LOCUS       sequence-263146-        3747 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 position 2 (T2_Xpr2).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3747)
  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 3747)
  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: GGE257
FEATURES             Location/Qualifiers
     source          1..3747
                     /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(635..654)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(636..654)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(661..678)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(661..677)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(670..692)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             815..1117
                     /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     996..1015
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1466..2260
                     /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             1466..2260
                     /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(1520..1539)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2130..2149
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      2460..2469
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2466..2840
                     /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      2978..3566
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3467..3486
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3720..3737
                     /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 gcccttgggg tctctggata tcggcccggg
      361 caaaccacac caacccttct cttctatcgc cagtcgaaat gtgacttttt aagtaatcaa
      421 ttcttgtaaa cacttgatcg tacgtagttg gatcatggcg cgacaaaata atttccaacc
      481 agtgtcactc actccactca gactgttgga gcacttgaca gatagatcac ccatcaagca
      541 gcaacttgga atgtcaagag accggaaggg cgaattctgc agatatccat cacactggcg
      601 gccgctcgag catgcatcta gagggcccaa ttcgccctat agtgagtcgt attacaattc
      661 actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg
      721 ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg
      781 cccttcccaa cagttgcgca gcctatacgt acggcagttt aaggtttaca cctataaaag
      841 agagagccgt tatcgtctgt ttgtggatgt acagagtgat attattgaca cgccggggcg
      901 acggatggtg atccccctgg ccagtgcacg tctgctgtca gataaagtct cccgtgaact
      961 ttacccggtg gtgcatatcg gggatgaaag ctggcgcatg atgaccaccg atatggccag
     1021 tgtgccggtc tccgttatcg gggaagaagt ggctgatctc agccaccgcg aaaatgacat
     1081 caaaaacgcc attaacctga tgttctgggg aatataaatg tcaggcatga gattatcaaa
     1141 aaggatcttc acctagatcc ttttcacgta gaaagccagt ccgcagaaac ggtgctgacc
     1201 ccggatgaat gtcagctact gggctatctg gacaagggaa aacgcaagcg caaagagaaa
     1261 gcaggtagct tgcagtgggc ttacatggcg atagctagac tgggcggttt tatggacagc
     1321 aagcgaaccg gaattgccag ctggggcgcc ctctggtaag gttgggaagc cctgcaaagt
     1381 aaactggatg gctttcttgc cgccaaggat ctgatggcgc aggggatcaa gctctgatca
     1441 agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc
     1501 ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc
     1561 tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga
     1621 cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt ggctggccac
     1681 gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct
     1741 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa
     1801 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc
     1861 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct
     1921 tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc
     1981 caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg
     2041 cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct
     2101 gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct
     2161 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca
     2221 gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga attattaacg cttacaattt
     2281 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tcaggtggca
     2341 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     2401 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatagcac gtgaggaggg
     2461 ccaccatggc caagttgacc agtgccgttc cggtgctcac cgcgcgcgac gtcgccggag
     2521 cggtcgagtt ctggaccgac cggctcgggt tctcccggga cttcgtggag gacgacttcg
     2581 ccggtgtggt ccgggacgac gtgaccctgt tcatcagcgc ggtccaggac caggtggtgc
     2641 cggacaacac cctggcctgg gtgtgggtgc gcggcctgga cgagctgtac gccgagtggt
     2701 cggaggtcgt gtccacgaac ttccgggacg cctccgggcc ggccatgacc gagatcggcg
     2761 agcagccgtg ggggcgggag ttcgccctgc gcgacccggc cggcaactgc gtgcacttcg
     2821 tggccgagga gcaggactga cacgtgctaa aacttcattt ttaatttaaa aggatctagg
     2881 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     2941 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     3001 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     3061 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     3121 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     3181 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     3241 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     3301 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     3361 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     3421 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     3481 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     3541 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     3601 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     3661 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     3721 gcgagtcagt gagcgaggaa gcggaag
//
