LOCUS       40924_34096        5887 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pPasscode2, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5887)
  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 5887)
  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 5887)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 5887)
  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..5887
                     /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..1135
                     /codon_start=1
                     /product="CelR-LacI"
                     /label=CelR-LacI
                     /protein_id="ALP32101.1"
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAIKEL
                     GYVPNRAARTLVTRRTDTVALVVSENNQKLFAEPFYAGIVLGVGVALSERGFQFVLATG
                     RSGIEHERLGGYLAGQHVDGVLLLSLHRDDPLPQMLDEAGVPYVYGGRPLGVPEEQVSY
                     VDIDNIGGGRQATQRLIETGHRRIATIAGPQDMVAGVERLQGYREALLAAGMEYDETLV
                     SYGDFTYDSGVAAMRELLDRAPDVDAVFAASDLMGLAALRVLRASGRRVPEDVAVVGYD
                     DSTVAEHAEPPMTSVNQPTELMGREMARLLVDRITGETTEPVRLVLETHLMVRESG"
     terminator      1156..1250
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     promoter        1299..1372
                     /label=PLtetO-1 promoter
                     /note="modified phage lambda PL promoter with tet operator
                     sites (Lutz and Bujard, 1997)"
     regulatory      1299..1346
                     /label=pLtetO-1
                     /note="pLtetO-1"
                     /regulatory_class="promoter"
     protein_bind    1299..1317
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1324..1342
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     regulatory      1353..1378
                     /label=RBS
                     /note="RBS"
                     /regulatory_class="ribosome_binding_site"
     CDS             1379..2458
                     /label=lacI
                     /note="lac repressor"
     regulatory      2482..2576
                     /label=T0
                     /note="T0"
                     /regulatory_class="terminator"
     terminator      2482..2576
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     regulatory      complement(2616..2710)
                     /label=T0
                     /note="T0"
                     /regulatory_class="terminator"
     terminator      2616..2710
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(2731..3783)
                     /codon_start=1
                     /product="GalR-ScrR"
                     /label=GalR-ScrR
                     /protein_id="ALP32103.1"
                     /translation="MKTKRVTIKDIAELAGVSKATASLVLNGRGKELRVAQETRERVLA
                     IMESLSYHPNANARALAQQTTETVGLVVGDVSDPFFGAMVKAVEQVAYHTGNFLLIGNG
                     YHNEQKERQAIEQLIRHRCAALVVHAKMIPDADLASLMKQMPGMVLINRILPGFENRCI
                     ALDDRYGAWLATRHLIQQGHTRIGYLCSNHSISDAEDRLQGYYDALAESGIAANDRLVT
                     FGEPDESGGEQAMTELLGRGRNFTAVACYNDSMAAGAMGVLNDNGIDVPGEISLIGFDD
                     VLVSRYVRPRLTTVRYPIVTMATQAAELALALADNRPLPEITNVFSPTLVRRHSVSTPS
                     LEASHHATSD"
     regulatory      complement(3784..3809)
                     /label=RBS
                     /note="RBS"
                     /regulatory_class="ribosome_binding_site"
     regulatory      complement(3842..3870)
                     /label=pLlacO
                     /note="pLlacO"
                     /regulatory_class="promoter"
     terminator      3906..3992
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     rep_origin      complement(4156..4744)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     terminator      complement(4832..4926)
