-monoclonal anti-SC35	Abcam	#ab11826; RRID: AB_298608
Mouse-monoclonal anti-PML	Abcam	#ab96051; RRID: AB_10679887
Mouse-monoclonal anti-Coilin	Abcam	# ab11822; RRID: AB_2081428
Mouse-monoclonal anti-CENPA	Abcam	# ab13939; RRID: AB_300766
Rabbit-polyclonal anti-RPA70 (RPA1)	Bethyl	# A300-241A; RRID: AB_2180681
Mouse-monoclonal anti-γ-tubulin	Sigma-Aldrich	#T6557; RRID: AB_477584
Mouse-monoclonal anti-LAP2β	BD Biosciences	# 611000; RRID: AB_398313
Rabbit-polyclonal anti-ATRIP	Millipore	# 07-625; RRID: AB_310761
Rabbit-polyclonal anti-Actin	Sigma-Aldrich	# A2066; RRID: AB_476693
Mouse-monoclonal anti-RPA, clone RPA34-19	Millipore	# MABE286; RRID: AB_11203661
Mouse-monoclonal anti-TOPBP1	BD Biosciences	# 611875; RRID: AB_399355
Rabbit-polyclonal anti-ETAA1	Novus Biologicals	#NBP1-90473; RRID: AB_11012531
Rabbit-polyclonal anti-ZFYVE19 (ANCHR)	Bethyl	# A301-808A; RRID: AB_1233064
Rabbit-polyclonal anti-MKLP1 (N-19)	SantaCruz Biotechnology	# sc-867; RRID :AB_631959
Rabbit-polyclonal anti-NUP153	Bethyl	# A301-788A; RRID: AB_1211259
Rabbit-polyclonal anti-CHK1 (FL-476)	SantaCruz Biotechnology	# sc-7898; RRID: AB_2229488
Rabbit-polyclonal anti-phospho-CHK1-S317p (D12H3)	Cell Signaling Technology	# 12302; RRID: AB_2783865
Rabbit-polyclonal anti-phospho-MCM2-S40/41p	Fitzgerald et al. (2014)	N/A
Rabbit-polyclonal anti-CHMP4C	Abcam	ab155668; RRID: AB_2892638
Goat anti-Mouse IgG-heavy and light chain cross-adsorbed Antibody DyLight® 550	Bethyl	# A90-516D3; RRID: AB_10634387
Goat anti-Rabbit IgG-heavy and light chain cross-adsorbed Antibody FITC Conjugated	Bethyl	# A120-201F; RRID: AB_67263
Goat anti-Mouse IgG (H + L) Secondary Antibody, HRP	ThermoFisher	# 31430; RRID: AB_228307
Peroxidase-AffiniPure Goat Anti-Rabbit IgG (H + L)	Jackson ImmunoResearch	# 111-035-144; RRID: AB_2307391
Bacterial and virus strains
One Shot™ TOP10 Chemically Competent E. coli	Invitrogen	#C404010
Chemicals, peptides, and recombinant proteins
ATRi (ETP-46464)	SelleckChem	#S8050
ATRi (VE821)	SelleckChem	#S8007
ATMi (KU55933)	SelleckChem	#S1092
DNA-PKi (NU7026)	SelleckChem	#S2893
CHK1i (MK-8776)	SelleckChem	#S2735
CDC7i (XL413)	SelleckChem	#S7547
CDC7i (PHA-767491)	SelleckChem	#S2742
AuroraBi (AZD1152)	SelleckChem	#S1147
Benzonase	Sigma	#E1014
Paraformaldehyde	EMS	#15710
VectraShield with DAPI	VectorLabs	#H-1200
Lambda Protein Phosphatase (Lambda PP)	NEB	#P0753S
Deposited data
Raw and analyzed data	This paper	This paper
Experimental models: Cell lines
HeLa	ATCC	#CCL-2
HeLa H2B-GFP/mCherry-tubulin	Kaczmarczyk and Sullivan (2014)	N/A
hTERT-RPE1	ATCC	#CRL-4000
Oligonucleotides
Silencer™ Negative Control No. 1 siRNA	Ambion	# AM4611
siLuciferase GL2: seq: 5′-CGUACGCGGAAUACUUCGA-3′	Eurofins	N/A
siATR; seq: 5′-CCUCCGUGAUGUUGCUUGA-3′	Eurofins	N/A
siATRIP; seq: 5′-AAGGUCCACAGAUUAUUAGAU-3′	Eurofins	N/A
siRPA2; seq: 5′-GCACCUUCUCAAGCCGAAA-3′	Eurofins	N/A
siRPA1; seq: 5′-AACUGGUUGACGAAAGUGGUG-3′	Eurofins	N/A
siTOPBP1	Dharmacon	# L-012358-00-0005
siETAA1; seq: 5′-UGACAAAGCAGUUAGGUAA-3′	Eurofins	N/A
siNUP153	Dharmacon	# L-005283-00-0005
siCHMP4B	Dharmacon	# L-018075-01-0010
siCHMP4C	Dharmacon	# L-015932-01-0010
Software and algorithms
ImageJ	Schindelin et al. (2012)	https://ImageJ.nih.gov/ij/
Deltavision SoftWoRx	SoftWoRx