LOCUS       sequence-402868-        6591 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses Bat-SL-CoVZC45 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 6591)
  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 6591)
  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: pAG416GAL-Bat-SL-CoVZC45_PLP
FEATURES             Location/Qualifiers
     source          1..6591
                     /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"
     primer_bind     complement(2579..2595)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2580..2596)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      2598..2845
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(2614..2632)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(2863..2882)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2864..2882)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2892..2909)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2892..2908)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2901..2923)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3049..3504
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3136..3155)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3346..3367
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3635..4438)
                     /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(4439..4654)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(4678..4697)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4797..4819
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4857..4875)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    4916..5419
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        5456..5560
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5561..5629,5630..6421)
                     /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(5779..5798)
                     /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 tacggaacat caaagtgttc actacagttg ataacattaa cctgcacact
     1621 caggttgtgg acatgagtat gacctatggt cagcagtttg gccctattta tctggatggt
     1681 gctgacgtta cgaagataaa gcctcataat agccacgagg gtaagacctt ttacgtgctc
     1741 ccaaatgatg acaccctccg cgcagaggcg tttgagtact accacactac cgacccgtca
     1801 tttctggggc gctatatgag tgcgttgaac cataccaaaa agtggaagta ccctcaagta
     1861 aacgggctga ctagcataaa gtgggctgat aacaactgtt acttggctac agccctcctg
     1921 actatacaac agatcgaact gaaatttaac cctccggctc ttcaagacgc ctattatcga
     1981 gcgcgcgcag gcgaggctgc aaacttttgt gcgctcatat tggcttactg taataagaca
     2041 gtaggcgagc tcggagacgt gcgcgaaaca atgaactact tgttccaaca cgcaaatctg
     2101 gactcatgca agagagtact taatgtcgtc tgcaagactt gcggccaaca gcagacgacc
     2161 ctgaaaggtg tggaggcagt aatgtacatg ggtacccttt cctacgagca actcaagaag
     2221 ggagtccaga tcccgtgcat gtgcggaaaa caagccacgc aatatctggt acagcaagag
     2281 agcccatttg taatgatgtc cgctcccccc gcgcaatatg aacttaaaca tgggacattc
     2341 gtatgtgcgt ctgaatatac cggcaactat cagtgtgggc attacaaaca tatcacttct
     2401 aaagagacgc tctattgtat cgacggcgcg ctcctcacga agtcatctga atacaaaggc
     2461 tccatcacgg acgtgttcta caaggagaac agctacacga cgacaattaa accagtctaa
     2521 aacccagctt tcttgtacta agtggtgatg ggctgcagga attcgatatc aagcttatcg
     2581 ataccgtcga cctcgagtca tgtaattagt tatgtcacgc ttacattcac gccctccccc
     2641 cacatccgct ctaaccgaaa aggaaggagt tagacaacct gaagtctagg tccctattta
     2701 tttttttata gttatgttag tattaagaac gttatttata tttcaaattt ttcttttttt
     2761 tctgtacaga cgcgtgtacg catgtaacat tatactgaaa accttgcttg agaaggtttt
     2821 gggacgctcg aaggctttaa tttgcggccg gtacccaatt cgccctatag tgagtcgtat
     2881 tacgcgcgct cactggccgt cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc
     2941 caacttaatc gccttgcagc acatccccct ttcgccagct ggcgtaatag cgaagaggcc
     3001 cgcaccgatc gcccttccca acagttgcgc agcctgaatg gcgaatggac gcgccctgta
     3061 gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca
     3121 gcgccctagc gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct
     3181 ttccccgtca agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc
     3241 acctcgaccc caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat
