LOCUS       62056_19145        6901 bp DNA     circular SYN 26-NOV-2019
DEFINITION  Expression vector pS238D1.M, complete sequence.
ACCESSION   MN393460
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6901)
  AUTHORS   Calles B, Goni-Moreno A, de Lorenzo V.
  TITLE     Digitalizing heterologous gene expression in Gram-negative bacteria 
            with a portable on/off module
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6901)
  AUTHORS   Calles B, Goni-Moreno A, de Lorenzo V.
  TITLE     Direct Submission
  JOURNAL   Submitted (28-AUG-2019) Systems Biology Program, Centro Nacional de 
            Biotecnologia, Darwin, 3, Cantoblanco, Madrid 28049, Spain
REFERENCE   3  (bases 1 to 6901)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (28-AUG-2019) Systems Biology Program, Centro Nacional de 
            Biotecnologia, Darwin, 3, Cantoblanco, Madrid 28049, Spain"
FEATURES             Location/Qualifiers
     source          1..6901
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      5..99
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             231..1043
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
     oriT            1213..1321
                     /label=oriT
                     /note="incP origin of transfer"
     CDS             complement(1335..1994)
                     /label=pBBR1 Rep
                     /note="replication protein for the broad-host-range plasmid
                     pBBR1 from Bordetella bronchiseptica"
     rep_origin      complement(1995..2766)
                     /direction=LEFT
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     terminator      complement(2881..2967)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     CDS             complement(2995..3957)
                     /label=XylS
                     /note="XylS regulator encoded by the Pseudomonas putida TOL
                     plasmid pWWO"
     promoter        4857..4937
                     /label=Pm promoter
                     /note="The bacterial Pm promoter is activated by XylS in
                     the presence of benzoate or m-toluate (Marques et al., 
                     1999)."
     regulatory      4972..4978
                     /regulatory_class="ribosome_binding_site"
     CDS             4987..5697
                     /label=lambda repressor (ts)
                     /note="temperature-sensitive variant of the phage lambda
                     repressor"
     CDS             5698..5715
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             5744..6454
                     /label=superfolder GFP
                     /note="GFP variant that folds robustly even when fused to
                     poorly folded proteins (Pedelacq et al., 2006)"
     misc_feature    6458..6514
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     regulatory      6547..6576
                     /label=synthetic T500 terminator
                     /note="synthetic T500 terminator"
                     /regulatory_class="terminator"
