LOCUS       sequence-334010-        8144 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Baculoviral transfer vector to express SARS-CoV-2 nsp14 in insect 
            cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8144)
  AUTHORS   Canal B, McClure AW, Curran JF, Wu M, Ulferts R, Weissmann F, Zeng 
            J, Bertolin AP, Milligan JC, Basu S, Drury LS, Deegan TD, Fujisawa 
            R, Roberts EL, Basier C, Labib K, Beale R, Howell M, Diffley JFX
  TITLE     Identifying SARS-CoV-2 antiviral compounds by screening for small 
            molecule inhibitors of nsp14/nsp10 exoribonuclease.
  JOURNAL   Biochem J. 2021 Jul 16;478(13):2445-2464. doi: 10.1042/BCJ20210198.
  PUBMED    34198326
REFERENCE   2  (bases 1 to 8144)
  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"; doi: "10"; journalName: 
            "Biochem J"; date: "2021-07-16- 16"; volume: "478"; issue: "13"; 
            pages: "2445-2464"
COMMENT     Sequence Label: pBIG1a_3xFlag-6His-nsp14 (SARS-CoV-2)
FEATURES             Location/Qualifiers
     source          1..8144
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             203..994
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      1035..1489
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1122..1141)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1331..1352
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1515..1619
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1620..1688,1689..2480)
                     /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     complement(1838..1857)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2651..3239
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3140..3159
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3393..3410
                     /label=L4440
                     /note="L4440 vector, forward primer"
     mobile_element  3544..3768
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     CDS             complement(3835..4368)
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(join(4557..4562,4563..4579,4580..4585))
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     protein_bind    complement(4981..5014)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(5060..5079)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5104..5152
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     promoter        5247..5338
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     5263..5280
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     regulatory      5382..5391
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             5391..5456
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             5463..5480
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     polyA_signal    7209..7343
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(7246..7265)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     7300..7319
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     mobile_element  7658..7823
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
ORIGIN
        1 gtccagaacc ttgaccgaac gcagcggtgg taacggcgca gtggcggttt tcatggcttg
       61 ttatgactgt ttttttgggg tacagtctat gcctcgggca tccaagcagc aagcgcgtta
      121 cgccgtgggt cgatgtttga tgttatggag cagcaacgat gttacgcagc agggcagtcg
      181 ccctaaaaca aagttaaaca tcatgaggga agcggtgatc gccgaagtat cgactcaact
      241 atcagaggta gttggcgtca tcgagcgcca tctcgaaccg acgttgctgg ccgtacattt
      301 gtacggctcc gcagtggatg gcggcctgaa gccacacagt gatattgatt tgctggttac
      361 ggtgaccgta aggcttgatg aaacaacgcg gcgagctttg atcaacgacc ttttggaaac
      421 ttcggcttcc cctggagaga gcgagattct ccgcgctgta gaagtcacca ttgttgtgca
      481 cgacgacatc attccgtggc gttatccagc taagcgcgaa ctgcaatttg gagaatggca
      541 gcgcaatgac attcttgcag gtatcttcga gccagccacg atcgacattg atctggctat
      601 cttgctgaca aaagcaagag aacatagcgt tgccttggta ggtccagcgg cggaggaact
      661 ctttgatccg gttcctgaac aggatctatt tgaggcgcta aatgaaacct taacgctatg
      721 gaactcgccg cccgactggg ctggcgatga gcgaaatgta gtgcttacgt tgtcccgcat
      781 ttggtacagc gcagtaaccg gcaaaatcgc gccgaaggat gtcgctgccg actgggcaat
      841 ggagcgcctg ccggcccagt atcagcccgt catacttgaa gctagacagg cttatcttgg
      901 acaagaagaa gatcgcttgg cctcgcgcgc agatcagttg gaagaatttg tccactacgt
      961 gaaaggcgag atcaccaagg tagtcggcaa ataatgtcta acaattcgtt caagccgacg
