LOCUS       sequence-402647-        6647 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses ZIKV NS2B-NS3 protease (with GS linker) from a GAL10 
            promoter with a URA3 marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6647)
  AUTHORS   Hong SJ, Resnick SJ, Iketani S, Cha JW, Albert BA, Fazekas CT, Chang
            CW, Liu H, Dagan S, Abagyan MR, Fajtova P, Culbertson B, Brace B, 
            Reddem ER, Forouhar F, Glickman JF, Balkovec JM, Stockwell BR, 
            Shapiro L, O'Donoghue AJ, Sabo Y, Freundlich JS, Ho DD, Chavez A
  TITLE     A multiplex method for rapidly identifying viral protease 
            inhibitors.
  JOURNAL   Mol Syst Biol. 2025 Feb;21(2):158-172. doi: 
            10.1038/s44320-024-00082-1. Epub 2025 Jan 6.
  PUBMED    39762652
REFERENCE   2  (bases 1 to 6647)
  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: 
            "10.1038/s44320-024-00082-1"; journalName: "Mol Syst Biol"; date: 
            "2025-02"; volume: "21"; issue: "2"; pages: "158-172"
COMMENT     Sequence Label: pAG416GAL10-ZIKV_NS2B-GS-NS3
FEATURES             Location/Qualifiers
     source          1..6647
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     743..760
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    877..898
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(913..918,919..936,937..943)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    951..967
                     /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)."
     primer_bind     956..978
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     975..991
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     975..991
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     1010..1030
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        1012..1030
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     promoter        complement(1108..1549)
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     primer_bind     complement(1114..1137)
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     protein_bind    1432..1549
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     protein_bind    1827..1851
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2635..2651)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2636..2652)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      2654..2901
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(2670..2688)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2919..2938)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2920..2938)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2948..2965)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2948..2964)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2957..2979)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3105..3560
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3192..3211)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3402..3423
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3691..4494)
                     /codon_start=1
                     /gene="S. cerevisiae URA3"
                     /product="orotidine-5'-phosphate decarboxylase, required 
                     for uracil biosynthesis"
                     /label=URA3
                     /note="yeast auxotrophic marker, counterselectable with 
                     5-fluoroorotic acid (5-FOA)"
                     /translation="MSKATYKERAATHPSPVAAKLFNIMHEKQTNLCASLDVRTTKELL
                     ELVEALGPKICLLKTHVDILTDFSMEGTVKPLKALSAKYNFLLFEDRKFADIGNTVKLQ
                     YSAGVYRIAEWADITNAHGVVGPGIVSGLKQAAEEVTKEPRGLLMLAELSCKGSLSTGE
                     YTKGTVDIAKSDKDFVIGFIAQRDMGGRDEGYDWLIMTPGVGLDDKGDALGQQYRTVDD
                     VVSTGSDIIIVGRGLFAKGRDAKVEGERYRKAGWEAYLRRCGQQN"
     promoter        complement(4495..4710)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4734..4753)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4853..4875
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4913..4931)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    4972..5475
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        5512..5616
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5617..5685,5686..6477)
                     /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(5835..5854)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
      841 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
      901 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
      961 ataacaattt cacacaggaa acagctatga ccatgattac gccaagcgcg caattaaccc
     1021 tcactaaagg gaacaaaagc tggagctcta tgaccatgat tacgccaagc ggggtgtgtt
     1081 gttggtgggt tgggtgtata gttttttctc cttgacgtta aagtatagag gtatattaac
     1141 aattttttgt tgatactttt atgacatttg aataagaagt aatacaaact gaaaatgttg
     1201 aaagtattag ttaaagtggt tatgcagctt ttccatttat atatctgtta atagatcaaa
     1261 aatcatcgct tcgctgatta attaccccag aaataaggct aaaaaactaa tcgcattatc
     1321 atcctatggt tgttaatttg attcgttaat ttgaaggttt gtggggccag gttactgcca
     1381 atttttcctc ttcataacca taaaagctag tattgtagaa tctttattgt tcggagcagt
     1441 gcggcgcgag gcacatctgc gtttcaggaa cgcgaccggt gaagacgagg acgcacggag
     1501 gagagtcttc cgtcggaggg ctgtcgcccg ctcggcggct tctaatccgt acttcaatat
     1561 agcaatgagc agttaagcgt attactgaaa gttccaaaga gaaggttttt ttaggctaag
     1621 ataatggggc tctttacatt tccacaacat ataagtaaga ttagatatgg atatgtatat
     1681 ggtggtaatg ccatgtaata tgattattaa acttctttgc gtccatccaa aaaaaaagta
     1741 agaatttttg aaaattcaat ataaatgtct aaaggtgaag aattattcag aattccagct
     1801 gaccaccgac tagtggatcc cccatcacaa gtttgtacaa aaaagcaggc tacaaaatgt
     1861 cgggaaagag tgtggacatg tacattgaaa gagcaggtga catcacatgg gaaaaggacg
     1921 cggaagtcac tggaaacagt cctcggcttg acgtggcact ggatgagagt ggtgacttct
