LOCUS       sequence-457092-        6461 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 6461)
  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 6461)
  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: pMT2076
FEATURES             Location/Qualifiers
     source          1..6461
                     /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(3291..3293,3294..3296,3297..4007)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     terminator      4085..4331
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     CDS             complement(4980..5774)
                     /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(5091..5110)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5701..5720
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     complement(6067..6084)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(6067..6083)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6076..6098)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(6292..6311)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6411..6433
                     /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 atcgggatta tagatgtctt tgtttgatag
     2281 agctttgtct tcgaaatttt tgggtgttat ccaatatatt agtcaagtag ccttcagaaa
     2341 tttcacgatc cttcaataac tctaaccgct cctgattcca tcccgataat agctcgatta
     2401 attgtgaatt aaaacctttc atttctttca gtatatcttc caactcttgt tccgacaaat
     2461 ggccgtcagt gtaatattgt tcggattgta agcgtctttc aatgttctgc acaatactat
     2521 cactttcctc ggtcaattgt ttggcccttt ctgggctaac ctctacgaat ggtatgtcgg
     2581 tggctaattt gtccgtgctg tcagaaaatg agtctttacc gtaggattta cttataccca
     2641 actggcgcgc ctcctttctc ttggatatag ctcatccaga tcaaagggaa tgcttaaatt
     2701 gagaaatagc tgtctcctta taatactttg tttccaattc aactgttgaa gattatttgg
     2761 ctcacttgat agttggttta acaatgccct agccctttca tcaagaaacg gtcgattttc
     2821 ctctatagcg ttgtttaggt cctctggttg cactcctagt ataccattca caatcttgac
     2881 aactttttca tcactcttac tatccgattg ttctaagtct tcaattgaga gagtactgaa
     2941 cgatctcgtt ggttcatcta gctctggcgc agcctcaaat tcctcaaaag aggcatcaga
     3001 aaacgagcca aactcttcat cttctttggc tttatttaca aggatcctat gtatgatacg
     3061 aaacgtaccg tatcgttaag gtagcgtcaa aatttaactg ttctaacccc tacttgacag
     3121 caatatataa acagaaggaa gctgccctgt cttaaacctt tttttttatc atcattatta
     3181 gcttactttc ataattgcga ctggttccaa ttgacaagct tttgatttta acgactttta
     3241 acgacaactt gagaagatca aaaaacaact aattattcga aacgactagt atggtttcta
     3301 agggtgaaga gttgttcact ggtgttgttc caatcttggt tgagttggac ggtgacgtta
     3361 acggacacaa gttctctgtt tctggtgaag gtgagggtga cgctacttac ggaaagttga
     3421 ctttgaagtt catctgtact actggtaagt tgccagttcc atggccaact ttggttacta
     3481 ctttgactta cggtgttcag tgtttctcca gatacccaga ccacatgaag cagcacgatt
     3541 tcttcaagtc tgctatgcca gagggttacg ttcaagagag aactatcttc ttcaaggacg
     3601 acggtaacta caagactaga gctgaggtta agttcgaggg tgacacattg gttaacagaa
     3661 tcgagttgaa gggtatcgac ttcaaagagg acggaaacat cttgggtcac aagttggagt
     3721 acaactacaa ctcccacaac gtttacatca tggctgacaa gcagaagaac ggtatcaagg
     3781 ttaacttcaa gatcagacac aacatcgagg acggttccgt tcaattggct gaccactacc
     3841 aacagaacac tccaattggt gacggtccag ttttgttgcc agacaaccac tacttgtcca
