LOCUS       40924_34101        5886 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pPasscode3, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5886)
  AUTHORS   Chan CT, Lee JW, Cameron DE, Bashor CJ, Collins JJ.
  TITLE     'Deadman' and 'Passcode' microbial kill switches for bacterial 
            containment
  JOURNAL   Nat. Chem. Biol. 12 (2), 82-86 (2016)
  PUBMED    26641934
REFERENCE   2  (bases 1 to 5886)
  AUTHORS   Chan CTY., Lee JW, Cameron DE, Bashor CJ, Collins JJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (12-OCT-2015) Department of Biological Engineering, 
            Massachusetts Institute of Technology, 77 Massachusetts Ave., 
            E25-302b, Cambridge, MA 02139, USA
REFERENCE   3  (bases 1 to 5886)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 5886)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat. Chem. 
            Biol."; date: "2016"; volume: "12"; issue: "2"; pages: "82-86"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (12-OCT-2015) Department of Biological Engineering, Massachusetts 
            Institute of Technology, 77 Massachusetts Ave., E25-302b, Cambridge,
            MA 02139, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..5886
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        42..115
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator
                     sites (Lutz and Bujard, 1997)"
     CDS             122..1159
                     /codon_start=1
                     /product="GalR-LacI"
                     /label=GalR-LacI
                     /protein_id="ALP32105.1"
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMESL
                     SYHPNANARALAQQTTETVGLVVGDVSDPFFGAMVKAVEQVAYHTGNFLLIGNGYHNEQ
                     KERQAIEQLIRHRCAALVVHAKMIPDADLASLMKQMPGMVLINRILPGFENRCIALDDR
                     YGAWLATRHLIQQGHTRIGYLCSNHSISDAEDRLQGYYDALAESGIAANDRLVTFGEPD
                     ESGGEQAMTELLGRGRNFTAVACYNDSMAAGAMGVLNDNGIDVPGEISLIGFDDVLVSR
                     YVRPRLTTVRYPIVTMATQAAELALALADNRPLPEITNVFSPTLVRRHSVSTPSLEASH
                     HATSD"
     terminator      1180..1274
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     promoter        1323..1396
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator
                     sites (Lutz and Bujard, 1997)"
     regulatory      1323..1370
                     /label=pLtetO-1
                     /note="pLtetO-1"
                     /regulatory_class="promoter"
     protein_bind    1323..1341
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1348..1366
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     regulatory      1377..1402
                     /label=RBS
                     /note="RBS"
                     /regulatory_class="ribosome_binding_site"
     CDS             1403..2482
                     /label=lacI
                     /note="lac repressor"
     regulatory      2506..2600
                     /label=T0
                     /note="T0"
                     /regulatory_class="terminator"
     terminator      2506..2600
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     regulatory      complement(2640..2734)
                     /label=T0
                     /note="T0"
                     /regulatory_class="terminator"
     terminator      2640..2734
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(2755..3783)
                     /codon_start=1
                     /product="CelR-ScrR"
                     /label=CelR-ScrR
                     /protein_id="ALP32107.1"
                     /translation="MKTKRVTIKDIAELAGVSKATASLVLNGRGKELRVAQETRERVLA
                     IIKELGYVPNRAARTLVTRRTDTVALVVSENNQKLFAEPFYAGIVLGVGVALSERGFQF
                     VLATGRSGIEHERLGGYLAGQHVDGVLLLSLHRDDPLPQMLDEAGVPYVYGGRPLGVPE
                     EQVSYVDIDNIGGGRQATQRLIETGHRRIATIAGPQDMVAGVERLQGYREALLAAGMEY
                     DETLVSYGDFTYDSGVAAMRELLDRAPDVDAVFAASDLMGLAALRVLRASGRRVPEDVA
                     VVGYDDSTVAEHAEPPMTSVNQPTELMGREMARLLVDRITGETTEPVRLVLETHLMVRE
                     SG"
