LOCUS       sequence-310560-        7454 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  see pDUAL_tacI: exchanged origin of replication (pBR322) and 
            introduced bom site.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7454)
  AUTHORS   Schwaiger KN, Voit A, Dobiasova H, Luley C, Wiltschi B, Nidetzky B
  TITLE     Plasmid Design for Tunable Two-Enzyme Co-Expression Promotes 
            Whole-Cell Production of Cellobiose.
  JOURNAL   Biotechnol J. 2020 Jul 15:e2000063. doi: 10.1002/biot.202000063.
  PUBMED    32668097
REFERENCE   2  (bases 1 to 7454)
  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: "Biotechnol 
            J. 2020 Jul 15:e2000063. doi: 10.1002/biot.202000063."
COMMENT     Sequence Label: pDUAL_2_tacI
FEATURES             Location/Qualifiers
     source          1..7454
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(12..17,18..33,34..40)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    48..64
                     /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     53..75
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     CDS             97..120
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     terminator      1648..1742
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     promoter        1749..1826
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     CDS             1827..2909
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     complement(1846..1865)
                     /label=LacI-R
                     /note="LacI, reverse primer"
     RBS             2919..2930
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     CDS             join(2937..3005,3006..3797)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(3155..3174)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     terminator      complement(3840..3887)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(6369..6386)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     RBS             6405..6427
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    6442..6466
                     /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     complement(6466..6485)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(6467..6485)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     rep_origin      6522..7110
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7011..7030
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7264..7281
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    7296..7436
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(7362..7381)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 gatccgagct gttgacaatt aatcatcggc tcgtataatg tgtggaattg tgagcggata
       61 acaatttcac acaggaaaca gaataccatg gcaagctgga gccacccgca gttcgaaaag
      121 ggtaccatga aaaataaagt ccaactgatt acctatgcgg atcgtctggg tgatggcacc
      181 attaaaagca tgacggacat cctgcgtacc cgctttgatg gcgtttatga cggtgtccac
      241 attctgccgt ttttcacccc gtttgatggc gccgacgcag gtttcgatcc gatcgaccat
      301 accaaagttg atgaacgtct gggtagctgg gatgacgtcg ccgaactgtc taaaacccac
      361 aacattatgg tggatgcaat cgttaatcac atgtcatggg aatcgaaaca gttccaagat
      421 gttctggcca aaggcgaaga atcagaatat tacccgatgt ttctgaccat gagcagcgtg
      481 ttcccgaacg gtgccacgga agaagacctg gcaggcattt atcgtccgcg tccgggtctg
      541 ccgtttaccc actacaaatt cgcaggcaaa acgcgtctgg tctgggtgtc ttttaccccg
      601 cagcaagtgg acatcgatac ggacagtgat aaaggttggg aatatctgat gtccattttc
      661 gaccaaatgg cggccagcca tgtgtcttat atccgcctgg atgcggttgg ctacggtgcc
      721 aaagaagcag gcaccagttg ctttatgacc ccgaaaacgt tcaaactgat ttcccgtctg
      781 cgcgaagaag gcgtgaaacg cggtctggaa attctgatcg aagtccacag ctattacaaa
      841 aaacaggtgg aaatcgcgtc taaagtggat cgcgtttatg actttgccct gccgccgctg
      901 ctgctgcatg ccctgagtac cggccacgtc gaaccggtgg cccattggac ggatattcgt
      961 ccgaacaatg ctgttaccgt cctggatacg catgacggca tcggcgttat tgatatcggt
     1021 tcagatcagc tggaccgctc gctgaaaggt ctggtgccgg acgaagatgt ggataacctg
