LOCUS       sequence-437144-        5381 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  pGAP-like vector encoding mutant of Neonothopanus nambi 
            hispidin-3-hydroxylase nnH3H_v2 under control of GAP promoter, for 
            yeast expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5381)
  AUTHORS   Shakhova ES, Karataeva TA, Markina NM, Mitiouchkina T, Palkina KA, 
            Perfilov MM, Wood MG, Hoang TT, Hall MP, Fakhranurova LI, Alekberova
            AE, Malyshevskaia AK, Gorbachev DA, Bugaeva EN, Pletneva LK, Babenko
            VV, Boldyreva DI, Gorokhovatsky AY, Balakireva AV, Gao F, Choob VV, 
            Encell LP, Wood KV, Yampolsky IV, Sarkisyan KS, Mishin AS
  TITLE     An improved pathway for autonomous bioluminescence imaging in 
            eukaryotes.
  JOURNAL   Nat Methods. 2024 Mar;21(3):406-410. doi: 
            10.1038/s41592-023-02152-y. Epub 2024 Jan 22.
  PUBMED    38253843
REFERENCE   2  (bases 1 to 5381)
  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"; doi: 
            "10.1038/s41592-023-02152-y"; journalName: "Nat Methods"; date: 
            "2024-03"; volume: "21"; issue: "3"; pages: "406-410"
COMMENT     Sequence Label: pNK5712
FEATURES             Location/Qualifiers
     source          1..5381
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1007..1483
                     /label=GAP promoter
                     /note="promoter for Pichia pastoris 
                     glyceraldehyde-3-phosphate dehydrogenase"
     CDS             2832..2861
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     CDS             2877..2894
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      2974..3219
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     promoter        3235..3648
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     promoter        3655..3702
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3759..4784
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTCYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     terminator      4883..5130
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(4899..4917)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     rep_origin      complement(join(5199..5381,1..406))
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5279..5298)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
       61 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
      121 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
      181 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
      241 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
      301 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
      361 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
      421 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
      481 gagatcagat ctctgctact ctggtcccaa gtgaaccacc ttttggaccc tattgaccgg
      541 accttaactt gccaaaccta aacgcttaat gcctcagacg ttttaatgcc tctcaacacc
      601 tccaaggttg ctttcttgag catgcctact aggaacttta acgaactgtg gggttgtaga
      661 cagtttcagg cgtgtcccga ccaatatggc ctactagact ctctgaaaaa tcacagtttt
      721 ccagtagttc cgatcaaatt accatcgaaa tggtcccata aacggacatt tgacatccgt
      781 tcctgaatta tagtcttcca ccgtggatca tggtgttcct ttttttccca aagaatatca
      841 gcatccctta actacgttag gtcagtgatg acaatggacc aaattgttgc aaggtttttc
      901 tttttctttc atcggcacat ttcagcctca catgcgacta ttatcgatca atgaaatcca
      961 tcaagattga aatcttaaaa ttgccccttt cacttgacag gatccttttt tgtagaaatg
     1021 tcttggtgtc ctcgtccaat caggtagcca tctctgaaat atctggctcc gttgcaactc
     1081 cgaacgacct gctggcaacg taaaattctc cggggtaaaa cttaaatgtg gagtaatgga
     1141 accagaaacg tctcttccct tctctctcct tccaccgccc gttaccgtcc ctaggaaatt
     1201 ttactctgct ggagagcttc ttctacggcc cccttgcagc aatgctcttc ccagcattac
     1261 gttgcgggta aaacggaggt cgtgtacccg acctagcagc ccagggatgg aaaagtcccg
     1321 gccgtcgctg gcaataatag cgggcggacg catgtcatga gattattgga aaccaccaga
     1381 atcgaatata aaaggcgaac acctttccca attttggttt ctcctgaccc aaagacttta
     1441 aatttaattt atttgtccct atttcaatca attgaacaac tatcaattcg aagaggaatt
     1501 caagcttcat atggctagca aatggcctca ttcgagaatt cattgtcagt gcttattgtt
     1561 ggtgctggtc ttggtggttt ggcagctgca atagcactta ggaggcaggg tcatgtggtc
     1621 aaaatatatg aatcttcatc attcaaagca gagttgggag caggtcttgc cgttcctcca
     1681 aataccctta ggtcattgca acaattggga tgtaatacag agaatttgaa tggagtcgat
     1741 aacttgtgct tcaccgctat gggatatgac ggttctgtcg gtatgatgaa caacatgaca
