LOCUS       sequence-248175-        6742 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  LacI-Mit1AD on pPICZ B expression vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6742)
  AUTHORS   Liu Y, Bai C, Liu Q, Xu Q, Qian Z, Peng Q, Yu J, Xu M, Zhou X, Zhang
            Y, Cai M
  TITLE     Engineered ethanol-driven biosynthetic system for improving 
            production of acetyl-CoA derived drugs in Crabtree-negative yeast.
  JOURNAL   Metab Eng. 2019 Jul;54:275-284. doi: 10.1016/j.ymben.2019.05.001. 
            Epub 2019 May 9.
  PUBMED    31077813
REFERENCE   2  (bases 1 to 6742)
  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: "Metab 
            Eng."; date: "2019-07"; volume: "54:275-284. doi"; pages: " 
            10.1016/j.ymben.2019.05.001. Epub 2019 May 9"
COMMENT     Sequence Label: pZ_P-ICL1-LacI-Mit1AD
FEATURES             Location/Qualifiers
     source          1..6742
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             4130..4147
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      4227..4473
                     /gene="Pichia pastoris AOX1"
                     /label=AOX1 terminator
                     /note="transcription terminator for AOX1"
     promoter        4488..4901
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     promoter        4908..4955
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             4974..5348
                     /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"
     terminator      5414..5661
                     /gene="S. cerevisiae CYC1"
                     /label=CYC1 terminator
                     /note="transcription terminator for CYC1"
     primer_bind     complement(5430..5448)
                     /label=CYC1
                     /note="CYC1 transcription termination signal, reverse 
                     primer"
     rep_origin      complement(5736..6324)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5816..5835)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 caaatgtgct gttccggtag cttgtaggaa gcggcatccg tagggcaata tacgactata
       61 gcttctaaag cgtagtacaa tgaaatgttc gaaggaacaa caaacggatt tgtttttcgt
      121 aggctcaacc cgttgaggtg taactcttta gcgaaagggt aagattgatt gttcgaagta
      181 gggcctcaaa gggaaagaga aaaaaaaaaa tacaccaaga gttacgtaag catatatttt
      241 ttacgtaaag catgattgaa tttcagcagt attgtttaac aaggctgatg tcgtgtgcca
      301 atcaaaacaa aagagattcg cataatgcca taattggggt gtgtgggcgc cccctaaaac
      361 gtctttctca tcatcatctg caacccccat cgaacctcat taaatcacat gacttgtgcg
      421 atcctcggtc aactcgttcc gtgcacccat tccaccccgg gctgaccaac gcaaggttct
      481 ccgagagtcc gctaccccag atttatatca gcaaccagtc acctttttcc gggcacgact
      541 ctatatgccc tggaaaaccg gagacgatga gcctgactgt aaaaggtgac agaaccccca
      601 actctggtta atctcttcaa caaatacttt attttctttc aattcaaaga acacagtatc
      661 aagtatatca agaatgggtg ttaagccagt tactttgtat gacgttgctg aatacgctgg
      721 agtttcctac caaactgtct ctagagttgt taatcaagct tctcatgtct ccgctaagac
      781 tagagagaag gttgaggctg ctatggctga attgaactat attccaaata gagttgctca
      841 gcagttggct ggaaagcaat ctttgttgat tggagtcgct acttcttctt tggctttgca
      901 tgctccatct cagattgttg ctgctattaa gtccagagct gaccagttgg gagcttctgt
      961 tgttgtttct atggttgaga gatctggagt tgaggcttgc aaggctgctg ttcataactt
     1021 gttggctcag agagtttctg gattgattat taattaccca ttggacgatc aagacgctat
     1081 tgccgttgag gccgcttgta ccaacgtccc agctttgttc ttggacgttt ccgatcaaac
