REAGENT or RESOURCE	SOURCE	IDENTIFIER
Antibodies
Mouse monoclonal antibody (mAb14)	Ku et al., 2021	N/A
Bacterial and virus strains
TransforMax EPI300 Chemically Competent E. coli	Lucigen Corporation	Cat#C300C105
NEB Stable Competent E. coli	New England Biolabs	Cat#C3040I
icSARS-CoV-2-mNG reporter virus	Xie et al., 2020a	N/A
icSARS-CoV-2 virus	Xie et al., 2020a	N/A
Biological samples
COVID-19 patient serum samples	UTMB Hospital	IRB#20-0070
Chemicals, peptides, and recombinant proteins
Puromycin	Thermo Fisher Scientific	Cat#A1113802
Doxycycline	Sigma-Aldrich	Cat#D3447
Polybrene	Sigma-Aldrich	Cat#TR-1003
Remdesivir	Gilead Sciences	N/A
TRIzol LS Reagent	Thermo Fisher Scientific	Cat#10296028
Critical commercial assays
Lenti-X Packaging Single Shots (VSV-G)	Takara	Cat#631275
iScript SYBR Green One-Step kit	Bio-Rad	Cat#170-8893
Ingenio Electroporation solution	Mirus Bio LLC	Cat#MIR 50117
T7 mMessage mMachine kit	Thermo Fisher Scientific	Cat#AM1344
QIAquick Gel Extraction Kit	QIAGEN	Cat#28706
SuperScript™ IV One-Step RT-PCR kit	Thermo Fisher Scientific	Cat#12594025
NEBuilder® HiFi DNA Assembly kit	New England Biolabs	Cat#E5520S
Deposited data
Raw image data for electron microscopy	This paper; and Mendeley Data	https://data.mendeley.com/api/datasets/s2dm94z42t/draft/files/5ec65493-deeb-4669-b292-efae0e6539be
Experimental models: cell lines
HEK293T cells	ATCC	Cat#CRL-3216, RRID:CVCL_0063
Vero CCL-81 cells	ATCC	Cat#CCL-81, RRID:CVCL_0059
Vero E6 cells	ATCC	Cat#CRL-1586, RRID: CVCL_0574
Calu-3 cells	ATCC	Cat#HTB-55, RRID:CVCL_0609
A549-hACE2 cells	Mossel et al., 2005	N/A
Vero-ORF3-E cells	This paper	N/A
Experimental models: organisms/strains
Hamster: HsdHan:AURA	Envigo	Cat#8902M
Mouse: 2B6.Cg-Tg(K18-ACE2)2Prlmn/J	Jackson Labs	Cat#034860
Oligonucleotides
Primer pcov-F56-F1 (TATACGAAGTTATA TTCGATGCGGCCGCGTCTCAGAGTGCT TTGGTTTATGATAATAAG)	Sigma-Aldrich	N/A
Primer pncov-R5 (TCGCACTAGAATA AACTCTGAACTC)	Sigma-Aldrich	N/A
Primer pncov-F6 (AGTTCAGAGT TTATTCTAGTGCGAATAATTG CACTTTTGAATATG)	Sigma-Aldrich	N/A
Primer pncov-R6 (ATGGCTAGTGTAACT AGCAAGAATACCAC)	Sigma-Aldrich	N/A
Primer pncov-F7 (GTATTCTTGCTAGTT ACACTAGCCATCCTTACTGCGCTTCG)	Sigma-Aldrich	N/A
Primer pncov-R8 (AGGTCGACTCTAGAGGATCC)	Sigma-Aldrich	N/A
Primer cov-21115-F (CATTTGTGGGTTTATACAACAAAAG)	Sigma-Aldrich	N/A
Primer TRS2-S-R (GAAAAACA AACATTATCCGGTTAGTTGTT AACAAG)	Sigma-Aldrich	N/A
Primer TRS2-S-F (CTTGTTAACAACTAA CCGGATAATGTTTGTTTTTC)	Sigma-Aldrich	N/A
Primer S-TRS2-M-R (GAAAAACTAATA TAATATTTAATCCGGTTATGTGTAA TGTAATTTGACTCCTTTGAGC)	Sigma-Aldrich	N/A
Primer TRS2-M-F (CCGGATTAAATATT ATATTAGTTTTTCTG)	Sigma-Aldrich	N/A
Primer M-TRS2-R (GTAATAAGAAAG CGTCCGGGATGTAGCAACAGTG)	Sigma-Aldrich	N/A
Primer M-TRS2-F (CACTGTTGCTAC ATCCCGGACGCTTTCTTATTAC)	Sigma-Aldrich	N/A
Primer ORF6-TRS2-mNG-R (CTTTGCTCA CCATATCCGGTTAATCAATCTCC)	Sigma-Aldrich	N/A
