REAGENT or RESOURCE	SOURCE	IDENTIFIER
Antibodies
Rabbit polyclonal anti-CA (1:1,000)	Welker et al., 2000	N/A
Mouse monoclonal anti-laminB (1:200)	Santa Cruz	Cat#sc-365962
Rabbit polyclonal anti-CPSF6 (1:250)	Merck	Cat#HPA039973
Sheep polyclonal anti-CA (1:5,000)	Müller et al., 2009	N/A
Rabbit polyclonal anti-IN (1:1,000)	Welker et al., 2000	N/A
Rabbit polyclonal anti-RT (1:1,000)	Müller et al., 2004	N/A
Rabbit polyclonal anti-GFP (1:1,000)	Müller et al., 2004	N/A
Mouse monoclonal anti-CA, FITC conjugated (KC57) (1:100)	Beckman Coulter	Cat#6604665
Donkey anti-rabbit IgG, Alexa Fluor 488 conjugated (1:1,000)	ThermoFisher Scientific	Cat#A-21206; RRID:AB_2535792
Donkey anti-mouse IgG, Alexa Fluor 647 conjugated (1:1,000)	ThermoFisher Scientific	Cat#A-31571; RRID:AB_162542
Donkey anti-rabbit IgG, IRDye 800CW conjugated (1:10,000)	LI-COR Biosciences	Cat#926-32213; RRID:AB_621848
Donkey anti-rabbit IgG, IRDye 680RD conjugated (1:10,000)	LI-COR Biosciences	Cat#926-68073 RRID:AB_10954442
Donkey anti-sheep IgG, IRDye 800CW conjugated (1:10,000)	LI-COR Biosciences	No longer available; RRID:AB_220181
Bacterial and virus strains
HIV-1NL4-3	Adachi et al., 1986	N/A
NNHIV (HIV-1NL4-3 IND64N/D116N tatΔ33-64bp)	This study	N/A
NNHIV-A77V (HIV-1NL4-3 IND64N/D116N tatΔ33-64bp CAA77V)	This study	N/A
AAV1P5	Börner et al., 2020	N/A
Chemicals, peptides, and recombinant proteins
mCLING.Atto647N	Synaptic Systems	Cat#710006AT1
Efavirenz	Division AIDS, NIAID	NIH AIDS Reagent Program Cat#4624
Enfuvirtide (T-20)	Roche	NIH AIDS Reagent Program Cat#9845
Retronectin	Takara Bio	Cat#T100B
TetraSpeck beads	ThermoFisher Scientific	Cat#T7279
Hoechst33258	Merck	Cat#94403
DNase I	Merck	Cat#DN25
Polyethylenimine (PEI)	Merck	Cat#408727
Deposited data
Cryo-EM density map of NPC from HIV-infected Human CD4+ T lymphoblasts	This study	EMDB: 11967
Cryo-EM density map of HIV-1 CA hexameric unit	Mattei et al., 2016	EMDB: 3465
Cryo-EM density map of full HIV-1 capsid	Mattei et al., 2016	N/A
Experimental models: cell lines
Human CD4+ T lymphoblast cells SupT1-R5	R. Doms (University of Pennsylvania, USA)	RRID:CVCL_WU89
Human embryonic kidney 293 T cells (HEK293T)	Pear et al., 1993	RRID:CVCL_1926; ATCC Cat#CRL-11268
Oligonucleotides
NL4-3 IND64N forward primer: GTAGCCCAGGAATATGGCA GCTAAACTGTACACATTTAGAAGGAAAAG	This study	N/A
NL4-3 IND64N reverse primer: CTTTTCCTTCTAAATGTGTAC AGTTTAGCTGCCATATTCCTGGGCTAC	This study	N/A
NL4-3 IND116N forward primer: GCAGGAAGATGGCCAGT AAAAACAGTACATACAAATAATGGCAGCAATTTCACCAG TACTACAGTTAAGG	This study	N/A
NL4-3 IND116N reverse primer: CCTTAACTGTAGTACT GGTGAAATTGCTGCCATTATTTGTATGTACTGTTTTT ACTGGCCATCTTCCTGC	This study	N/A
mScarlet forward primer: AGGACGAGGACCGGGATCCA CCGGTCGCCACCATGGTG	This study	N/A
mScarlet reverse primer: TGATTATGATCTAGAGTC GCTTACTTGTACAGCTCGTCCATGCC	This study	N/A
AAV eGFP forward primer: GAGCGCACCATCTTCTTCAAG	Michler et al., 2016	N/A
AAV eGFP reverse primer: TGTCGCCCTCGAACTTCAC	Michler et al., 2016	N/A
