LOCUS       sequence-402644-        6518 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses SV-A 3C protease from a GAL10 promoter with a URA3 marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6518)
  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 6518)
  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-SV-A_3C
FEATURES             Location/Qualifiers
     source          1..6518
                     /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(2506..2522)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2507..2523)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      2525..2772
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(2541..2559)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2790..2809)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2791..2809)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2819..2836)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2819..2835)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2828..2850)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2976..3431
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3063..3082)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3273..3294
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3562..4365)
                     /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(4366..4581)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4605..4624)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4724..4746
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4784..4802)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    4843..5346
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        5383..5487
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5488..5556,5557..6348)
                     /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(5706..5725)
                     /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 tacaaaatgg
     1861 gtttcgaccc cgccgtcatg aagatcatgg gcaatgtgga ctcttttgtc accctctcgg
     1921 gtactaaacc catctggact atgtcctgcc tctggattgg cggtcgcaat ctgattgctc
     1981 cttcccacgc attcgtctcc gacgagtacg agatcaccca catccgcgtt ggatcgcgca
     2041 cgcttgacgt gtcgcgtgtt acgcgggtgg atgacggtga gctgtctcta ctctcggtgc
     2101 cggatgggcc ggagcataag agcttgatcc gctacatccg atctgcttcc cccaaatctg
     2161 gtatactagc ctccaaattc tctgacactc ctgtctttgt ttctttctgg aatggcaagt
     2221 cccactccac ccccctcccc ggggtcgtgg acgagaaaga ctcgttcacg taccgctgct
     2281 catctttcca aggcttgtgc ggttcgccga tgattgccac tgatcctggc ggcttgggta
     2341 tcctcggtat ccacgtcgct ggggtggctg gctacaacgg cttctccgca cgcctcaccc
     2401 ccgagcgtgt ccaagctttc ctttcccacc tggctacacc ccaataaaac ccagctttct
     2461 tgtactaagt ggtgatgggc tgcaggaatt cgatatcaag cttatcgata ccgtcgacct
     2521 cgagtcatgt aattagttat gtcacgctta cattcacgcc ctccccccac atccgctcta
     2581 accgaaaagg aaggagttag acaacctgaa gtctaggtcc ctatttattt ttttatagtt
     2641 atgttagtat taagaacgtt atttatattt caaatttttc ttttttttct gtacagacgc
     2701 gtgtacgcat gtaacattat actgaaaacc ttgcttgaga aggttttggg acgctcgaag
     2761 gctttaattt gcggccggta cccaattcgc cctatagtga gtcgtattac gcgcgctcac
     2821 tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc
     2881 ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc
     2941 cttcccaaca gttgcgcagc ctgaatggcg aatggacgcg ccctgtagcg gcgcattaag
     3001 cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc
     3061 cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc cccgtcaagc
     3121 tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc tcgaccccaa
     3181 aaaacttgat tagggtgatg gttcacgtag tgggccatcg ccctgataga cggtttttcg
     3241 ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa ctggaacaac
     3301 actcaaccct atctcggtct attcttttga tttataaggg attttgccga tttcggccta
     3361 ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca aaatattaac
