LOCUS       Exported                4049 bp ds-DNA     circular SYN 12-MAY-2021
DEFINITION  Part of GoldenGate Yarrowia lipolytica toolkit for assembling an 
            expression plasmid for Y. lipolytic yeast. Inducible promoter for 
            gene expression in position 3 (P3_EYK300-5AB).
ACCESSION   .
VERSION     .
KEYWORDS    GGE259
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4049)
  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 4049)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 12, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..4049
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     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        143..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(937..956)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(938..956)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(963..980)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(963..979)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(972..994)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             1117..1419
                     /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     1298..1317
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1768..2562
                     /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(1822..1841)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2432..2451
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      2762..2771
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             2768..3142
                     /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      3280..3868
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3769..3788
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4022..4039
                     /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 tctctgtcac gatacgcgta
      361 tcgatgcatc tacttttctc tatactgtac gtttcaatct ggggaagcgg aatcccaaaa
      421 gggaaagccg ccgcattaag ctccacagcg ggaagcggaa tcccaaaagg gaaagccgcc
      481 gcattaagct ccacagcggg aagcggaatc ccaaaaggga aagccgccgc attaagctcc
      541 acagcgggaa gcggaatccc aaaagggaaa gccgccgcat taagctccac agcgggaagc
      601 ggaatcccaa aagggaaagc cgccgcatta agctccacag ccttgcataa tccgatgacc
      661 tgactagtgc ggacaaagac tattatttcg aggcaaggcc accacgtacc gcggtcccaa
      721 acttttgcaa agctgaaaac agcgtggggg tcaacgtgga tcagaaagag gggcagatca
      781 gcttctataa gaagctcctt tccccacaat tggcccacac gacacttcta cacacttaca
      841 catctactgg atccccacag agaccggaag ggcgaattct gcagatatcc atcacactgg
      901 cggccgctcg agcatgcatc tagagggccc aattcgccct atagtgagtc gtattacaat
      961 tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat
     1021 cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat
     1081 cgcccttccc aacagttgcg cagcctatac gtacggcagt ttaaggttta cacctataaa
     1141 agagagagcc gttatcgtct gtttgtggat gtacagagtg atattattga cacgccgggg
     1201 cgacggatgg tgatccccct ggccagtgca cgtctgctgt cagataaagt ctcccgtgaa
     1261 ctttacccgg tggtgcatat cggggatgaa agctggcgca tgatgaccac cgatatggcc
     1321 agtgtgccgg tctccgttat cggggaagaa gtggctgatc tcagccaccg cgaaaatgac
     1381 atcaaaaacg ccattaacct gatgttctgg ggaatataaa tgtcaggcat gagattatca
     1441 aaaaggatct tcacctagat ccttttcacg tagaaagcca gtccgcagaa acggtgctga
     1501 ccccggatga atgtcagcta ctgggctatc tggacaaggg aaaacgcaag cgcaaagaga
     1561 aagcaggtag cttgcagtgg gcttacatgg cgatagctag actgggcggt tttatggaca
     1621 gcaagcgaac cggaattgcc agctggggcg ccctctggta aggttgggaa gccctgcaaa
     1681 gtaaactgga tggctttctt gccgccaagg atctgatggc gcaggggatc aagctctgat
     1741 caagagacag gatgaggatc gtttcgcatg attgaacaag atggattgca cgcaggttct
     1801 ccggccgctt gggtggagag gctattcggc tatgactggg cacaacagac aatcggctgc
     1861 tctgatgccg ccgtgttccg gctgtcagcg caggggcgcc cggttctttt tgtcaagacc
     1921 gacctgtccg gtgccctgaa tgaactgcaa gacgaggcag cgcggctatc gtggctggcc
     1981 acgacgggcg ttccttgcgc agctgtgctc gacgttgtca ctgaagcggg aagggactgg
     2041 ctgctattgg gcgaagtgcc ggggcaggat ctcctgtcat ctcaccttgc tcctgccgag
     2101 aaagtatcca tcatggctga tgcaatgcgg cggctgcata cgcttgatcc ggctacctgc
     2161 ccattcgacc accaagcgaa acatcgcatc gagcgagcac gtactcggat ggaagccggt
     2221 cttgtcgatc aggatgatct ggacgaagag catcaggggc tcgcgccagc cgaactgttc
     2281 gccaggctca aggcgagcat gcccgacggc gaggatctcg tcgtgaccca tggcgatgcc
     2341 tgcttgccga atatcatggt ggaaaatggc cgcttttctg gattcatcga ctgtggccgg
     2401 ctgggtgtgg cggaccgcta tcaggacata gcgttggcta cccgtgatat tgctgaagag
     2461 cttggcggcg aatgggctga ccgcttcctc gtgctttacg gtatcgccgc tcccgattcg
     2521 cagcgcatcg ccttctatcg ccttcttgac gagttcttct gaattattaa cgcttacaat
     2581 ttcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg catcaggtgg
     2641 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     2701 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatagc acgtgaggag
     2761 ggccaccatg gccaagttga ccagtgccgt tccggtgctc accgcgcgcg acgtcgccgg
     2821 agcggtcgag ttctggaccg accggctcgg gttctcccgg gacttcgtgg aggacgactt
     2881 cgccggtgtg gtccgggacg acgtgaccct gttcatcagc gcggtccagg accaggtggt
     2941 gccggacaac accctggcct gggtgtgggt gcgcggcctg gacgagctgt acgccgagtg
     3001 gtcggaggtc gtgtccacga acttccggga cgcctccggg ccggccatga ccgagatcgg
     3061 cgagcagccg tgggggcggg agttcgccct gcgcgacccg gccggcaact gcgtgcactt
     3121 cgtggccgag gagcaggact gacacgtgct aaaacttcat ttttaattta aaaggatcta
     3181 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     3241 ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt tttttctgcg
     3301 cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt gtttgccgga
     3361 tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc agataccaaa
     3421 tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg tagcaccgcc
     3481 tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg ataagtcgtg
     3541 tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt cgggctgaac
     3601 ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac tgagatacct
     3661 acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc
     3721 ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg gaaacgcctg
     3781 gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg
     3841 ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt tacggttcct
     3901 ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg attctgtgga
     3961 taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     4021 cagcgagtca gtgagcgagg aagcggaag
//
