LOCUS       sequence-402753-        7423 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses SARS-CoV-2 papain-like protease from a GAL promoter with a
            URA3 marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7423)
  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 7423)
  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: pAG426GAL-SARS-CoV-2_PLP
FEATURES             Location/Qualifiers
     source          1..7423
                     /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        1054..1495
                     /gene="S. cerevisiae GAL1"
                     /label=GAL1 promoter
                     /note="inducible promoter, regulated by Gal4"
     protein_bind    1054..1171
                     /label=UAS
                     /bound_moiety="Gal4"
                     /note="upstream activating sequence mediating 
                     Gal4-dependent induction"
     primer_bind     1466..1489
                     /label=GAL1
                     /note="S. cerevisiae GAL1 promoter, forward primer"
     primer_bind     1510..1526
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     1510..1526
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     protein_bind    1534..1558
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     protein_bind    complement(2525..2549)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(2582..2598)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2583..2599)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      2601..2848
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(2617..2635)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2866..2885)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2867..2885)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2895..2912)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2895..2911)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2904..2926)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3052..3507
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3139..3158)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3349..3370
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3638..4441)
                     /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(4442..4657)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4681..4700)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4800..4822
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4860..4878)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     rep_origin      4919..6261
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    5251..5298
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     promoter        6288..6392
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6393..6461,6462..7253)
                     /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(6611..6630)
                     /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 gtacggatta gaagccgccg agcgggtgac
     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 accccggatt ctagaactag tggatccccc atcacaagtt tgtacaaaaa agcaggctac
     1561 aaaatggaag tgaggactat taaggtgttt acaacagtag acaacattaa cctccacacg
     1621 caagttgtgg acatgtcaat gacatatgga caacagtttg gtccaactta tttggatgga
     1681 gctgatgtta ctaaaataaa acctcataat tcacatgaag gtaaaacatt ttatgtttta
     1741 cctaatgatg acactctacg tgttgaggct tttgagtact accacacaac tgatcctagt
     1801 tttctgggta ggtacatgtc agcattaaat cacactaaaa agtggaaata cccacaagtt
     1861 aatggtttaa cttctattaa atgggcagat aacaactgtt atcttgccac tgcattgtta
     1921 acactccaac aaatagagtt gaagtttaat ccacctgctc tacaagatgc ttattacaga
     1981 gcaagggctg gtgaagctgc taacttttgt gcacttatct tagcctactg taataagaca
     2041 gtaggtgagt taggtgatgt tagagaaaca atgagttact tgtttcaaca tgccaattta
     2101 gattcttgca aaagagtctt gaacgtggtg tgtaaaactt gtggacaaca gcagacaacc
     2161 cttaagggtg tagaagctgt tatgtacatg ggcacacttt cttatgaaca atttaagaaa
     2221 ggtgttcaga taccttgtac gtgtggtaaa caagctacaa aatatctagt acaacaggag
     2281 tcaccttttg ttatgatgtc agcaccacct gctcagtatg aacttaagca tggtacattt
     2341 acttgtgcta gtgagtacac tggtaattac cagtgtggtc actataaaca tataacttct
     2401 aaagaaactt tgtattgcat agacggtgct ttacttacaa agtcctcaga atacaaaggt
     2461 cctattacgg atgttttcta caaagaaaac agttacacaa caaccataaa accagttact
     2521 taaaacccag ctttcttgta caaagtggtg atgggctgca ggaattcgat atcaagctta
     2581 tcgataccgt cgacctcgag tcatgtaatt agttatgtca cgcttacatt cacgccctcc
     2641 ccccacatcc gctctaaccg aaaaggaagg agttagacaa cctgaagtct aggtccctat
     2701 ttattttttt atagttatgt tagtattaag aacgttattt atatttcaaa tttttctttt
     2761 ttttctgtac agacgcgtgt acgcatgtaa cattatactg aaaaccttgc ttgagaaggt
     2821 tttgggacgc tcgaaggctt taatttgcgg ccggtaccca attcgcccta tagtgagtcg
     2881 tattacgcgc gctcactggc cgtcgtttta caacgtcgtg actgggaaaa ccctggcgtt
     2941 acccaactta atcgccttgc agcacatccc cctttcgcca gctggcgtaa tagcgaagag
     3001 gcccgcaccg atcgcccttc ccaacagttg cgcagcctga atggcgaatg gacgcgccct
     3061 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     3121 ccagcgccct agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     3181 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     3241 ggcacctcga ccccaaaaaa cttgattagg gtgatggttc acgtagtggg ccatcgccct
