LOCUS       sequence-263092-        4407 bp    DNA     circular SYN 15-APR-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_4UASxpr-TEF).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4407)
  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 4407)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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: GGE294
FEATURES             Location/Qualifiers
     source          1..4407
                     /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(1295..1314)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1296..1314)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1321..1338)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(1321..1337)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1330..1352)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             1475..1777
                     /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     1656..1675
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             2126..2920
                     /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             2126..2920
                     /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(2180..2199)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2790..2809
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      3120..3129
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3126..3500
                     /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      3638..4226
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4127..4146
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4380..4397
                     /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 gcccttcccg gtctctgtgg ggatccttcg
      361 ggtgtgagtt gacaaggaga gagagaaaag aagaggaaag gtaattcggg gacggtggtc
      421 ttttataccc ttggctaaag tcccaaccac aaagcaaaaa aattttcagt agtctatttt
      481 gcgtccggca tgggttaccc ggatggccag acaaagaaac tagtacaaag tctgaacaag
      541 cgtagattcc agactgcagt accctacgcc cttaacggca agtgtgggaa ccgggggagg
      601 tttgatatgt ggggtgaagg gggctctcgc cggggttggg tccgctactg ggtcaatttg
      661 gggtcaattg gggcaattgg ggctgttttt tgggacacaa atgcgccgcc aaccccctct
      721 ctatcgaacg cacttttgcc cgtgatcagt gcatgccgga tcgaggtggg cgggaacaac
      781 cctctcgata atgtacgtac actacacaca aagagaagca agtgggggcg ggactgcacg
      841 gcacttgtga gacacctcag catgccggat cgaggtgggc gggaacaacc ctctcgataa
      901 tgtacgtaca ctacacacaa agagaagcaa gtgggggcgg gactgcacgg cacttgtgag
      961 acacctcagc atgccggatc gaggtgggcg ggaacaaccc tctcgataat gtacgtacac
     1021 tacacacaaa gagaagcaag tgggggcggg actgcacggc acttgtgaga cacctcagca
     1081 tgccggatcg aggtgggcgg gaacaaccct ctcgataatg tacgtacact acacacaaag
     1141 agaagcaagt gggggcggga ctgcacggca cttgtgagac acctcagcat gcacgcgtat
     1201 cgatacgcgt atcgtgacag agaccaaggg cgaattctgc agatatccat cacactggcg
     1261 gccgctcgag catgcatcta gagggcccaa ttcgccctat agtgagtcgt attacaattc
     1321 actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc aacttaatcg
     1381 ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc gcaccgatcg
     1441 cccttcccaa cagttgcgca gcctatacgt acggcagttt aaggtttaca cctataaaag
     1501 agagagccgt tatcgtctgt ttgtggatgt acagagtgat attattgaca cgccggggcg
     1561 acggatggtg atccccctgg ccagtgcacg tctgctgtca gataaagtct cccgtgaact
     1621 ttacccggtg gtgcatatcg gggatgaaag ctggcgcatg atgaccaccg atatggccag
     1681 tgtgccggtc tccgttatcg gggaagaagt ggctgatctc agccaccgcg aaaatgacat
     1741 caaaaacgcc attaacctga tgttctgggg aatataaatg tcaggcatga gattatcaaa
     1801 aaggatcttc acctagatcc ttttcacgta gaaagccagt ccgcagaaac ggtgctgacc
     1861 ccggatgaat gtcagctact gggctatctg gacaagggaa aacgcaagcg caaagagaaa
     1921 gcaggtagct tgcagtgggc ttacatggcg atagctagac tgggcggttt tatggacagc
     1981 aagcgaaccg gaattgccag ctggggcgcc ctctggtaag gttgggaagc cctgcaaagt
     2041 aaactggatg gctttcttgc cgccaaggat ctgatggcgc aggggatcaa gctctgatca
     2101 agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc
     2161 ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc
     2221 tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg tcaagaccga
     2281 cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt ggctggccac
     2341 gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa gggactggct
     2401 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa
     2461 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc
     2521 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct
     2581 tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc
     2641 caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg gcgatgcctg
     2701 cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct
     2761 gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct
     2821 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca
     2881 gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga attattaacg cttacaattt
     2941 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tcaggtggca
     3001 cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata
     3061 tgtatccgct catgagacaa taaccctgat aaatgcttca ataatagcac gtgaggaggg
     3121 ccaccatggc caagttgacc agtgccgttc cggtgctcac cgcgcgcgac gtcgccggag
     3181 cggtcgagtt ctggaccgac cggctcgggt tctcccggga cttcgtggag gacgacttcg
     3241 ccggtgtggt ccgggacgac gtgaccctgt tcatcagcgc ggtccaggac caggtggtgc
     3301 cggacaacac cctggcctgg gtgtgggtgc gcggcctgga cgagctgtac gccgagtggt
     3361 cggaggtcgt gtccacgaac ttccgggacg cctccgggcc ggccatgacc gagatcggcg
     3421 agcagccgtg ggggcgggag ttcgccctgc gcgacccggc cggcaactgc gtgcacttcg
     3481 tggccgagga gcaggactga cacgtgctaa aacttcattt ttaatttaaa aggatctagg
     3541 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     3601 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     3661 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     3721 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     3781 ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     3841 catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc
     3901 ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg
     3961 ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac
     4021 agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg
     4081 taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt
     4141 atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct
     4201 cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg
     4261 ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat tctgtggata
     4321 accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg accgagcgca
     4381 gcgagtcagt gagcgaggaa gcggaag
//
