LOCUS       sequence-333998-        7319 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Baculoviral transfer vector to express nsp7-His6-nsp8 fusion protein
            (SARS-CoV-2) in insect cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7319)
  AUTHORS   Bertolin AP, Weissmann F, Zeng J, Posse V, Milligan JC, Canal B, 
            Ulferts R, Wu M, Drury LS, Howell M, Beale R, Diffley JFX
  TITLE     Identifying SARS-CoV-2 antiviral compounds by screening for small 
            molecule inhibitors of nsp12/7/8 RNA-dependent RNA polymerase.
  JOURNAL   Biochem J. 2021 Jul 16;478(13):2425-2443. doi: 10.1042/BCJ20210200.
  PUBMED    34198323
REFERENCE   2  (bases 1 to 7319)
  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: "2425-2443"
COMMENT     Sequence Label: pBIG1b_nsp7-His6-nsp8 (SARS-CoV-2)
FEATURES             Location/Qualifiers
     source          1..7319
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             215..748
                     /codon_start=1
                     /gene="aacC1"
                     /product="gentamycin acetyltransferase"
                     /label=GmR
                     /note="confers resistance to gentamycin"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     mobile_element  815..1039
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7R
                     /note="mini-Tn7 element (right end of the Tn7 transposon)"
     primer_bind     complement(1173..1190)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1344..1932)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1424..1443)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2103..2894,2895..2963))
                     /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     2726..2745
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2964..3068)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3094..3548)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3231..3252)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3442..3461
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     CDS             complement(3589..4380)
                     /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"
     mobile_element  4904..5069
                     /mobile_element_type="transposon:Tn7"
                     /label=Tn7L
                     /note="mini-Tn7 element (left end of the Tn7 transposon)"
     polyA_signal    5384..5518
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5408..5427)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5462..5481
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(6245..6262)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     promoter        complement(6564..6655)
                     /gene="polh from Autographa californica"
                     /label=polyhedrin promoter
                     /note="promoter for the baculovirus polyhedrin gene"
     primer_bind     complement(6622..6639)
                     /label=Polyhedrin-F
                     /note="Polyhedrin promoter, forward primer"
     polyA_signal    6750..6798
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     primer_bind     6823..6842
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     protein_bind    6888..6921
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        join(7317..7319,1..3,4..20,21..26)
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
ORIGIN
        1 acataagcct gttcggttcg taaactgtaa tgcaagtagc gtatgcgctc acgcaactgg
       61 tccagaacct tgaccgaacg cagcggtggt aacggcgcag tggcggtttt catggcttgt
      121 tatgactgtt tttttgtaca gtctatgcct cgggcatcca agcagcaagc gcgttacgcc
      181 gtgggtcgat gtttgatgtt atggagcagc aacgatgtta cgcagcagca acgatgttac
      241 gcagcagggc agtcgcccta aaacaaagtt aggtggctca agtatgggca tcattcgcac
      301 atgtaggctc ggccctgacc aagtcaaatc catgcgggct gctcttgatc ttttcggtcg
      361 tgagttcgga gacgtagcca cctactccca acatcagccg gactccgatt acctcgggaa
      421 cttgctccgt agtaagacat tcatcgcgct tgctgccttc gaccaagaag cggttgttgg
      481 cgctctcgcg gcttacgttc tgcccaagtt tgagcagccg cgtagtgaga tctatatcta
      541 tgatctcgca gtctccggcg agcaccggag gcagggcatt gccaccgcgc tcatcaatct
