LOCUS       40924_5549        8265 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pBAC-TC, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8265)
  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 8265)
  AUTHORS   Chan CTY., Lee JW, Cameron DE, Bashor CJ, Collins JJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (12-OCT-2015) Biological engineering, MIT, 45 Carleton St,
            MIT Building E25-Rm302, Cambridge, MA 02142, USA
REFERENCE   3  (bases 1 to 8265)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8265)
  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) Biological engineering, MIT, 45 Carleton St, MIT 
            Building E25-Rm302, Cambridge, MA 02142, 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..8265
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(105..1073)
                     /codon_start=1
                     /label=sopB
                     /note="partitioning protein for the bacterial F plasmid"
                     /translation="MKRAPVIPKHTLNTQPVEDTSLSTPAAPMVDSLIARVGVMARGNA
                     ITLPVCGRDVKFTLEVLRGDSVEKTSRVWSGNERDQELLTEDALDDLIPSFLLTGQQTP
                     AFGRRVSGVIEIADGSRRRKAAALTESDYRVLVGELDDEQMAALSRLGNDYRPTSAYER
                     GQRYASRLQNEFAGNISALADAENISRKIITRCINTAKLPKSVVALFSHPGELSARSGD
                     ALQKAFTDKEELLKQQASNLHEQKKAGVIFEAEEVITLLTSVLKTSSASRTSLSSRHQF
                     APGATVLYKGDKMVLNLDRSRVPTECIEKIEAILKELEKPAP"
     CDS             complement(1076..2248)
                     /codon_start=1
                     /label=sopA
                     /note="partitioning protein for the bacterial F plasmid"
                     /translation="MFRMKLMETLNQCINAGHEMTKAIAIAQFNDDSPEARKITRRWRI
                     GEAADLVGVSSQAIRDAEKAGRLPHPDMEIRGRVEQRVGYTIEQINHMRDVFGTRLRRA
                     EDVFPPVIGVAAHKGGVYKTSVSVHLAQDLALKGLRVLLVEGNDPQGTASMYHGWVPDL
                     HIHAEDTLLPFYLGEKDDVTYAIKPTCWPGLDIIPSCLALHRIETELMGKFDEGKLPTD
                     PHLMLRLAIETVAHDYDVIVIDSAPNLGIGTINVVCAADVLIVPTPAELFDYTSALQFF
                     DMLRDLLKNVDLKGFEPDVRILLTKYSNSNGSQSPWMEEQIRDAWGSMVLKNVVRETDE
                     VGKGQIRMRTVFEQAIDQRSSTGAWRNALSIWEPVCNEIFDRLIKPRWEIR"
     misc_feature    complement(2574..2824)
                     /label=incC
                     /note="incompatibility region of the bacterial F plasmid"
     CDS             complement(2830..3582)
                     /codon_start=1
                     /label=repE
                     /note="replication initiation protein for the bacterial F 
                     plasmid"
                     /translation="MAETAVINHKKRKNSPRIVQSNDLTEAAYSLSRDQKRMLYLFVDQ
                     IRKSDGTLQEHDGICEIHVAKYAEIFGLTSAEASKDIRQALKSFAGKEVVFYRPEEDAG
                     DEKGYESFPWFIKRAHSPSRGLYSVHINPYLIPFFIGLQNRFTQFRLSETKEITNPYAM
                     RLYESLCQYRKPDGSGIVSLKIDWIIERYQLPQSYQRMPDFRRRFLQVCVNEINSRTPM
                     RLSYIEKKKGRQTTHIVFSFRDITSMTTG"
     rep_origin      complement(3673..3892)
                     /direction=LEFT
                     /label=ori2
                     /note="secondary origin of replication for the bacterial F 
                     plasmid; also known as oriS"
     rep_origin      complement(3968..4582)
                     /direction=LEFT
                     /label=oriV
                     /note="origin of replication for the bacterial F plasmid"
     CDS             5606..6397
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     promoter        6632..6661
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and
                     lac UV5 promoters"
     protein_bind    6669..6685
                     /label=lac operator
                     /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)."