     1141 tccaattaat tctattattt tttctcacga ggatggaact agattgggag ttgaacactt
     1201 ggttgctttg ggacatcaac agattgcttt gttggctgga ccattgtctt ccgtttctgc
     1261 tagattgaga ttggccggat ggcacaagta cttgaccaga aaccagattc aaccaattgc
     1321 tgagagagag ggagattggt ctgctatgtc tggattccag cagactatgc agatgttgaa
     1381 cgaaggaatt gtcccaaccg ctatgttggt cgctaatgac caaatggctt tgggagctat
     1441 gagagctatt actgaatctg gattgagagt cggagctgac atttctgttg ttggatatga
     1501 tgacactgag gattcttctt gctacattcc accattgact actattaagc aagacttcag
     1561 attgttggga cagacttctg ttgatagatt gttgcagttg tcccaaggac aagctgttaa
     1621 aggaaaccaa ttgttgccag tttctttggt taagagaaag actactttgg ctccaaacac
     1681 tcagactgct tccccaagag ctttggctga ctctttgatg caattggcta gacaagtctc
     1741 tagattggag tctggacaag gtggcggcgg ctctgttaac aactccatga aggatttctt
     1801 aggcaagaaa acggtggatg gagctgatag tctcaatttg gccgtgaatc tgcaacaaca
     1861 gcagagttca aacacaattg ccaatcaatc gctttcctca attggattgg aaagttttgg
     1921 ttacggctct ggtatcaaaa acgagtttaa cttccaagac ttgataggtt caaactctgg
     1981 cagttcagat ccgacatttt cagtagacgc tgacgaggcc caaaaactcg acatttccaa
     2041 caagaacagt cgtaagagac agaaactagg tttgctgccg gtcagcaatg caacttccca
     2101 tttgaacggt ttcaatggaa tgtccaatgg aaagtcacac tctttctctt caccgtctgg
     2161 gactaatgac gatgaactaa gtggcttgat gttcaactca ccaagcttca accccctcac
     2221 agttaacgat tctaccaaca acagcaacca caatataggt ttgtctccga tgtcatgctt
     2281 attttctaca gttcaagaag catctcaaaa aaagcatgga aattccagta gacacttttc
     2341 atacccatct gggccggagg acctttggtt caatgagttc caaaaacagg ccctcacagc
     2401 caatggagaa aatgctgtcc aacagggaga tgatgcttct aagaacaaca cagccattcc
     2461 taaggaccag tcttcgaact catcgatttt cagttcacgt tctagtgcag cttctagcaa
     2521 ctcaggagac gatattggaa ggatgggccc attctccaaa ggaccagaga ttgagttcaa
     2581 ctacgattct tttttggaat cgttgaaggc agagtcaccc tcttcttcaa agtacaatct
     2641 gccggaaact ttgaaagagt acatgaccct tagttcgtct catctgaata gtcaacactc
     2701 cgacactttg gcaaatggca ctaacggtaa ctattctagc accgtttcca acaacttgag
     2761 cttaagtttg aactccttct ctttctctga caagttctca ttgagtccac caacaatcac
     2821 tgacgccgaa aagttttcat tgatgagaaa cttcattgac aacatctcgc catggtttga
     2881 cacttttgac aataccaaac agtttggaac aaaaattcca gttctggcca aaaaatgttc
     2941 ttcattgtac tatgccattc tggctatatc ttctcgtcaa agagaaagga taaagaaaga
     3001 gcacaatgaa aaaacattgc aatgctacca atactcacta caacagctca tccctactgt
     3061 tcaaagctca aataatattg agtacattat cacatgtatt ctcctgagtg tgttccacat
     3121 catgtctagt gaaccttcaa cccagaggga catcattgtg tcattggcaa aatacattca
     3181 agcatgcaac ataaacggat ttacatctaa tgacaaactg gaaaagagta ttttctggaa
     3241 ctatgtcaat ttggatttgg ctacttgtgc aatcggtgaa gagtcaatgg tcattccttt
     3301 tagctactgg gttaaagaga caactgacta caagaccatt caagatgtga agccattttt
     3361 caccaagaag actagcacga caactgacga tgacttggac gatatgtatg ccatctacat
     3421 gctgtacatt agtggtagaa tcattaacct gttgaactgc agagatgcga agctcaattt
     3481 tgagcccaag tgggagtttt tgtggaatga actcaatgaa tgggaattga acaaaccctt
     3541 gacctttcaa agtattgttc agttcaaggc caatgacgaa tcgcagggcg gatcaacttt
     3601 tccaactgtt ctattctcca actctcgaag ctgttacagt aaccagctgt atcatatgag
     3661 ctacatcatc ttagtgcaga ataaaccacg attatacaaa atccccttta ctacagtttc
     3721 tgcttcaatg tcatctccat cggacaacaa agctgggatg tctgcttcca gcacacctgc
     3781 ttcagaccac cacgcttctg gtgatcattt gtctccaaga agtgtagagc cctctctttc
     3841 gacaacgttg agccctccgc ctaatgcaaa cggtgcaggt aacaagttcc gctctacgct
