LOCUS       sequence-263183-        3943 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Part of GoldenGate Yarrowia lipolytica toolkit for assembling an 
            expression plasmid for Y. lipolytic yeast. Sequence for integration 
            into Y. lipolytica genome, randomly or at zeta platforms. 
            Linearization of cassette by NotI restriction enzyme after the 
            assembly.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3943)
  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 3943)
  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: GGE038
FEATURES             Location/Qualifiers
     source          1..3943
                     /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(831..850)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(832..850)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(857..874)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(857..873)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(866..888)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             1011..1313
                     /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     1192..1211
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1662..2456
                     /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             1662..2456
                     /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(1716..1735)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2326..2345
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      2656..2665
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2662..3036
                     /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      3174..3762
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3663..3682
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3916..3933
                     /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 tctgagtcat gtgtaacact
      361 cgctctggag agttagtcat ccgacagggt aactctaatc tcccaacacc ttattaactc
      421 tgcgtaactg taactcttct tgccacgtcg atcttactca attttcctgc tcatcatctg
      481 ctggattgtt gtctatcgtc tggctctaat acatttattg tttattgccc aaacaacttt
      541 cattgcacgt aagtgaattg ttttataaca gcgttcgcca attgctgcgc catcgtcgtc
      601 cggctgtcct accgttaggg gtagtgtgtc tcacactacc gaggttacta gagttgggaa
      661 agcgatactg cctcggacac accacctggg tcttacgact gcagagagaa tcggcgttac
      721 ctctctcaca aagcccttca gtgcggccgc tgcgagagac caagggcgaa ttctgcagat
      781 atccatcaca ctggcggccg ctcgagcatg catctagagg gcccaattcg ccctatagtg
      841 agtcgtatta caattcactg gccgtcgttt tacaacgtcg tgactgggaa aaccctggcg
      901 ttacccaact taatcgcctt gcagcacatc cccctttcgc cagctggcgt aatagcgaag
      961 aggcccgcac cgatcgccct tcccaacagt tgcgcagcct atacgtacgg cagtttaagg
     1021 tttacaccta taaaagagag agccgttatc gtctgtttgt ggatgtacag agtgatatta
     1081 ttgacacgcc ggggcgacgg atggtgatcc ccctggccag tgcacgtctg ctgtcagata
     1141 aagtctcccg tgaactttac ccggtggtgc atatcgggga tgaaagctgg cgcatgatga
     1201 ccaccgatat ggccagtgtg ccggtctccg ttatcgggga agaagtggct gatctcagcc
     1261 accgcgaaaa tgacatcaaa aacgccatta acctgatgtt ctggggaata taaatgtcag
     1321 gcatgagatt atcaaaaagg atcttcacct agatcctttt cacgtagaaa gccagtccgc
     1381 agaaacggtg ctgaccccgg atgaatgtca gctactgggc tatctggaca agggaaaacg
     1441 caagcgcaaa gagaaagcag gtagcttgca gtgggcttac atggcgatag ctagactggg
     1501 cggttttatg gacagcaagc gaaccggaat tgccagctgg ggcgccctct ggtaaggttg
     1561 ggaagccctg caaagtaaac tggatggctt tcttgccgcc aaggatctga tggcgcaggg
     1621 gatcaagctc tgatcaagag acaggatgag gatcgtttcg catgattgaa caagatggat
     1681 tgcacgcagg ttctccggcc gcttgggtgg agaggctatt cggctatgac tgggcacaac
     1741 agacaatcgg ctgctctgat gccgccgtgt tccggctgtc agcgcagggg cgcccggttc
     1801 tttttgtcaa gaccgacctg tccggtgccc tgaatgaact gcaagacgag gcagcgcggc
     1861 tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt gctcgacgtt gtcactgaag
     1921 cgggaaggga ctggctgcta ttgggcgaag tgccggggca ggatctcctg tcatctcacc
     1981 ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat gcggcggctg catacgcttg
     2041 atccggctac ctgcccattc gaccaccaag cgaaacatcg catcgagcga gcacgtactc
     2101 ggatggaagc cggtcttgtc gatcaggatg atctggacga agagcatcag gggctcgcgc
     2161 cagccgaact gttcgccagg ctcaaggcga gcatgcccga cggcgaggat ctcgtcgtga
     2221 cccatggcga tgcctgcttg ccgaatatca tggtggaaaa tggccgcttt tctggattca
     2281 tcgactgtgg ccggctgggt gtggcggacc gctatcagga catagcgttg gctacccgtg
     2341 atattgctga agagcttggc ggcgaatggg ctgaccgctt cctcgtgctt tacggtatcg
     2401 ccgctcccga ttcgcagcgc atcgccttct atcgccttct tgacgagttc ttctgaatta
     2461 ttaacgctta caatttcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     2521 accgcatcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc
     2581 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     2641 tagcacgtga ggagggccac catggccaag ttgaccagtg ccgttccggt gctcaccgcg
     2701 cgcgacgtcg ccggagcggt cgagttctgg accgaccggc tcgggttctc ccgggacttc
     2761 gtggaggacg acttcgccgg tgtggtccgg gacgacgtga ccctgttcat cagcgcggtc
     2821 caggaccagg tggtgccgga caacaccctg gcctgggtgt gggtgcgcgg cctggacgag
     2881 ctgtacgccg agtggtcgga ggtcgtgtcc acgaacttcc gggacgcctc cgggccggcc
     2941 atgaccgaga tcggcgagca gccgtggggg cgggagttcg ccctgcgcga cccggccggc
     3001 aactgcgtgc acttcgtggc cgaggagcag gactgacacg tgctaaaact tcatttttaa
     3061 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     3121 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     3181 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     3241 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     3301 gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca cttcaagaac
     3361 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt
     3421 ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag
     3481 cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc
     3541 gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag
     3601 gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca
     3661 gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt
     3721 cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc
     3781 tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc tgcgttatcc
     3841 cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc tcgccgcagc
     3901 cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aag
//