     1081 gtgaatacga ttcatgcgaa cacccacggc gaatcacagg cagctaccgg tgcggcggca
     1141 tcgaacctgg atctgtacca agttaatagt acctactact ccgctctggg ctgtaacgat
     1201 cagcactaca tcgctgcgcg tgcggtgcag tttttcctgc cgggtgttcc gcaagtctat
     1261 tacgtgggcg ccctggcagg taaaaatgat atggaactgc tgcgcaaaac caacaatggt
     1321 cgtgacatta accgccatta ttacagcacg gccgaaatcg atgaaaatct gaaacgtccg
     1381 gtggttaaag cactgaacgc tctggcgaaa tttcgcaatg aactggatgc ttttgacggc
     1441 accttctcat ataccacgga tgacgatacg agtatttcct ttacctggcg tggtgaaacg
     1501 tcgcaggcca ccctgacgtt cgaaccgaaa cgcggcctgg gtgttgataa taccacgccg
     1561 gtcgcaatgc tggaatggga agacagtgct ggtgaccatc gctccgacga cctgatcgct
     1621 aatccgccgg ttgttgcgta aaagcttgac tcctgttgat agatccagta atgacctcag
     1681 aactccatct ggatttgttc agaacgctcg gttgccgccg ggcgtttttt attggtgaga
     1741 atgaattcga caccatcgaa tggtgcaaaa cctttcgcgg tatggcatga tagcgcccgg
     1801 aagagagtca attcagggtg gtgaatgtga aaccagtaac gttatacgat gtcgcagagt
     1861 atgccggtgt ctcttatcag accgtttccc gcgtggtgaa ccaggccagc cacgtttctg
     1921 cgaaaacgcg ggaaaaagtg gaagcggcga tggcggagct gaattacatt cccaaccgcg
     1981 tggcacaaca actggcgggc aaacagtcgt tgctgattgg cgttgccacc tccagtctgg
     2041 ccctgcacgc gccgtcgcaa attgtcgcgg cgattaaatc tcgcgccgat caactgggtg
     2101 ccagcgtggt ggtgtcgatg gtagaacgaa gcggcgtcga agcctgtaaa gcggcggtgc
     2161 acaatcttct cgcgcaacgc gtcagtgggc tgatcattaa ctatccgctg gatgaccagg
     2221 atgccattgc tgtggaagct gcctgcacta atgttccggc gttatttctt gatgtctctg
     2281 accagacacc catcaacagt attattttct cccatgagga cggtacgcga ctgggcgtgg
     2341 agcatctggt cgcattgggt caccagcaaa tcgcgctgtt agcgggccca ttaagttctg
     2401 tctcggcgcg tctgcgtctg gctggctggc ataaatatct cactcgcaat caaattcagc
     2461 cgatagcgga acgggaaggc gactggagtg ccatgtccgg ttttcaacaa accatgcaaa
     2521 tgctgaatga gggcatcgtt cccactgcga tgctggttgc caacgatcag atggcgctgg
     2581 gcgcaatgcg cgccattacc gagtccgggc tgcgcgttgg tgcggatatc tcggtagtgg
     2641 gatacgacga taccgaggac agctcatgtt atatcccgcc gttaaccacc atcaaacagg
     2701 attttcgcct gctggggcaa accagcgtgg accgcttgct gcaactctct cagggccagg
     2761 cggtgaaggg caatcagctg ttgcccgtct cactggtgaa aagaaaaacc accctggcgc
     2821 ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag ctggcacgac
     2881 aggtttcccg actggaaagc gggcagtgaa gatctattaa agaggagaaa ttaagcatga
     2941 gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc cttcctgttt
     3001 ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg ggtgcacgag
     3061 tgggttacat cgaactggat ctcaacagcg gtaagatcct tgagagtttt cgccccgaag
     3121 aacgttttcc aatgatgagc acttttaaag ttctgctatg tggcgcggta ttatcccgta
     3181 ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta ttctcagaat gacttggttg
     3241 agtactcacc agtcacagaa aagcatctta cggatggcat gacagtaaga gaattatgca
     3301 gtgctgccat aaccatgagt gataacactg cggccaactt acttctgaca acgatcggag
     3361 gaccgaagga gctaaccgct tttttgcaca acatggggga tcatgtaact cgccttgatc
     3421 gttgggaacc ggagctgaat gaagccatac caaacgacga gcgtgacacc acgatgcctg
     3481 tagcaatggc aacaacgttg cgcaaactat taactggcga actacttact ctagcttccc
     3541 ggcaacaatt aatagactgg atggaggcgg ataaagttgc aggaccactt ctgcgctcgg
     3601 cccttccggc tggctggttt attgctgata aatctggagc cggtgagcgt gggtctcgcg
     3661 gtatcattgc agcactgggg ccagatggta agccctcccg tatcgtagtt atctacacga
     3721 cggggagtca ggcaactatg gatgaacgaa atagacagat cgctgagata ggtgcctcac
     3781 tgattaagca ttggtaactg cagatcctta gcgaaagcta aggatttttt ttactcgagc
     3841 aaaaaacccc tcaagacccg tttagaggcc ccaaggggtt atgctagtgt acactacaat
     3901 gtcacatcga cacgaactgt tgaccctgcg ggggcataag gaaccaagtt gccttctaca
     3961 ggtgttccat caacgacaag gcgaccgcgt gatccgtcgg tcccactgtt agtgactgtt
     4021 atctcgtaag tggctccacg ggcgacacgc gtaaccgtaa agctaggtac gtcaggtccg
     4081 atctgcgggt ctaccaccaa gccgtcgtac tctgggcgaa ccccaagaag gtactgtgac
     4141 acggtcacaa agttccaagc cgcagtcccg gtaagccagc tgttctttgc ttcaccgtga