     1981 ccttggtaga ggaagatggt ccacccatga gagagggagg tggaggttca ggggggggtg
     2041 gaagtggcgc cctctgggac gtgcctgctc ccaaagaagt gaagaaagga gagaccacag
     2101 atggagtgta cagagtgatg actcgcagac tgctaggttc aacacaggtt ggagtgggag
     2161 tcatgcaaga gggagtcttc cacaccatgt ggcacgttac aaaaggagcc gcactgagga
     2221 gcggtgaggg aagacttgat ccatactggg gggatgtcaa gcaggacttg gtgtcatact
     2281 gtgggccttg gaagttggat gcagcttggg atggactcag cgaggtacag cttttggccg
     2341 tacctcccgg agagagggcc agaaacattc agaccctgcc tggaatattc aagacaaagg
     2401 acggggacat cggagcagtt gctctggact accctgcagg gacctcagga tctccgatcc
     2461 tagacaaatg tggaagagtg ataggactct atggcaatgg ggttgtgatc aagaatggaa
     2521 gctatgttag tgctataacc cagggaaaga gggaggagga gactccggtt gaataaaacc
     2581 cagctttctt gtactaagtg gtgatgggct gcaggaattc gatatcaagc ttatcgatac
     2641 cgtcgacctc gagtcatgta attagttatg tcacgcttac attcacgccc tccccccaca
     2701 tccgctctaa ccgaaaagga aggagttaga caacctgaag tctaggtccc tatttatttt
     2761 tttatagtta tgttagtatt aagaacgtta tttatatttc aaatttttct tttttttctg
     2821 tacagacgcg tgtacgcatg taacattata ctgaaaacct tgcttgagaa ggttttggga
     2881 cgctcgaagg ctttaatttg cggccggtac ccaattcgcc ctatagtgag tcgtattacg
     2941 cgcgctcact ggccgtcgtt ttacaacgtc gtgactggga aaaccctggc gttacccaac
     3001 ttaatcgcct tgcagcacat ccccctttcg ccagctggcg taatagcgaa gaggcccgca
     3061 ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga atggacgcgc cctgtagcgg
     3121 cgcattaagc gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc
     3181 cctagcgccc gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc
     3241 ccgtcaagct ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct
     3301 cgaccccaaa aaacttgatt agggtgatgg ttcacgtagt gggccatcgc cctgatagac
     3361 ggtttttcgc cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac
     3421 tggaacaaca ctcaacccta tctcggtcta ttcttttgat ttataaggga ttttgccgat
     3481 ttcggcctat tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa
     3541 aatattaacg cttacaattt cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     3601 tcacaccgca tagggtaata actgatataa ttaaattgaa gctctaattt gtgagtttag
     3661 tatacatgca tttacttata atacagtttt ttagttttgc tggccgcatc ttctcaaata
     3721 tgcttcccag cctgcttttc tgtaacgttc accctctacc ttagcatccc ttccctttgc
     3781 aaatagtcct cttccaacaa taataatgtc agatcctgta gagaccacat catccacggt
     3841 tctatactgt tgacccaatg cgtctccctt gtcatctaaa cccacaccgg gtgtcataat
     3901 caaccaatcg taaccttcat ctcttccacc catgtctctt tgagcaataa agccgataac
     3961 aaaatctttg tcgctcttcg caatgtcaac agtaccctta gtatattctc cagtagatag
     4021 ggagcccttg catgacaatt ctgctaacat caaaaggcct ctaggttcct ttgttacttc
     4081 ttctgccgcc tgcttcaaac cgctaacaat acctgggccc accacaccgt gtgcattcgt
     4141 aatgtctgcc cattctgcta ttctgtatac acccgcagag tactgcaatt tgactgtatt
     4201 accaatgtca gcaaattttc tgtcttcgaa gagtaaaaaa ttgtacttgg cggataatgc
     4261 ctttagcggc ttaactgtgc cctccatgga aaaatcagtc aagatatcca catgtgtttt
     4321 tagtaaacaa attttgggac ctaatgcttc aactaactcc agtaattcct tggtggtacg
     4381 aacatccaat gaagcacaca agtttgtttg cttttcgtgc atgatattaa atagcttggc
     4441 agcaacagga ctaggatgag tagcagcacg ttccttatat gtagctttcg acatgattta
     4501 tcttcgtttc ctgcaggttt ttgttctgtg cagttgggtt aagaatactg ggcaatttca
     4561 tgtttcttca acactacata tgcgtatata taccaatcta agtctgtgct ccttccttcg
     4621 ttcttccttc tgttcggaga ttaccgaatc aaaaaaattt caaggaaacc gaaatcaaaa
     4681 aaaagaataa aaaaaaaatg atgaattgaa aaggtggtat ggtgcactct cagtacaatc
     4741 tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc
     4801 tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc
     4861 tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg
     4921 atacgcctat ttttataggt taatgtcatg ataataatgg tttcttagac ggatcgcttg
     4981 cctgtaactt acacgcgcct cgtatctttt aatgatggaa taatttggga atttactctg
     5041 tgtttattta tttttatgtt ttgtatttgg attttagaaa gtaaataaag aaggtagaag
     5101 agttacggaa tgaagaaaaa aaaataaaca aaggtttaaa aaatttcaac aaaaagcgta
     5161 ctttacatat atatttatta gacaagaaaa gcagattaaa tagatataca ttcgattaac
     5221 gataagtaaa atgtaaaatc acaggatttt cgtgtgtggt cttctacaca gacaagatga
     5281 aacaattcgg cattaatacc tgagagcagg aagagcaaga taaaaggtag tatttgttgg
     5341 cgatccccct agagtctttt acatcttcgg aaaacaaaaa ctattttttc tttaatttct
     5401 ttttttactt tctattttta atttatatat ttatattaaa aaatttaaat tataattatt
     5461 tttatagcac gtgatgaaaa ggacccaggt ggcacttttc ggggaaatgt gcgcggaacc
     5521 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     5581 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
     5641 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
     5701 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
     5761 ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc
     5821 acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa
     5881 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
     5941 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
     6001 gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct
     6061 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
     6121 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
     6181 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
     6241 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
     6301 attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg
     6361 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
     6421 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg
     6481 tcagaccaag tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     6541 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     6601 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttc
//
