LOCUS       sequence-402867-        7299 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Gateway-compatible destination vector for yeast expression with GALL
            promoter/6stop mutation/URA3 marker.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7299)
  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 7299)
  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-ccdB-6stop
FEATURES             Location/Qualifiers
     source          1..7299
                     /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..1674
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(1699..1704,1705..1722,1723..1729)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1783..2442
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1849..1868)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2784..3089
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VSRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     2968..2987
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     primer_bind     complement(3287..3303)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3288..3304)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     terminator      3306..3553
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(3322..3340)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(3571..3590)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3572..3590)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3600..3617)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3600..3616)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3609..3631)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      3757..4212
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3844..3863)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4054..4075
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(4343..5146)
                     /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(5147..5362)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(5386..5405)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5505..5527
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5565..5583)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    5624..6127
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        6164..6268
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6269..6337,6338..7129)
                     /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(6487..6506)
                     /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 caaaaaagct gaacgagaaa cgtaaaatga tataaatatc aatatattaa attagatttt
     1621 gcataaaaaa cagactacat aatactgtaa aacacaacat atccagtcac tatggcggcc
     1681 gcattaggca ccccaggctt tacactttat gcttccggct cgtataatgt gtggattttg
     1741 agttaggatc cgtcgagatt ttcaggagct aaggaagcta aaatggagaa aaaaatcact
     1801 ggatatacca ccgttgatat atcccaatgg catcgtaaag aacattttga ggcatttcag
     1861 tcagttgctc aatgtaccta taaccagacc gttcagctgg atattacggc ctttttaaag
     1921 accgtaaaga aaaataagca caagttttat ccggccttta ttcacattct tgcccgcctg
     1981 atgaatgctc atccggaatt ccgtatggca atgaaagacg gtgagctggt gatatgggat
     2041 agtgttcacc cttgttacac cgttttccat gagcaaactg aaacgttttc atcgctctgg
     2101 agtgaatacc acgacgattt ccggcagttt ctacacatat attcgcaaga tgtggcgtgt
     2161 tacggtgaaa acctggccta tttccctaaa gggtttattg agaatatgtt tttcgtctca
     2221 gccaatccct gggtgagttt caccagtttt gatttaaacg tggccaatat ggacaacttc
     2281 ttcgcccccg ttttcaccat gggcaaatat tatacgcaag gcgacaaggt gctgatgccg
     2341 ctggcgattc aggttcatca tgccgtttgt gatggcttcc atgtcggcag aatgcttaat
     2401 gaattacaac agtactgcga tgagtggcag ggcggggcgt aaacgcgtgg atccggctta
     2461 ctaaaagcca gataacagta tgcgtatttg cgcgctgatt tttgcggtat aagaatatat
     2521 actgatatgt atacccgaag tatgtcaaaa agaggtatgc tatgaagcag cgtattacag
     2581 tgacagttga cagcgacagc tatcagttgc tcaaggcata tatgatgtca atatctccgg
     2641 tctggtaagc acaaccatgc agaatgaagc ccgtcgtctg cgtgccgaac gctggaaagc
     2701 ggaaaatcag gaagggatgg ctgaggtcgc ccggtttatt gaaatgaacg gctcttttgc
     2761 tgacgagaac aggggctggt gaaatgcagt ttaaggttta cacctataaa agagagagcc
     2821 gttatcgtct gtttgtggat gtacagagtg atattattga cacgcccggg cgacggatgg
     2881 tgatccccct ggccagtgca cgtctgctgt cagataaagt ctcccgtgaa ctttacccgg
     2941 tggtgcatat cggggatgaa agctggcgca tgatgaccac cgatatggcc agtgtgccgg
     3001 tctccgttat cggggaagaa gtggctgatc tcagccaccg cgaaaatgac atcaaaaacg
     3061 ccattaacct gatgttctgg ggaatataaa tgtcaggctc ccttatacac agccagtctg
     3121 caggtcgacc atagtgactg gatatgttgt gttttacagt attatgtagt ctgtttttta
     3181 tgcaaaatct aatttaatat attgatattt atatcatttt acgtttctcg ttcagctttc
     3241 ttgtactaag tggtgatggg ctgcaggaat tcgatatcaa gcttatcgat accgtcgacc
     3301 tcgagtcatg taattagtta tgtcacgctt acattcacgc cctcccccca catccgctct