     3901 ctggcatgcc aagcagatct gtgggatttc tatcaacaac aaccacaaca gcaatctagc
     3961 agccaaagtg aactcattgc aaccattgtg gcacgctgga aagctaatta gttccaagtc
     4021 tgaacataca cagttgctga aactgttgaa caaccttgag tgtgcaacag gctggcctat
     4081 gaactggaag ggcaaggagt taattgacta ctggaatgtt gaagaataac atcatcatca
     4141 tcatcattga gtttgtagcc ttagacatga ctgttcctca gttcaagttg ggcacttacg
     4201 agaagaccgg tcttgctaga ttctaatcaa gaggatgtca gaatgccatt tgcctgagag
     4261 atgcaggctt catttttgat acttttttat ttgtaaccta tatagtatag gatttttttt
     4321 gtcattttgt ttcttctcgt acgagcttgc tcctgatcag cctatctcgc agctgatgaa
     4381 tatcttgtgg taggggtttg ggaaaatcat tcgagtttga tgtttttctt ggtatttccc
     4441 actcctcttc agagtacaga agattaagtg agaccttcgt ttgtgcggat cccccacaca
     4501 ccatagcttc aaaatgtttc tactcctttt ttactcttcc agattttctc ggactccgcg
     4561 catcgccgta ccacttcaaa acacccaagc acagcatact aaatttcccc tctttcttcc
     4621 tctagggtgt cgttaattac ccgtactaaa ggtttggaaa agaaaaaaga gaccgcctcg
     4681 tttctttttc ttcgtcgaaa aaggcaataa aaatttttat cacgtttctt tttcttgaaa
     4741 attttttttt ttgatttttt tctctttcga tgacctccca ttgatattta agttaataaa
     4801 cggtcttcaa tttctcaagt ttcagtttca tttttcttgt tctattacaa ctttttttac
     4861 ttcttgctca ttagaaagaa agcatagcaa tctaatctaa gggcggtgtt gacaattaat
     4921 catcggcata gtatatcggc atagtataat acgacaaggt gaggaactaa accatggcca
     4981 agttgaccag tgccgttccg gtgctcaccg cgcgcgacgt cgccggagcg gtcgagttct
     5041 ggaccgaccg gctcgggttc tcccgggact tcgtggagga cgacttcgcc ggtgtggtcc
     5101 gggacgacgt gaccctgttc atcagcgcgg tccaggacca ggtggtgccg gacaacaccc
     5161 tggcctgggt gtgggtgcgc ggcctggacg agctgtacgc cgagtggtcg gaggtcgtgt
     5221 ccacgaactt ccgggacgcc tccgggccgg ccatgaccga gatcggcgag cagccgtggg
     5281 ggcgggagtt cgccctgcgc gacccggccg gcaactgcgt gcacttcgtg gccgaggagc
     5341 aggactgaca cgtccgacgc ggcccgacgg gtccgaggcc tcggagatcc gtcccccttt
     5401 tcctttgtcg atatcatgta attagttatg tcacgcttac attcacgccc tccccccaca
     5461 tccgctctaa ccgaaaagga aggagttaga caacctgaag tctaggtccc tatttatttt
     5521 tttatagtta tgttagtatt aagaacgtta tttatatttc aaatttttct tttttttctg
     5581 tacagacgcg tgtacgcatg taacattata ctgaaaacct tgcttgagaa ggttttggga
     5641 cgctcgaagg ctttaatttg caagctggag accaacatgt gagcaaaagg ccagcaaaag
     5701 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     5761 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     5821 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     5881 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     5941 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     6001 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     6061 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     6121 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     6181 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     6241 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     6301 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     6361 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gatcagatct tcatctaaca
     6421 ctttgtatag cacatcgtac cagtttatca aaatccaaca agttttcctc tgggtactga
     6481 aattcgatta gtaataaacg ggcgcacttg gtgatctcct catcctgttc actagccttc
     6541 ttttcgctta acttcatcaa cgagatcaac tcgagtctgc gagcagagaa ctgtttccaa
     6601 agagtttcat cgctgaaatt cagcttcttt cgaataaaat gtgtcactcc cactttatta
     6661 ctcgtggtgt tatttgtttt ccacgaacta gttgaatcgc catctttact accgtcctgg
     6721 gtgacatccg gcgaattcgg ga
//
