LOCUS       sequence-458297-        5928 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 5928)
  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 5928)
  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: pMT2095
FEATURES             Location/Qualifiers
     source          1..5928
                     /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(2758..2760,2761..2763,2764..3474)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     terminator      3552..3798
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     CDS             complement(4447..5241)
                     /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(4558..4577)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5168..5187
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     complement(5534..5551)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5534..5550)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5543..5565)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(5759..5778)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5878..5900
                     /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 atccatcgat ttgtatgtga aatagctgaa
     2281 attcgaaaat ttcattatgg ctgtatctac tttagcgtat taggcatttg agcattggct
     2341 tgaacaatgc gggctgtagt gtgtcaccaa agaaaccatt cgggttcgga tctggaagtc
     2401 ctcatcacgt gatgccgatc tcgtgtattt tattttcaga taacacctga agacttttgg
     2461 gtcggaggac tggctctttc cgatcaaatt ggaatggaaa attgctccgg atcctatgta
     2521 tgatacgaaa cgtaccgtat cgttaaggta gcgtcaaaat ttaactgttc taacccctac
     2581 ttgacagcaa tatataaaca gaaggaagct gccctgtctt aaaccttttt ttttatcatc
     2641 attattagct tactttcata attgcgactg gttccaattg acaagctttt gattttaacg
     2701 acttttaacg acaacttgag aagatcaaaa aacaactaat tattcgaaac gactagtatg
     2761 gtttctaagg gtgaagagtt gttcactggt gttgttccaa tcttggttga gttggacggt
     2821 gacgttaacg gacacaagtt ctctgtttct ggtgaaggtg agggtgacgc tacttacgga
     2881 aagttgactt tgaagttcat ctgtactact ggtaagttgc cagttccatg gccaactttg
     2941 gttactactt tgacttacgg tgttcagtgt ttctccagat acccagacca catgaagcag
     3001 cacgatttct tcaagtctgc tatgccagag ggttacgttc aagagagaac tatcttcttc
     3061 aaggacgacg gtaactacaa gactagagct gaggttaagt tcgagggtga cacattggtt
     3121 aacagaatcg agttgaaggg tatcgacttc aaagaggacg gaaacatctt gggtcacaag
     3181 ttggagtaca actacaactc ccacaacgtt tacatcatgg ctgacaagca gaagaacggt
     3241 atcaaggtta acttcaagat cagacacaac atcgaggacg gttccgttca attggctgac
     3301 cactaccaac agaacactcc aattggtgac ggtccagttt tgttgccaga caaccactac
     3361 ttgtccactc aatccgcttt gtccaaggac ccaaacgaga agagagatca catggttttg
     3421 ttggagttcg ttactgctgc tggtatcact ttgggtatgg acgagttgta caagtaactc
     3481 gagccttaga catgactgtt cctcagttca agttgggcac ttacgagaag accggtcttg
     3541 ctagattcta atcaagagga tgtcagaatg ccatttgcct gagagatgca ggcttcattt
     3601 ttgatacttt tttatttgta acctatatag tataggattt tttttgtcat tttgtttctt
     3661 ctcgtacgag cttgctcctg atcagcctat ctcgcagctg atgaatatct tgtggtaggg
     3721 gtttgggaaa atcattcgag tttgatgttt ttcttggtat ttcccactcc tcttcagagt
     3781 acagaagatt aagtgagacg ttcgtttgtg caagcttaga tcttctagag acaataagaa
     3841 gaaaaaaaaa gaaaagcggt gggggaggga ttattaaata aggattatgt aaccccaggg
     3901 taccgttcta tacatattta aggattattt aggacaatcg atgaaatcgg catcaaactg
     3961 gatgggagta tagtgtccgg ataatcggat aaatcatctt gcgaggagcc gcttggttgg
     4021 ttggtgagag gagtgaaata tgtgtctcct cacccaagaa tcgcgatatc agcaccctgt
     4081 gggggacact attggcctcc ctcccaaacc ttcgatgtgg tagtgcttta ttatattgat
     4141 tacattgatt acatagctaa accctgcctg gttgcaagtt gagctccgaa ttccaatatt
     4201 agtaaaatgc ctgcaagata acctcggtat ggcgtccgac cccgcttaat tattttaact
     4261 cctttccaac gaggacttcg taatttttga ttagggagtt gagaaacggg gggtcttgat
     4321 acctcctcga tttcagatcc caccccctct cagtcccaag tgggaccccc ctcggccgtg
     4381 aaatgcgcgc actttagttt ttttcgcatg taaacgccgg tgtccgtcat ctagagaatt
     4441 ccccgctcag aagaactcgt caagaaggcg atagaaggcg atgcgctgcg aatcgggagc
     4501 ggcgataccg taaagcacga ggaagcggtc agcccattcg ccgccaagct cttcagcaat
     4561 atcacgggta gccaacgcta tgtcctgata gcggtccgcc acacccagcc ggccacagtc
     4621 gatgaatcca gaaaagcggc cattttccac catgatattc ggcaagcagg catcgccatg
     4681 ggtcacgacg agatcctcgc cgtcgggcat gcgcgccttg agcctggcga acagttcggc
     4741 tggcgcgagc ccctgatgct cttcgtccag atcatcctga tcgacaagac cggcttccat
     4801 ccgagtacgt gctcgctcga tgcgatgttt cgcttggtgg tcgaatgggc aggtagccgg
     4861 atcaagcgta tgcagccgcc gcattgcatc agccatgatg gatactttct cggcaggagc
     4921 aaggtgagat gacaggagat cctgccccgg cacttcgccc aatagcagcc agtcccttcc
     4981 cgcttcagtg acaacgtcga gcacagctgc gcaaggaacg cccgtcgtgg ccagccacga
     5041 tagccgcgct gcctcgtcct gcagttcatt cagggcaccg gacaggtcgg tcttgacaaa
     5101 aagaaccggg cgcccctgcg ctgacagccg gaacacggcg gcatcagagc agccgattgt
     5161 ctgttgtgcc cagtcatagc cgaatagcct ctccacccaa gcggccggag aacctgcgtg
     5221 caatccatct tgttcaatca tatggtttct atattatctt tgtactaaag agcaattgat
     5281 aatgtgcgag aaaaactggt ccttatatac cgtttgcagc actccctccc gaactttacg
     5341 aaaagtcgtg cgccacctga ttttcatcac gccaaaaacc tacacgtatg actactccgg
     5401 gccagtgttc accacgagct atatagtgtt aattaattac cttattggtt agctctgcat
     5461 gtaagggtgg tgtgagccgg gaattgggtc tactctagcg ttcagtaagg tgatataaag
     5521 ctctgtagaa ttcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta
     5581 cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg
     5641 cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg cgcctgatgc
     5701 ggtattttct ccttacgcat ctgtgcggta tttcacaccg catatggtgc actctcagta
     5761 caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg
     5821 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
     5881 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcga
//
