LOCUS       sequence-263144-        3661 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_Lip2).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3661)
  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 3661)
  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: GGE015
FEATURES             Location/Qualifiers
     source          1..3661
                     /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(549..568)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(550..568)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(575..592)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(575..591)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(584..606)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             729..1031
                     /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     910..929
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1380..2174
                     /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             1380..2174
                     /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(1434..1453)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2044..2063
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      2374..2383
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             2380..2754
                     /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      2892..3480
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3381..3400
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3634..3651
                     /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 tctggattgt ctgtggtatc
      361 taagctattt atcactcttt acaacttcta cctcaactat ctactttaat aaatgaatat
      421 cgtttattct ctatgattac tgtatatgcg ttcctctaag acaaatcggt caagagacca
      481 agggcgaatt ctgcagatat ccatcacact ggcggccgct cgagcatgca tctagagggc
      541 ccaattcgcc ctatagtgag tcgtattaca attcactggc cgtcgtttta caacgtcgtg
      601 actgggaaaa ccctggcgtt acccaactta atcgccttgc agcacatccc cctttcgcca
      661 gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctat
      721 acgtacggca gtttaaggtt tacacctata aaagagagag ccgttatcgt ctgtttgtgg
      781 atgtacagag tgatattatt gacacgccgg ggcgacggat ggtgatcccc ctggccagtg
      841 cacgtctgct gtcagataaa gtctcccgtg aactttaccc ggtggtgcat atcggggatg
      901 aaagctggcg catgatgacc accgatatgg ccagtgtgcc ggtctccgtt atcggggaag
      961 aagtggctga tctcagccac cgcgaaaatg acatcaaaaa cgccattaac ctgatgttct
     1021 ggggaatata aatgtcaggc atgagattat caaaaaggat cttcacctag atccttttca
     1081 cgtagaaagc cagtccgcag aaacggtgct gaccccggat gaatgtcagc tactgggcta
     1141 tctggacaag ggaaaacgca agcgcaaaga gaaagcaggt agcttgcagt gggcttacat
     1201 ggcgatagct agactgggcg gttttatgga cagcaagcga accggaattg ccagctgggg
     1261 cgccctctgg taaggttggg aagccctgca aagtaaactg gatggctttc ttgccgccaa
     1321 ggatctgatg gcgcagggga tcaagctctg atcaagagac aggatgagga tcgtttcgca
     1381 tgattgaaca agatggattg cacgcaggtt ctccggccgc ttgggtggag aggctattcg
     1441 gctatgactg ggcacaacag acaatcggct gctctgatgc cgccgtgttc cggctgtcag
     1501 cgcaggggcg cccggttctt tttgtcaaga ccgacctgtc cggtgccctg aatgaactgc
     1561 aagacgaggc agcgcggcta tcgtggctgg ccacgacggg cgttccttgc gcagctgtgc
     1621 tcgacgttgt cactgaagcg ggaagggact ggctgctatt gggcgaagtg ccggggcagg
     1681 atctcctgtc atctcacctt gctcctgccg agaaagtatc catcatggct gatgcaatgc
     1741 ggcggctgca tacgcttgat ccggctacct gcccattcga ccaccaagcg aaacatcgca
     1801 tcgagcgagc acgtactcgg atggaagccg gtcttgtcga tcaggatgat ctggacgaag
     1861 agcatcaggg gctcgcgcca gccgaactgt tcgccaggct caaggcgagc atgcccgacg
     1921 gcgaggatct cgtcgtgacc catggcgatg cctgcttgcc gaatatcatg gtggaaaatg
     1981 gccgcttttc tggattcatc gactgtggcc ggctgggtgt ggcggaccgc tatcaggaca
     2041 tagcgttggc tacccgtgat attgctgaag agcttggcgg cgaatgggct gaccgcttcc
     2101 tcgtgcttta cggtatcgcc gctcccgatt cgcagcgcat cgccttctat cgccttcttg
     2161 acgagttctt ctgaattatt aacgcttaca atttcctgat gcggtatttt ctccttacgc
     2221 atctgtgcgg tatttcacac cgcatcaggt ggcacttttc ggggaaatgt gcgcggaacc
     2281 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     2341 tgataaatgc ttcaataata gcacgtgagg agggccacca tggccaagtt gaccagtgcc
     2401 gttccggtgc tcaccgcgcg cgacgtcgcc ggagcggtcg agttctggac cgaccggctc
     2461 gggttctccc gggacttcgt ggaggacgac ttcgccggtg tggtccggga cgacgtgacc
     2521 ctgttcatca gcgcggtcca ggaccaggtg gtgccggaca acaccctggc ctgggtgtgg
     2581 gtgcgcggcc tggacgagct gtacgccgag tggtcggagg tcgtgtccac gaacttccgg
     2641 gacgcctccg ggccggccat gaccgagatc ggcgagcagc cgtgggggcg ggagttcgcc
     2701 ctgcgcgacc cggccggcaa ctgcgtgcac ttcgtggccg aggagcagga ctgacacgtg
     2761 ctaaaacttc atttttaatt taaaaggatc taggtgaaga tcctttttga taatctcatg
     2821 accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt agaaaagatc
     2881 aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa
     2941 ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct ttttccgaag
     3001 gtaactggct tcagcagagc gcagatacca aatactgttc ttctagtgta gccgtagtta
     3061 ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct aatcctgtta
     3121 ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg ggttggactc aagacgatag
     3181 ttaccggata aggcgcagcg gtcgggctga acggggggtt cgtgcacaca gcccagcttg
     3241 gagcgaacga cctacaccga actgagatac ctacagcgtg agctatgaga aagcgccacg
     3301 cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg gcagggtcgg aacaggagag
     3361 cgcacgaggg agcttccagg gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc
     3421 cacctctgac ttgagcgtcg atttttgtga tgctcgtcag gggggcggag cctatggaaa
     3481 aacgccagca acgcggcctt tttacggttc ctggcctttt gctggccttt tgctcacatg
     3541 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
     3601 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
     3661 g
//