      601 cctcaagcat gaggccaacg cgcttggtgc ttatgtgatc tacgtgcaag cagattacgg
      661 tgacgatccc gcagtggctc tctatacaaa gttgggcata cgggaagaag tgatgcactt
      721 tgatatcgac ccaagtaccg ccacctaaca attcgttcaa gccgagatcg gcttcccggc
      781 cgcggagttg ttcggtaaat tgtcacaacg ccgcgaatat agtctttacc atgcccttgg
      841 ccacgcccct ctttaatacg acgggcaatt tgcacttcag aaaatgaaga gtttgcttta
      901 gccataacaa aagtccagta tgctttttca cagcataact ggactgattt cagtttacaa
      961 ctattctgtc tagtttaaga ctttattgtc atagtttaga tctattttgt tcagtttaag
     1021 actttattgt ccgcccacac ccgcttacgc agggcatcca tttattactc aaccgtaacc
     1081 gattttgcca ggttacgcgg ctggtctatg cggtgtgaaa taccgcacag atgcgtaagg
     1141 agaaaatacc gcatcaggcg ctcttccgct tcctcgctca ctgactcgct gcgctcggtc
     1201 gttcggctgc ggcgagcggt atcagctcac tcaaaggcgg taatacggtt atccacagaa
     1261 tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc agcaaaaggc caggaaccgt
     1321 aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga gcatcacaaa
     1381 aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata ccaggcgttt
     1441 ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac cggatacctg
     1501 tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg taggtatctc
     1561 agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc cgttcagccc
     1621 gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag acacgactta
     1681 tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt aggcggtgct
     1741 acagagttct tgaagtggtg gcctaactac ggctacacta gaagaatagt atttggtatc
     1801 tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg atccggcaaa
     1861 caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac gcgcagaaaa
     1921 aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca gtggaacgaa
     1981 aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt
     2041 ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac
     2101 agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc
     2161 atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc
     2221 cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata
     2281 aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc
     2341 cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc
     2401 aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca
     2461 ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa
     2521 gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca
     2581 ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt
     2641 tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt
     2701 tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg
     2761 ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga
     2821 tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc
     2881 agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg
     2941 acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag
     3001 ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg
     3061 gttccgcgca catttccccg aaaagtgcca cctaaattgt aagcgttaat attttgttaa
     3121 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     3181 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     3241 acaagagtcc actattaaga acgtggactc caacgtcaaa gggcgaaaaa ccgtctatca
     3301 gggcgatggc ccactacgtg aaccatcacc ctaatcaagt tttttggggt cgaggtgccg
     3361 taaagcacta aatcggaacc ctaaagggag cccccgattt agagcttgac ggggaaagcc
     3421 ggcgaacgtg gcgagaaagg aagggaagaa agcgaaagga gcgggcgcta gggcgctggc
     3481 aagtgtagcg gtcacgctgc gcgtaaccac cacacccgcc gcgcttaatg cgccgctaca
     3541 gggcgcgtcc cattcgccaa tccgtcggct tgaacgaatt gttagacatt atttgccgac
     3601 taccttggtg atctcgcctt tcacgtagtg gacaaattct tccaactgat ctgcgcgcga
     3661 ggccaagcga tcttcttctt gtccaagata agcctgtcta gcttcaagta tgacgggctg
     3721 atactgggcc ggcaggcgct ccattgccca gtcggcagcg acatccttcg gcgcgatttt
     3781 gccggttact gcgctgtacc aaatgcggga caacgtaagc actacatttc gctcatcgcc
     3841 agcccagtcg ggcggcgagt tccatagcgt taaggtttca tttagcgcct caaatagatc
     3901 ctgttcagga accggatcaa agagttcctc cgccgctgga cctaccaagg caacgctatg
     3961 ttctcttgct tttgtcagca agatagccag atcaatgtcg atcgtggctg gctcgaagat