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     CDS             complement(4960..5751)
                     /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 ggcgatcaaa gagctgggct acgtgccgaa ccgcgcagcc cgcaccctgg tcacccgacg
      301 taccgacacc gtagccctgg tggtgtcgga aaacaaccag aagctcttcg ccgaaccctt
      361 ctatgccggg atcgtgctcg gcgtgggggt tgctctgtcc gaacggggat tccagttcgt
      421 cctggccacg ggccgctccg ggatagagca tgagcggctg ggcggctacc tggccggaca
      481 gcacgtcgac ggggtcctcc tgctgtcgct ccaccgcgac gacccgctgc cgcagatgct
      541 ggacgaggcc ggggtgccgt acgtctacgg cggccgtccg ctcggcgtcc ccgaagaaca
      601 ggtgtcctat gtcgatatcg acaacatcgg cgggggacgc caggccaccc agcggctgat
      661 cgagaccggg caccggcgga tcgctacgat cgcgggcccg caggacatgg tcgctggtgt
      721 ggaacgcctc caggggtatc gcgaagcact gctcgccgcg gggatggagt acgacgagac
      781 gctggtgagc tacggtgact tcacctacga cagcggggtg gccgcgatgc gggagctgct
      841 ggatcgggcc cccgacgtgg acgccgtgtt cgcggcctcc gacttgatgg ggctggccgc
      901 gctgcgggtg ctgcgtgctt cgggacgccg cgtgcccgag gatgtggcgg tggtcggcta
      961 cgacgactcg accgtagccg agcacgccga accgccgatg accagcgtca accagcccac
     1021 cgagctgatg ggccgggaga tggcccggct gctcgtggac cggatcaccg gggagaccac
     1081 cgaaccggtg cggctggtgc tggagaccca tttgatggtg cgggaatccg ggtgatgtac
     1141 acgagaattg gcttggactc ctgttgatag atccagtaat gacctcagaa ctccatctgg
     1201 atttgttcag aacgctcggt tgccgccggg cgttttttat tggtgagaat ccaagcagta
     1261 gtcaccgcga tctgaagtat ccatggaacg ggcggccgtc cctatcagtg atagagattg
     1321 acatccctat cagtgataga gatactgagc acatcagcag gacgcactga ccggatccat
     1381 gaaaccagta acgttatacg atgtcgcaga gtatgccggt gtctcttatc agaccgtttc
     1441 ccgcgtggtg aaccaggcca gccacgtttc tgcgaaaacg cgggaaaaag tggaagcggc
     1501 gatggcggag ctgaattaca ttcccaaccg cgtggcacaa caactggcgg gcaaacagtc
     1561 gttgctgatt ggcgttgcca cctccagtct ggccctgcac gcgccgtcgc aaattgtcgc
     1621 ggcgattaaa tctcgcgccg atcaactggg tgccagcgtg gtggtgtcga tggtagaacg
     1681 aagcggcgtc gaagcctgta aagcggcggt gcacaatctt ctcgcgcaac gcgtcagtgg
     1741 gctgatcatt aactatccgc tggatgacca ggatgccatt gctgtggaag ctgcctgcac
     1801 taatgttccg gcgttatttc ttgatgtctc tgaccagaca cccatcaaca gtattatttt
     1861 ctcccatgaa gacggtacgc gactgggcgt ggagcatctg gtcgcattgg gtcaccagca
     1921 aatcgcgctg ttagcgggcc cattaagttc tgtctcggcg cgtctgcgtc tggctggctg
     1981 gcataaatat ctcactcgca atcaaattca gccgatagcg gaacgggaag gcgactggag
     2041 tgccatgtcc ggttttcaac aaaccatgca aatgctgaat gagggcatcg ttcccactgc
     2101 gatgctggtt gccaacgatc agatggcgct gggcgcaatg cgcgccatta ccgagtccgg
     2161 gctgcgcgtt ggtgcggata tctcggtagt gggatacgac gataccgaag acagctcatg
     2221 ttatatcccg ccgttaacca ccatcaaaca ggattttcgc ctgctggggc aaaccagcgt
     2281 ggaccgcttg ctgcaactct ctcagggcca ggcggtgaag ggcaatcagc tgttgcccgt
     2341 ctcactggtg aaaagaaaaa ccaccctggc gcccaatacg caaaccgcct ctccccgcgc
     2401 gttggccgat tcattaatgc agctggcacg acaggtttcc cgactggaaa gcgggcagtg
     2461 atgtacacga gaattggctt ggactcctgt tgatagatcc agtaatgacc tcagaactcc
     2521 atctggattt gttcagaacg ctcggttgcc gccgggcgtt ttttattggt gagaatccaa
     2581 gcagtagtca ccgcgctgca gtgactactg cttggattct caccaataaa aaacgcccgg
     2641 cggcaaccga gcgttctgaa caaatccaga tggagttctg aggtcattac tggatctatc
     2701 aacaggagtc caagccaatt ctcgcccggg ttagtcgctg gttgcatgat gacttgcctc
     2761 cagcgacgga gttgacactg aatgacgacg taccagcgtc ggactaaaga cattagtgat
     2821 ttccgggaga gggcgattat ccgccagcgc caaagccagt tcggcagcct gggtcgccat
     2881 cgtcacgatt gggtaacgca cggtggtcag gcgcggacgc acatagcgtg acaccagcac