Primer ORF6-TRS2-mNG-F (GGAGATTGA TTAACCGGATATGGTGAGCAAAG)	Sigma-Aldrich	N/A
Primer ORF7-TRS2-ORF8-R (CAAGAAA TTTCATATCCGGTTAGGCGTGACAAG)	Sigma-Aldrich	N/A
Primer ORF7-TRS2-ORF8-F (CTTGTCAC GCCTAACCGGATATGAAATTTCTTG)	Sigma-Aldrich	N/A
Primer ORF8-TRS2-N-R (CATTATCAG ACATTTTAGTTTATCCGGTTAGAT GAAATCTAAAACAACACGAACGTC)	Sigma-Aldrich	N/A
Primer TRS2-N-F (CCGGATAAAC TAAAATGTCTGATAATGG)	Sigma-Aldrich	N/A
Primer cov-28501-R (GGTGTTAATT GGAACGCCTTGTCC)	Sigma-Aldrich	N/A
Primer M-T130N-F (CCATGGCACTA TTCTGAACAGACCGCTTCTAGAAAG)	Sigma-Aldrich	N/A
Primer M-T130N-R (CTTTCTAGAAGCG GTCTGTTCAGAATAGTGCCATGG)	Sigma-Aldrich	N/A
Primer 5ʹUTR-TRS2-F (GATCTGTTCTCTAA CCGGATTTTAAAATCTGTGTG)	Sigma-Aldrich	N/A
Primer 5ʹUTR-TRS2-R (CACACAGATTTTA AAATCCGGTTAGAGAACAGATC)	Sigma-Aldrich	N/A
Primer EcoR1-mCherry-F (CACTTCCTAC CCTCGTAAAGAATTCGCCACCATGG TGAGCAAGGGCGAGGAG)	Sigma-Aldrich	N/A
Primer F2A-optE-R (GACACAAAAGA ATACATTGGCCCAGGGTTGGACTCGAC)	Sigma-Aldrich	N/A
Primer F2A-optE-F (CCCTGGGCCAA TGTATTCTTTTGTGTCTGAAG)	Sigma-Aldrich	N/A
Primer EcoR1-Cov-optE-R (GGGGAGGGA GAGGGGCGGGAATTCCTACACCAGCAG GTCGGGGACC)	Sigma-Aldrich	N/A
Primer EcoR1-IRES-F (TAGGAATTCCC GCCCCTCTCCCTCCCCCC)	Sigma-Aldrich	N/A
Primer EMCV-IRES-R (ATTATCATCG TGTTTTTCAAAGGAAAACCACG)	Sigma-Aldrich	N/A
Primer IRES-optORF3-F (GTTTTCCTTT GAAAAACACGATGATAATATGGA CCTGTTCATGAGAATC)	Sigma-Aldrich	N/A
Primer BamH1-Cov-optORF3-R (CTCGCAGGGGAGGTGGTCTGGATCC CTCACAGAGGAACAGATGTGGTGG)	Sigma-Aldrich	N/A
Primer CoV-T7-N-F (ACTGTAATACGA CTCACTATAGGATGTCTGATAATGGA CCCCAAAATC)	Sigma-Aldrich	N/A
Primer polyT-N-R ([T]37AGGCCTGAGTTGAGTCAGCAC)	Sigma-Aldrich	N/A
Primer CoV19-N2-F (TTACAAACATTGGCCGCAAA)	Sigma-Aldrich	N/A
Primer CoV19-N2-R (GCGCGACATTCCGAAGAA)	Sigma-Aldrich	N/A
Recombinant DNA
pUC57-CoV2-F1	Xie et al., 2020a	N/A
pCC1-CoV2-F2	Xie et al., 2020a	N/A
pCC1-CoV2-F3	Xie et al., 2020a	N/A
pUC57-CoV2-F4	Xie et al., 2020a	N/A
pUC57-CoV2-F5	Xie et al., 2020a	N/A
pUC57-CoV2-F6	Xie et al., 2020a	N/A
pCC1-CoV2-F7-mNG	Xie et al., 2020a	N/A
pCC1-CoV2-F567-mNG	This paper	N/A
pCC1-CoV2-F567-mNG-ΔORF3-E	This paper	N/A
pLVX-TetOne-Puro	Takara	Cat#631849
pLVX-ORF3-E	This paper	N/A
Synthesized SARS-CoV-2 ORF3 gene (sequence-optimized)	This paper	N/A
Synthesized SARS-CoV-2 E gene (sequence-optimized)	This paper	N/A
Software and algorithms
Geneious	Biomatters	N/A
ImageJ	NIH	N/A
Prism 9.0 software	GraphPad	N/A
Illustrator CC	Adobe	N/A
Image Lab	Bio-rad	N/A
BioRender	BioRender	N/A
QuantStudio Software v1.3	ThermoFisher Scientific	N/A
Other
T4 DNA Ligase	New England Biolabs	Cat#M0202S
BsaI restriction endonuclease	New England Biolabs	Cat#R3733S
Esp3I restriction endonuclease	New England Biolabs	Cat#R0734S
EcoRI restriction endonuclease	New England Biolabs	Cat#R3101S
BamHI restriction endonuclease	New England Biolabs	Cat#R3136S
Platinum SuperFi II DNA Polymerase	Thermo Fisher Scientific	Cat#12361010