     1801 gattacagag aggcttatgg tacatcatgg atcatggtgc acagagtcga tttgcacaat
     1861 gaacttatga gggttgcctt ggatccagga ggtttgggac cccctgctac attgcacttg
     1921 aaccacaggg tcaccttttg tgatgttgac gcttgcaccg ttacattcac aaatggtaca
     1981 acccaatcag ctgatttgat cgtgggagcc gatggaataa ggtcaacaat tagaagattt
     2041 gtgcttgaag aggacgtcac aatccctgcc tcaggtatcg ttggttttag gtggcttgtc
     2101 caagccgatg ctcttgatcc ttacccagaa ttggattgga tagttaagaa accaccactt
     2161 ggagccagac ttatatctac ccctcagaac ccacagtctg gtgttggttt ggctgacagg
     2221 agaactatta taatatatgc atgtagggga ggtactatgg ttaacgttct tgctgttcac
     2281 gatgatgaga gagatcagaa cacagcagat tggtcagtgc cagcatcaaa agatgacctt
     2341 tttagggtgt tccatgacta tcacccgagg ttcaggaggc ttttggagtt ggcacaggac
     2401 ataaatcttt ggcaaatgag ggttgtgcca gtgttgaaaa aatgggttaa caagagggtg
     2461 tgccttcttg gcgacgcagc acatgcaatg ttgcctacat tgggtcaagg tttcggaatg
     2521 ggattggagg acgcagtcgc ccttggtaca cttcttccaa agggtacaac tgcatctcag
     2581 attgaaacca ggcttgctgt gtacgaacaa ttgaggaaag atagggcaga gtttgtggct
     2641 gccgagtctt acgaagaaca atacgtgcct gaaaaaagag gtctttatct taggtctaag
     2701 gaacttaggg atagagttat gggttacgac atcaaggttg agtctgaaaa ggtgttggaa
     2761 accttgttgc gttcttcaaa ctcagcaggt agttaactag cttagctaaa taagcggccg
     2821 ccagctttct agaacaaaaa ctcatctcag aagaggatct gaatagcgcc gtcgaccatc
     2881 atcatcatca tcattgagtt tgtagcctta gacatgactg ttcctcagtt caagttgggc
     2941 acttacgaga agaccggtct tgctagattc taatcaagag gatgtcagaa tgccatttgc
     3001 ctgagagatg caggcttcat ttttgatact tttttatttg taacctatat agtataggat
     3061 ttttttgtca ttttgtttct tctcgtacga gcttgctcct gatcagccta tctcgcagct
     3121 gatgaatatc ttgtggtagg ggtttgggaa aatcattcga gtttgatgtt tttcttggta
     3181 tttcccactc ctcttcagag tacagaagat taagtgagaa ccttcgtttg tgcggatccc
     3241 ccacacacca tagcttcaaa atgtttctac tcctttttta ctcttccaga ttttctcgga
     3301 ctccgcgcat cgccgtacca cttcaaaaca cccaagcaca gcatactaaa tttcccctct
     3361 ttcttcctct agggtgtcgt taattacccg tactaaaggt ttggaaaaga aaaaagagac
     3421 cgcctcgttt ctttttcttc gtcgaaaaag gcaataaaaa tttttatcac gtttcttttt
     3481 cttgaaaatt tttttttttg atttttttct ctttcgacga cctcccattg atatttaagt
     3541 taataaacgg tcttcaattt ctcaagtttc agtttcattt ttcttgttct attacaactt
     3601 tttttacttc ttgctcatta gaaagaaagc atagcaatct aatctaaggg cggtgttgac
     3661 aattaatcat cggcatagta tatcggcata gtataatacg acaaggtgag gaactaaacc
     3721 atggatgtct ctattcaacg ggaaacgtct tgctcgagat gaaaaagcct gaactcaccg
     3781 cgacgtctgt cgagaagttt ctgatcgaaa agttcgacag cgtctccgac ctgatgcagc
     3841 tctcggaggg cgaagaatct cgtgctttca gcttcgatgt aggagggcgt ggatatgtcc
     3901 tgcgggtaaa tagctgcgcc gatggtttct acaaagatcg ttatgtttat cggcactttg
     3961 catcggccgc gctcccgatt ccggaagtgc ttgacattgg ggaattcagc gagagcctga
     4021 cctattgcat ctcccgccgt gcacagggtg tcacgttgca agacctgcct gaaaccgaac
     4081 tgcccgctgt tctgcagccg gtcgcggagg ccatggatgc gatcgctgcg gccgatctta
     4141 gccagacgag cgggttcggc ccattcggac cgcaaggaat cggtcaatac actacatggc
     4201 gtgatttcat atgcgcgatt gctgatcccc atgtgtatca ctggcaaact gtgatggacg
     4261 acaccgtcag tgcgtccgtc gcgcaggctc tcgatgagct gatgctttgg gccgaggact
     4321 gccccgaagt ccggcacctc gtgcacgcgg atttcggctc caacaatgtc ctgacggaca
     4381 atggccgcat aacagcggtc attgactgga gcgaggcgat gttcggggat tcccaatacg
     4441 aggtcgccaa catcttcttc tggaggccgt ggttggcttg tatggagcag cagacgtgct
     4501 acttcgagcg gaggcatccg gagcttgcag gatcgccgcg gctccgggcg tatatgctcc
     4561 gcattggtct tgaccaactc tatcagagct tggttgacgg caatttcgat gatgcagctt
     4621 gggcgcaggg tcgatgcgac gcaatcgtcc gatccggagc cgggactgtc gggcgtacac
     4681 aaatcgcccg cagaagcgcg gccgtctgga ccgatggctg tgtagaagta ctcgccgata
     4741 gtggaaaccg acgccccagc actcgtccga gggcaaagga ataaactagt cacttcgtgg
     4801 ccgaggagca ggactgacac gtccgacgcg gcccgacggg tccgaggcct cggagatccg
     4861 tccccctttt cctttgtcga tatcatgtaa ttagttatgt cacgcttaca ttcacgccct
     4921 ccccccacat ccgctctaac cgaaaaggaa ggagttagac aacctgaagt ctaggtccct
     4981 atttattttt ttatagttat gttagtatta agaacgttat ttatatttca aatttttctt
     5041 ttttttctgt acagacgcgt gtacgcatgt aacattatac tgaaaacctt gcttgagaag
     5101 gttttgggac gctcgaaggc tttaatttgc aagctgaaca tgtgagcaaa aggccagcaa
     5161 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     5221 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     5281 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     5341 cttaccggat acctgtccgc ctttctccct tcgggaagcg t
//
