LOCUS       sequence-402754-        6313 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses HAV 3C protease from a GALL promoter with a URA3 marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6313)
  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 6313)
  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: pAG416GALL-HAV_3C
FEATURES             Location/Qualifiers
     source          1..6313
                     /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"
     primer_bind     1466..1489
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     protein_bind    1550..1574
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2301..2317)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2302..2318)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      2320..2567
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(2336..2354)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2585..2604)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2586..2604)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2614..2631)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2614..2630)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2623..2645)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2771..3226
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2858..2877)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3068..3089
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3357..4160)
                     /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(4161..4376)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4400..4419)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4519..4541
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4579..4597)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    4638..5141
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        5178..5282
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5283..5351,5352..6143)
                     /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(5501..5520)
                     /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 ggctcgtgac
     1081 agccctccga aggaagactc tcctccgtgc gtcctcgtct tcaccggtcg cgttcctgaa
     1141 acgcagatgt gcctcgcgcc gcactgctcc gaacaataaa gattctacaa tactagcttt
     1201 tatggttatg aagaggaaaa attggcagta acctggcccc acaaaccttc aaatgaacga
     1261 atcaaattaa caaccatagg atgataatgc gattagtttt ttagccttat ttctggggta
     1321 attaatcagc gaagcgatga tttttgatct attaacagat atataaatgc aaaaactgca
     1381 taaccacttt aactaatact ttcaacattt tcggtttgta ttacttctta ttcaaatgta
     1441 ataaaagtat caacaaaaaa ttgttaatat acctctatac tttaacgtca aggagaaaaa
     1501 accccggatc cggaattcca gctgaccacc gactagtgga tcccccatca caagtttgta
     1561 caaaaaagca ggctacaaaa tgtcaacttt ggaaatagca ggactggtta ggaagaactt
     1621 ggttcagttt ggagttggag agaagaatgg atgtgtgaga tgggttatga atgccttggg
     1681 agtgaaagat gattggctgc ttgtgccttc ccatgcttat aaatttgaga aagattatga
     1741 aatgatggag ttttatttta atagaggtgg aacttactat tcaatttcag ctggtaatgt
     1801 tgttattcaa tctttggatg tgggattcca ggatgttgtt ctgatgaagg ttcctacaat
     1861 tcctaagttt agagatatta ctcagcattt tattaagaaa ggggatgtgc ctagagcttt
     1921 gaatcgcctg gcaacattag tgacaactgt aaatggaacc cctatgttaa tttctgaggg
     1981 cccactaaag atggaagaga aagctactta tgttcataag aaaaatgatg gtacaacagt
     2041 tgatttaact gtggatcagg catggagagg aaaaggcgaa ggtcttcctg gaatgtgtgg
     2101 tggggccttg gtttcatcga atcaatctat acagaatgca atcttgggca tccatgttgc
     2161 tggaggaaat tcaattcttg ttgcaaaatt ggttactcaa gaaatgttcc aaaatattga
     2221 taagaaaatt gaaagtcagt aaaacccagc tttcttgtac taagtggtga tgggctgcag
     2281 gaattcgata tcaagcttat cgataccgtc gacctcgagt catgtaatta gttatgtcac
     2341 gcttacattc acgccctccc cccacatccg ctctaaccga aaaggaagga gttagacaac
     2401 ctgaagtcta ggtccctatt tattttttta tagttatgtt agtattaaga acgttattta
     2461 tatttcaaat ttttcttttt tttctgtaca gacgcgtgta cgcatgtaac attatactga
     2521 aaaccttgct tgagaaggtt ttgggacgct cgaaggcttt aatttgcggc cggtacccaa
     2581 ttcgccctat agtgagtcgt attacgcgcg ctcactggcc gtcgttttac aacgtcgtga
     2641 ctgggaaaac cctggcgtta cccaacttaa tcgccttgca gcacatcccc ctttcgccag
     2701 ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc caacagttgc gcagcctgaa
