LOCUS       sequence-259344-        4045 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Part of GoldenGate Yarrowia lipolytica toolkit for assembling an 
            expression plasmid for Y. lipolytic yeast. Inducible promoter for 
            gene expression in position1 (P1_EYK300-5AB).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4045)
  AUTHORS   Larroude M, Park YK, Soudier P, Kubiak M, Nicaud JM, Rossignol T
  TITLE     A modular Golden Gate toolkit for Yarrowia lipolytica synthetic 
            biology.
  JOURNAL   Microb Biotechnol. 2019 May 31. doi: 10.1111/1751-7915.13427.
  PUBMED    31148366
REFERENCE   2  (bases 1 to 4045)
  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: "Microb 
            Biotechnol. 2019 May 31. doi: 10.1111/1751-7915.13427."
COMMENT     Sequence Label: GGE250
FEATURES             Location/Qualifiers
     source          1..4045
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(143..148,149..166,167..173)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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     186..208
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     205..221
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        239..257
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     239..256
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(933..952)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(934..952)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(959..976)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(959..975)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(968..990)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             1113..1415
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
                     SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     1294..1313
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1764..2558
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             1764..2558
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1818..1837)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2428..2447
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      2758..2767
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2764..3138
                     /codon_start=1
                     /gene="Sh ble from Streptoalloteichus hindustanus"
                     /product="antibiotic-binding protein"
                     /label=BleoR
                     /note="confers resistance to bleomycin, phleomycin, and 
                     Zeocin(TM)"
                     /translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
                     VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
                     EFALRDPAGNCVHFVAEEQD"
     rep_origin      3276..3864
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3765..3784
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4018..4035
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 agcgcccaat acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc
       61 acgacaggtt tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc
      121 tcactcatta ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa
      181 ttgtgagcgg ataacaattt cacacaggaa acagctatga ccatgattac gccaagctat
      241 ttaggtgaca ctatagaata ctcaagctat gcatcaagct tggtaccgag ctcggatcca
      301 ctagtaacgg ccgccagtgt gctggaattc gcccttggtc tctcattgga tccagtagat
      361 gtgtaagtgt gtagaagtgt cgtgtgggcc aattgtgggg aaaggagctt cttatagaag
      421 ctgatctgcc cctctttctg atccacgttg acccccacgc tgttttcagc tttgcaaaag
      481 tttgggaccg cggtacgtgg tggccttgcc tcgaaataat agtctttgtc cgcactagtc
      541 aggtcatcgg attatgcaag gctgtggagc ttaatgcggc ggctttccct tttgggattc
      601 cgcttcccgc tgtggagctt aatgcggcgg ctttcccttt tgggattccg cttcccgctg
      661 tggagcttaa tgcggcggct ttcccttttg ggattccgct tcccgctgtg gagcttaatg
      721 cggcggcttt cccttttggg attccgcttc ccgctgtgga gcttaatgcg gcggctttcc
      781 cttttgggat tccgcttccc cagattgaaa cgtacagtat agagaaaagt agatgcatcg
      841 atacgcgtat cgccgtagag accaagggcg aattctgcag atatccatca cactggcggc
      901 cgctcgagca tgcatctaga gggcccaatt cgccctatag tgagtcgtat tacaattcac
      961 tggccgtcgt tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc
     1021 ttgcagcaca tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc
     1081 cttcccaaca gttgcgcagc ctatacgtac ggcagtttaa ggtttacacc tataaaagag
     1141 agagccgtta tcgtctgttt gtggatgtac agagtgatat tattgacacg ccggggcgac
     1201 ggatggtgat ccccctggcc agtgcacgtc tgctgtcaga taaagtctcc cgtgaacttt
     1261 acccggtggt gcatatcggg gatgaaagct ggcgcatgat gaccaccgat atggccagtg
     1321 tgccggtctc cgttatcggg gaagaagtgg ctgatctcag ccaccgcgaa aatgacatca
     1381 aaaacgccat taacctgatg ttctggggaa tataaatgtc aggcatgaga ttatcaaaaa
     1441 ggatcttcac ctagatcctt ttcacgtaga aagccagtcc gcagaaacgg tgctgacccc
     1501 ggatgaatgt cagctactgg gctatctgga caagggaaaa cgcaagcgca aagagaaagc
     1561 aggtagcttg cagtgggctt acatggcgat agctagactg ggcggtttta tggacagcaa
     1621 gcgaaccgga attgccagct ggggcgccct ctggtaaggt tgggaagccc tgcaaagtaa
     1681 actggatggc tttcttgccg ccaaggatct gatggcgcag gggatcaagc tctgatcaag
     1741 agacaggatg aggatcgttt cgcatgattg aacaagatgg attgcacgca ggttctccgg
     1801 ccgcttgggt ggagaggcta ttcggctatg actgggcaca acagacaatc ggctgctctg
     1861 atgccgccgt gttccggctg tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc
     1921 tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg gctatcgtgg ctggccacga
     1981 cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga agcgggaagg gactggctgc
     2041 tattgggcga agtgccgggg caggatctcc tgtcatctca ccttgctcct gccgagaaag
     2101 tatccatcat ggctgatgca atgcggcggc tgcatacgct tgatccggct acctgcccat
     2161 tcgaccacca agcgaaacat cgcatcgagc gagcacgtac tcggatggaa gccggtcttg
     2221 tcgatcagga tgatctggac gaagagcatc aggggctcgc gccagccgaa ctgttcgcca
     2281 ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt gacccatggc gatgcctgct
     2341 tgccgaatat catggtggaa aatggccgct tttctggatt catcgactgt ggccggctgg
     2401 gtgtggcgga ccgctatcag gacatagcgt tggctacccg tgatattgct gaagagcttg
     2461 gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat cgccgctccc gattcgcagc
     2521 gcatcgcctt ctatcgcctt cttgacgagt tcttctgaat tattaacgct tacaatttcc
     2581 tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcatc aggtggcact
     2641 tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca ttcaaatatg
     2701 tatccgctca tgagacaata accctgataa atgcttcaat aatagcacgt gaggagggcc
     2761 accatggcca agttgaccag tgccgttccg gtgctcaccg cgcgcgacgt cgccggagcg
     2821 gtcgagttct ggaccgaccg gctcgggttc tcccgggact tcgtggagga cgacttcgcc
     2881 ggtgtggtcc gggacgacgt gaccctgttc atcagcgcgg tccaggacca ggtggtgccg
     2941 gacaacaccc tggcctgggt gtgggtgcgc ggcctggacg agctgtacgc cgagtggtcg
     3001 gaggtcgtgt ccacgaactt ccgggacgcc tccgggccgg ccatgaccga gatcggcgag
     3061 cagccgtggg ggcgggagtt cgccctgcgc gacccggccg gcaactgcgt gcacttcgtg
     3121 gccgaggagc aggactgaca cgtgctaaaa cttcattttt aatttaaaag gatctaggtg
     3181 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     3241 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     3301 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     3361 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     3421 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     3481 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     3541 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     3601 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     3661 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     3721 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     3781 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     3841 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     3901 ttttgctggc cttttgctca catgttcttt cctgcgttat cccctgattc tgtggataac
     3961 cgtattaccg cctttgagtg agctgatacc gctcgccgca gccgaacgac cgagcgcagc
     4021 gagtcagtga gcgaggaagc ggaag
//
