LOCUS       sequence-352569-        6794 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses yeGFP under the control of a synthetic promoter (PGAL7-B).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6794)
  AUTHORS   Kwak S, Mahmud B, Dantas G
  TITLE     A Tunable and Expandable Transactivation System in Probiotic Yeast 
            Saccharomyces boulardii.
  JOURNAL   ACS Synth Biol. 2022 Jan 21;11(1):508-514. doi: 
            10.1021/acssynbio.1c00384. Epub 2021 Dec 23.
  PUBMED    34939781
REFERENCE   2  (bases 1 to 6794)
  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: 
            "10.1021/acssynbio.1c00384"; journalName: "ACS Synth Biol"; date: 
            "2022-01-21- 21"; volume: "11"; issue: "1"; pages: "508-514"
COMMENT     Sequence Label: pRS426-PGAL7-B_GFP_TCYC1
FEATURES             Location/Qualifiers
     source          1..6794
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(23..41)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     rep_origin      82..1424
                     /label=2u ori
                     /note="yeast 2u plasmid origin of replication"
     protein_bind    414..461
                     /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        1451..1555
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1556..1624,1625..2416)
                     /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(1774..1793)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2587..3175
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3076..3095
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3329..3346
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    3463..3484
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(3499..3504,3505..3522,3523..3529)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3537..3553
                     /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     3542..3564
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     3561..3577
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     3561..3577
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     3596..3616
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        3598..3616
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     CDS             3835..4551
                     /codon_start=1
                     /product="yeast-enhanced green fluorescent protein"
                     /label=yeGFP
                     /note="codons optimized for translation in S. cerevisiae 
                     and C. albicans"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     CDS             3835..4548
                     /codon_start=1
                     /product="green fluorescent protein"
                     /label=GFP
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITHGMDELYK"
     terminator      4558..4805
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(4574..4592)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     primer_bind     complement(4823..4842)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(4824..4842)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(4852..4869)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(4852..4868)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4861..4883)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      5009..5464
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5096..5115)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5306..5327
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(5595..6398)
                     /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(6399..6614)
                     /gene="S. cerevisiae URA3"
                     /label=URA3 promoter
     primer_bind     complement(6638..6657)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6757..6779
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg
       61 ataataatgg tttcttagta tgatccaata tcaaaggaaa tgatagcatt gaaggatgag
      121 actaatccaa ttgaggagtg gcagcatata gaacagctaa agggtagtgc tgaaggaagc
      181 atacgatacc ccgcatggaa tgggataata tcacaggagg tactagacta cctttcatcc
      241 tacataaata gacgcatata agtacgcatt taagcataaa cacgcactat gccgttcttc
      301 tcatgtatat atatatacag gcaacacgca gatataggtg cgacgtgaac agtgagctgt
      361 atgtgcgcag ctcgcgttgc attttcggaa gcgctcgttt tcggaaacgc tttgaagttc
      421 ctattccgaa gttcctattc tctagaaagt ataggaactt cagagcgctt ttgaaaacca
      481 aaagcgctct gaagacgcac tttcaaaaaa ccaaaaacgc accggactgt aacgagctac
      541 taaaatattg cgaataccgc ttccacaaac attgctcaaa agtatctctt tgctatatat
      601 ctctgtgcta tatccctata taacctaccc atccaccttt cgctccttga acttgcatct
      661 aaactcgacc tctacatttt ttatgtttat ctctagtatt actctttaga caaaaaaatt
      721 gtagtaagaa ctattcatag agtgaatcga aaacaatacg aaaatgtaaa catttcctat
      781 acgtagtata tagagacaaa atagaagaaa ccgttcataa ttttctgacc aatgaagaat
      841 catcaacgct atcactttct gttcacaaag tatgcgcaat ccacatcggt atagaatata
      901 atcggggatg cctttatctt gaaaaaatgc acccgcagct tcgctagtaa tcagtaaacg
      961 cgggaagtgg agtcaggctt tttttatgga agagaaaata gacaccaaag tagccttctt
     1021 ctaaccttaa cggacctaca gtgcaaaaag ttatcaagag actgcattat agagcgcaca
     1081 aaggagaaaa aaagtaatct aagatgcttt gttagaaaaa tagcgctctc gggatgcatt
     1141 tttgtagaac aaaaaagaag tatagattct ttgttggtaa aatagcgctc tcgcgttgca
     1201 tttctgttct gtaaaaatgc agctcagatt ctttgtttga aaaattagcg ctctcgcgtt
     1261 gcatttttgt tttacaaaaa tgaagcacag attcttcgtt ggtaaaatag cgctttcgcg
     1321 ttgcatttct gttctgtaaa aatgcagctc agattctttg tttgaaaaat tagcgctctc
     1381 gcgttgcatt tttgttctac aaaatgaagc acagatgctt cgttcaggtg gcacttttcg
     1441 gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa atatgtatcc
     1501 gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga agagtatgag
     1561 tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc ttcctgtttt
     1621 tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg gtgcacgagt
     1681 gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc gccccgaaga
     1741 acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat tatcccgtat