     2761 tggcgaatgg acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     2821 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     2881 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     2941 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     3001 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     3061 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     3121 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     3181 caaaaattta acgcgaattt taacaaaata ttaacgctta caatttcctg atgcggtatt
     3241 ttctccttac gcatctgtgc ggtatttcac accgcatagg gtaataactg atataattaa
     3301 attgaagctc taatttgtga gtttagtata catgcattta cttataatac agttttttag
     3361 ttttgctggc cgcatcttct caaatatgct tcccagcctg cttttctgta acgttcaccc
     3421 tctaccttag catcccttcc ctttgcaaat agtcctcttc caacaataat aatgtcagat
     3481 cctgtagaga ccacatcatc cacggttcta tactgttgac ccaatgcgtc tcccttgtca
     3541 tctaaaccca caccgggtgt cataatcaac caatcgtaac cttcatctct tccacccatg
     3601 tctctttgag caataaagcc gataacaaaa tctttgtcgc tcttcgcaat gtcaacagta
     3661 cccttagtat attctccagt agatagggag cccttgcatg acaattctgc taacatcaaa
     3721 aggcctctag gttcctttgt tacttcttct gccgcctgct tcaaaccgct aacaatacct
     3781 gggcccacca caccgtgtgc attcgtaatg tctgcccatt ctgctattct gtatacaccc
     3841 gcagagtact gcaatttgac tgtattacca atgtcagcaa attttctgtc ttcgaagagt
     3901 aaaaaattgt acttggcgga taatgccttt agcggcttaa ctgtgccctc catggaaaaa
     3961 tcagtcaaga tatccacatg tgtttttagt aaacaaattt tgggacctaa tgcttcaact
     4021 aactccagta attccttggt ggtacgaaca tccaatgaag cacacaagtt tgtttgcttt
     4081 tcgtgcatga tattaaatag cttggcagca acaggactag gatgagtagc agcacgttcc
     4141 ttatatgtag ctttcgacat gatttatctt cgtttcctgc aggtttttgt tctgtgcagt
     4201 tgggttaaga atactgggca atttcatgtt tcttcaacac tacatatgcg tatatatacc
     4261 aatctaagtc tgtgctcctt ccttcgttct tccttctgtt cggagattac cgaatcaaaa
     4321 aaatttcaag gaaaccgaaa tcaaaaaaaa gaataaaaaa aaaatgatga attgaaaagg
     4381 tggtatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc cagccccgac
     4441 acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca tccgcttaca
     4501 gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg tcatcaccga
     4561 aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat gtcatgataa
     4621 taatggtttc ttagacggat cgcttgcctg taacttacac gcgcctcgta tcttttaatg
     4681 atggaataat ttgggaattt actctgtgtt tatttatttt tatgttttgt atttggattt
     4741 tagaaagtaa ataaagaagg tagaagagtt acggaatgaa gaaaaaaaaa taaacaaagg
     4801 tttaaaaaat ttcaacaaaa agcgtacttt acatatatat ttattagaca agaaaagcag
     4861 attaaataga tatacattcg attaacgata agtaaaatgt aaaatcacag gattttcgtg
     4921 tgtggtcttc tacacagaca agatgaaaca attcggcatt aatacctgag agcaggaaga
     4981 gcaagataaa aggtagtatt tgttggcgat ccccctagag tcttttacat cttcggaaaa
     5041 caaaaactat tttttcttta atttcttttt ttactttcta tttttaattt atatatttat
     5101 attaaaaaat ttaaattata attattttta tagcacgtga tgaaaaggac ccaggtggca
     5161 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     5221 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga
     5281 gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc
     5341 ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg
     5401 cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc
     5461 ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat
     5521 cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact
     5581 tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat
     5641 tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga
     5701 tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc
     5761 ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga
     5821 tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag
     5881 cttcccggca acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc
     5941 gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt
     6001 ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct
     6061 acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg
     6121 cctcactgat taagcattgg taactgtcag accaagttta ctcatatata ctttagattg
     6181 atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     6241 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     6301 tcaaaggatc ttc
//
