LOCUS       sequence-418210-        7768 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses SARS-CoV-2 NSP2 in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7768)
  AUTHORS   Li J, Gui Q, Liang FX, Sall J, Zhang Q, Duan Y, Dhabaria A, Askenazi
            M, Ueberheide B, Stapleford KA, Pagano M
  TITLE     The REEP5/TRAM1 complex binds SARS-CoV-2 NSP3 and promotes virus 
            replication.
  JOURNAL   J Virol. 2023 Sep 28:e0050723. doi: 10.1128/jvi.00507-23.
  PUBMED    37768083
REFERENCE   2  (bases 1 to 7768)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "J Virol. 
            2023 Sep 28:e0050723. doi: 10.1128/jvi.00507-23."
COMMENT     Sequence Label: NSP2-Flag
FEATURES             Location/Qualifiers
     source          1..7768
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(14..33)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     133..155
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     promoter        279..383
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(384..452,453..1244)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEDRFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(602..621)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1415..2003
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1904..1923
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2157..2174
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        complement(2248..2352)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        2462..2765
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        2766..2969
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     2920..2940
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     2966..2990
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             3082..3105
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     protein_bind    3112..3136
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(5090..5114)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(5146..5165)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(5147..5165)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(5176..5193)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5176..5192)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5185..5207)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     polyA_signal    5597..5731
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5634..5653)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5688..5707
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      5857..6312
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5944..5963)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6154..6175
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6425..6782
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6425..6616
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6448..6468)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6633..6768
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6833..7627
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6887..6906)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7497..7516
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    7693..7741
                     /note="synthetic polyadenylation signal"
ORIGIN
        1 ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc
       61 caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag
      121 ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg
      181 cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg ataataatgg
      241 tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct atttgtttat
      301 ttttctaaat acattcagat atgtatccgc tcatgagaca ataaccctga taaatgcttc
      361 aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc cttattccct
      421 tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg aaagtaaaag
      481 atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc aacagcggta
      541 agatccttga gagttttcgc cccgaagacc gttttccaat gatgagcact tttaaagttc
      601 tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc ggtcgccgca
      661 tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag catcttacgg
      721 atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat aacactgcgg
      781 ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt ttgcacaaca
