LOCUS       sequence-457096-        6033 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive rPhlTA mutant expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6033)
  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 6033)
  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: pYI2206
FEATURES             Location/Qualifiers
     source          1..6033
                     /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"
     promoter        2256..2732
                     /label=GAP promoter
                     /note="promoter for Pichia pastoris 
                     glyceraldehyde-3-phosphate dehydrogenase"
     CDS             3485..3505
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     terminator      3584..3830
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     primer_bind     complement(5639..5656)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(5639..5655)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5648..5670)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     complement(5864..5883)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5983..6005
                     /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 atcctttttt gtagaaatgt cttggtgtcc
     2281 tcgtccaatc aggtagccat ctctgaaata tctggctccg ttgcaactcc gaacgacctg
     2341 ctggcaacgt aaaattctcc ggggtaaaac ttaaatgtgg agtaatggaa ccagaaacgt
     2401 ctcttccctt ctctctcctt ccaccgcccg ttaccgtccc taggaaattt tactctgctg
     2461 gagagcttct tctacggccc ccttgcagca atgctcttcc cagcattacg ttgcgggtaa
     2521 aacggaggtc gtgtacccga cctagcagcc cagggatgga aaagtcccgg ccgtcgctgg
     2581 caataatagc gggcggacgc atgtcatgag attattggaa accaccagaa tcgaatataa
     2641 aaggcgaaca cctttcccaa ttttggtttc tcctgaccca aagactttaa atttaattta
     2701 tttgtcccta tttcaatcaa ttgaacaact atcaaaacac aactagtgcg tcgacatggc
     2761 tagaacttca cccagatcat ctattggttc tctgagatca ccacatactc ataaggctat
     2821 tttgacctcc accatcgaaa tcttgaaaga atgtggttac tccggtttgt ccattgaatc
     2881 tgttgctaga cgtgctggtg cttctaaacc tactatctac agatggtgga ctaacaaggc
     2941 tgctttgatt gctgaagttt acgaaaacga atccgaacag gttagaaagt ttccagattt
     3001 gggttccttt acggccgatt tggatttctt gttgagaaac ttgtggaagg tttggagaga
     3061 aactatttgt ggtgaagccc tcagatgcgt tattgcagaa gctcgattag atccagctac
     3121 tttgactcaa ttgaaggacc aattcatgga aagacgtaga gaaatgccaa aaaagttggt
     3181 taaaaacgcc atctctaacg gtgaattgcc aaaggataca aaccgtgagt tgttgttgga
     3241 tatgattttc ggtttttgct ggtacaggtt gttgactgaa caattgactg ttgaacaaga
     3301 catcgaagag ttcacgttct tgttaatcaa tggtgtttgt ccaggtactc aaagaggtcc
     3361 agcagacgct ttggatgatt ttgatttgga tatgttgcct gctgatgcct tggatgactt
     3421 cgacttagac atgttaccag ccgatgcatt agacgatttc gaccttgata tgttacctgg
     3481 tccaccaaaa aagaagagaa aggtctaacc tcgagcctta gacatgactg ttcctcagtt
     3541 caagttgggc acttacgaga agaccggtct tgctagattc taatcaagag gatgtcagaa
     3601 tgccatttgc ctgagagatg caggcttcat ttttgatact tttttatttg taacctatat
     3661 agtataggat tttttttgtc attttgtttc ttctcgtacg agcttgctcc tgatcagcct
     3721 atctcgcagc tgatgaatat cttgtggtag gggtttggga aaatcattcg agtttgatgt
     3781 ttttcttggt atttcccact cctcttcaga gtacagaaga ttaagtgaga agttcgtttg
     3841 tgcaagcttt ctagattgct tagttctgga agacaagcaa ccatctaccc ttttcggtaa
     3901 cgtatttttg gacaatctac atatcttgcc atttttcctg gaagcacttc aagcgtggca
     3961 aagaaattct gtctcatatt ggtagatggt ttttggttaa ttatcgtagc ttagtgtatg
     4021 accaaaatat atcaaattga atgtatgcga ctattacaat cttagtactt tcactttgat
     4081 cactctacta ctatgatctg tcatagttga agcattctaa ctgtacgaca ggatgtcggt
     4141 tcgaaatgtt gtaaacgtac caaactttga tcatttatat ttttttctga atgtcgtaag
     4201 cagtttgaaa ttttagtgtc ttcgattata ccaccatatt cgaactattt tgatggcgcg
     4261 ccatcttcaa ttgcagagaa ttctgcactc aactatactg tgcaatcggc cctgttagat
     4321 tgagtatgta atatatcaga cagattacca ggcgagcgat taatggaagt gttttcagac
     4381 tgcagaattc ccatgctaac agagcagtga gagtatgaag atggatttta atatcttcat
     4441 gaaggcacaa atatgttcct ctctttcgtt ttgcgttacc gtaaatgtca aaaattctag
     4501 agattcaaaa gaattggcta taagaaagct tgaatgtcgt tcaactttgt gaaatcgtcg
     4561 tttcagtcaa aaggtagagg gagcgagcat aatcgaatag gaactaatat tcacgaataa
     4621 gtgtactgac aataaacctt ggagatcatc tagcgactta cttcgctaga gaattcgccg
     4681 taccacttca aaacacccaa gcacagcata ctaaatttcc cctctttctt cctctagggt
     4741 gtcgttaatt acccgtacta aaggtttgga aaagaaaaaa gagaccgcct cgtttctttt
     4801 tcttcgtcga aaaaggcaat aaaaattttt atcacgtttc tttttcttga aaattttttt
     4861 ttttgatttt tttctctttc gatgacctcc cattgatatt taagttaata aacggtcttc
     4921 aatttctcaa gtttcagttt catttttctt gttctattac aacttttttt acttcttgct
     4981 cattagaaag aaagcatagc aatctaatct aagttttaat tacaaaatga agactttcaa
     5041 catctcccag caggacttgg agttggttga agttgctact gagaagatca ctatgttgta
     5101 cgaggacaac aagcaccacg ttggagctgc tatcagaact aagactggtg agatcatctc
     5161 cgctgttcac atcgaggctt acatcggtag agttactgtt tgtgctgagg ctatcgctat
     5221 cggttctgct gtttctaacg gtcagaagga cttcgacact atcgttgctg ttagacaccc
     5281 atactctgac gaggttgaca gatccatcag agttgtttcc ccatgtggta tgtgtagaga
     5341 gttgatctct gactacgctc cagactgttt cgttttgatc gagatgaacg gaaagttggt
     5401 taagactact atcgaggaat tgatcccatt gaagtacact agaaactaag tattaagaac
     5461 gttatttata tttcaaattt ttcttttttt tctctacaga cgctgtacgc atgtaacatt
     5521 atactgaaaa ccttgcttga gaaggttttg ggacgctcga agatccagct gcattaatga
     5581 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttgaattcac
     5641 tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc
     5701 ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc
     5761 cttcccaaca gttgcgcagc ctgaatggcg aatggcgcct gatgcggtat tttctcctta
     5821 cgcatctgtg cggtatttca caccgcatat ggtgcactct cagtacaatc tgctctgatg
     5881 ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt
     5941 gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc
     6001 agaggttttc accgtcatca ccgaaacgcg cga
//