     1021 gattggcgaa tgggacgcgc cctgtagcgg cgcattaagc gcggcgggtg tggtggttac
     1081 gcgcagcgtg accgctacac ttgccagcgc cctagcgccc gctcctttcg ctttcttccc
     1141 ttcctttctc gccacgttcg ccggctttcc ccgtcaagct ctaaatcggg ggctcccttt
     1201 agggttccga tttagtgctt tacggcacct cgaccccaaa aaacttgatt agggtgatgg
     1261 ttcacgtagt gggccatcgc cctgatagac ggtttttcgc cctttgacgt tggagtccac
     1321 gttcttaata gtggactctt gttccaaact ggaacaacac tcaaccctat ctcggtctat
     1381 tcttttgatt tataagggat tttgccgatt tcggcctatt ggttaaaaaa tgagctgatt
     1441 taacaaaaat ttaacgcgaa ttttaacaaa atattaacgc ttacaattta ggtggcactt
     1501 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     1561 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     1621 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     1681 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     1741 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     1801 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     1861 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     1921 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     1981 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     2041 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     2101 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     2161 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     2221 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     2281 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     2341 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     2401 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     2461 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     2521 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     2581 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     2641 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     2701 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     2761 taactggctt cagcagagcg cagataccaa atactattct tctagtgtag ccgtagttag
     2821 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     2881 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     2941 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     3001 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     3061 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     3121 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     3181 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     3241 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     3301 tctttcctgc gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg
     3361 ataccgctcg ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag
     3421 agcgcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatagacc
     3481 agccgcgtaa cctggcaaaa tcggttacgg ttgagtaata aatggatgcc ctgcgtaagc
     3541 gggtgtgggc ggacaataaa gtcttaaact gaacaaaata gatctaaact atgacaataa
     3601 agtcttaaac tagacagaat agttgtaaac tgaaatcagt ccagttatgc tgtgaaaaag
     3661 catactggac ttttgttatg gctaaagcaa actcttcatt ttctgaagtg caaattgccc
     3721 gtcgtattaa agaggggcgt ggccaagggc atggtaaaga ctatattcgc ggcgttgtga
     3781 caatttaccg aacaactccg cggccgggaa gccgatctcg gcttgaacga attgttaggt
     3841 ggcggtactt gggtcgatat caaagtgcat cacttcttcc cgtatgccca actttgtata
     3901 gagagccact gcgggatcgt caccgtaatc tgcttgcacg tagatcacat aagcaccaag
     3961 cgcgttggcc tcatgcttga ggagattgat gagcgcggtg gcaatgccct gcctccggtg
     4021 ctcgccggag actgcgagat catagatata gatctcacta cgcggctgct caaacttggg
     4081 cagaacgtaa gccgcgagag cgccaacaac cgcttcttgg tcgaaggcag caagcgcgat
     4141 gaatgtctta ctacggagca agttcccgag gtaatcggag tccggctgat gttgggagta
     4201 ggtggctacg tctccgaact cacgaccgaa aagatcaaga gcagcccgca tggatttgac
     4261 ttggtcaggg ccgagcctac atgtgcgaat gatgcccata cttgagccac ctaactttgt
     4321 tttagggcga ctgccctgct gcgtaacatc gttgctgctg cgtaacatcg ttgctgctcc
     4381 ataacatcaa acatcgaccc acggcgtaac gcgcttgctg cttggatgcc cgaggcatag
     4441 actgtacaaa aaaacagtca taacaagcca tgaaaaccgc cactgcgccg ttaccaccgc
     4501 tgcgttcggt caaggttctg gaccagttgc gtgagcgcat acgctacttg cattacagtt
     4561 tacgaaccga acaggcttat gtcaactggg tttgtgcctt catccgtttc cacggtgtgc
     4621 gtcacccggc aaccttgggc agcagcgaag tcgaggcatt tctgtcctgg ctggcgaacg
     4681 agcgcaaggt ttcggtctcc acgcatcgtc aggcattggc ggccttgctg ttcttctacg