     terminator      complement(6583..6610)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(6650..6696)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     regulatory      complement(6711..6819)
                     /note="anti CI sRNA; MicC-based"
                     /regulatory_class="other"
     regulatory      complement(6820..6901)
                     /label=PL-s1con
                     /note="PL-s1con"
                     /regulatory_class="promoter"
ORIGIN
        1 cttggactcc tgttgataga tccagtaatg acctcagaac tccatctgga tttgttcaga
       61 acgctcggtt gccgccgggc gttttttatt ggtgagaatc caggggtccc caataattac
      121 gatttaaatt tgtgtctcaa aatctctgat gttacattgc acaagataaa aatatatcat
      181 catgaacaat aaaactgtct gcttacataa acagtaatac aaggggtgtt atgagccata
      241 ttcagcgtga aacgagctgt agccgtccgc gtctgaacag caacatggat gcggatctgt
      301 atggctataa atgggcgcgt gataacgtgg gtcagagcgg cgcgaccatt tatcgtctgt
      361 atggcaaacc ggatgcgccg gaactgtttc tgaaacatgg caaaggcagc gtggcgaacg
      421 atgtgaccga tgaaatggtg cgtctgaact ggctgaccga atttatgccg ctgccgacca
      481 ttaaacattt tattcgcacc ccggatgatg cgtggctgct gaccaccgcg attccgggca
      541 aaaccgcgtt tcaggtgctg gaagaatatc cggatagcgg cgaaaacatt gtggatgcgc
      601 tggccgtgtt tctgcgtcgt ctgcatagca ttccggtgtg caactgcccg tttaacagcg
      661 atcgtgtgtt tcgtctggcc caggcgcaga gccgtatgaa caacggcctg gtggatgcga
      721 gcgattttga tgatgaacgt aacggctggc cggtggaaca ggtgtggaaa gaaatgcata
      781 aactgctgcc gtttagcccg gatagcgtgg tgacccacgg cgattttagc ctggataacc
      841 tgattttcga tgaaggcaaa ctgattggct gcattgatgt gggccgtgtg ggcattgcgg
      901 atcgttatca ggatctggcc attctgtgga actgcctggg cgaatttagc ccgagcctgc
      961 aaaaacgtct gtttcagaaa tatggcattg ataatccgga tatgaacaaa ctgcaatttc
     1021 atctgatgct ggatgaattt ttctaataat taattggacc gcggtccgcg cgttgtcctt
     1081 ttccgctgca taaccctgct tcggggtcat tatagcgatt ttttcggtat atccatcctt
     1141 tttcgcacga tatacaggat tttgccaaag ggttcgtgta gactttcctt ggtgtatcca
     1201 acggcgtcag ccgggcagga taggtgaagt aggcccaccc gcgagcgggt gttccttctt
     1261 cactgtccct tattcgcacc tggcggtgct caacgggaat cctgctctgc gaggctggcc
     1321 gtaggccggc cctaccggcg cggcagcgtt acccgtgtcg gcggctccaa cggctcgcca
     1381 tcgtccagaa aacacggctc atcgggcatc ggcaggcgct gctgcccgcg ccgttcccat
     1441 tcctccgttt cggtcaaggc tggcaggtct ggttccatgc ccggaatgcc gggctggctg
     1501 ggcggctcct cgccggggcc ggtcggtagt tgctgctcgc ccggatacag ggtcgggatg
     1561 cggcgcaggt cgccatgccc caacagcgat tcgtcctggt cgtcgtgatc aaccaccacg
     1621 gcggcactga acaccgacag gcgcaactgg tcgcggggct ggccccacgc cacgcggtca
     1681 ttgaccacgt aggccgacac ggtgccgggg ccgttgagct tcacgacgga gatccagcgc
     1741 tcggccacca agtccttgac tgcgtattgg accgtccgca aagaacgtcc gatgagcttg
     1801 gaaagtgtct tctggctgac caccacggcg ttctggtggc ccatctgcgc cacgaggtga
     1861 tgcagcagca ttgccgccgt gggtttcctc gcaataagcc cggcccacgc ctcatgcgct
     1921 ttgcgttccg tttgcaccca gtgaccgggc ttgttcttgg cttgaatgcc gatttctctg
     1981 gactgcgtgg ccatgcttat ctccatgcgg taggggtgcc gcacggttgc ggcaccatgc