     3301 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     3361 tccaaactgg aacaacactc aaccctatct cggtctattc ttttgattta taagggattt
     3421 tgccgatttc ggcctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     3481 ttaacaaaat attaacgctt acaatttcct gatgcggtat tttctcctta cgcatctgtg
     3541 cggtatttca caccgcatag ggtaataact gatataatta aattgaagct ctaatttgtg
     3601 agtttagtat acatgcattt acttataata cagtttttta gttttgctgg ccgcatcttc
     3661 tcaaatatgc ttcccagcct gcttttctgt aacgttcacc ctctacctta gcatcccttc
     3721 cctttgcaaa tagtcctctt ccaacaataa taatgtcaga tcctgtagag accacatcat
     3781 ccacggttct atactgttga cccaatgcgt ctcccttgtc atctaaaccc acaccgggtg
     3841 tcataatcaa ccaatcgtaa ccttcatctc ttccacccat gtctctttga gcaataaagc
     3901 cgataacaaa atctttgtcg ctcttcgcaa tgtcaacagt acccttagta tattctccag
     3961 tagataggga gcccttgcat gacaattctg ctaacatcaa aaggcctcta ggttcctttg
     4021 ttacttcttc tgccgcctgc ttcaaaccgc taacaatacc tgggcccacc acaccgtgtg
     4081 cattcgtaat gtctgcccat tctgctattc tgtatacacc cgcagagtac tgcaatttga
     4141 ctgtattacc aatgtcagca aattttctgt cttcgaagag taaaaaattg tacttggcgg
     4201 ataatgcctt tagcggctta actgtgccct ccatggaaaa atcagtcaag atatccacat
     4261 gtgtttttag taaacaaatt ttgggaccta atgcttcaac taactccagt aattccttgg
     4321 tggtacgaac atccaatgaa gcacacaagt ttgtttgctt ttcgtgcatg atattaaata
     4381 gcttggcagc aacaggacta ggatgagtag cagcacgttc cttatatgta gctttcgaca
     4441 tgatttatct tcgtttcctg caggtttttg ttctgtgcag ttgggttaag aatactgggc
     4501 aatttcatgt ttcttcaaca ctacatatgc gtatatatac caatctaagt ctgtgctcct
     4561 tccttcgttc ttccttctgt tcggagatta ccgaatcaaa aaaatttcaa ggaaaccgaa
     4621 atcaaaaaaa agaataaaaa aaaaatgatg aattgaaaag gtggtatggt gcactctcag
     4681 tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa cacccgctga
     4741 cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg tgaccgtctc
     4801 cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga gacgaaaggg
     4861 cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt cttagtatga
     4921 tccaatatca aaggaaatga tagcattgaa ggatgagact aatccaattg aggagtggca
     4981 gcatatagaa cagctaaagg gtagtgctga aggaagcata cgataccccg catggaatgg
     5041 gataatatca caggaggtac tagactacct ttcatcctac ataaatagac gcatataagt
     5101 acgcatttaa gcataaacac gcactatgcc gttcttctca tgtatatata tatacaggca
     5161 acacgcagat ataggtgcga cgtgaacagt gagctgtatg tgcgcagctc gcgttgcatt
     5221 ttcggaagcg ctcgttttcg gaaacgcttt gaagttccta ttccgaagtt cctattctct
     5281 agaaagtata ggaacttcag agcgcttttg aaaaccaaaa gcgctctgaa gacgcacttt
     5341 caaaaaacca aaaacgcacc ggactgtaac gagctactaa aatattgcga ataccgcttc
     5401 cacaaacatt gctcaaaagt atctctttgc tatatatctc tgtgctatat ccctatataa
     5461 cctacccatc cacctttcgc tccttgaact tgcatctaaa ctcgacctct acatttttta
     5521 tgtttatctc tagtattact ctttagacaa aaaaattgta gtaagaacta ttcatagagt
     5581 gaatcgaaaa caatacgaaa atgtaaacat ttcctatacg tagtatatag agacaaaata
     5641 gaagaaaccg ttcataattt tctgaccaat gaagaatcat caacgctatc actttctgtt
     5701 cacaaagtat gcgcaatcca catcggtata gaatataatc ggggatgcct ttatcttgaa
     5761 aaaatgcacc cgcagcttcg ctagtaatca gtaaacgcgg gaagtggagt caggcttttt
     5821 ttatggaaga gaaaatagac accaaagtag ccttcttcta accttaacgg acctacagtg
     5881 caaaaagtta tcaagagact gcattataga gcgcacaaag gagaaaaaaa gtaatctaag
     5941 atgctttgtt agaaaaatag cgctctcggg atgcattttt gtagaacaaa aaagaagtat
     6001 agattctttg ttggtaaaat agcgctctcg cgttgcattt ctgttctgta aaaatgcagc
     6061 tcagattctt tgtttgaaaa attagcgctc tcgcgttgca tttttgtttt acaaaaatga
     6121 agcacagatt cttcgttggt aaaatagcgc tttcgcgttg catttctgtt ctgtaaaaat
     6181 gcagctcaga ttctttgttt gaaaaattag cgctctcgcg ttgcattttt gttctacaaa
     6241 atgaagcaca gatgcttcgt tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     6301 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     6361 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     6421 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     6481 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     6541 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     6601 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     6661 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     6721 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     6781 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     6841 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     6901 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca
     6961 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     7021 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     7081 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     7141 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     7201 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     7261 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     7321 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     7381 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttc
//
