LOCUS       sequence-348196-        6658 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Transient mammalian expression of codon optimized SARS-CoV-2 
            Nucleocapsid (P80R). 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 Apr 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-P80R
FEATURES             Location/Qualifiers
     source          1..6658
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        123..502
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        503..706
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     657..677
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     751..770
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        751..769
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2137..2154)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2143..2367
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2413..2841
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2500..2519)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2710..2731
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2850..2870)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2855..3184
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3035..3170
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3097..3116
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3251..4045
                     /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(3305..3324)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3915..3934
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4219..4340
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4256..4275)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4310..4329
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4389..4405)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4389..4405)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4402..4424)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4413..4429
                     /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(4437..4443,4444..4461,4462..4467))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4482..4503
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4620..4637)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4791..5379)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4871..4890)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5550..6341,6342..6410))
                     /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     6173..6192
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6411..6515)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6590..6609)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gtagtgcgcg agcaaaattt aagctacaac aaggcaaggc ttgaccgaca attgcatgaa
       61 gaatctgctt agggttaggc gttttgcgct gcttcgcgat gtacgggcca gatatacgcg
      121 ttgacattga ttattgacta gttattaata gtaatcaatt acggggtcat tagttcatag
      181 cccatatatg gagttccgcg ttacataact tacggtaaat ggcccgcctg gctgaccgcc
      241 caacgacccc cgcccattga cgtcaataat gacgtatgtt cccatagtaa cgccaatagg
      301 gactttccat tgacgtcaat gggtggagta tttacggtaa actgcccact tggcagtaca
      361 tcaagtgtat catatgccaa gtacgccccc tattgacgtc aatgacggta aatggcccgc
      421 ctggcattat gcccagtaca tgaccttatg ggactttcct acttggcagt acatctacgt
      481 attagtcatc gctattacca tggtgatgcg gttttggcag tacatcaatg ggcgtggata
      541 gcggtttgac tcacggggat ttccaagtct ccaccccatt gacgtcaatg ggagtttgtt
      601 ttggcaccaa aatcaacggg actttccaaa atgtcgtaac aactccgccc cattgacgca
      661 aatgggcggt aggcgtgtac ggtgggaggt ctatataagc agagctctct ggctaactag
      721 agaacccact gcttactggc ttatcgaaat taatacgact cactataggg agacccaagc
      781 tggctagcct tccatttcag gtgtcgtgag gatctatttc cggtgaattc gccgccacca
      841 tgagcgataa cggcccccaa aaccaacgga atgcccctcg gataacattt ggtggcccta
      901 gtgattccac aggaagtaac caaaacggag aacggtctgg agcccgctct aaacagaggc
      961 gaccacaggg attgccaaat aacacagcgt catggtttac agccctgact caacatggaa
     1021 aggaggactt gaaatttcca cgaggacagg gtgtaccaat aaacacaaac agtagtcggg
     1081 acgatcagat cgggtattat agaagggcca cgagacgcat cagaggtgga gatggtaaaa
     1141 tgaaagatct cagtccgcgc tggtactttt attatttggg aaccggccca gaagcgggcc
     1201 tcccatatgg agccaataag gacgggatta tatgggtggc tacagaggga gccctcaata
     1261 cgccaaaaga tcacatcgga acaagaaacc ctgctaataa tgccgcaatc gtgctgcagt
     1321 tgcctcaggg gacgacgctt cctaaaggct tttacgcaga aggatcacgc ggcggcagcc
     1381 aagcatcctc aaggtctagt tccagaagtc gaaacagctc cagaaactcc acaccagggt
     1441 ccagtagggg cacaagtccg gcgcggatgg cgggcaacgg cggagacgcc gcactcgctc
     1501 tgttgctcct ggaccggctc aaccaacttg aatccaagat gagtggtaag ggacagcagc
     1561 aacaaggtca aaccgtaacc aagaaaagcg ctgcagaagc ttctaaaaaa cctcgacaaa
     1621 aacggaccgc tacgaaggca tataacgtta ctcaagcctt cgggagacga gggccggagc
     1681 aaacccaggg gaattttgga gatcaggagc tcatccggca agggacagat tacaaacatt
     1741 ggcctcaaat cgctcagttc gcgccgtccg caagtgcttt tttcggaatg tctcgcatcg
     1801 gtatggaagt aacaccatcc gggacatggc tcacttatac cggcgccatc aagctggacg
     1861 acaaagatcc aaatttcaaa gatcaggtga ttctcttgaa taaacacata gacgcgtata
     1921 agacgtttcc acccaccgaa cccaagaagg acaaaaagaa aaaagctgat gaaacacaag
     1981 cgttgcccca gcgacagaag aagcagcaaa ccgtcaccct cctgccagcc gctgatctcg
     2041 atgacttctc caagcaactt caacagtcca tgagctcagc cgattctact caggcgtaac