     2041 gcaatcagct gcaacttttc ggcagcgcga caacaattat gcgttgcgta aaagtggcag
     2101 tcaattacag attttcttta acctacgcaa tgagctattg cggggggtgc cgcaatgagc
     2161 tgttgcgtac cccccttttt taagttgttg atttttaagt ctttcgcatt tcgccctata
     2221 tctagttctt tggtgcccaa agaagggcac ccctgcgggg ttcccccacg ccttcggcgc
     2281 ggctccccct ccggcaaaaa gtggcccctc cggggcttgt tgatcgactg cgcggccttc
     2341 ggccttgccc aaggtggcgc tgcccccttg gaacccccgc actcgccgcc gtgaggctcg
     2401 gggggcaggc gggcgggctt cgcccttcga ctgcccccac tcgcataggc ttgggtcgtt
     2461 ccaggcgcgt caaggccaag ccgctgcgcg gtcgctgcgc gagccttgac ccgccttcca
     2521 cttggtgtcc aaccggcaag cgaagcgcgc aggccgcagg ccggaggctt ttccccagag
     2581 aaaattaaaa aaattgatgg ggcaaggccg caggccgcgc agttggagcc ggtgggtatg
     2641 tggtcgaagg ctgggtagcc ggtgggcaat ccctgtggtc aagctcgtgg gcaggcgcag
     2701 cctgtccatc agcttgtcca gcagggttgt ccacgggccg agcgaagcga gccagccggt
     2761 ggccgctcgc ggccatcgtc cacatatcca cgggctggca agggagcgca gcgaccgcgc
     2821 agggcgaagc ccggagagca agcccgtagg gggggcgcgc ccagctgtct agggcggcgg
     2881 atttgtccta ctcaggagag cgttcaccga caaacaacag ataaaacgaa aggcccagtc
     2941 tttcgactga gcctttcgtt ttatttgatg cctttaatta aacgttcgta atcaagccac
     3001 ttcctttttg cattgacgca gggtgtcgga aggcaactcg ccgaacgcgc tcctatagtt
     3061 ttcagcgaag cgtcccaaat gtaagaagcc gtagtctagg gctatctcag ttatactacg
     3121 cacattggca ctgggatcgt tcaagcaggc gcggatgctt tcgagcttgc ggttgcggat
     3181 gtagttcttc ggcgtggtgc cggcgtgctt ctcgaacaaa ttgtagagcg agcgtggact
     3241 catcatcgcc agctccgcta accgctcaag gctgatattc cgtttgagat tctcctcaat
     3301 gaattgaacg actcgctcga aagacgggtt acctttgctg aaaatttcac ggctgacatt
     3361 gctgcccagc atttcgagca gcttggaagc gatgatcccc gcatagtgct cttggacccg
     3421 aggcatcgac tttgtatgtt ccgcttcgtc acaaactaac ccgagtagat tgataaagcc
     3481 atcgagttgc tggagattgt gtcgcgcggc gaaacggata ccctccctcg gcttgtgcca
     3541 attgttgtca ctgcacgccc gatcaaggac cactgagggc aatttaacga taaatttctc
     3601 gcaatcttct gaataggtca ggtcggcttg gtcatccgga ttgagcagca atagttcgcc
     3661 cggcgcaaaa tagtgctcct ggccatggcc acgccacagg caatggcctt tgagtattat
     3721 ttgcagatga taacaggtct ctaatccagg cgagattacc ctcacgctac cgccgtagct
     3781 gattcgacac agatcgaggc atccgaagat tctgtggtgc agcctgcctg ccgggcgccc
     3841 gcccttgggc aggcgaatag agtgcgtacc gacatactgg ttaacataat cggagactgc
     3901 atagggctcg gcgtggacga agatctgact tttctcgttc aataagcaaa aatccatagt
     3961 tcacggttct cttattttaa tgtgggctgc ttggtgtgat gtagaaaggc gccaagtcga
     4021 tgaaaatgca tctcgacgtg atgcgtatac gggttacccc cattgccacg ttgcgccatc
     4081 ctttttgcaa tcagtgacca cttttccaag caaaaataac gccaagcaga acgaagacgt
     4141 tctttttaag aagcgagaac accagaagtt cgtgctgtcg gggcatgggg cgacgaattg
     4201 gcggataaag gggatctgct ggatattacg gcctttttaa agaccgtaaa gaaaaataag
     4261 cacaagtttt atccggcctt tattcacatt cttgcccgcc tgatgaatgc tcatccgtaa
     4321 ttacgtatgg caatgaaaga cggtgagctg gtgatatggg atagtgttca cccttgttac
     4381 accgttttcc atgagcaaac tgaaacgttt tcatcgctct ggagtgaata ccacgacgat
     4441 ttccggcagt ttctacacat atattcgcaa gatgtggcgt gttacggtga aaacctggcc