     4741 gcaaggtgct gtgcacggat ctgcccttgc ttcaggagat cggtagacct cggccgtcgc
     4801 ggcgcttgcc ggtggtgctg accccggatg aagtggttcg catcctcggt tttctggaag
     4861 gcgagcatcg tttgttcgcc caggactcta gctatagttc tagtggttgg ctacagcttt
     4921 gtttgtacta tcaacaggtt gaactgctga tcaacagatc ctctacgcgg ccgcggtacc
     4981 ataacttcgt atagcataca ttatacgaag ttatctgggt ttaaacgctc tatggtctaa
     5041 agtttttatc gaggctaact gaaacacgga aggagacaat accggaagga acccgcgcta
     5101 tgacggcaat aaaaagacag aataaaacgc acgggtgttg ggtcgtttgt tcataaacgc
     5161 ggggttcggt cccagggctg gcactctgtc aagtgaacgc tctatggtct aaagatttaa
     5221 atcgacctac tccggaatat taatagatca tggagataat taaaatgata accatctcgc
     5281 aaataaataa gtattttact gttttcgtaa cagttttgta ataaaaaaac ctataaatat
     5341 tccggattat tcataccgtc ccaccatcgg gcgcggatcc agccaccatg gactacaagg
     5401 accacgacgg cgattacaag gatcatgaca tcgactacaa agacgacgat gacaagggtt
     5461 cccaccatca ccaccatcac tctgctgtgc tgcaggctga gaacgtgacc ggcctgttca
     5521 aggactgctc caaagtgatc accggactgc accccactca ggctcctact cacctgtctg
     5581 tggacaccaa gttcaagacc gagggcctgt gcgtggacat ccctggtatc cctaaggaca
     5641 tgacctaccg tcgtctgatc tccatgatgg gcttcaagat gaactaccaa gtgaacggct
     5701 accccaacat gttcatcacc cgtgaagagg ctatccgtca cgtgcgcgct tggatcggtt
     5761 tcgacgttga aggttgccac gctaccaggg aagctgtggg tactaacctg cctctgcagc
     5821 tgggattctc caccggtgtt aacctggtgg ctgtgcccac tggttacgtg gacaccccta
     5881 acaacaccga cttctcccgt gtgtccgcta agcctcctcc tggcgaccaa ttcaagcact
     5941 tgatccctct gatgtacaag ggcctgcctt ggaacgtcgt gcgtatcaag atcgtgcaga
     6001 tgctgtctga caccctgaag aacctgtccg accgtgtggt gttcgtgctg tgggctcacg
     6061 gtttcgagct gacctccatg aagtactttg tgaagatcgg tcccgagagg acctgctgct
     6121 tgtgcgatag acgtgctacc tgcttctcta ccgcttccga cacctacgct tgctggcacc
     6181 actccatcgg cttcgactac gtgtacaacc ccttcatgat cgacgtgcag cagtggggtt
     6241 tcaccggcaa cctgcagtct aaccacgacc tgtactgcca ggtgcacggc aacgctcacg
     6301 tggcctcttg tgacgctatc atgacccgtt gcctggctgt gcacgagtgc ttcgtgaagc
     6361 gtgtggactg gaccatcgag taccccatca tcggcgacga gctgaagatc aacgctgctt
     6421 gcagaaaggt gcagcacatg gtggtcaagg ctgctctgct ggctgacaag ttccctgtcc
     6481 tgcacgacat cggcaacccc aaggctatta agtgcgtgcc ccaggctgac gtcgagtgga
     6541 agttctacga cgctcagccc tgctccgaca aggcttacaa gatcgaggaa ctgttctact
     6601 cctacgctac ccactcagac aagttcaccg acggtgtctg cctgttctgg aactgcaatg
     6661 tggatcgtta ccccgctaac tccatcgtgt gccgtttcga cacccgtgtg ctgtccaacc
     6721 tgaacctgcc tggttgcgac ggtggttccc tgtacgtgaa caagcacgct ttccacactc
     6781 cagctttcga caagtccgct ttcgtgaacc tgaagcagct gcccttcttc tactactccg
     6841 actctccctg cgagtcccac ggaaagcaag tggtgtccga catcgattac gtgcccctga
     6901 agtccgctac ctgcatcacc agatgcaacc tcggtggtgc tgtgtgccgt caccacgcta
     6961 acgagtaccg tctgtacctg gacgcttaca acatgatgat ctccgctggt ttctccctgt
     7021 gggtgtacaa gcagttcgac acttacaacc tgtggaacac cttcaccagg ctgcagtaaa
     7081 agcttgtcga gaagtactag aggatcataa tcagccatac cacatttgta gaggttttac
     7141 ttgctttaaa aaacctccca cacctccccc tgaacctgaa acataaaatg aatgcaattg
     7201 ttgttgttaa cttgtttatt gcagcttata atggttacaa ataaagcaat agcatcacaa
     7261 atttcacaaa taaagcattt ttttcactgc attctagttg tggtttgtcc aaactcatca
     7321 atgtatctta tcatgtctgg atctgatcac tgcttgagcc tagaagatcc ggctgctaac
     7381 aaagcccgaa aggaagctga gttggctgct gccaccgctg agcaataact atcataacca
     7441 tttaaataaa tctatatctc aatcggggtt acgtcgatac cccaccgaga ccccattggg
     7501 accaatacgc ccgcgtttct tccttttccc caccccaacc cccaagttcg ggtgaaggcc
     7561 cagggctcgc agccaacgtc ggggcggcaa gccctgccat agccactacg ggttaattaa
     7621 aaactatatc tcaatcgggg tttaaacagg agatccgaac cagataagtg aaatctagtt
     7681 ccaaactatt ttgtcatttt taattttcgt attagcttac gacgctacac ccagttccca
     7741 tctattttgt cactcttccc taaataatcc ttaaaaactc catttccacc cctcccagtt
     7801 cccaactatt ttgtccgccc acagcggggc atttttcttc ctgttatgtt tttaatcaaa
     7861 catcctgcca actccatgtg acaaaccgtc atcttcggct actttttctc tgtcacagaa
     7921 tgaaaatttt tctgtcatct cttcgttatt aatgtttgta attgactgaa tatcaacgct
     7981 tatttgcagc ctgaactggg aggcagaata aatgatcata tcgtcaatta ttacctccac
     8041 ggggagagcc tgagcaaact ggcctcaggc atttgagaag cacacggtca cactgcttcc
     8101 ggtagtcaat aaaccggtaa gtagcgtatg cgctcacgca actg
//