     3421 gcttacaatt tcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     3481 atagggtaat aactgatata attaaattga agctctaatt tgtgagttta gtatacatgc
     3541 atttacttat aatacagttt tttagttttg ctggccgcat cttctcaaat atgcttccca
     3601 gcctgctttt ctgtaacgtt caccctctac cttagcatcc cttccctttg caaatagtcc
     3661 tcttccaaca ataataatgt cagatcctgt agagaccaca tcatccacgg ttctatactg
     3721 ttgacccaat gcgtctccct tgtcatctaa acccacaccg ggtgtcataa tcaaccaatc
     3781 gtaaccttca tctcttccac ccatgtctct ttgagcaata aagccgataa caaaatcttt
     3841 gtcgctcttc gcaatgtcaa cagtaccctt agtatattct ccagtagata gggagccctt
     3901 gcatgacaat tctgctaaca tcaaaaggcc tctaggttcc tttgttactt cttctgccgc
     3961 ctgcttcaaa ccgctaacaa tacctgggcc caccacaccg tgtgcattcg taatgtctgc
     4021 ccattctgct attctgtata cacccgcaga gtactgcaat ttgactgtat taccaatgtc
     4081 agcaaatttt ctgtcttcga agagtaaaaa attgtacttg gcggataatg cctttagcgg
     4141 cttaactgtg ccctccatgg aaaaatcagt caagatatcc acatgtgttt ttagtaaaca
     4201 aattttggga cctaatgctt caactaactc cagtaattcc ttggtggtac gaacatccaa
     4261 tgaagcacac aagtttgttt gcttttcgtg catgatatta aatagcttgg cagcaacagg
     4321 actaggatga gtagcagcac gttccttata tgtagctttc gacatgattt atcttcgttt
     4381 cctgcaggtt tttgttctgt gcagttgggt taagaatact gggcaatttc atgtttcttc
     4441 aacactacat atgcgtatat ataccaatct aagtctgtgc tccttccttc gttcttcctt
     4501 ctgttcggag attaccgaat caaaaaaatt tcaaggaaac cgaaatcaaa aaaaagaata
     4561 aaaaaaaaat gatgaattga aaaggtggta tggtgcactc tcagtacaat ctgctctgat
     4621 gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct
     4681 tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt
     4741 cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta
     4801 tttttatagg ttaatgtcat gataataatg gtttcttaga cggatcgctt gcctgtaact
     4861 tacacgcgcc tcgtatcttt taatgatgga ataatttggg aatttactct gtgtttattt
     4921 atttttatgt tttgtatttg gattttagaa agtaaataaa gaaggtagaa gagttacgga
     4981 atgaagaaaa aaaaataaac aaaggtttaa aaaatttcaa caaaaagcgt actttacata
     5041 tatatttatt agacaagaaa agcagattaa atagatatac attcgattaa cgataagtaa
     5101 aatgtaaaat cacaggattt tcgtgtgtgg tcttctacac agacaagatg aaacaattcg
     5161 gcattaatac ctgagagcag gaagagcaag ataaaaggta gtatttgttg gcgatccccc
     5221 tagagtcttt tacatcttcg gaaaacaaaa actatttttt ctttaatttc tttttttact
     5281 ttctattttt aatttatata tttatattaa aaaatttaaa ttataattat ttttatagca
     5341 cgtgatgaaa aggacccagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt
     5401 ttatttttct aaatacattc aaatatgtat ccgctcatga gacaataacc ctgataaatg
     5461 cttcaataat attgaaaaag gaagagtatg agtattcaac atttccgtgt cgcccttatt
     5521 cccttttttg cggcattttg ccttcctgtt tttgctcacc cagaaacgct ggtgaaagta
     5581 aaagatgctg aagatcagtt gggtgcacga gtgggttaca tcgaactgga tctcaacagc
     5641 ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc caatgatgag cacttttaaa
     5701 gttctgctat gtggcgcggt attatcccgt attgacgccg ggcaagagca actcggtcgc
     5761 cgcatacact attctcagaa tgacttggtt gagtactcac cagtcacaga aaagcatctt
     5821 acggatggca tgacagtaag agaattatgc agtgctgcca taaccatgag tgataacact
     5881 gcggccaact tacttctgac aacgatcgga ggaccgaagg agctaaccgc ttttttgcac
     5941 aacatggggg atcatgtaac tcgccttgat cgttgggaac cggagctgaa tgaagccata
     6001 ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg caacaacgtt gcgcaaacta
     6061 ttaactggcg aactacttac tctagcttcc cggcaacaat taatagactg gatggaggcg
     6121 gataaagttg caggaccact tctgcgctcg gcccttccgg ctggctggtt tattgctgat
     6181 aaatctggag ccggtgagcg tgggtctcgc ggtatcattg cagcactggg gccagatggt
     6241 aagccctccc gtatcgtagt tatctacacg acggggagtc aggcaactat ggatgaacga
     6301 aatagacaga tcgctgagat aggtgcctca ctgattaagc attggtaact gtcagaccaa
     6361 gtttactcat atatacttta gattgattta aaacttcatt tttaatttaa aaggatctag
     6421 gtgaagatcc tttttgataa tctcatgacc aaaatccctt aacgtgagtt ttcgttccac
     6481 tgagcgtcag accccgtaga aaagatcaaa ggatcttc
//