     4501 tatttcccta aagggtttat tgagaatatg tttttcgtct cagccaatcc ctgggtgagt
     4561 ttcaccagtt ttgatttaaa cgtggccaat atggacaact tcttcgcccc cgttttcacc
     4621 atgcatggga attagcttga tctgaccaac gaccggtagc ggagctatcc aacggcggta
     4681 taccaggaaa acacacagca ggtacatcag aacagtacca tgactgaaga acaaatagtt
     4741 ttttcctgat ccataaagca gaacggcctg ctccatgaca aatctggctc cccaactaat
     4801 gccccatgca gccagcataa ccagcataaa gtgcagtgtc cggtttgata gggataagtc
     4861 cagccttgca agaagcggat acaggagtgc aaaaaatggc tatctctagt aaggcctacc
     4921 ccttaggctt tatgcaacag aaacaataat aatggagtca tgaccatgcc taggaggaaa
     4981 aaacatatga gcacaaaaaa gaaaccatta acacaagagc agcttgagga tgcacgtcgc
     5041 cttaaagcaa tttatgaaaa aaagaaaaat gaacttggct tatcccagga atctgtcgca
     5101 gacaagatgg ggatggggca gtcaggcgtt ggtgctttat ttaatggcat caatgcatta
     5161 aatgcttata acgccgcatt gcttacaaaa attctcaaag ttagcgttga agaatttagc
     5221 ccttcaatcg ccagagaaat ctacgagatg tatgaagcgg ttagtatgca gccgtcactt
     5281 agaagtgagt atgagtaccc tgttttttct catgttcagg cagggatgtt ctcacctaag
     5341 ctgagaacct ttaccaaagg tgatgcggag agatgggtaa gcacaaccaa aaaagccagt
     5401 gattctgcat tctggcttga ggttgaaggt aattccatga ccgcaccaac aggctccaag
     5461 ccgagctttc ctgacggaat gttaattctc gttgaccctg agcaggctgt tgagccaggt
     5521 gatttctgca tagccagact tgggggtgat gagtttacct tcaagaaact gatcagggat
     5581 agcggtcagg tgtttttaca accactaaac ccacagtacc caatgatccc atgcaatgag
     5641 agttgttccg ttgtggggaa agttatcgct agtcagtggc ctgaagagac gtttggccat
     5701 catcaccacc atcattagga tggtggtgat gataatggct agccgtaaag gtgaagaact
     5761 gttcaccggt gttgttccga tcctggttga actggatggt gatgttaacg gccacaaatt
     5821 ctctgttcgt ggtgaaggtg aaggtgatgc aaccaacggt aaactgaccc tgaaattcat
     5881 ctgcactacc ggtaaactgc cggttccatg gccgactctg gtgactaccc tgacctatgg
     5941 tgttcagtgt ttttctcgtt acccggatca catgaagcag catgatttct tcaaatctgc
     6001 aatgccggaa ggttatgtac aggagcgcac catttctttc aaagacgatg gcacctacaa
     6061 aacccgtgca gaggttaaat ttgaaggtga tactctggtg aaccgtattg aactgaaagg
     6121 cattgatttc aaagaggacg gcaacatcct gggccacaaa ctggaatata acttcaactc
     6181 ccataacgtt tacatcaccg cagacaaaca gaagaacggt atcaaagcta acttcaaaat
     6241 tcgccataac gttgaagacg gtagcgtaca gctggcggac cactaccagc agaacactcc
     6301 gatcggtgat ggtccggttc tgctgccgga taaccactac ctgtccaccc agtctaaact
     6361 gtccaaagac ccgaacgaaa agcgcgacca catggtgctg ctggagttcg ttactgccgc
     6421 aggtatcacg cacggcatgg atgaactcta caaataagaa ttcgagctcg gtacccgggg
     6481 atcctctaga gtcgacctgc aggcatgcaa gcttgcggcc gcgtcgtgac tgggaaaacc
     6541 ctggcgcaaa gcccgccgaa aggcgggctt ttctgttata aacgcagaaa ggcccacccg
     6601 aaggtgagcc agtgtgactc tagtagagag cgttcaccga caaaaacaga taaaacgaaa
     6661 ggcccagtct ttcgactgag cctttcgttt tatttgatgc ctggctcgag ctcgagaaaa
     6721 aaagcccgga cgactgttcg ggcttgtctt tttatatgtt ggaaaatcag tggcaatgca
     6781 atggcccaac agaaaatgag cacaaaaaag aaaccattat gtgctcatta taaccgccag
     6841 tggtatttat gtcaacaccg ccagagataa tttatcaccg cagatggtta tctgtactag
     6901 t
//