     2101 tcgagtctag agggcccgtt taaacccgct gatcagcctc gactgtgcct tctagttgcc
     2161 agccatctgt tgtttgcccc tcccccgtgc cttccttgac cctggaaggt gccactccca
     2221 ctgtcctttc ctaataaaat gaggaaattg catcgcattg tctgagtagg tgtcattcta
     2281 ttctgggggg tggggtgggg caggacagca agggggagga ttgggaagac aatagcaggc
     2341 atgctgggga tgcggtgggc tctatggctt ctgaggcgga aagaaccagc tggggctcta
     2401 gggggtatcc ccacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
     2461 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
     2521 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
     2581 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     2641 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     2701 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     2761 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     2821 aacaaaaatt taacgcgaat taattctgtg gaatgtgtgt cagttagggt gtggaaagtc
     2881 cccaggctcc ccagcaggca gaagtatgca aagcatgcat ctcaattagt cagcaaccag
     2941 gtgtggaaag tccccaggct ccccagcagg cagaagtatg caaagcatgc atctcaatta
     3001 gtcagcaacc atagtcccgc ccctaactcc gcccatcccg cccctaactc cgcccagttc
     3061 cgcccattct ccgccccatg gctgactaat tttttttatt tatgcagagg ccgaggccgc
     3121 ctctgcctct gagctattcc agaagtagtg aggaggcttt tttggaggcc taggcttttg
     3181 caaaaagctc ccgggagctt gtatatccat tttcggatct gatcaagaga caggatgagg
     3241 atcgtttcgc atgattgaac aagatggatt gcacgcaggt tctccggccg cttgggtgga
     3301 gaggctattc ggctatgact gggcacaaca gacaatcggc tgctctgatg ccgccgtgtt
     3361 ccggctgtca gcgcaggggc gcccggttct ttttgtcaag accgacctgt ccggtgccct
     3421 gaatgaactg caggacgagg cagcgcggct atcgtggctg gccacgacgg gcgttccttg
     3481 cgcagctgtg ctcgacgttg tcactgaagc gggaagggac tggctgctat tgggcgaagt
     3541 gccggggcag gatctcctgt catctcacct tgctcctgcc gagaaagtat ccatcatggc
     3601 tgatgcaatg cggcggctgc atacgcttga tccggctacc tgcccattcg accaccaagc
     3661 gaaacatcgc atcgagcgag cacgtactcg gatggaagcc ggtcttgtcg atcaggatga
     3721 tctggacgaa gagcatcagg ggctcgcgcc agccgaactg ttcgccaggc tcaaggcgcg
     3781 catgcccgac ggcgaggatc tcgtcgtgac ccatggcgat gcctgcttgc cgaatatcat
     3841 ggtggaaaat ggccgctttt ctggattcat cgactgtggc cggctgggtg tggcggaccg
     3901 ctatcaggac atagcgttgg ctacccgtga tattgctgaa gagcttggcg gcgaatgggc
     3961 tgaccgcttc ctcgtgcttt acggtatcgc cgctcccgat tcgcagcgca tcgccttcta
     4021 tcgccttctt gacgagttct tctgagcggg actctggggt tcgaaatgac cgaccaagcg
     4081 acgcccaacc tgccatcacg agatttcgat tccaccgccg ccttctatga aaggttgggc
     4141 ttcggaatcg ttttccggga cgccggctgg atgatcctcc agcgcgggga tctcatgctg
     4201 gagttcttcg cccaccccaa cttgtttatt gcagcttata atggttacaa ataaagcaat
     4261 agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc
     4321 aaactcatca atgtatctta tcatgtctgt ataccgtcga cctctagcta gagcttggcg
     4381 taatcatggt catagctgtt tcctgtgtga aattgttatc cgctcacaat tccacacaac
     4441 atacgagccg gaagcataaa gtgtaaagcc tggggtgcct aatgagtgag ctaactcaca
     4501 ttaattgcgt tgcgctcact gcccgctttc cagtcgggaa acctgtcgtg ccagctgcat
     4561 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     4621 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     4681 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     4741 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     4801 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     4861 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     4921 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     4981 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     5041 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     5101 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     5161 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     5221 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     5281 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     5341 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     5401 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     5461 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     5521 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     5581 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     5641 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     5701 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     5761 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     5821 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     5881 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     5941 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     6001 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     6061 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     6121 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     6181 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     6241 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     6301 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     6361 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     6421 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     6481 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     6541 gacgtcgacg gatcgggaga tctcccgatc ccctatggtg cactctcagt acaatctgct
     6601 ctgatgccgc atagttaagc cagtatctgc tccctgcttg tgtgttggag gtcgctga
//