      841 tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa gccataccaa
      901 acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc aaactattaa
      961 ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg gaggcggata
     1021 aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt gctgataaat
     1081 ctggagctgg tgagcgtggg tctcgcggta tcattgcagc actggggcca gatggtaagc
     1141 cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat gaacgaaata
     1201 gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca gaccaagttt
     1261 actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg atctaggtga
     1321 agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg ttccactgag
     1381 cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa
     1441 tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg ccggatcaag
     1501 agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata ccaaatactg
     1561 ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca ccgcctacat
     1621 acctcgctct gctaatcctg ttaccagtgg ctgctgccgg tggcgataag tcgtgtctta
     1681 ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc tgaacggggg
     1741 gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga tacctacagc
     1801 gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg tatccggtaa
     1861 gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac gcctggtatc
     1921 tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg tgatgctcgt
     1981 caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct
     2041 tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct gtggataacc
     2101 gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg
     2161 agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt
     2221 ggccgattca ttaatgcaga gcttgcaatt cgcgcttttt caatattatt gaagcattta
     2281 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     2341 aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa ccattattat
     2401 catgacatta acctataaaa ataggcgtat tacgaggccc tttcacgcat tagatgcatg
     2461 tcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac ccccgcccat
     2521 tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc cattgacgtc
     2581 aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg tatcatatgc
     2641 caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat tatgcccagt
     2701 acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc atcgctatta
     2761 ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt gactcacggg
     2821 gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac caaaatcaac
     2881 gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc ggtaggcgtg
     2941 tacggtggga ggtctatata agcagagctc gtttagtgaa ccgtcagatc gcctggagac
     3001 gccatccacg ctgttttgac ctccatagaa gacaccggga ccgatccagc ctccggactc
     3061 tagcctaggc cgcggaccat ggactacaaa gacgatgacg acaaggaagg aacaagtttg
     3121 tacaaaaaag caggcttgaa ggaattcggt accatggcag tcactcgcta tgtcgacaac
     3181 aatttctgtg gcccagatgg gtaccctctt gattgcatca aagattttct cgcacgcgcg
     3241 ggcaagtcaa tgtgcactct ttccgaacaa cttgattaca tcgagtcgaa gagaggtgtc
     3301 tactgctgcc gtgaccatga gcatgaaatt gcctggttca ctgagcgctc tgataagagc
     3361 tacgagcacc agacaccctt cgaaattaag agtgccaaga aatttgacac tttcaaaggg
     3421 gaatgcccaa agtttgtgtt tcctcttaac tcaaaagtca aagtcattca accacgtgtt
     3481 gaaaagaaaa agactgaggg tttcatgggg cgtatacgct ctgtgtaccc tgttgcatct
     3541 ccacaggagt gtaacaatat gcacttgtct accttgatga aatgtaatca ttgcgatgaa
     3601 gtttcatggc agacgtgcga ctttctgaaa gccacttgtg aacattgtgg cactgaaaat
     3661 ttagttattg aaggacctac tacatgtggg tacctaccta ctaatgctgt agtgaaaatg
     3721 ccatgtcctg cctgtcaaga cccagagatt ggacctgagc atagtgttgc agattatcac
     3781 aaccactcaa acattgaaac tcgactccgc aagggaggta ggactagatg ttttggaggc
     3841 tgtgtgtttg cctatgttgg ctgctataat aagcgtgcct actgggttcc tcgtgctagt
     3901 gctgatattg gctcaggcca tactggcatt actggtgaca atgtggagac cttgaatgag
     3961 gatctccttg agatactgag tcgtgaacgt gttaacatta acattgttgg cgattttcat
     4021 ttgaatgaag aggttgccat cattttggca tctttctctg cttctacaag tgcctttatt
     4081 gacactataa agagtcttga ttacaagtct ttcaaaacca ttgttgagtc ctgcggtaac
     4141 tataaagtta ccaagggaaa gcccgtaaaa ggtgcttgga acattggaca acagagatca
     4201 gttttaacac cactgtgtgg ttttccctca caggctgctg gtgttatcag atcaattttt