     3901 ctcaatccgc tttgtccaag gacccaaacg agaagagaga tcacatggtt ttgttggagt
     3961 tcgttactgc tgctggtatc actttgggta tggacgagtt gtacaagtaa ctcgagcctt
     4021 agacatgact gttcctcagt tcaagttggg cacttacgag aagaccggtc ttgctagatt
     4081 ctaatcaaga ggatgtcaga atgccatttg cctgagagat gcaggcttca tttttgatac
     4141 ttttttattt gtaacctata tagtatagga ttttttttgt cattttgttt cttctcgtac
     4201 gagcttgctc ctgatcagcc tatctcgcag ctgatgaata tcttgtggta ggggtttggg
     4261 aaaatcattc gagtttgatg tttttcttgg tatttcccac tcctcttcag agtacagaag
     4321 attaagtgag acgttcgttt gtgcaagctt agatcttcta gagacaataa gaagaaaaaa
     4381 aaagaaaagc ggtgggggag ggattattaa ataaggatta tgtaacccca gggtaccgtt
     4441 ctatacatat ttaaggatta tttaggacaa tcgatgaaat cggcatcaaa ctggatggga
     4501 gtatagtgtc cggataatcg gataaatcat cttgcgagga gccgcttggt tggttggtga
     4561 gaggagtgaa atatgtgtct cctcacccaa gaatcgcgat atcagcaccc tgtgggggac
     4621 actattggcc tccctcccaa accttcgatg tggtagtgct ttattatatt gattacattg
     4681 attacatagc taaaccctgc ctggttgcaa gttgagctcc gaattccaat attagtaaaa
     4741 tgcctgcaag ataacctcgg tatggcgtcc gaccccgctt aattatttta actcctttcc
     4801 aacgaggact tcgtaatttt tgattaggga gttgagaaac ggggggtctt gatacctcct
     4861 cgatttcaga tcccaccccc tctcagtccc aagtgggacc cccctcggcc gtgaaatgcg
     4921 cgcactttag tttttttcgc atgtaaacgc cggtgtccgt catctagaga attccccgct
     4981 cagaagaact cgtcaagaag gcgatagaag gcgatgcgct gcgaatcggg agcggcgata
     5041 ccgtaaagca cgaggaagcg gtcagcccat tcgccgccaa gctcttcagc aatatcacgg
     5101 gtagccaacg ctatgtcctg atagcggtcc gccacaccca gccggccaca gtcgatgaat
     5161 ccagaaaagc ggccattttc caccatgata ttcggcaagc aggcatcgcc atgggtcacg
     5221 acgagatcct cgccgtcggg catgcgcgcc ttgagcctgg cgaacagttc ggctggcgcg
     5281 agcccctgat gctcttcgtc cagatcatcc tgatcgacaa gaccggcttc catccgagta
     5341 cgtgctcgct cgatgcgatg tttcgcttgg tggtcgaatg ggcaggtagc cggatcaagc
     5401 gtatgcagcc gccgcattgc atcagccatg atggatactt tctcggcagg agcaaggtga
     5461 gatgacagga gatcctgccc cggcacttcg cccaatagca gccagtccct tcccgcttca
     5521 gtgacaacgt cgagcacagc tgcgcaagga acgcccgtcg tggccagcca cgatagccgc
     5581 gctgcctcgt cctgcagttc attcagggca ccggacaggt cggtcttgac aaaaagaacc
     5641 gggcgcccct gcgctgacag ccggaacacg gcggcatcag agcagccgat tgtctgttgt
     5701 gcccagtcat agccgaatag cctctccacc caagcggccg gagaacctgc gtgcaatcca
     5761 tcttgttcaa tcatatggtt tctatattat ctttgtacta aagagcaatt gataatgtgc
     5821 gagaaaaact ggtccttata taccgtttgc agcactccct cccgaacttt acgaaaagtc
     5881 gtgcgccacc tgattttcat cacgccaaaa acctacacgt atgactactc cgggccagtg
     5941 ttcaccacga gctatatagt gttaattaat taccttattg gttagctctg catgtaaggg
     6001 tggtgtgagc cgggaattgg gtctactcta gcgttcagta aggtgatata aagctctgta
     6061 gaattcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg ttacccaact
     6121 taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag aggcccgcac
     6181 cgatcgccct tcccaacagt tgcgcagcct gaatggcgaa tggcgcctga tgcggtattt
     6241 tctccttacg catctgtgcg gtatttcaca ccgcatatgg tgcactctca gtacaatctg
     6301 ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg
     6361 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
     6421 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg a
//