     regulatory      complement(3784..3809)
                     /label=RBS
                     /note="RBS"
                     /regulatory_class="ribosome_binding_site"
     regulatory      complement(3841..3869)
                     /label=pLlacO
                     /note="pLlacO"
                     /regulatory_class="promoter"
     terminator      3905..3991
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     rep_origin      complement(4155..4743)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(4831..4925)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(4959..5750)
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
ORIGIN
        1 gacgtctgtg caagtactag tatccatgga acgggcggcc gtccctatca gtgatagaga
       61 ttgacatccc tatcagtgat agagatactg agcacatcag caggacgcac tgaccggatc
      121 catgaaacca gtaacgttat acgatgtcgc agagtatgcc ggtgtctctt atcagaccgt
      181 ttcccgcgtg gtgaaccagg ccagccacgt ttctgcgaaa acgcgggaaa aagtggaagc
      241 ggcgatggag tctcttagct atcacccgaa cgccaacgcc cgtgcgctgg cgcagcagac
      301 cactgaaacg gtcggtctgg tcgttggtga tgtttccgat ccgtttttcg gtgcaatggt
      361 gaaagcggtc gaacaggtgg cttatcacac cggtaatttt ttattgattg gcaacggtta
      421 ccacaacgaa caaaaagagc gtcaggccat tgagcaactg atccgccatc gctgtgctgc
      481 gttggtcgtc catgccaaaa tgatcccgga tgctgattta gcctcattaa tgaaacaaat
      541 gcccggtatg gtgctgatca accgtatcct gcctggcttt gaaaaccgtt gtattgctct
      601 ggacgatcgt tacggtgcct ggctggcaac gcgtcattta attcagcaag gtcatacccg
      661 cattggttat ctgtgctcta accactctat ttctgacgcc gaagatcgtc tgcaagggta
      721 ttacgatgcc cttgctgaaa gtggtattgc ggccaatgac cggctggtga catttggcga
      781 accagacgaa agcggcggcg aacaggcaat gaccgagctt ttgggacgag gaagaaattt
      841 cactgcggta gcctgttata acgattcaat ggcggcgggt gcgatgggcg ttctcaatga
      901 taatggtatt gatgtaccgg gtgagatttc gttaattggc tttgatgatg tgctggtgtc
      961 acgctatgtg cgtccgcgcc tgaccaccgt gcgttaccca atcgtgacga tggcgaccca
     1021 ggctgccgaa ctggctttgg cgctggcgga taatcgccct ctcccggaaa tcactaatgt
     1081 ctttagtccg acgctggtac gtcgtcattc agtgtcaact ccgtcgctgg aggcaagtca
     1141 tcatgcaacc agcgactaat gtacacgaga attggcttgg actcctgttg atagatccag
     1201 taatgacctc agaactccat ctggatttgt tcagaacgct cggttgccgc cgggcgtttt
     1261 ttattggtga gaatccaagc agtagtcacc gcgatctgaa gtatccatgg aacgggcggc
     1321 cgtccctatc agtgatagag attgacatcc ctatcagtga tagagatact gagcacatca
     1381 gcaggacgca ctgaccggat ccatgaaacc agtaacgtta tacgatgtcg cagagtatgc
     1441 cggtgtctct tatcagaccg tttcccgcgt ggtgaaccag gccagccacg tttctgcgaa
     1501 aacgcgggaa aaagtggaag cggcgatggc ggagctgaat tacattccca accgcgtggc
     1561 acaacaactg gcgggcaaac agtcgttgct gattggcgtt gccacctcca gtctggccct
     1621 gcacgcgccg tcgcaaattg tcgcggcgat taaatctcgc gccgatcaac tgggtgccag
     1681 cgtggtggtg tcgatggtag aacgaagcgg cgtcgaagcc tgtaaagcgg cggtgcacaa
     1741 tcttctcgcg caacgcgtca gtgggctgat cattaactat ccgctggatg accaggatgc
     1801 cattgctgtg gaagctgcct gcactaatgt tccggcgtta tttcttgatg tctctgacca
     1861 gacacccatc aacagtatta ttttctccca tgaagacggt acgcgactgg gcgtggagca
     1921 tctggtcgca ttgggtcacc agcaaatcgc gctgttagcg ggcccattaa gttctgtctc
     1981 ggcgcgtctg cgtctggctg gctggcataa atatctcact cgcaatcaaa ttcagccgat
     2041 agcggaacgg gaaggcgact ggagtgccat gtccggtttt caacaaacca tgcaaatgct
     2101 gaatgagggc atcgttccca ctgcgatgct ggttgccaac gatcagatgg cgctgggcgc
     2161 aatgcgcgcc attaccgagt ccgggctgcg cgttggtgcg gatatctcgg tagtgggata
     2221 cgacgatacc gaagacagct catgttatat cccgccgtta accaccatca aacaggattt
     2281 tcgcctgctg gggcaaacca gcgtggaccg cttgctgcaa ctctctcagg gccaggcggt
     2341 gaagggcaat cagctgttgc ccgtctcact ggtgaaaaga aaaaccaccc tggcgcccaa
     2401 tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgg cacgacaggt
     2461 ttcccgactg gaaagcgggc agtgatgtac acgagaattg gcttggactc ctgttgatag
     2521 atccagtaat gacctcagaa ctccatctgg atttgttcag aacgctcggt tgccgccggg
     2581 cgttttttat tggtgagaat ccaagcagta gtcaccgcgc tgcagtgact actgcttgga
     2641 ttctcaccaa taaaaaacgc ccggcggcaa ccgagcgttc tgaacaaatc cagatggagt
