LOCUS       sequence-457112-        6653 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  2,4-Diacetylphloroglucinol (DAPG)-regulatable EGFP expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6653)
  AUTHORS   Tominaga M, Shima Y, Nozaki K, Ito Y, Someda M, Shoya Y, Hashii N, 
            Obata C, Matsumoto-Kitano M, Suematsu K, Matsukawa T, Hosoya K, 
            Hashiba N, Kondo A, Ishii J
  TITLE     Designing strong inducible synthetic promoters in yeasts.
  JOURNAL   Nat Commun. 2024 Dec 19;15(1):10653. doi: 
            10.1038/s41467-024-54865-z.
  PUBMED    39702268
REFERENCE   2  (bases 1 to 6653)
  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"; journalName: "Nat 
            Commun."; date: "2024-12-19"; volume: "15(1):10653. doi"; pages: " 
            10.1038/s41467-024-54865-z"
COMMENT     Sequence Label: pMT950
FEATURES             Location/Qualifiers
     source          1..6653
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(10..28)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        96..200
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(201..269,270..1061)
                     /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(419..438)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      1232..1820
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1721..1740
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     1974..1991
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    2108..2129
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(2144..2149,2150..2167,2168..2174)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2182..2198
                     /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     2187..2209
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     2206..2222
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     2206..2222
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             join(3483..3485,3486..3488,3489..4199)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     terminator      4277..4523
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     CDS             complement(5172..5966)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5283..5302)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5893..5912
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     complement(6259..6276)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6259..6275)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6268..6290)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(6484..6503)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6603..6625
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
ORIGIN
        1 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
       61 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
      121 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
      181 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
      241 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
      301 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
      361 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
      421 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
      481 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
      541 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
      601 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
      661 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
      721 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
      781 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
      841 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
      901 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
      961 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     1021 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     1081 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     1141 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     1201 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     1261 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     1321 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     1381 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     1441 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     1501 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     1561 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     1621 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     1681 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     1741 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     1801 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     1861 gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg gataaccgta
     1921 ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag cgcagcgagt
     1981 cagtgagcga ggaagcggaa gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc
     2041 cgattcatta atgcagctgg cacgacaggt ttcccgactg gaaagcgggc agtgagcgca
     2101 acgcaattaa tgtgagttag ctcactcatt aggcacccca ggctttacac tttatgcttc
     2161 cggctcgtat gttgtgtgga attgtgagcg gataacaatt tcacacagga aacagctatg
     2221 accatgatta cgccaagctt gcggccgcgg atctcatcaa gagagctgaa caggaaatag
     2281 acgtcttgat gcccggttat actcacttgc aaagagctca accaatcaga tggtctcact
     2341 ggttgagcat gtatgctacc tatttcactg aagattatga gagactgaat caaatcgtta