     4261 gcgcgcacac ttgatgcagc aaaccactca attcctgatt tgcaaagagc agctgtcacc
     4321 atacttgatg gtatttctga acagtcatta cgtcttgtcg acgccatggt ttatacttca
     4381 gacctgctca ccaacagtgt cattattatg gcatatgtaa ctggtggtct tgtacaacag
     4441 acttctcagt ggttgtctaa tcttttgggc actactgttg aaaaactcag gcctatcttt
     4501 gaatggattg aggcgaaact tagtgcagga gttgaatttc tcaaggatgc ttgggagatt
     4561 ctcaaatttc tcattacagg tgtttttgac atcgtcaagg gtcaaataca ggttgcttca
     4621 gataacatca aggattgtgt aaaatgcttc attgatgttg ttaacaaggc actcgaaatg
     4681 tgcattgatc aagtcactat cgctggcgca aagttgcgat cactcaactt aggtgaagtc
     4741 ttcatcgctc aaagcaaggg actttaccgt cagtgtatac gtggcaagga gcagctgcaa
     4801 ctactcatgc ctcttaaggc accaaaagaa gtaacctttc ttgaaggtga ttcacatgac
     4861 acagtactta cctctgagga ggttgttctc aagaacggtg aactcgaagc actcgagacg
     4921 cccgttgata gcttcacaaa tggagctatc gttggcacac cagtctgtgt aaatggcctc
     4981 atgctcttag agattaagga caaagaacaa tactgcgcat tgtctcctgg tttactggct
     5041 acaaacaatg tctttcgctt aaaagggggt tagctcgagt gcggccgcaa cccagctttc
     5101 ttgtacaaag tggttgatcg cgtgcatgcg acgtcatagc tctctcccta tagtgagtcg
     5161 tattataagc taggcactgg ccgtcgtttt acaacgtcgt gactgggaaa actgctagct
     5221 tgggatcttt gtgaaggaac cttacttctg tggtgtgaca taattggaca aactacctac
     5281 agagatttaa agctctaagg taaatataaa atttttaagt gtataatgtg ttaaactagc
     5341 tgcatatgct tgctgcttga gagttttgct tactgagtat gatttatgaa aatattatac
     5401 acaggagcta gtgattctaa ttgtttgtgt attttagatt cacagtccca aggctcattt
     5461 caggcccctc agtcctcaca gtctgttcat gatcataatc agccatacca catttgtaga
     5521 ggttttactt gctttaaaaa acctcccaca cctccccctg aacctgaaac ataaaatgaa
     5581 tgcaattgtt gttgttaact tgtttattgc agcttataat ggttacaaat aaagcaatag
     5641 catcacaaat ttcacaaata aagcattttt ttcactgcat tctagttgtg gtttgtccaa
     5701 actcatcaat gtatcttatc atgtctggat cgatcctgca ttaatgaatc ggccaacgcg
     5761 cggggagagg cggtttgcgt attggctggc gtaatagcga agaggcccgc accgatcgcc
     5821 cttcccaaca gttgcgcagc ctgaatggcg aatgggacgc gccctgtagc ggcgcattaa
     5881 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     5941 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ctccgtcaag
     6001 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     6061 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     6121 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     6181 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     6241 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     6301 cgcttacaat ttcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     6361 catacgcgga tctgcgcagc accatggcct gaaataacct ctgaaagagg aacttggtta
     6421 gcttctgagg cggaaagaac cagctgtgga atgtgtgtca gttagggtgt ggaaagtccc
     6481 caggctcccc agcaggcaga agtatgcaaa gcatgcatct caattagtca gcaaccaggt
     6541 gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt
     6601 cagcaaccat agtcccgccc ctaactccgc ccatcccgcc cctaactccg cccagttccg
     6661 cccattctcc gccccatggc tgactaattt tttttattta tgcagaggcc gaggccgcct
     6721 cggcctctga gctattccag aagtagtgag gaggcttttt tggaggccta ggcttttgca
     6781 aaaagcttga ttcttctgac acaacagtct cgaacttaag gctagagcca ccatgattga
     6841 acaagatgga ttgcacgcag gttctccggc cgcttgggtg gagaggctat tcggctatga
     6901 ctgggcacaa cagacaatcg gctgctctga tgccgccgtg ttccggctgt cagcgcaggg
     6961 gcgcccggtt ctttttgtca agaccgacct gtccggtgcc ctgaatgaac tgcaggacga
     7021 ggcagcgcgg ctatcgtggc tggccacgac gggcgttcct tgcgcagctg tgctcgacgt
     7081 tgtcactgaa gcgggaaggg actggctgct attgggcgaa gtgccggggc aggatctcct
     7141 gtcatctcac cttgctcctg ccgagaaagt atccatcatg gctgatgcaa tgcggcggct
     7201 gcatacgctt gatccggcta cctgcccatt cgaccaccaa gcgaaacatc gcatcgagcg
     7261 agcacgtact cggatggaag ccggtcttgt cgatcaggat gatctggacg aagagcatca
     7321 ggggctcgcg ccagccgaac tgttcgccag gctcaaggcg cgcatgcccg acggcgagga
     7381 tctcgtcgtg acccatggcg atgcctgctt gccgaatatc atggtggaaa atggccgctt
     7441 ttctggattc atcgactgtg gccggctggg tgtggcggac cgctatcagg acatagcgtt
     7501 ggctacccgt gatattgctg aagagcttgg cggcgaatgg gctgaccgct tcctcgtgct
     7561 ttacggtatc gccgctcccg attcgcagcg catcgccttc tatcgccttc ttgacgagtt
     7621 cttctgagcg ggactctggg gttcgcgaaa tgaccgacca agcgacgccc aacctgccat
     7681 cacgatggcc gcaataaaat atctttattt tcattacatc tgtgtgttgg ttttttgtgt
     7741 ggatcgatag cgataaggat ccgcgcat
//
