LOCUS       sequence-348197-        6658 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transient mammalian expression of codon optimized SARS-CoV-2 
            Nucleocapsid (P13L). 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-P13L
FEATURES             Location/Qualifiers
     source          1..6658
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        142..521
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        522..725
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     676..696
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     770..789
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        770..788
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2156..2173)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2162..2386
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2432..2860
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2519..2538)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2729..2750
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2869..2889)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2874..3203
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3054..3189
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3116..3135
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3270..4064
                     /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(3324..3343)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3934..3953
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4238..4359
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4275..4294)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4329..4348
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4408..4424)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4408..4424)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4421..4443)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4432..4448
                     /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(4456..4462,4463..4480,4481..4486))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4501..4522
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4639..4656)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4810..5398)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4890..4909)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5569..6360,6361..6429))
                     /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     6192..6211
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6430..6534)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6609..6628)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gtgtgttgga ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct
       61 tgaccgacaa ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg
      121 tacgggccag atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta
      181 cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg
      241 gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc
      301 ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa
      361 ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca
      421 atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta
      481 cttggcagta catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt
      541 acatcaatgg gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg
      601 acgtcaatgg gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca
      661 actccgcccc attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca
      721 gagctctctg gctaactaga gaacccactg cttactggct tatcgaaatt aatacgactc
      781 actataggga gacccaagct ggctagcctt ccatttcagg tgtcgtgagg atctatttcc
      841 ggtgaattcg ccgccaccat gagcgataac ggcccccaaa accaacggaa tgccctgcgg
      901 ataacatttg gtggccctag tgattccaca ggaagtaacc aaaacggaga acggtctgga
      961 gcccgctcta aacagaggcg accacaggga ttgccaaata acacagcgtc atggtttaca
     1021 gccctgactc aacatggaaa ggaggacttg aaatttccac gaggacaggg tgtaccaata
     1081 aacacaaaca gtagtccaga cgatcagatc gggtattata gaagggccac gagacgcatc
     1141 agaggtggag atggtaaaat gaaagatctc agtccgcgct ggtactttta ttatttggga
     1201 accggcccag aagcgggcct cccatatgga gccaataagg acgggattat atgggtggct
     1261 acagagggag ccctcaatac gccaaaagat cacatcggaa caagaaaccc tgctaataat
     1321 gccgcaatcg tgctgcagtt gcctcagggg acgacgcttc ctaaaggctt ttacgcagaa
     1381 ggatcacgcg gcggcagcca agcatcctca aggtctagtt ccagaagtcg aaacagctcc
     1441 agaaactcca caccagggtc cagtaggggc acaagtccgg cgcggatggc gggcaacggc
     1501 ggagacgccg cactcgctct gttgctcctg gaccggctca accaacttga atccaagatg
     1561 agtggtaagg gacagcagca acaaggtcaa accgtaacca agaaaagcgc tgcagaagct
     1621 tctaaaaaac ctcgacaaaa acggaccgct acgaaggcat ataacgttac tcaagccttc
     1681 gggagacgag ggccggagca aacccagggg aattttggag atcaggagct catccggcaa
     1741 gggacagatt acaaacattg gcctcaaatc gctcagttcg cgccgtccgc aagtgctttt
     1801 ttcggaatgt ctcgcatcgg tatggaagta acaccatccg ggacatggct cacttatacc
     1861 ggcgccatca agctggacga caaagatcca aatttcaaag atcaggtgat tctcttgaat
     1921 aaacacatag acgcgtataa gacgtttcca cccaccgaac ccaagaagga caaaaagaaa
     1981 aaagctgatg aaacacaagc gttgccccag cgacagaaga agcagcaaac cgtcaccctc
     2041 ctgccagccg ctgatctcga tgacttctcc aagcaacttc aacagtccat gagctcagcc
     2101 gattctactc aggcgtaact cgagtctaga gggcccgttt aaacccgctg atcagcctcg
     2161 actgtgcctt ctagttgcca gccatctgtt gtttgcccct cccccgtgcc ttccttgacc
     2221 ctggaaggtg ccactcccac tgtcctttcc taataaaatg aggaaattgc atcgcattgt
     2281 ctgagtaggt gtcattctat tctggggggt ggggtggggc aggacagcaa gggggaggat
