LOCUS       62056_19140        7255 bp DNA     circular SYN 26-NOV-2019
DEFINITION  Expression vector pS238D.NIa, complete sequence.
ACCESSION   MN393462
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7255)
  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  JOURNAL   Unpublished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 7255)
  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 7255)
  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            "Unpublished"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..7255
                     /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             4990..6069
                     /label=lacI
                     /note="lac repressor"
     CDS             6070..6087
                     /label=6xHis
                     /note="6xHis affinity tag"
     gene            6107..6847
                     /gene="nIa"
                     /label=nIa
     CDS             6107..6847
                     /codon_start=1
                     /transl_table=11
                     /gene="nIa"
                     /product="NIa"
                     /label=nIa
                     /note="specific protease"
                     /protein_id="QGN03492.1"
                     /translation="MASSKSLFRGLRDYNPIASSICQLNNSSGARQSVMFGLGFGGLIV
                     TNQHLFKRNDGELTIRSHHGEFVVKDTKTLKLLPCKGRDIVIIRLPKDFPPFPKRLQFR
                     TPTTEDRVCLIGSNFQTKSISSTMSETSATYPVDNSHFWKHWISTKDGHCGLPIVSTRD
                     GSILGLHSLANSTNTQNFYAAFPDNFETTYLSNQDNDNWIKQWRYNPDEVCWGSLQLKR
                     DIPQSPFTICKLLTDLDGEFVYTQ"
     regulatory      6925..6954
                     /label=synthetic T500 terminator
                     /note="synthetic T500 terminator"
                     /regulatory_class="terminator"
     terminator      complement(6961..6988)
                     /label=T7Te terminator
                     /note="phage T7 early transcription terminator"
     terminator      complement(7028..7074)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     regulatory      complement(7090..7197)
                     /note="anti LacI sRNA; MicC-based"
                     /regulatory_class="other"
     regulatory      complement(7198..7255)
                     /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 aaacatatgg tgaaaccagt aacgttatac gatgtcgcag agtatgccgg tgtctcttat
     5041 cagaccgttt cccgcgtggt gaaccaggcc agccacgttt ctgcgaaaac gcgggaaaaa
     5101 gtggaagcgg cgatggcgga gctgaattac attcccaacc gcgtggcaca acaactggcg
     5161 ggcaaacagt cgttgctgat tggcgttgcc acctccagtc tggccctgca cgcgccgtcg
     5221 caaattgtcg cggcgattaa atctcgcgcc gatcaactgg gtgccagcgt ggtggtgtcg
     5281 atggtagaac gaagcggcgt cgaagcctgt aaagcggcgg tgcacaatct tctcgcgcaa
     5341 cgcgtcagtg ggctgatcat taactatccg ctggatgacc aggatgccat tgctgtggaa
     5401 gctgcctgca ctaatgttcc ggcgttattt cttgatgtct ctgaccagac acccatcaac
     5461 agtattattt tctcccatga agacggtacg cgactgggcg tggagcatct ggtcgcattg
     5521 ggtcaccagc aaatcgcgct gttagcgggc ccattaagtt ctgtctcggc gcgtctgcgt
     5581 ctggctggct ggcataaata tctcactcgc aatcaaattc agccgatagc ggaacgggaa
     5641 ggcgactgga gtgccatgtc cggttttcaa caaaccatgc aaatgctgaa tgagggcatc
     5701 gttcccactg cgatgctggt tgccaacgat cagatggcgc tgggcgcaat gcgcgccatt
     5761 accgagtccg ggctgcgcgt tggtgcggat atctcggtag tgggatacga cgataccgaa
     5821 gacagctcat gttatatccc gccgttaacc accatcaaac aggattttcg cctgctgggg
     5881 caaaccagcg tggaccgctt gctgcaactc tctcagggcc aggcggtgaa gggcaatcag
     5941 ctgttgcccg tctcactggt gaaaagaaaa accaccctgg cgcccaatac gcaaaccgcc
     6001 tctccccgcg cgttggccga ttcattaatg cagctggcac gacaggtttc ccgactggaa
     6061 agcgggcagc atcatcacca ccatcattag gatggtggtg atgataatgg ctagcagtaa
     6121 atcactgttt agaggcctga gagactataa tccaatcgca agctcaatat gccaattgaa
     6181 taactcatct ggtgctagac aaagtgtaat gtttggactt ggctttgggg gtttaattgt
     6241 cacgaatcag catttgttca aaaggaatga cggagagcta acaatccgat cgcatcatgg
     6301 ggaattcgta gtgaaggaca caaaaactct caaactgctt ccttgcaaag gtcgagacat
     6361 agtgatcatc agattaccaa aggatttccc tccttttccg aagaggttgc agttccgcac
     6421 cccgacgact gaggacagag tttgtttaat tggttcaaat ttccaaacga agagcatttc
     6481 aagcaccatg tcggaaacaa gcgcaacata tccagttgat aacagtcatt tctggaaaca
     6541 ctggattagc acgaaggatg gtcattgcgg attacccatc gtgagcactc gagatggcag
     6601 tattcttggg ctacacagtc ttgcaaattc aacgaacacc cagaatttct atgcagcttt
     6661 ccctgacaac ttcgagacca catacttgtc aaatcaagac aatgataact ggataaagca
     6721 gtggcgatac aacccggatg aagtttgctg gggatcccta caactcaaga gggacattcc
     6781 acagagtccg tttacaattt gtaaactgct aacggatctt gatggggaat ttgtttacac
     6841 tcagtaaggt acccggggat cctctagagt cgacctgcag gcatgcaagc ttgcggccgc
     6901 gtcgtgactg ggaaaaccct ggcgcaaagc ccgccgaaag gcgggctttt ctgttataaa
     6961 cgcagaaagg cccacccgaa ggtgagccag tgtgactcta gtagagagcg ttcaccgaca
     7021 aaaacagata aaacgaaagg cccagtcttt cgactgagcc tttcgtttta tttgatgcct
     7081 ggctcgagct cgagaaaaaa agcccggacg actgttcggg cttgtctttt tatatgttgg
     7141 aaaatcagtg gcaatgcaat ggcccaacag aaaatggtga aaccagtaac gttatactga
     7201 atctaagtat caatgtgagc gctcacaagt caacactctt tttgataaaa ctagt
//