     4201 cggaccgcct ccttccctgc tatcatttgt gcatagacat agggttcaag acggtgaacg
     4261 tcgcttatgt cctcgcgata cgcgggagta attctcttgt agtaatcaaa tgcccgtccg
     4321 cctcttccga cgacagtttc cgctatgata acccaaggat tattgtggca aaatatacct
     4381 ccattctctt tatacccggg tggataagtg gatacttcac ccatgtgtac ttggtatgta
     4441 gtgtatgctg ggtattgaag gaccataccg tgatcggttg ccaacatttc gttcacagaa
     4501 tcaagcgcct taatagcggg agcgtccgta tcttgaggcc cttcgcctac accgactcca
     4561 gccatcaccg cgaacccttg cggttcgatc caaatcttcc cttcgtcatg tgcgtcggtg
     4621 ccgatagggt ttccataata gtcataagct ctcaagaacc agctgccgtc ccacccgtcg
     4681 gtcagaagtg cgtctctcat ttcagccaca tgccctcttg ctctatcggc tacgtcggcc
     4741 aacccgcgcc gcgctgccaa ctcggcatac tgctccccat ataaaacgaa ttgcgcggct
     4801 atgaacgttg actccgctac gccaccggct tggttctccg ttgtctgaaa tgactcgccc
     4861 ggggttgtac tgaagcaatt aagattcaaa caatcgttcc aatctgcgcg gccaataaga
     4921 ggcaaaccgt ggggaccccg atgggttacg gtaaactcaa aactacgggt caggtgctca
     4981 aataacggaa cttctgaccc cggttcgtta tcaaaaggta cgggctcatc taatatggag
     5041 aagtcaccag tttctttgat gtaggcggca gttcccgcga ttaaccaaag aggatcgtcg
     5101 ttaaacccag agccaatgtc gttgttaccg cgcttcgtta aaggttggta ttggtgataa
     5161 gcggaaccgt ccgcgaactg ggttgacgct atgtcaatga tgcgctcacg cgcccgctct
     5221 gggattaagt gaacaaagcc cagaaggtct tggttgctat cacgaaaccc catccctctc
     5281 ccaataccgg tctcgaaaaa gctcgccgag cgactcatgt taaaggttac catgcattgg
     5341 tactgattcc atatattaac catccggtcc aatttttcgt cattggaact aacggaatac
     5401 gtcgataaca aatccgtcca ataatccttc aaagcggcga aagctgcgtc agtctgttcg
     5461 gacgtagcaa atcttgataa taaagcgtga gctctttctt tattcacgac ctgttttgcg
     5521 tcgtctgccc acttctcctc atcaggattt tccacgtacc caagcacata aaccaattct
     5581 ctggattctc caggcgctaa agacactgcc acgctatgag agccaatagg ataccatcca
     5641 cttgcaacgc tatttgcact ttctccctta aggggaactg cggcttcacc aagcgagtta
     5701 taggctccca caaatgtgtc gcgatctgta tcgaaaccct cggcctgcgt attaacggcg
     5761 aataccgcgt agtgatcgcg tctctcccgg tactctgtac ggtgataaat tgcactgcca
     5821 tgcggagatt cttgttccac ttcaacctct cctatgctca gattacgctg gtagttagtt
     5881 tgatcgtctt gcgcgttcca aagacaaaat tcgacaaaag aaaacaacgt cagggattta
     5941 tacgagtcag aggtattagt gaccgtaact ttctggacct ccgcgttctc accgactggg
     6001 acgaaaaaca aggtctcaac ccgtacccca tttctttcac ctgtaatggt actgtagcca
     6061 agaccatgtc tggcttcaaa atgatctaag tctgctttaa caggaagcca cgatggcgtc
     6121 caaacgtccc caccgtcgtt tacataaaga taccggcctc ccgcgtctgc tggtatgttg
     6181 ttgtatctgt atcttgtcaa tctacgcatc ttggcatcgc gataaaacga atagccgcca
     6241 gcctggtgcg acaaaagact gaagaactgt tccgaaccca gatagttgat ccaagggtac
     6301 ggagtatgcg gtgtggtgat aacgtactcg cgtgcttcat cgtcaaaatg accatagcgg
     6361 gtcgacccgt gatggtgatg gtgatggctg ctgcccatat gatatctcct tcttaaagtt
     6421 aaacaaaatt atttctagag gggaattgtt atccgctcac aattccccta tagtgagtcg
     6481 tattaatttc gcgggatcga ctagtagatc aaaggatctt cttgagatcc tttttttctg
     6541 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     6601 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     6661 aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     6721 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     6781 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     6841 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     6901 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     6961 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     7021 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     7081 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     7141 ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc tgattctgtg
     7201 gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg aacgaccgag
     7261 cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt tctccttacg
     7321 catctgtgcg gtatttcaca ccgcatatgg tgcactctca gtacaatctg ctctgatgcc
     7381 gcatagttaa gccagtatac actccgctat cgctacgtga ctgggtcatg gctgcgcccc
     7441 gacacccgcc aacg
//
