LOCUS       sequence-457111-        5668 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 5668)
  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 5668)
  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: pKN108
FEATURES             Location/Qualifiers
     source          1..5668
                     /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(2498..2500,2501..2503,2504..3214)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     terminator      3292..3538
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     CDS             complement(4187..4981)
                     /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(4298..4317)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     4908..4927
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     complement(5274..5291)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5274..5290)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5283..5305)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(5499..5518)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5618..5640
                     /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 atcctatgta tgatacgaaa cgtaccgtat
     2281 cgttaaggta gcgtcaaaat ttaactgttc taacccctac ttgacagcaa tatataaaca
     2341 gaaggaagct gccctgtctt aaaccttttt ttttatcatc attattagct tactttcata
     2401 attgcgactg gttccaattg acaagctttt gattttaacg acttttaacg acaacttgag
     2461 aagatcaaaa aacaactaat tattcgaaac gactagtatg gtttctaagg gtgaagagtt
     2521 gttcactggt gttgttccaa tcttggttga gttggacggt gacgttaacg gacacaagtt
     2581 ctctgtttct ggtgaaggtg agggtgacgc tacttacgga aagttgactt tgaagttcat
     2641 ctgtactact ggtaagttgc cagttccatg gccaactttg gttactactt tgacttacgg
     2701 tgttcagtgt ttctccagat acccagacca catgaagcag cacgatttct tcaagtctgc
     2761 tatgccagag ggttacgttc aagagagaac tatcttcttc aaggacgacg gtaactacaa
     2821 gactagagct gaggttaagt tcgagggtga cacattggtt aacagaatcg agttgaaggg
     2881 tatcgacttc aaagaggacg gaaacatctt gggtcacaag ttggagtaca actacaactc
     2941 ccacaacgtt tacatcatgg ctgacaagca gaagaacggt atcaaggtta acttcaagat
     3001 cagacacaac atcgaggacg gttccgttca attggctgac cactaccaac agaacactcc
     3061 aattggtgac ggtccagttt tgttgccaga caaccactac ttgtccactc aatccgcttt
     3121 gtccaaggac ccaaacgaga agagagatca catggttttg ttggagttcg ttactgctgc
     3181 tggtatcact ttgggtatgg acgagttgta caagtaactc gagccttaga catgactgtt
     3241 cctcagttca agttgggcac ttacgagaag accggtcttg ctagattcta atcaagagga
     3301 tgtcagaatg ccatttgcct gagagatgca ggcttcattt ttgatacttt tttatttgta
     3361 acctatatag tataggattt tttttgtcat tttgtttctt ctcgtacgag cttgctcctg
     3421 atcagcctat ctcgcagctg atgaatatct tgtggtaggg gtttgggaaa atcattcgag
     3481 tttgatgttt ttcttggtat ttcccactcc tcttcagagt acagaagatt aagtgagacg
     3541 ttcgtttgtg caagcttaga tcttctagag acaataagaa gaaaaaaaaa gaaaagcggt
     3601 gggggaggga ttattaaata aggattatgt aaccccaggg taccgttcta tacatattta
     3661 aggattattt aggacaatcg atgaaatcgg catcaaactg gatgggagta tagtgtccgg
     3721 ataatcggat aaatcatctt gcgaggagcc gcttggttgg ttggtgagag gagtgaaata
     3781 tgtgtctcct cacccaagaa tcgcgatatc agcaccctgt gggggacact attggcctcc
     3841 ctcccaaacc ttcgatgtgg tagtgcttta ttatattgat tacattgatt acatagctaa
     3901 accctgcctg gttgcaagtt gagctccgaa ttccaatatt agtaaaatgc ctgcaagata
     3961 acctcggtat ggcgtccgac cccgcttaat tattttaact cctttccaac gaggacttcg
     4021 taatttttga ttagggagtt gagaaacggg gggtcttgat acctcctcga tttcagatcc
     4081 caccccctct cagtcccaag tgggaccccc ctcggccgtg aaatgcgcgc actttagttt
     4141 ttttcgcatg taaacgccgg tgtccgtcat ctagagaatt ccccgctcag aagaactcgt
     4201 caagaaggcg atagaaggcg atgcgctgcg aatcgggagc ggcgataccg taaagcacga
     4261 ggaagcggtc agcccattcg ccgccaagct cttcagcaat atcacgggta gccaacgcta
     4321 tgtcctgata gcggtccgcc acacccagcc ggccacagtc gatgaatcca gaaaagcggc
     4381 cattttccac catgatattc ggcaagcagg catcgccatg ggtcacgacg agatcctcgc
     4441 cgtcgggcat gcgcgccttg agcctggcga acagttcggc tggcgcgagc ccctgatgct
     4501 cttcgtccag atcatcctga tcgacaagac cggcttccat ccgagtacgt gctcgctcga
     4561 tgcgatgttt cgcttggtgg tcgaatgggc aggtagccgg atcaagcgta tgcagccgcc
     4621 gcattgcatc agccatgatg gatactttct cggcaggagc aaggtgagat gacaggagat
     4681 cctgccccgg cacttcgccc aatagcagcc agtcccttcc cgcttcagtg acaacgtcga
     4741 gcacagctgc gcaaggaacg cccgtcgtgg ccagccacga tagccgcgct gcctcgtcct
     4801 gcagttcatt cagggcaccg gacaggtcgg tcttgacaaa aagaaccggg cgcccctgcg
     4861 ctgacagccg gaacacggcg gcatcagagc agccgattgt ctgttgtgcc cagtcatagc
     4921 cgaatagcct ctccacccaa gcggccggag aacctgcgtg caatccatct tgttcaatca
     4981 tatggtttct atattatctt tgtactaaag agcaattgat aatgtgcgag aaaaactggt
     5041 ccttatatac cgtttgcagc actccctccc gaactttacg aaaagtcgtg cgccacctga
     5101 ttttcatcac gccaaaaacc tacacgtatg actactccgg gccagtgttc accacgagct
     5161 atatagtgtt aattaattac cttattggtt agctctgcat gtaagggtgg tgtgagccgg
     5221 gaattgggtc tactctagcg ttcagtaagg tgatataaag ctctgtagaa ttcactggcc
     5281 gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta cccaacttaa tcgccttgca
     5341 gcacatcccc ctttcgccag ctggcgtaat agcgaagagg cccgcaccga tcgcccttcc
     5401 caacagttgc gcagcctgaa tggcgaatgg cgcctgatgc ggtattttct ccttacgcat
     5461 ctgtgcggta tttcacaccg catatggtgc actctcagta caatctgctc tgatgccgca
     5521 tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg
     5581 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg
     5641 ttttcaccgt catcaccgaa acgcgcga
//
