LOCUS       sequence-457107-        5947 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 5947)
  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 5947)
  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: pMT2096
FEATURES             Location/Qualifiers
     source          1..5947
                     /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(2777..2779,2780..2782,2783..3493)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     terminator      3571..3817
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     CDS             complement(4466..5260)
                     /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(4577..4596)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     5187..5206
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     complement(5553..5570)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5553..5569)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5562..5584)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(5778..5797)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5897..5919
                     /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 atctagaagg ggagttccat tcagagaaac
     2281 tcaccacatt tctggtgaat gtgtcagaca agccgaggag ttgaaccttt ctggtattga
     2341 tcagttgtcc ctcgaacaat tgaaatccat tgactcccgt tttgaggctg atgtggcttc
     2401 aacgtttgac tttgaagcca gtgttgaaaa aagaactgcc accggaggaa cttctaagac
     2461 tgctgtttta aagcaattgg atgcactgaa tgaaaagcta gagtcttgac tgcaggtcga
     2521 ctctagagga tcctatgtat gatacgaaac gtaccgtatc gttaaggtag cgtcaaaatt
     2581 taactgttct aacccctact tgacagcaat atataaacag aaggaagctg ccctgtctta
     2641 aacctttttt tttatcatca ttattagctt actttcataa ttgcgactgg ttccaattga
     2701 caagcttttg attttaacga cttttaacga caacttgaga agatcaaaaa acaactaatt
     2761 attcgaaacg actagtatgg tttctaaggg tgaagagttg ttcactggtg ttgttccaat
     2821 cttggttgag ttggacggtg acgttaacgg acacaagttc tctgtttctg gtgaaggtga
     2881 gggtgacgct acttacggaa agttgacttt gaagttcatc tgtactactg gtaagttgcc
     2941 agttccatgg ccaactttgg ttactacttt gacttacggt gttcagtgtt tctccagata
     3001 cccagaccac atgaagcagc acgatttctt caagtctgct atgccagagg gttacgttca
     3061 agagagaact atcttcttca aggacgacgg taactacaag actagagctg aggttaagtt
     3121 cgagggtgac acattggtta acagaatcga gttgaagggt atcgacttca aagaggacgg
     3181 aaacatcttg ggtcacaagt tggagtacaa ctacaactcc cacaacgttt acatcatggc
     3241 tgacaagcag aagaacggta tcaaggttaa cttcaagatc agacacaaca tcgaggacgg
     3301 ttccgttcaa ttggctgacc actaccaaca gaacactcca attggtgacg gtccagtttt
     3361 gttgccagac aaccactact tgtccactca atccgctttg tccaaggacc caaacgagaa
     3421 gagagatcac atggttttgt tggagttcgt tactgctgct ggtatcactt tgggtatgga
     3481 cgagttgtac aagtaactcg agccttagac atgactgttc ctcagttcaa gttgggcact
     3541 tacgagaaga ccggtcttgc tagattctaa tcaagaggat gtcagaatgc catttgcctg
     3601 agagatgcag gcttcatttt tgatactttt ttatttgtaa cctatatagt ataggatttt
     3661 ttttgtcatt ttgtttcttc tcgtacgagc ttgctcctga tcagcctatc tcgcagctga
     3721 tgaatatctt gtggtagggg tttgggaaaa tcattcgagt ttgatgtttt tcttggtatt
     3781 tcccactcct cttcagagta cagaagatta agtgagacgt tcgtttgtgc aagcttagat
     3841 cttctagaga caataagaag aaaaaaaaag aaaagcggtg ggggagggat tattaaataa
     3901 ggattatgta accccagggt accgttctat acatatttaa ggattattta ggacaatcga
     3961 tgaaatcggc atcaaactgg atgggagtat agtgtccgga taatcggata aatcatcttg
     4021 cgaggagccg cttggttggt tggtgagagg agtgaaatat gtgtctcctc acccaagaat
     4081 cgcgatatca gcaccctgtg ggggacacta ttggcctccc tcccaaacct tcgatgtggt
     4141 agtgctttat tatattgatt acattgatta catagctaaa ccctgcctgg ttgcaagttg
     4201 agctccgaat tccaatatta gtaaaatgcc tgcaagataa cctcggtatg gcgtccgacc
     4261 ccgcttaatt attttaactc ctttccaacg aggacttcgt aatttttgat tagggagttg
     4321 agaaacgggg ggtcttgata cctcctcgat ttcagatccc accccctctc agtcccaagt
     4381 gggacccccc tcggccgtga aatgcgcgca ctttagtttt tttcgcatgt aaacgccggt
     4441 gtccgtcatc tagagaattc cccgctcaga agaactcgtc aagaaggcga tagaaggcga
     4501 tgcgctgcga atcgggagcg gcgataccgt aaagcacgag gaagcggtca gcccattcgc
     4561 cgccaagctc ttcagcaata tcacgggtag ccaacgctat gtcctgatag cggtccgcca
     4621 cacccagccg gccacagtcg atgaatccag aaaagcggcc attttccacc atgatattcg
     4681 gcaagcaggc atcgccatgg gtcacgacga gatcctcgcc gtcgggcatg cgcgccttga
     4741 gcctggcgaa cagttcggct ggcgcgagcc cctgatgctc ttcgtccaga tcatcctgat
     4801 cgacaagacc ggcttccatc cgagtacgtg ctcgctcgat gcgatgtttc gcttggtggt
     4861 cgaatgggca ggtagccgga tcaagcgtat gcagccgccg cattgcatca gccatgatgg
     4921 atactttctc ggcaggagca aggtgagatg acaggagatc ctgccccggc acttcgccca
     4981 atagcagcca gtcccttccc gcttcagtga caacgtcgag cacagctgcg caaggaacgc
     5041 ccgtcgtggc cagccacgat agccgcgctg cctcgtcctg cagttcattc agggcaccgg
     5101 acaggtcggt cttgacaaaa agaaccgggc gcccctgcgc tgacagccgg aacacggcgg
     5161 catcagagca gccgattgtc tgttgtgccc agtcatagcc gaatagcctc tccacccaag
     5221 cggccggaga acctgcgtgc aatccatctt gttcaatcat atggtttcta tattatcttt
     5281 gtactaaaga gcaattgata atgtgcgaga aaaactggtc cttatatacc gtttgcagca
     5341 ctccctcccg aactttacga aaagtcgtgc gccacctgat tttcatcacg ccaaaaacct
     5401 acacgtatga ctactccggg ccagtgttca ccacgagcta tatagtgtta attaattacc
     5461 ttattggtta gctctgcatg taagggtggt gtgagccggg aattgggtct actctagcgt
     5521 tcagtaaggt gatataaagc tctgtagaat tcactggccg tcgttttaca acgtcgtgac
     5581 tgggaaaacc ctggcgttac ccaacttaat cgccttgcag cacatccccc tttcgccagc
     5641 tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc aacagttgcg cagcctgaat
     5701 ggcgaatggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat ttcacaccgc
     5761 atatggtgca ctctcagtac aatctgctct gatgccgcat agttaagcca gccccgacac
     5821 ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc tcccggcatc cgcttacaga
     5881 caagctgtga ccgtctccgg gagctgcatg tgtcagaggt tttcaccgtc atcaccgaaa
     5941 cgcgcga
//