     3301 agacggtttt tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc
     3361 aaactggaac aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc
     3421 cgatttcggc ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaatttta
     3481 acaaaatatt aacgcttaca atttcctgat gcggtatttt ctccttacgc atctgtgcgg
     3541 tatttcacac cgcatagggt aataactgat ataattaaat tgaagctcta atttgtgagt
     3601 ttagtataca tgcatttact tataatacag ttttttagtt ttgctggccg catcttctca
     3661 aatatgcttc ccagcctgct tttctgtaac gttcaccctc taccttagca tcccttccct
     3721 ttgcaaatag tcctcttcca acaataataa tgtcagatcc tgtagagacc acatcatcca
     3781 cggttctata ctgttgaccc aatgcgtctc ccttgtcatc taaacccaca ccgggtgtca
     3841 taatcaacca atcgtaacct tcatctcttc cacccatgtc tctttgagca ataaagccga
     3901 taacaaaatc tttgtcgctc ttcgcaatgt caacagtacc cttagtatat tctccagtag
     3961 atagggagcc cttgcatgac aattctgcta acatcaaaag gcctctaggt tcctttgtta
     4021 cttcttctgc cgcctgcttc aaaccgctaa caatacctgg gcccaccaca ccgtgtgcat
     4081 tcgtaatgtc tgcccattct gctattctgt atacacccgc agagtactgc aatttgactg
     4141 tattaccaat gtcagcaaat tttctgtctt cgaagagtaa aaaattgtac ttggcggata
     4201 atgcctttag cggcttaact gtgccctcca tggaaaaatc agtcaagata tccacatgtg
     4261 tttttagtaa acaaattttg ggacctaatg cttcaactaa ctccagtaat tccttggtgg
     4321 tacgaacatc caatgaagca cacaagtttg tttgcttttc gtgcatgata ttaaatagct
     4381 tggcagcaac aggactagga tgagtagcag cacgttcctt atatgtagct ttcgacatga
     4441 tttatcttcg tttcctgcag gtttttgttc tgtgcagttg ggttaagaat actgggcaat
     4501 ttcatgtttc ttcaacacta catatgcgta tatataccaa tctaagtctg tgctccttcc
     4561 ttcgttcttc cttctgttcg gagattaccg aatcaaaaaa atttcaagga aaccgaaatc
     4621 aaaaaaaaga ataaaaaaaa aatgatgaat tgaaaaggtg gtatggtgca ctctcagtac
     4681 aatctgctct gatgccgcat agttaagcca gccccgacac ccgccaacac ccgctgacgc
     4741 gccctgacgg gcttgtctgc tcccggcatc cgcttacaga caagctgtga ccgtctccgg
     4801 gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct
     4861 cgtgatacgc ctatttttat aggttaatgt catgataata atggtttctt agacggatcg
     4921 cttgcctgta acttacacgc gcctcgtatc ttttaatgat ggaataattt gggaatttac
     4981 tctgtgttta tttattttta tgttttgtat ttggatttta gaaagtaaat aaagaaggta
     5041 gaagagttac ggaatgaaga aaaaaaaata aacaaaggtt taaaaaattt caacaaaaag
     5101 cgtactttac atatatattt attagacaag aaaagcagat taaatagata tacattcgat
     5161 taacgataag taaaatgtaa aatcacagga ttttcgtgtg tggtcttcta cacagacaag
     5221 atgaaacaat tcggcattaa tacctgagag caggaagagc aagataaaag gtagtatttg
     5281 ttggcgatcc ccctagagtc ttttacatct tcggaaaaca aaaactattt tttctttaat
     5341 ttcttttttt actttctatt tttaatttat atatttatat taaaaaattt aaattataat
     5401 tatttttata gcacgtgatg aaaaggaccc aggtggcact tttcggggaa atgtgcgcgg
     5461 aacccctatt tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata
     5521 accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg
     5581 tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac
     5641 gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact
     5701 ggatctcaac agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat
     5761 gagcactttt aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga
     5821 gcaactcggt cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac
     5881 agaaaagcat cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat
     5941 gagtgataac actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac
     6001 cgcttttttg cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct
     6061 gaatgaagcc ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac
     6121 gttgcgcaaa ctattaactg gcgaactact tactctagct tcccggcaac aattaataga
     6181 ctggatggag gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg
     6241 gtttattgct gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact
     6301 ggggccagat ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac
     6361 tatggatgaa cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta
     6421 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
     6481 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
     6541 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt c
//