     2401 aaaggttgaa caaatcccca ttgggagctg gagctttggc tggtcatcct tatggaattg
     2461 atcgtgaata cattgctgag agattagggt ttgattctgt tattggtaat tctttggccg
     2521 ctgtttcaga cagagatttt gtagtcgaaa ccatgttctg gtcttcgttg tttatgaatc
     2581 atatttctcg attctcagaa gatttgatca tttactccac tggagagttt ggatttatca
     2641 agttggcaga tgcttattct actggatctt ctctgatgcc tcaaaaaaaa aacccagact
     2701 ctttggagtt attgaggggt aaatctggta gatgttttgg ggccttggct ggtttcctca
     2761 tgtctattaa gtccattccg tcaacctata acaaagatat gcaagaggat aaggagcctt
     2821 tatttgatac tctaatcact gtagagcact cgattttgat agcatccggt gtagtttcta
     2881 ccttgaacat tgatgccgaa cgaatgaaga atgctctaac tatggatatg ctggctacag
     2941 atcttgccga ctatttagtt agaaggggag ttccattcag agaaactcac cacatttctg
     3001 gtgaatgtgt cagacaagcc gaggagttga acctttctgg tattgatcag ttgtccctcg
     3061 aacaattgaa atccattgac tcccgttttg aggctgatgt ggcttcaacg tttgactttg
     3121 aagccagtgt tgaaaaaaga actgccaccg gaggaacttc taagactgct gttttaaagc
     3181 aattggatgc actgaatgaa aagctagagt cttgactgca ggtcgactct agaggatcct
     3241 atgtatgata cgaaacgtac cgtatcgtta aggtagcgtc aaaatttaac tgttctaacc
     3301 cctacttgac agcaatatat aaacagaagg aagctgccct gtcttaaacc ttttttttta
     3361 tcatcattat tagcttactt tcataattgc gactggttcc aattgacaag cttttgattt
     3421 taacgacttt taacgacaac ttgagaagat caaaaaacaa ctaattattc gaaacgacta
     3481 gtatggtttc taagggtgaa gagttgttca ctggtgttgt tccaatcttg gttgagttgg
     3541 acggtgacgt taacggacac aagttctctg tttctggtga aggtgagggt gacgctactt
     3601 acggaaagtt gactttgaag ttcatctgta ctactggtaa gttgccagtt ccatggccaa
     3661 ctttggttac tactttgact tacggtgttc agtgtttctc cagataccca gaccacatga
     3721 agcagcacga tttcttcaag tctgctatgc cagagggtta cgttcaagag agaactatct
     3781 tcttcaagga cgacggtaac tacaagacta gagctgaggt taagttcgag ggtgacacat
     3841 tggttaacag aatcgagttg aagggtatcg acttcaaaga ggacggaaac atcttgggtc
     3901 acaagttgga gtacaactac aactcccaca acgtttacat catggctgac aagcagaaga
     3961 acggtatcaa ggttaacttc aagatcagac acaacatcga ggacggttcc gttcaattgg
     4021 ctgaccacta ccaacagaac actccaattg gtgacggtcc agttttgttg ccagacaacc
     4081 actacttgtc cactcaatcc gctttgtcca aggacccaaa cgagaagaga gatcacatgg
     4141 ttttgttgga gttcgttact gctgctggta tcactttggg tatggacgag ttgtacaagt
     4201 aactcgagcc ttagacatga ctgttcctca gttcaagttg ggcacttacg agaagaccgg
     4261 tcttgctaga ttctaatcaa gaggatgtca gaatgccatt tgcctgagag atgcaggctt
     4321 catttttgat acttttttat ttgtaaccta tatagtatag gatttttttt gtcattttgt
     4381 ttcttctcgt acgagcttgc tcctgatcag cctatctcgc agctgatgaa tatcttgtgg
     4441 taggggtttg ggaaaatcat tcgagtttga tgtttttctt ggtatttccc actcctcttc
     4501 agagtacaga agattaagtg agacgttcgt ttgtgcaagc ttagatcttc tagagacaat
     4561 aagaagaaaa aaaaagaaaa gcggtggggg agggattatt aaataaggat tatgtaaccc
     4621 cagggtaccg ttctatacat atttaaggat tatttaggac aatcgatgaa atcggcatca
     4681 aactggatgg gagtatagtg tccggataat cggataaatc atcttgcgag gagccgcttg
     4741 gttggttggt gagaggagtg aaatatgtgt ctcctcaccc aagaatcgcg atatcagcac
     4801 cctgtggggg acactattgg cctccctccc aaaccttcga tgtggtagtg ctttattata
     4861 ttgattacat tgattacata gctaaaccct gcctggttgc aagttgagct ccgaattcca
     4921 atattagtaa aatgcctgca agataacctc ggtatggcgt ccgaccccgc ttaattattt
     4981 taactccttt ccaacgagga cttcgtaatt tttgattagg gagttgagaa acggggggtc
     5041 ttgatacctc ctcgatttca gatcccaccc cctctcagtc ccaagtggga cccccctcgg
     5101 ccgtgaaatg cgcgcacttt agtttttttc gcatgtaaac gccggtgtcc gtcatctaga
     5161 gaattccccg ctcagaagaa ctcgtcaaga aggcgataga aggcgatgcg ctgcgaatcg
     5221 ggagcggcga taccgtaaag cacgaggaag cggtcagccc attcgccgcc aagctcttca
     5281 gcaatatcac gggtagccaa cgctatgtcc tgatagcggt ccgccacacc cagccggcca
     5341 cagtcgatga atccagaaaa gcggccattt tccaccatga tattcggcaa gcaggcatcg
     5401 ccatgggtca cgacgagatc ctcgccgtcg ggcatgcgcg ccttgagcct ggcgaacagt
     5461 tcggctggcg cgagcccctg atgctcttcg tccagatcat cctgatcgac aagaccggct
     5521 tccatccgag tacgtgctcg ctcgatgcga tgtttcgctt ggtggtcgaa tgggcaggta
     5581 gccggatcaa gcgtatgcag ccgccgcatt gcatcagcca tgatggatac tttctcggca
     5641 ggagcaaggt gagatgacag gagatcctgc cccggcactt cgcccaatag cagccagtcc
     5701 cttcccgctt cagtgacaac gtcgagcaca gctgcgcaag gaacgcccgt cgtggccagc
     5761 cacgatagcc gcgctgcctc gtcctgcagt tcattcaggg caccggacag gtcggtcttg
     5821 acaaaaagaa ccgggcgccc ctgcgctgac agccggaaca cggcggcatc agagcagccg
     5881 attgtctgtt gtgcccagtc atagccgaat agcctctcca cccaagcggc cggagaacct
     5941 gcgtgcaatc catcttgttc aatcatatgg tttctatatt atctttgtac taaagagcaa
     6001 ttgataatgt gcgagaaaaa ctggtcctta tataccgttt gcagcactcc ctcccgaact
     6061 ttacgaaaag tcgtgcgcca cctgattttc atcacgccaa aaacctacac gtatgactac
     6121 tccgggccag tgttcaccac gagctatata gtgttaatta attaccttat tggttagctc
     6181 tgcatgtaag ggtggtgtga gccgggaatt gggtctactc tagcgttcag taaggtgata
     6241 taaagctctg tagaattcac tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg
     6301 cgttacccaa cttaatcgcc ttgcagcaca tccccctttc gccagctggc gtaatagcga
     6361 agaggcccgc accgatcgcc cttcccaaca gttgcgcagc ctgaatggcg aatggcgcct
     6421 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatat ggtgcactct
     6481 cagtacaatc tgctctgatg ccgcatagtt aagccagccc cgacacccgc caacacccgc
     6541 tgacgcgccc tgacgggctt gtctgctccc ggcatccgct tacagacaag ctgtgaccgt
     6601 ctccgggagc tgcatgtgtc agaggttttc accgtcatca ccgaaacgcg cga
//