     4021 acctgcaaga atgtcattgc gctgccattc tccaaattgc agttcgcgct tagctggata
     4081 acgccacgga atgatgtcgt cgtgcacaac aatggtgact tctacagcgc ggagaatctc
     4141 gctctctcca ggggaagccg aagtttccaa aaggtcgttg atcaaagctc gccgcgttgt
     4201 ttcatcaagc cttacggtca ccgtaaccag caaatcaata tcactgtgtg gcttcaggcc
     4261 gccatccact gcggagccgt acaaatgtac ggccagcaac gtcggttcga gatggcgctc
     4321 gatgacgcca actacctctg atagttgagt cgatacttcg gcgatcaccg cttccctcat
     4381 gatgtttaac tttgttttag ggcgactgcc ctgctgcgta acatcgttgc tgctccataa
     4441 catcaaacat cgacccacgg cgtaacgcgc ttgctgcttg gatgcccgag gcatagactg
     4501 taccccaaaa aaacagtcat aacaagccat gaaaaccgcc actgcgccgt taccaccgct
     4561 gcgttcggtc aaggttctgg accagttgcg tgagcgcata cgctacttac cggtttattg
     4621 actaccggaa gcagtgtgac cgtgtgcttc tcaaatgcct gaggccagtt tgctcaggct
     4681 ctccccgtgg aggtaataat tgacgatatg atcatttatt ctgcctccca gttcaggctg
     4741 caaataagcg ttgatattca gtcaattaca aacattaata acgaagagat gacagaaaaa
     4801 ttttcattct gtgacagaga aaaagtagcc gaagatgacg gtttgtcaca tggagttggc
     4861 aggatgtttg attaaaaaca taacaggaag aaaaatgccc cgctgtgggc ggacaaaata
     4921 gttgggaact gggaggggtg gaaatggagt ttttaaggat tatttaggga agagtgacaa
     4981 aatagatggg aactgggtgt agcgtcgtaa gctaatacga aaattaaaaa tgacaaaata
     5041 gtttggaact agatttcact tatctggttc ggatctcctg tttaaacgtg caatagtatc
     5101 cagtttttaa ttaacccgta gtggctatgg cagggcttgc cgccccgacg ttggctgcga
     5161 gccctgggcc ttcacccgaa cttgggggtt ggggtgggga aaaggaagaa acgcgggcgt
     5221 attggtccca atggggtctc ggtggggtat cgacgtaacc ccgattgaga tatagattta
     5281 tttaaatggt tatgatagtt attgctcagc ggtggcagca gccaactcag cttcctttcg
     5341 ggctttgtta gcagccggat cttctaggct caagcagtga tcagatccag acatgataag
     5401 atacattgat gagtttggac aaaccacaac tagaatgcag tgaaaaaaat gctttatttg
     5461 tgaaatttgt gatgctattg ctttatttgt aaccattata agctgcaata aacaagttaa
     5521 caacaacaat tgcattcatt ttatgtttca ggttcagggg gaggtgtggg aggtttttta
     5581 aagcaagtaa aacctctaca aatgtggtat ggctgattat gatcctctag tacttctcga
     5641 caagctttta ctgcagcttc acagcggagt tagcacgcag agcagtcacg atcaggggcc
     5701 aagccaggtt gggagagttg tccatggaga tctcggacag ctgcacgatc ttggagtcag
     5761 cgtccaccac ttgctggatc tcccacagag cggaagcgta ggtgaaggtg gtgccgtcgc
     5821 aagtgttctt gtaggtgttg tagtcgggga tgaccaccat cagcttagca gcggtggtca
     5881 gagggatgat gttcaggggc acgcaaccgt cacgagcgtt gttgatgatg ttgttcagag
     5941 cgtcgttgtc cagcttgcgg agcatggtga acagcatggt ctgcatagcg gaggtcactt
     6001 tagcacgctt gtcctcggaa cgagcctgct tgtacatctg ggtcatagcc tggtcagcca
     6061 tcttctcgag cttgcgctgc atagcagcgt cacggtcgaa ctcggacttg gccacgttca
     6121 gggacttttt cagcttcttc agcaccacct cagagtcgcc gttagccaca gcctgctcgt
     6181 aagcctcttg agcggtagcg aaagcagcgt aggagggcag agaggagaac tcggaagcga
     6241 tagcgtgatg gtggtgatgg tgctgcaggg tagcacggtt gtccagcatc tcttcgcaca
     6301 gcttgttgat gtccacagca ccttgcatag acagcaggac ggacagcagg gacaccatct
     6361 tctcgaaagc ctcggtggtg tccttagcca gcaggatgtc gttgtgcagt tggacgcact
     6421 gagcccacag cttgctggag gattccacac gcagctgttg cagcacagac agcagcacca
     6481 cggaggtgca cttcacgtcg gacatcttgg acatggtggc tggatccgcg cccgatggtg
     6541 ggacggtatg aataatccgg aatatttata ggttttttta ttacaaaact gttacgaaaa
     6601 cagtaaaata cttatttatt tgcgagatgg ttatcatttt aattatctcc atgatctatt
     6661 aatattccgg agtaggtcga tttaaatctt tagaccatag agcgttcact tgacagagtg
     6721 ccagccctgg gaccgaaccc cgcgtttatg aacaaacgac ccaacacccg tgcgttttat
     6781 tctgtctttt tattgccgtc atagcgcggg ttccttccgg tattgtctcc ttccgtgttt
     6841 cagttagcct cgataaaaac cccgattgag atatagttta aacccagata acttcgtata
     6901 atgtatgcta tacgaagtta tggtaccgcg gccgcgtaga ggatctgttg atcagcagtt
     6961 caacctgttg atagtacaaa caaagctgta gccaaccact agaactatag ctagagtcct
     7021 gggcgaacaa acgatgctcg ccttccagaa aaccgaggat gcgaaccact tcatccgggg
     7081 tcagcaccac cggcaagcgc cgcgacggcc gaggtctacc gatctcctga agcaagggca
     7141 gatccgtgca cagcaccttg ccgtagaaga acagcaaggc cgccaatgcc tgacgatgcg
     7201 tggagaccga aaccttgcgc tcgttcgcca gccaggacag aaatgcctcg acttcgctgc
     7261 tgcccaaggt tgccgggtga cgcacaccgt ggaaacggat gaaggcacaa acccagttg
//