     2341 tgggaagaca atagcaggca tgctggggat gcggtgggct ctatggcttc tgaggcggaa
     2401 agaaccagct ggggctctag ggggtatccc cacgcgccct gtagcggcgc attaagcgcg
     2461 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgccct agcgcccgct
     2521 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     2581 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     2641 cttgattagg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     2701 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     2761 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     2821 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt aattctgtgg aatgtgtgtc
     2881 agttagggtg tggaaagtcc ccaggctccc cagcaggcag aagtatgcaa agcatgcatc
     2941 tcaattagtc agcaaccagg tgtggaaagt ccccaggctc cccagcaggc agaagtatgc
     3001 aaagcatgca tctcaattag tcagcaacca tagtcccgcc cctaactccg cccatcccgc
     3061 ccctaactcc gcccagttcc gcccattctc cgccccatgg ctgactaatt ttttttattt
     3121 atgcagaggc cgaggccgcc tctgcctctg agctattcca gaagtagtga ggaggctttt
     3181 ttggaggcct aggcttttgc aaaaagctcc cgggagcttg tatatccatt ttcggatctg
     3241 atcaagagac aggatgagga tcgtttcgca tgattgaaca agatggattg cacgcaggtt
     3301 ctccggccgc ttgggtggag aggctattcg gctatgactg ggcacaacag acaatcggct
     3361 gctctgatgc cgccgtgttc cggctgtcag cgcaggggcg cccggttctt tttgtcaaga
     3421 ccgacctgtc cggtgccctg aatgaactgc aggacgaggc agcgcggcta tcgtggctgg
     3481 ccacgacggg cgttccttgc gcagctgtgc tcgacgttgt cactgaagcg ggaagggact
     3541 ggctgctatt gggcgaagtg ccggggcagg atctcctgtc atctcacctt gctcctgccg
     3601 agaaagtatc catcatggct gatgcaatgc ggcggctgca tacgcttgat ccggctacct
     3661 gcccattcga ccaccaagcg aaacatcgca tcgagcgagc acgtactcgg atggaagccg
     3721 gtcttgtcga tcaggatgat ctggacgaag agcatcaggg gctcgcgcca gccgaactgt
     3781 tcgccaggct caaggcgcgc atgcccgacg gcgaggatct cgtcgtgacc catggcgatg
     3841 cctgcttgcc gaatatcatg gtggaaaatg gccgcttttc tggattcatc gactgtggcc
     3901 ggctgggtgt ggcggaccgc tatcaggaca tagcgttggc tacccgtgat attgctgaag
     3961 agcttggcgg cgaatgggct gaccgcttcc tcgtgcttta cggtatcgcc gctcccgatt
     4021 cgcagcgcat cgccttctat cgccttcttg acgagttctt ctgagcggga ctctggggtt
     4081 cgaaatgacc gaccaagcga cgcccaacct gccatcacga gatttcgatt ccaccgccgc
     4141 cttctatgaa aggttgggct tcggaatcgt tttccgggac gccggctgga tgatcctcca
     4201 gcgcggggat ctcatgctgg agttcttcgc ccaccccaac ttgtttattg cagcttataa
     4261 tggttacaaa taaagcaata gcatcacaaa tttcacaaat aaagcatttt tttcactgca
     4321 ttctagttgt ggtttgtcca aactcatcaa tgtatcttat catgtctgta taccgtcgac
     4381 ctctagctag agcttggcgt aatcatggtc atagctgttt cctgtgtgaa attgttatcc
     4441 gctcacaatt ccacacaaca tacgagccgg aagcataaag tgtaaagcct ggggtgccta
     4501 atgagtgagc taactcacat taattgcgtt gcgctcactg cccgctttcc agtcgggaaa
     4561 cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg gggagaggcg gtttgcgtat
     4621 tgggcgctct tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg
     4681 agcggtatca gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc
     4741 aggaaagaac atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt
     4801 gctggcgttt ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag
     4861 tcagaggtgg cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc
     4921 cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc
     4981 ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt
     5041 cgttcgctcc aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt
     5101 atccggtaac tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc
     5161 agccactggt aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa
     5221 gtggtggcct aactacggct acactagaag aacagtattt ggtatctgcg ctctgctgaa
     5281 gccagttacc ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg
     5341 tagcggtggt ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga
     5401 agatcctttg atcttttcta cggggtctga cgctcagtgg aacgaaaact cacgttaagg
     5461 gattttggtc atgagattat caaaaaggat cttcacctag atccttttaa attaaaaatg
     5521 aagttttaaa tcaatctaaa gtatatatga gtaaacttgg tctgacagtt accaatgctt
     5581 aatcagtgag gcacctatct cagcgatctg tctatttcgt tcatccatag ttgcctgact
     5641 ccccgtcgtg tagataacta cgatacggga gggcttacca tctggcccca gtgctgcaat
     5701 gataccgcga gacccacgct caccggctcc agatttatca gcaataaacc agccagccgg
     5761 aagggccgag cgcagaagtg gtcctgcaac tttatccgcc tccatccagt ctattaattg
     5821 ttgccgggaa gctagagtaa gtagttcgcc agttaatagt ttgcgcaacg ttgttgccat
     5881 tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg gcttcattca gctccggttc
     5941 ccaacgatca aggcgagtta catgatcccc catgttgtgc aaaaaagcgg ttagctcctt
     6001 cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg ttatcactca tggttatggc
     6061 agcactgcat aattctctta ctgtcatgcc atccgtaaga tgcttttctg tgactggtga
     6121 gtactcaacc aagtcattct gagaatagtg tatgcggcga ccgagttgct cttgcccggc
     6181 gtcaatacgg gataataccg cgccacatag cagaacttta aaagtgctca tcattggaaa
     6241 acgttcttcg gggcgaaaac tctcaaggat cttaccgctg ttgagatcca gttcgatgta
     6301 acccactcgt gcacccaact gatcttcagc atcttttact ttcaccagcg tttctgggtg
     6361 agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata agggcgacac ggaaatgttg
     6421 aatactcata ctcttccttt ttcaatatta ttgaagcatt tatcagggtt attgtctcat
     6481 gagcggatac atatttgaat gtatttagaa aaataaacaa ataggggttc cgcgcacatt
     6541 tccccgaaaa gtgccacctg acgtcgacgg atcgggagat ctcccgatcc cctatggtgc
     6601 actctcagta caatctgctc tgatgccgca tagttaagcc agtatctgct ccctgctt
//