     2941 atcatcaaag ccaattaacg aaatctcacc cggtacatca ataccattat cattgagaac
     3001 gcccatcgca cccgccgcca ttgaatcgtt ataacaggct accgcagtga aatttcttcc
     3061 tcgtcccaaa agctcggtca ttgcctgttc gccgccgctt tcgtctggtt cgccaaatgt
     3121 caccagccgg tcattggccg caataccact ttcagcaagg gcatcgtaat acccttgcag
     3181 acgatcttcg gcgtcagaaa tagagtggtt agagcacaga taaccaatgc gggtatgacc
     3241 ttgctgaatt aaatgacgcg ttgccagcca ggcaccgtaa cgatcgtcca gagcaataca
     3301 acggttttca aagccaggca ggatacggtt gatcagcacc ataccgggca tttgtttcat
     3361 taatgaggct aaatcagcat ccgggatcat tttggcatgg acgaccaacg cagcacagcg
     3421 atggcggatc agttgctcaa tggcctgacg ctctttttgt tcgttgtggt aaccgttgcc
     3481 aatcaataaa aaattaccgg tgtgataagc cacctgttcg accgctttca ccattgcacc
     3541 gaaaaacgga tcggaaacat caccaacgac cagaccgacc gtttcagtgg tctgctgcgc
     3601 cagcgcacgg gcgttggcgt tcgggtgata gctaagagac tccatgatcg ccagcacgcg
     3661 ctcgcgcgtc tcctgcgcca cgcgcagctc tttgccacgg ccgttgagca ccagactggc
     3721 ggtcgctttg gagacgcccg ccagttcggc gatatctttg atagttacgc gtttggtttt
     3781 catcttaagc ctcctcttga ctcttgaccc ctaggggtca gtgcgtcctg ctgatgtgct
     3841 cagtatcttg tgagcgctca caattgtcaa ctcgagagaa tcaatattaa tccaacgcgt
     3901 ggcatcaaat aaaacgaaag gctcagtcga aagactgggc ctttcgtttt atctgttgtt
     3961 tgtcggtgaa cgctctcctg agtaggacaa atccgccgcc ctagacttag gcgttcggct
     4021 gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga
     4081 taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc
     4141 cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg
     4201 ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg
     4261 aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt
     4321 tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt
     4381 gtaggtcgtt cgctccaagc tgggctgtct gcacgaaccc cccgttcagc ccgaccgctg
     4441 cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact
     4501 ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt
     4561 cttgaagtgg tggcctaact acggctacac tagaaggaca gtatttggta tctgcgctct
     4621 gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac
     4681 cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc
     4741 tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg
     4801 ttaagggatt ttggtcatgg ctagtgcttg gattctcacc aataaaaaac gcccggcggc
     4861 aaccgagcgt tctgaacaaa tccagatgga gttctgaggt cattactgga tctatcaaca
     4921 ggagtccaag ccaattctcg aaccccagag tcccgctcag aagaactcgt caagaaggcg
     4981 atagaaggcg atgcgctgcg aatcgggagc ggcgataccg taaagcacga ggaagcggtc
     5041 agcccattcg ccgccaagct cttcagcaat atcacgggta gccaacgcta tgtcctgata
     5101 gcggtccgcc acacccagcc ggccacagtc gatgaatcca gaaaagcggc cattttccac
     5161 catgatattc ggcaagcagg catcgccgtg ggtcacgacg agatcctcgc cgtcgggcat
     5221 acgcgccttg agcctggcga acagttcggc tggcgcgagc ccctgatgct cttcgtccag
     5281 atcatcctga tcgacaagac cggcttccat ccgagtacgt gctcgctcga tgcgatgttt
     5341 cgcttggtgg tcgaatgggc aggtagccgg atcaagcgta tgcagccgcc gcattgcatc
     5401 agccatgatg gatactttct cggcaggagc aaggtgagat gacaggagat cctgccccgg
     5461 cacttcgccc aatagcagcc agtcccttcc cgcttcagtg acaacgtcga gcacagccgc
     5521 gcaaggaacg cccgtcgtgg ccagccacga tagccgcgct gcctcgtctt gcagttcatt
     5581 cagggcaccg gacaggtcgg tcttgacaaa aagaaccggg cgaccctgcg ctgacagccg
     5641 gaacacggcg gcatcagagc agccgattgt ctgttgtgcc cagtcatagc cgaatagcct
     5701 ctccacccaa gccgccggag aacctgcgtg caatccatct tgttcaatca tgcgaaacga
     5761 tcctcatcct gtctcttgat cagatcttga tcccctgcgc catcagatcc ttggcggcaa
     5821 gaaagccatc cagtttactt tgcagggctt cccaacctta ccagagggcg gcccaactgg
     5881 caattcc
//
