LOCUS       sequence-348099-        6658 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transient mammalian expression of codon optimized SARS-CoV-2 
            Nucleocapsid (original Wuhan Hu1 sequence). Used for generating RNA 
            packaging virus-like particles..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6658)
  AUTHORS   Syed AM, Taha TY, Tabata T, Chen IP, Ciling A, Khalid MM, Sreekumar 
            B, Chen PY, Hayashi JM, Soczek KM, Ott M, Doudna JA
  TITLE     Rapid assessment of SARS-CoV-2 evolved variants using virus-like 
            particles.
  JOURNAL   Science. 2021 Nov 4:eabl6184. doi: 10.1126/science.abl6184.
  PUBMED    34735219
REFERENCE   2  (bases 1 to 6658)
  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: "Science. 
            2021 Nov 4:eabl6184. doi: 10.1126/science.abl6184."
COMMENT     Sequence Label: CoV2-N-WT-Hu1
FEATURES             Location/Qualifiers
     source          1..6658
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        72..451
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        452..655
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     606..626
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     700..719
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        700..718
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2086..2103)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2092..2316
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2362..2790
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2449..2468)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2659..2680
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2799..2819)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2804..3133
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2984..3119
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3046..3065
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3200..3994
                     /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(3254..3273)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3864..3883
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4168..4289
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4205..4224)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4259..4278
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4338..4354)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4338..4354)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4351..4373)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4362..4378
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(4386..4392,4393..4410,4411..4416))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4431..4452
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4569..4586)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4740..5328)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4820..4839)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5499..6290,6291..6359))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6122..6141
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6360..6464)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6539..6558)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag
       61 atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt
      121 agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg
      181 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
      241 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
      301 ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa
      361 atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta
      421 catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg
      481 gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg
      541 gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc
      601 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctctctg
      661 gctaactaga gaacccactg cttactggct tatcgaaatt aatacgactc actataggga
      721 gacccaagct ggctagcctt ccatttcagg tgtcgtgagg atctatttcc ggtgaattcg
      781 ccgccaccat gagcgataac ggcccccaaa accaacggaa tgcccctcgg ataacatttg
      841 gtggccctag tgattccaca ggaagtaacc aaaacggaga acggtctgga gcccgctcta
      901 aacagaggcg accacaggga ttgccaaata acacagcgtc atggtttaca gccctgactc
      961 aacatggaaa ggaggacttg aaatttccac gaggacaggg tgtaccaata aacacaaaca
     1021 gtagtccaga cgatcagatc gggtattata gaagggccac gagacgcatc agaggtggag
     1081 atggtaaaat gaaagatctc agtccgcgct ggtactttta ttatttggga accggcccag
     1141 aagcgggcct cccatatgga gccaataagg acgggattat atgggtggct acagagggag
     1201 ccctcaatac gccaaaagat cacatcggaa caagaaaccc tgctaataat gccgcaatcg
     1261 tgctgcagtt gcctcagggg acgacgcttc ctaaaggctt ttacgcagaa ggatcacgcg
     1321 gcggcagcca agcatcctca aggtctagtt ccagaagtcg aaacagctcc agaaactcca
     1381 caccagggtc cagtaggggc acaagtccgg cgcggatggc gggcaacggc ggagacgccg
     1441 cactcgctct gttgctcctg gaccggctca accaacttga atccaagatg agtggtaagg
     1501 gacagcagca acaaggtcaa accgtaacca agaaaagcgc tgcagaagct tctaaaaaac
     1561 ctcgacaaaa acggaccgct acgaaggcat ataacgttac tcaagccttc gggagacgag
     1621 ggccggagca aacccagggg aattttggag atcaggagct catccggcaa gggacagatt
     1681 acaaacattg gcctcaaatc gctcagttcg cgccgtccgc aagtgctttt ttcggaatgt
     1741 ctcgcatcgg tatggaagta acaccatccg ggacatggct cacttatacc ggcgccatca
     1801 agctggacga caaagatcca aatttcaaag atcaggtgat tctcttgaat aaacacatag
     1861 acgcgtataa gacgtttcca cccaccgaac ccaagaagga caaaaagaaa aaagctgatg
     1921 aaacacaagc gttgccccag cgacagaaga agcagcaaac cgtcaccctc ctgccagccg
     1981 ctgatctcga tgacttctcc aagcaacttc aacagtccat gagctcagcc gattctactc
     2041 aggcgtaact cgagtctaga gggcccgttt aaacccgctg atcagcctcg actgtgcctt
     2101 ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg
     2161 ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt
     2221 gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca
     2281 atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa agaaccagct
     2341 ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg
     2401 tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt
     2461 tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc
     2521 tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgattagg
     2581 gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg
     2641 agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct
     2701 cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg
     2761 agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc agttagggtg
     2821 tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc tcaattagtc
     2881 agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca
     2941 tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc
     3001 gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc
     3061 cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct
     3121 aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcaagagac
     3181 aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt ctccggccgc
     3241 ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct gctctgatgc
     3301 cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc
     3361 cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg ccacgacggg
     3421 cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact ggctgctatt
     3481 gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg agaaagtatc
     3541 catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct gcccattcga
     3601 ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg gtcttgtcga
     3661 tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt tcgccaggct
     3721 caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc
     3781 gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc ggctgggtgt
     3841 ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag agcttggcgg
     3901 cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat
     3961 cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt cgaaatgacc
     4021 gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc cttctatgaa
     4081 aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca gcgcggggat
     4141 ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa tggttacaaa
     4201 taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca ttctagttgt
     4261 ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac ctctagctag
     4321 agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc gctcacaatt
     4381 ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta atgagtgagc
     4441 taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc
     4501 cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct
     4561 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     4621 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     4681 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     4741 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     4801 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     4861 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     4921 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     4981 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     5041 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     5101 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     5161 aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa gccagttacc
     5221 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     5281 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     5341 atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg gattttggtc
     5401 atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg aagttttaaa
     5461 tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt aatcagtgag
     5521 gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact ccccgtcgtg
     5581 tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat gataccgcga
     5641 gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg aagggccgag
     5701 cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg ttgccgggaa
     5761 gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat tgctacaggc
     5821 atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc ccaacgatca
     5881 aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg
     5941 atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc agcactgcat
     6001 aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga gtactcaacc
     6061 aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc gtcaatacgg
     6121 gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa acgttcttcg
     6181 gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta acccactcgt
     6241 gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg agcaaaaaca
     6301 ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg aatactcata
     6361 ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat gagcggatac
     6421 atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt tccccgaaaa
     6481 gtgccacctg acgtcgacgg atcgggagat ctcccgatcc cctatggtgc actctcagta
     6541 caatctgctc tgatgccgca tagttaagcc agtatctgct ccctgcttgt gtgttggagg
     6601 tcgctgagta gtgcgcgagc aaaatttaag ctacaacaag gcaaggcttg accgacaa
//