     1801 tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg acttggttga
     1861 gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag aattatgcag
     1921 tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa cgatcggagg
     1981 accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc gccttgatcg
     2041 ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca cgatgcctgt
     2101 agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc tagcttcccg
     2161 gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc
     2221 ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg
     2281 tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac
     2341 ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact
     2401 gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttaaa
     2461 acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc tcatgaccaa
     2521 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     2581 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     2641 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     2701 tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca
     2761 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     2821 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     2881 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     2941 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     3001 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     3061 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     3121 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     3181 cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt
     3241 tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt gagctgatac
     3301 cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg
     3361 cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca gctggcacga
     3421 caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga gttagctcac
     3481 tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt gtggaattgt
     3541 gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca agcgcgcaat
     3601 taaccctcac taaagggaac aaaagctgga gctccgcgaa ctcgagaccg aagggggact
     3661 ggctatacag tgttcacaaa atagccaagc tgaaaataat gtgtagctat gttcagttag
     3721 tttggctagc aaagatataa aagcaggtcg gaaatattta tgggcattat tatgcagagc
     3781 atcaacatga taaaaaaaaa cagttgaata ttccctcaaa aaactagtaa aacaatgtct
     3841 aaaggtgaag aattattcac tggtgttgtc ccaattttgg ttgaattaga tggtgatgtt
     3901 aatggtcaca aattttctgt ctccggtgaa ggtgaaggtg atgctactta cggtaaattg
     3961 accttaaaat ttatttgtac tactggtaaa ttgccagttc catggccaac cttagtcact
     4021 actttaactt atggtgttca atgtttttct agatacccag atcatatgaa acaacatgac
     4081 tttttcaagt ctgccatgcc agaaggttat gttcaagaaa gaactatttt tttcaaagat
     4141 gacggtaact acaagaccag agctgaagtc aagtttgaag gtgatacctt agttaataga
     4201 atcgaattaa aaggtattga ttttaaagaa gatggtaaca ttttaggtca caaattggaa
     4261 tacaactata actctcacaa tgtttacatc atggctgaca aacaaaagaa tggtatcaaa
     4321 gttaacttca aaattagaca caacattgaa gatggttctg ttcaattagc tgaccattat
     4381 caacaaaata ctccaattgg tgatggtcca gtcttgttac cagacaacca ttacttatcc
     4441 actcaatctg ccttatccaa agatccaaac gaaaagagag accacatggt cttgttagaa
     4501 tttgttactg ctgctggtat tacccatggt atggatgaat tgtacaaata actcgagtca
     4561 tgtaattagt tatgtcacgc ttacattcac gccctccccc cacatccgct ctaaccgaaa
     4621 aggaaggagt tagacaacct gaagtctagg tccctattta tttttttata gttatgttag
     4681 tattaagaac gttatttata tttcaaattt ttcttttttt tctgtacaga cgcgtgtacg
     4741 catgtaacat tatactgaaa accttgcttg agaaggtttt gggacgctcg aaggctttaa
     4801 tttgcggccg gtacccaatt cgccctatag tgagtcgtat tacgcgcgct cactggccgt
     4861 cgttttacaa cgtcgtgact gggaaaaccc tggcgttacc caacttaatc gccttgcagc
     4921 acatccccct ttcgccagct ggcgtaatag cgaagaggcc cgcaccgatc gcccttccca
     4981 acagttgcgc agcctgaatg gcgaatggac gcgccctgta gcggcgcatt aagcgcggcg
     5041 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc gcccgctcct
     5101 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     5161 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     5221 gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     5281 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     5341 cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc ctattggtta
     5401 aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgcttaca
     5461 atttcctgat gcggtatttt ctccttacgc atctgtgcgg tatttcacac cgcatagggt
     5521 aataactgat ataattaaat tgaagctcta atttgtgagt ttagtataca tgcatttact
     5581 tataatacag ttttttagtt ttgctggccg catcttctca aatatgcttc ccagcctgct
     5641 tttctgtaac gttcaccctc taccttagca tcccttccct ttgcaaatag tcctcttcca
     5701 acaataataa tgtcagatcc tgtagagacc acatcatcca cggttctata ctgttgaccc
     5761 aatgcgtctc ccttgtcatc taaacccaca ccgggtgtca taatcaacca atcgtaacct
     5821 tcatctcttc cacccatgtc tctttgagca ataaagccga taacaaaatc tttgtcgctc
     5881 ttcgcaatgt caacagtacc cttagtatat tctccagtag atagggagcc cttgcatgac
     5941 aattctgcta acatcaaaag gcctctaggt tcctttgtta cttcttctgc cgcctgcttc
     6001 aaaccgctaa caatacctgg gcccaccaca ccgtgtgcat tcgtaatgtc tgcccattct
     6061 gctattctgt atacacccgc agagtactgc aatttgactg tattaccaat gtcagcaaat
     6121 tttctgtctt cgaagagtaa aaaattgtac ttggcggata atgcctttag cggcttaact
     6181 gtgccctcca tggaaaaatc agtcaagata tccacatgtg tttttagtaa acaaattttg
     6241 ggacctaatg cttcaactaa ctccagtaat tccttggtgg tacgaacatc caatgaagca
     6301 cacaagtttg tttgcttttc gtgcatgata ttaaatagct tggcagcaac aggactagga
     6361 tgagtagcag cacgttcctt atatgtagct ttcgacatga tttatcttcg tttcctgcag
     6421 gtttttgttc tgtgcagttg ggttaagaat actgggcaat ttcatgtttc ttcaacacta
     6481 catatgcgta tatataccaa tctaagtctg tgctccttcc ttcgttcttc cttctgttcg
     6541 gagattaccg aatcaaaaaa atttcaagga aaccgaaatc aaaaaaaaga ataaaaaaaa
     6601 aatgatgaat tgaaaaggtg gtatggtgca ctctcagtac aatctgctct gatgccgcat
     6661 agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc
     6721 tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt
     6781 tttcaccgtc atca
//
