LOCUS       Exported                3997 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Part of GoldenGate Yarrowia lipolytica toolkit for assembling an 
            expression plasmid for Y. lipolytic yeast. Inducible promoter for 
            gene expression in position1 (P1_EYK300-4AB).
ACCESSION   .
VERSION     .
KEYWORDS    GGE132
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3997)
  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 3997)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..3997
                     /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(885..904)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(886..904)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(911..928)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(911..927)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(920..942)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     CDS             1065..1367
                     /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     1246..1265
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     CDS             1716..2510
                     /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(1770..1789)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2380..2399
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     regulatory      2710..2719
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             2716..3090
                     /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      3228..3816
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3717..3736
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     3970..3987
                     /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 tctcattgga tccagtagat
      361 gtgtaagtgt gtagaagtgt cgtgtgggcc aattgtgggg aaaggagctt cttatagaag
      421 ctgatctgcc cctctttctg atccacgttg acccccacgc tgttttcagc tttgcaaaag
      481 tttgggaccg cggtacgtgg tggccttgcc tcgaaataat agtctttgtc cgcactagtc
      541 aggtcatcgg attatgcaag gctgtggagc ttaatgcggc ggctttccct tttgggattc
      601 cgcttcccgc tgtggagctt aatgcggcgg ctttcccttt tgggattccg cttcccgctg
      661 tggagcttaa tgcggcggct ttcccttttg ggattccgct tcccgctgtg gagcttaatg
      721 cggcggcttt cccttttggg attccgcttc cccagattga aacgtacagt atagagaaaa
      781 gtagatgcat cgatacgcgt atcgccgtag agaccaaggg cgaattctgc agatatccat
      841 cacactggcg gccgctcgag catgcatcta gagggcccaa ttcgccctat agtgagtcgt
      901 attacaattc actggccgtc gttttacaac gtcgtgactg ggaaaaccct ggcgttaccc
      961 aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc gaagaggccc
     1021 gcaccgatcg cccttcccaa cagttgcgca gcctatacgt acggcagttt aaggtttaca
     1081 cctataaaag agagagccgt tatcgtctgt ttgtggatgt acagagtgat attattgaca
     1141 cgccggggcg acggatggtg atccccctgg ccagtgcacg tctgctgtca gataaagtct
     1201 cccgtgaact ttacccggtg gtgcatatcg gggatgaaag ctggcgcatg atgaccaccg
     1261 atatggccag tgtgccggtc tccgttatcg gggaagaagt ggctgatctc agccaccgcg
     1321 aaaatgacat caaaaacgcc attaacctga tgttctgggg aatataaatg tcaggcatga
     1381 gattatcaaa aaggatcttc acctagatcc ttttcacgta gaaagccagt ccgcagaaac
     1441 ggtgctgacc ccggatgaat gtcagctact gggctatctg gacaagggaa aacgcaagcg
     1501 caaagagaaa gcaggtagct tgcagtgggc ttacatggcg atagctagac tgggcggttt
     1561 tatggacagc aagcgaaccg gaattgccag ctggggcgcc ctctggtaag gttgggaagc
     1621 cctgcaaagt aaactggatg gctttcttgc cgccaaggat ctgatggcgc aggggatcaa
     1681 gctctgatca agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg
     1741 caggttctcc ggccgcttgg gtggagaggc tattcggcta tgactgggca caacagacaa
     1801 tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca ggggcgcccg gttctttttg
     1861 tcaagaccga cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt
     1921 ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga cgttgtcact gaagcgggaa
     1981 gggactggct gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc
     2041 ctgccgagaa agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg
     2101 ctacctgccc attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg
     2161 aagccggtct tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg
     2221 aactgttcgc caggctcaag gcgagcatgc ccgacggcga ggatctcgtc gtgacccatg
     2281 gcgatgcctg cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact
     2341 gtggccggct gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg
     2401 ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc
     2461 ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga attattaacg
     2521 cttacaattt cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca
     2581 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     2641 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatagcac
     2701 gtgaggaggg ccaccatggc caagttgacc agtgccgttc cggtgctcac cgcgcgcgac
     2761 gtcgccggag cggtcgagtt ctggaccgac cggctcgggt tctcccggga cttcgtggag
     2821 gacgacttcg ccggtgtggt ccgggacgac gtgaccctgt tcatcagcgc ggtccaggac
     2881 caggtggtgc cggacaacac cctggcctgg gtgtgggtgc gcggcctgga cgagctgtac
     2941 gccgagtggt cggaggtcgt gtccacgaac ttccgggacg cctccgggcc ggccatgacc
     3001 gagatcggcg agcagccgtg ggggcgggag ttcgccctgc gcgacccggc cggcaactgc
     3061 gtgcacttcg tggccgagga gcaggactga cacgtgctaa aacttcattt ttaatttaaa
     3121 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     3181 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     3241 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     3301 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     3361 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     3421 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     3481 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     3541 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     3601 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     3661 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     3721 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     3781 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     3841 cggttcctgg ccttttgctg gccttttgct cacatgttct ttcctgcgtt atcccctgat
     3901 tctgtggata accgtattac cgcctttgag tgagctgata ccgctcgccg cagccgaacg
     3961 accgagcgca gcgagtcagt gagcgaggaa gcggaag
//
