LOCUS       sequence-263088-        4516 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Part of GoldenGate Yarrowia lipolytica toolkit for assembling an 
            expression plasmid for Y. lipolytic yeast. Promoter for gene 
            expression in position 3 (P3_GAPdh).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4516)
  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 4516)
  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: GGE008
FEATURES             Location/Qualifiers
     source          1..4516
                     /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(1404..1423)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1405..1423)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1430..1447)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1430..1446)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1439..1461)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             1584..1886
                     /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     1765..1784
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             2235..3029
                     /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             2235..3029
                     /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(2289..2308)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2899..2918
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      3229..3238
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3235..3609
                     /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      3747..4335
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4236..4255
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4489..4506
                     /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 tctgtggtgt tgatgtgtgt
      361 ttaattcaag aatgaatata gagaagagaa gaagaaaaaa gattcaattg agccggcgat
      421 gcagaccctt atataaatgt tgccttggac agacggagca agcccgccca aacctacgtt
      481 cggtataata tgttaagctt tttaacacaa aggtttggct tggggtaacc tgatgtggtg
      541 caaaagaccg ggcgttggcg agccattgcg cgggcgaatg gggccgtgac tcgtctcaaa
      601 ttcgagggcg tgcctcaatt cgtgcccccg tggctttttc ccgccgtttc cgccccgttt
      661 gcaccactgc agccgcttct ttggttcgga caccttgctg cgagctaggt gccttgtgct
      721 acttaaaaag tggcctccca acaccaacat gacatgagtg cgtgggccaa gacacgttgg
      781 cggggtcgca gtcggctcaa tggcccggaa aaaacgctgc tggagctggt tcggacgcag
      841 tccgccgcgg cgtatcgata tccgcaaggt tccatggcgc cattgccctc cgtcggcgtc
      901 tatcccgcaa cctctaaata gagcgggaat ataacccaag cttctttttt tttcctttaa
      961 cacgcacacc cccaactatc atgttgctgc tgctgtttga ctctactctg tggaggggtg
     1021 ctcccaccca acccaaccta caggtggatc cggcgctgtg attggctgat aagtctccta
     1081 tccggactaa ttctgaccaa tgggacatgc gcgcaggacc caaatgccgc aattacgtaa
     1141 ccccaacgaa atgcctaccc ctctttggag cccagcggcc ccaaatcccc ccaagcagcc
     1201 cggttctacc ggcttccatc tccaagcacc cctttctcca caccccacaa aaagacccgt
     1261 gcaggacatc ctactgcgtc cggtttatgc tcgccggccg gtcccggctc tttccgacta
     1321 ccgtgacaga gaccaagggc gaattctgca gatatccatc acactggcgg ccgctcgagc
     1381 atgcatctag agggcccaat tcgccctata gtgagtcgta ttacaattca ctggccgtcg
     1441 ttttacaacg tcgtgactgg gaaaaccctg gcgttaccca acttaatcgc cttgcagcac
     1501 atcccccttt cgccagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac
     1561 agttgcgcag cctatacgta cggcagttta aggtttacac ctataaaaga gagagccgtt
     1621 atcgtctgtt tgtggatgta cagagtgata ttattgacac gccggggcga cggatggtga
     1681 tccccctggc cagtgcacgt ctgctgtcag ataaagtctc ccgtgaactt tacccggtgg
     1741 tgcatatcgg ggatgaaagc tggcgcatga tgaccaccga tatggccagt gtgccggtct
     1801 ccgttatcgg ggaagaagtg gctgatctca gccaccgcga aaatgacatc aaaaacgcca
     1861 ttaacctgat gttctgggga atataaatgt caggcatgag attatcaaaa aggatcttca
     1921 cctagatcct tttcacgtag aaagccagtc cgcagaaacg gtgctgaccc cggatgaatg
     1981 tcagctactg ggctatctgg acaagggaaa acgcaagcgc aaagagaaag caggtagctt
     2041 gcagtgggct tacatggcga tagctagact gggcggtttt atggacagca agcgaaccgg
     2101 aattgccagc tggggcgccc tctggtaagg ttgggaagcc ctgcaaagta aactggatgg
     2161 ctttcttgcc gccaaggatc tgatggcgca ggggatcaag ctctgatcaa gagacaggat
     2221 gaggatcgtt tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg
     2281 tggagaggct attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg
     2341 tgttccggct gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg
     2401 ccctgaatga actgcaagac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc
     2461 cttgcgcagc tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg
     2521 aagtgccggg gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca
     2581 tggctgatgc aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc
     2641 aagcgaaaca tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg
     2701 atgatctgga cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg
     2761 cgagcatgcc cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata
     2821 tcatggtgga aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg
     2881 accgctatca ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat
     2941 gggctgaccg cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct
     3001 tctatcgcct tcttgacgag ttcttctgaa ttattaacgc ttacaatttc ctgatgcggt
     3061 attttctcct tacgcatctg tgcggtattt cacaccgcat caggtggcac ttttcgggga
     3121 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     3181 atgagacaat aaccctgata aatgcttcaa taatagcacg tgaggagggc caccatggcc
     3241 aagttgacca gtgccgttcc ggtgctcacc gcgcgcgacg tcgccggagc ggtcgagttc
     3301 tggaccgacc ggctcgggtt ctcccgggac ttcgtggagg acgacttcgc cggtgtggtc
     3361 cgggacgacg tgaccctgtt catcagcgcg gtccaggacc aggtggtgcc ggacaacacc
     3421 ctggcctggg tgtgggtgcg cggcctggac gagctgtacg ccgagtggtc ggaggtcgtg
     3481 tccacgaact tccgggacgc ctccgggccg gccatgaccg agatcggcga gcagccgtgg
     3541 gggcgggagt tcgccctgcg cgacccggcc ggcaactgcg tgcacttcgt ggccgaggag
     3601 caggactgac acgtgctaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     3661 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     3721 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     3781 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     3841 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta
     3901 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     3961 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     4021 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     4081 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     4141 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     4201 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     4261 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     4321 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     4381 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     4441 gcctttgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg
     4501 agcgaggaag cggaag
//