     3361 aaccgaaaag gaaggagtta gacaacctga agtctaggtc cctatttatt tttttatagt
     3421 tatgttagta ttaagaacgt tatttatatt tcaaattttt cttttttttc tgtacagacg
     3481 cgtgtacgca tgtaacatta tactgaaaac cttgcttgag aaggttttgg gacgctcgaa
     3541 ggctttaatt tgcggccggt acccaattcg ccctatagtg agtcgtatta cgcgcgctca
     3601 ctggccgtcg ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc
     3661 cttgcagcac atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc
     3721 ccttcccaac agttgcgcag cctgaatggc gaatggacgc gccctgtagc ggcgcattaa
     3781 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc
     3841 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     3901 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     3961 aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     4021 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     4081 cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg atttcggcct
     4141 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     4201 cgcttacaat ttcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     4261 catagggtaa taactgatat aattaaattg aagctctaat ttgtgagttt agtatacatg
     4321 catttactta taatacagtt ttttagtttt gctggccgca tcttctcaaa tatgcttccc
     4381 agcctgcttt tctgtaacgt tcaccctcta ccttagcatc ccttcccttt gcaaatagtc
     4441 ctcttccaac aataataatg tcagatcctg tagagaccac atcatccacg gttctatact
     4501 gttgacccaa tgcgtctccc ttgtcatcta aacccacacc gggtgtcata atcaaccaat
     4561 cgtaaccttc atctcttcca cccatgtctc tttgagcaat aaagccgata acaaaatctt
     4621 tgtcgctctt cgcaatgtca acagtaccct tagtatattc tccagtagat agggagccct
     4681 tgcatgacaa ttctgctaac atcaaaaggc ctctaggttc ctttgttact tcttctgccg
     4741 cctgcttcaa accgctaaca atacctgggc ccaccacacc gtgtgcattc gtaatgtctg
     4801 cccattctgc tattctgtat acacccgcag agtactgcaa tttgactgta ttaccaatgt
     4861 cagcaaattt tctgtcttcg aagagtaaaa aattgtactt ggcggataat gcctttagcg
     4921 gcttaactgt gccctccatg gaaaaatcag tcaagatatc cacatgtgtt tttagtaaac
     4981 aaattttggg acctaatgct tcaactaact ccagtaattc cttggtggta cgaacatcca
     5041 atgaagcaca caagtttgtt tgcttttcgt gcatgatatt aaatagcttg gcagcaacag
     5101 gactaggatg agtagcagca cgttccttat atgtagcttt cgacatgatt tatcttcgtt
     5161 tcctgcaggt ttttgttctg tgcagttggg ttaagaatac tgggcaattt catgtttctt
     5221 caacactaca tatgcgtata tataccaatc taagtctgtg ctccttcctt cgttcttcct
     5281 tctgttcgga gattaccgaa tcaaaaaaat ttcaaggaaa ccgaaatcaa aaaaaagaat
     5341 aaaaaaaaaa tgatgaattg aaaaggtggt atggtgcact ctcagtacaa tctgctctga
     5401 tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc cctgacgggc
     5461 ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga gctgcatgtg
     5521 tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg tgatacgcct
     5581 atttttatag gttaatgtca tgataataat ggtttcttag acggatcgct tgcctgtaac
     5641 ttacacgcgc ctcgtatctt ttaatgatgg aataatttgg gaatttactc tgtgtttatt
     5701 tatttttatg ttttgtattt ggattttaga aagtaaataa agaaggtaga agagttacgg
     5761 aatgaagaaa aaaaaataaa caaaggttta aaaaatttca acaaaaagcg tactttacat
     5821 atatatttat tagacaagaa aagcagatta aatagatata cattcgatta acgataagta
     5881 aaatgtaaaa tcacaggatt ttcgtgtgtg gtcttctaca cagacaagat gaaacaattc
     5941 ggcattaata cctgagagca ggaagagcaa gataaaaggt agtatttgtt ggcgatcccc
     6001 ctagagtctt ttacatcttc ggaaaacaaa aactattttt tctttaattt ctttttttac
     6061 tttctatttt taatttatat atttatatta aaaaatttaa attataatta tttttatagc
     6121 acgtgatgaa aaggacccag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     6181 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     6241 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     6301 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     6361 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     6421 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     6481 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     6541 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     6601 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     6661 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     6721 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     6781 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     6841 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     6901 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     6961 taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg ggccagatgg
     7021 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     7081 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     7141 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     7201 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     7261 ctgagcgtca gaccccgtag aaaagatcaa aggatcttc
//