AAV eGFP probe: FAM-ACGACGGCAACTACA-BHQ1	Michler et al., 2016	N/A
Gag forward primer: CATGTTTTCAGCATTATCAGAAGGA	Palmer et al., 2003	N/A
Gag reverse primer: TGCTTGATGTCCCCCCACT	Palmer et al., 2003	N/A
Gag probe, FAM-CCACCCCACAAGATTTAAACACCAT GCTAA-BHQ1	Palmer et al., 2003	N/A
2-LTR forward primer: CTAACTAGGGAACCCACTGCT	Puertas et al., 2018	N/A
2-LTR reverse primer: GTAGTTCTGCCAATCAGGGAA	Puertas et al., 2018	N/A
2-LTR probe: FAM-AGCCTCAATAAAGCTTGCCTTGA GTGC-BHQ1	Puertas et al., 2018	N/A
RPP30 forward primer: GATTTGGACCTGCGAGCG	Hindson et al., 2011	N/A
RPP30 reverse primer: GCGGCTGTCTCCACAAGT	Hindson et al., 2011	N/A
RPP30 probe: FAM-CTGACCTGAAGGCTCT-BHQ1	Hindson et al., 2011	N/A
Recombinant DNA
pNLC4-3	Bohne and Kräusslich, 2004	N/A
pNNHIV (pNLC4-3 IND64N/D116N tatΔ33-64bp)	This study	N/A
pUC19 NL4-35’CA-3’Vpr	J. Daecke (University of Heidelberg, Germany)	N/A
pNLC4-3 tatΔ33-64bp	Bejarano et al., 2019	N/A
pNNHIV-A77V	This study	N/A
pNL4-3-A77V	Bejarano et al., 2019	N/A
pVpr.IN.eGFP	A. Cereseto (CIBIO, Mattareo, Italy); Albanese et al., 2008	N/A
pmScarlet C1	Bindels et al., 2017	Addgene Cat#85042; RRID:Addgene_85042
pVpr.IND64N/D116N.eGFP	D. A. Bejarano (University of Heidelberg, Germany)	N/A
pVpr.IN.mScarlet	This study	N/A
pVpr.INNN.mScarlet (Vpr.IND64N/D116N.mScarlet)	This study	N/A
AAV helper plasmid encoding rep and 1P5 cap gene	Börner et al., 2020	N/A
CPSF6 triple shRNA	Bejarano et al., 2019	N/A
NS control AAV	Börner et al., 2010	N/A
AAV helper plasmid providing helper functions for AAV production	Matsushita et al., 1998	N/A
Software and algorithms
FIJI (v 2.1.0/1.53c)	Schindelin et al., 2012	RRID:SCR_002285
IMOD (v 4.9.2 and v 4.9.4)	Kremer et al., 1996	RRID:SCR_003297
Icy (v. 2.0.3.0)	de Chaumont et al., 2012	RRID:SCR_010587
ec-CLEM (Icy plugin; v 1.0.1.5)	Paul-Gilloteaux et al., 2017	http://icy.bioimageanalysis.org/plugin/ec-clem/
SerialEM (v 3.7.9)	Mastronarde, 2005	RRID:SCR_017293
MATLAB (v 2016b)	MathWorks	RRID:SCR_001622
MATLAB scripts for NPC diameter measurement	C. E. Zimmerli (EMBL, Heidelberg, Germany)	https://doi.org/10.5281/zenodo.4266419
novaSTA package	B. Turoňová (EMBL, Heidelberg, Germany)	https://github.com/turonova/novaSTA
novaCTF	Turoňová et al., 2017	https://github.com/turonova/novaCTF
gCTF	Zhang, 2016	https://www2.mrc-lmb.cam.ac.uk/research/locally-developed-software/zhang-software/
UCSF Chimera (v 1.14)	Pettersen et al., 2004	RRID:SCR_004097
Place Object (UCSF Chimera plug-in)	Qu et al., 2018	https://www2.mrc-lmb.cam.ac.uk/groups/briggs/resources/place-object/
Amira (v 2019.3 and v 2020.1)	ThermoFisher Scientific	RRID:SCR_007353
Imaris (v 9.3)	Oxford Instruments	RRID:SCR_007370
Volocity (v 6.3)	Perkin Elmer	RRID:SCR_002668
Flowjo (v 10.5.0)	FlowJo, BD Biosciences	RRID:SCR_008520
LI-COR Image Studio (v 5.2.5)	LI-COR Biosciences	RRID:SCR_015795
Prism (v 5.01 and v 8.4.3)	GraphPad Software	RRID:SCR_002798
Other
anti-HIV-1 p24 Hybridoma (183-H12-5C)	B. Chesebro (NIAID, Montana, USA)	NIH AIDS Reagent Program Cat#1513