     2701 tctgaggtca ttactggatc tatcaacagg agtccaagcc aattctcgcc cgggttaccc
     2761 actttcgcga accattaaat gagtttcgag taccaaacga accggttccg tagtttcgcc
     2821 tgtaatacga tctacaagca aacgagccat ctcacgaccc attaattctg tcggctggtt
     2881 gacgctcgtc atggggggtt ccgcgtgttc cgccaccgtc gaatcgtcgt acccaacgac
     2941 cgcgacatct tcaggcacac ggcgccccga tgcacgcagt acacgtaacg ccgccagccc
     3001 cataagatca gatgctgcga atacggcatc cacatccggt gcgcggtcca gaagttcgcg
     3061 catggcggcg acgcccgaat catacgtaaa gtcaccatat gatactaatg tttcatcgta
     3121 ttccatgccc gcggctaata atgcctcacg gtatccttgc agacgctcta ctcctgccac
     3181 catatcttgc gggccggcaa tagtcgcgat acgacgatga cccgtttcga tcagacgctg
     3241 ggtagcctgg cgcccgccac caatattgtc gatatcgaca tagctgacct gctcttccgg
     3301 cacacccagc gggcggccac catatacata cggtactcca gcttcgtcca gcatctgcgg
     3361 cagcggatcg tcgcggtgca aactaagaag cagcacgccg tccacatgct ggcctgccag
     3421 atagcccccc aggcgttcgt gctcaatgcc cgaacgaccc gtagccagga cgaactggaa
     3481 accacgttca ctcagagcca caccgacacc cagcacgata cctgcataaa acggttccgc
     3541 aaacagtttc tggttatttt cggacaccac caaagccacg gtgtccgtgc ggcgtgtaac
     3601 caatgtgcgt gccgcgcgat tcgggacgta accgagttct ttgataatag ccagcacgcg
     3661 ctcacgggtt tcctgggcga cacgcagttc tttaccacga ccattaagaa caagacttgc
     3721 cgttgcttta ctcaccccag ctaattccgc gatatcttta atggttacgc gtttcgtctt
     3781 catcttaaga tcctataaaa tggctagagc taggggtcag tgcgtcctgc tgatgtgctc
     3841 agtatcttgt gagcgctcac aattgtcaac tcgagagaat caatattaat ccaacgcgtg
     3901 gcatcaaata aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt
     3961 gtcggtgaac gctctcctga gtaggacaaa tccgccgccc tagacttagg cgttcggctg
     4021 cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga atcaggggat
     4081 aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg taaaaaggcc
     4141 gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa aaatcgacgc
     4201 tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctgga
     4261 agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct gtccgccttt
     4321 ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct cagttcggtg
     4381 taggtcgttc gctccaagct gggctgtctg cacgaacccc ccgttcagcc cgaccgctgc
     4441 gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt atcgccactg
     4501 gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc tacagagttc
     4561 ttgaagtggt ggcctaacta cggctacact agaaggacag tatttggtat ctgcgctctg
     4621 ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa acaaaccacc
     4681 gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa aaaaggatct
     4741 caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga aaactcacgt
     4801 taagggattt tggtcatggc tagtgcttgg attctcacca ataaaaaacg cccggcggca
     4861 accgagcgtt ctgaacaaat ccagatggag ttctgaggtc attactggat ctatcaacag
     4921 gagtccaagc caattctcga accccagagt cccgctcaga agaactcgtc aagaaggcga
     4981 tagaaggcga tgcgctgcga atcgggagcg gcgataccgt aaagcacgag gaagcggtca
     5041 gcccattcgc cgccaagctc ttcagcaata tcacgggtag ccaacgctat gtcctgatag
     5101 cggtccgcca cacccagccg gccacagtcg atgaatccag aaaagcggcc attttccacc
     5161 atgatattcg gcaagcaggc atcgccgtgg gtcacgacga gatcctcgcc gtcgggcata
     5221 cgcgccttga gcctggcgaa cagttcggct ggcgcgagcc cctgatgctc ttcgtccaga
     5281 tcatcctgat cgacaagacc ggcttccatc cgagtacgtg ctcgctcgat gcgatgtttc
     5341 gcttggtggt cgaatgggca ggtagccgga tcaagcgtat gcagccgccg cattgcatca
     5401 gccatgatgg atactttctc ggcaggagca aggtgagatg acaggagatc ctgccccggc
     5461 acttcgccca atagcagcca gtcccttccc gcttcagtga caacgtcgag cacagccgcg
     5521 caaggaacgc ccgtcgtggc cagccacgat agccgcgctg cctcgtcttg cagttcattc
     5581 agggcaccgg acaggtcggt cttgacaaaa agaaccgggc gaccctgcgc tgacagccgg
     5641 aacacggcgg catcagagca gccgattgtc tgttgtgccc agtcatagcc gaatagcctc
     5701 tccacccaag ccgccggaga acctgcgtgc aatccatctt gttcaatcat gcgaaacgat
     5761 cctcatcctg tctcttgatc agatcttgat cccctgcgcc atcagatcct tggcggcaag
     5821 aaagccatcc agtttacttt gcagggcttc ccaaccttac cagagggcgg cccaactggc
     5881 aattcc
//
