LOCUS       sequence-263162-        3663 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Part of GoldenGate Yarrowia lipolytica toolkit for assembling an 
            expression plasmid for Y. lipolytic yeast. Terminator of gene 
            expression for junction of position1-3. When using the GG platform 
            for expressing only one gene (T1-3_Lip2).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3663)
  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 3663)
  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: GGE020
FEATURES             Location/Qualifiers
     source          1..3663
                     /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(551..570)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(552..570)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(577..594)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(577..593)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(586..608)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             731..1033
                     /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     912..931
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1382..2176
                     /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             1382..2176
                     /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(1436..1455)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2046..2065
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      2376..2385
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2382..2756
                     /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      2894..3482
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3383..3402
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3636..3653
                     /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 gccctttgta caggtctctt ctagtgtctg
      361 tggtatctaa gctatttatc actctttaca acttctacct caactatcta ctttaataaa
      421 tgaatatcgt ttattctcta tgattactgt atatgcgttc ctctaagaca gagtagagac
      481 caagggcgaa ttctgcagat atccatcaca ctggcggccg ctcgagcatg catctagagg
      541 gcccaattcg ccctatagtg agtcgtatta caattcactg gccgtcgttt tacaacgtcg
      601 tgactgggaa aaccctggcg ttacccaact taatcgcctt gcagcacatc cccctttcgc
      661 cagctggcgt aatagcgaag aggcccgcac cgatcgccct tcccaacagt tgcgcagcct
      721 atacgtacgg cagtttaagg tttacaccta taaaagagag agccgttatc gtctgtttgt
      781 ggatgtacag agtgatatta ttgacacgcc ggggcgacgg atggtgatcc ccctggccag
      841 tgcacgtctg ctgtcagata aagtctcccg tgaactttac ccggtggtgc atatcgggga
      901 tgaaagctgg cgcatgatga ccaccgatat ggccagtgtg ccggtctccg ttatcgggga
      961 agaagtggct gatctcagcc accgcgaaaa tgacatcaaa aacgccatta acctgatgtt
     1021 ctggggaata taaatgtcag gcatgagatt atcaaaaagg atcttcacct agatcctttt
     1081 cacgtagaaa gccagtccgc agaaacggtg ctgaccccgg atgaatgtca gctactgggc
     1141 tatctggaca agggaaaacg caagcgcaaa gagaaagcag gtagcttgca gtgggcttac
     1201 atggcgatag ctagactggg cggttttatg gacagcaagc gaaccggaat tgccagctgg
     1261 ggcgccctct ggtaaggttg ggaagccctg caaagtaaac tggatggctt tcttgccgcc
     1321 aaggatctga tggcgcaggg gatcaagctc tgatcaagag acaggatgag gatcgtttcg
     1381 catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg agaggctatt
     1441 cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt tccggctgtc
     1501 agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc tgaatgaact
     1561 gcaagacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt gcgcagctgt
     1621 gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag tgccggggca
     1681 ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg ctgatgcaat
     1741 gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag cgaaacatcg
     1801 catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg atctggacga
     1861 agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcga gcatgcccga
     1921 cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca tggtggaaaa
     1981 tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc gctatcagga
     2041 catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg ctgaccgctt
     2101 cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct atcgccttct
     2161 tgacgagttc ttctgaatta ttaacgctta caatttcctg atgcggtatt ttctccttac
     2221 gcatctgtgc ggtatttcac accgcatcag gtggcacttt tcggggaaat gtgcgcggaa
     2281 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     2341 cctgataaat gcttcaataa tagcacgtga ggagggccac catggccaag ttgaccagtg
     2401 ccgttccggt gctcaccgcg cgcgacgtcg ccggagcggt cgagttctgg accgaccggc
     2461 tcgggttctc ccgggacttc gtggaggacg acttcgccgg tgtggtccgg gacgacgtga
     2521 ccctgttcat cagcgcggtc caggaccagg tggtgccgga caacaccctg gcctgggtgt
     2581 gggtgcgcgg cctggacgag ctgtacgccg agtggtcgga ggtcgtgtcc acgaacttcc
     2641 gggacgcctc cgggccggcc atgaccgaga tcggcgagca gccgtggggg cgggagttcg
     2701 ccctgcgcga cccggccggc aactgcgtgc acttcgtggc cgaggagcag gactgacacg
     2761 tgctaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     2821 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     2881 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     2941 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     3001 aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt
     3061 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     3121 taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat
     3181 agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct
     3241 tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca
     3301 cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag
     3361 agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc
     3421 gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga
     3481 aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca
     3541 tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag
     3601 ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg
     3661 aag
//