     CDS             6732..7352
                     /codon_start=1
                     /label=TetR
                     /note="tetracycline repressor TetR"
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     CDS             7353..8060
                     /codon_start=1
                     /label=mCherry
                     /note="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     terminator      8123..8209
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
ORIGIN
        1 aagtggggcc cagagagaat attcaggcca gttatgcttt ctggcctgta acaaaggaca
       61 ttaagtaaag acagataaac gtagactaaa acgtggtcgc atcagggtgc tggcttttca
      121 agttccttaa gaatggcctc aattttctct atacactcag ttggaacacg ggacctgtcc
      181 aggttaagca ccattttatc gcccttatac aatactgtcg ctccaggagc aaactgatgt
      241 cgtgagctta aactagttct tgatgcagat gacgttttaa gcacagaagt taaaagagtg
      301 ataacttctt cagcttcaaa tatcacccca gcttttttct gctcatgaag gttagatgcc
      361 tgctgcttaa gtaattcctc tttatctgta aaggcttttt gaagtgcatc acctgaccgg
      421 gcagatagtt caccggggtg agaaaaaaga gcaacaactg atttaggcaa tttggcggtg
      481 ttgatacagc gggtaataat cttacgtgaa atattttccg catcagccag cgcagaaata
      541 tttccagcaa attcattctg caatcggctt gcataacgct gaccacgttc ataagcactt
      601 gttgggcgat aatcgttacc caatctggat aatgcagcca tctgctcatc atccagctcg
      661 ccaaccagaa cacgataatc actttcggta agtgcagcag ctttacgacg gcgactccca
      721 tcggcaattt ctatgacacc agatactctt cgaccgaacg ccggtgtctg ttgaccagtc
      781 agtagaaaag aagggatgag atcatccagt gcgtcctcag taagcagctc ctggtcacgt
      841 tcattacctg accatacccg agaggtcttc tcaacactat caccccggag cacttcaaga
      901 gtaaacttca catcccgacc acatacaggc aaagtaatgg cattaccgcg agccattact
      961 cctacgcgcg caattaacga atccaccatc ggggcagctg gtgtcgataa cgaagtatct
     1021 tcaaccggtt gagtattgag cgtatgtttt ggaataacag gcgcacgctt cattatctaa
     1081 tctcccagcg tggtttaatc agacgatcga aaatttcatt gcagacaggt tcccaaatag
     1141 aaagagcatt tctccaggca ccagttgaag agcgttgatc aatggcctgt tcaaaaacag
     1201 ttctcatccg gatctgacct ttaccaactt catccgtttc acgtacaaca ttttttagaa
     1261 ccatgcttcc ccaggcatcc cgaatttgct cctccatcca cggggactga gagccattac
     1321 tattgctgta tttggtaagc aaaatacgta catcaggctc gaacccttta agatcaacgt
     1381 tcttgagcag atcacgaagc atatcgaaaa actgcagtgc ggaggtgtag tcaaacaact
     1441 cagcaggcgt gggaacaatc agcacatcag cagcacatac gacattaatc gtgccgatac
     1501 ccaggttagg cgcgctgtca ataactatga catcatagtc atgagcaaca gtttcaatgg
     1561 ccagtcggag catcaggtgt ggatcggtgg gcagtttacc ttcatcaaat ttgcccatta
     1621 actcagtttc aatacggtgc agagccagac aggaaggaat aatgtcaagc cccggccagc
     1681 aagtgggctt tattgcataa gtgacatcgt ccttttcccc aagatagaaa ggcaggagag
     1741 tgtcttctgc atgaatatga agatctggta cccatccgtg atacattgag gctgttccct
     1801 gggggtcgtt accttccacg agcaaaacac gtagcccctt cagagccaga tcctgagcaa
     1861 gatgaacaga aactgaggtt ttgtaaacgc cacctttatg ggcagcaacc ccgatcaccg
     1921 gtggaaatac gtcttcagca cgtcgcaatc gcgtaccaaa cacatcacgc atatgattaa
     1981 tttgttcaat tgtataacca acacgttgct caacccgtcc tcgaatttcc atatccgggt
     2041 gcggtagtcg ccctgctttc tcggcatctc tgatagcctg agaagaaacc ccaactaaat
     2101 ccgctgcttc acctattctc cagcgccggg ttattttcct cgcttccggg ctgtcatcat
     2161 taaactgtgc aatggcgata gccttcgtca tttcatgacc agcgtttatg cactggttaa
     2221 gtgtttccat gagtttcatt ctgaacatcc tttaatcatt gctttgcgtt tttttattaa
     2281 atcttgcaat ttactgcaaa gcaacaacaa aatcgcaaag tcatcaaaaa accgcaaagt
     2341 tgtttaaaat aagagcaaca ctacaaaagg agataagaag agcacatacc tcagtcactt
     2401 attatcacta gcgctcgccg cagccgtgta accgagcata gcgagcgaac tggcgaggaa
     2461 gcaaagaaga actgttctgt cagatagctc ttacgctcag cgcaagaaga aatatccacc
     2521 gtgggaaaaa ctccaggtag aggtacacac gcggatagcc aattcagagt aataaactgt
     2581 gataatcaac cctcatcaat gatgacgaac taacccccga tatcaggtca catgacgaag
     2641 ggaaagagaa ggaaatcaac tgtgacaaac tgccctcaaa tttggcttcc ttaaaaatta
     2701 cagttcaaaa agtatgagaa aatccatgca ggctgaagga aacagcaaaa ctgtgacaaa
     2761 ttaccctcag taggtcagaa caaatgtgac gaaccaccct caaatctgtg acagataacc
     2821 ctcagactat cctgtcgtca tggaagtgat atcgcggaag gaaaatacga tatgagtcgt
     2881 ctggcggcct ttctttttct caatgtatga gaggcgcatt ggagttctgc tgttgatctc
     2941 attaacacag acctgcagga agcggcggcg gaagtcaggc atacgctggt aactttgagg
     3001 cagctggtaa cgctctatga tccagtcgat tttcagagag acgatgcctg agccatccgg
     3061 cttacgatac tgacacaggg attcgtataa acgcatggca tacggattgg tgatttcttt
     3121 tgtttcacta agccgaaact gcgtaaaccg gttctgtaac ccgataaaga agggaatgag
     3181 atatgggttg atatgtacac tgtaaagccc tctggatgga ctgtgcgcac gtttgataaa
     3241 ccaaggaaaa gattcatagc ctttttcatc gccggcatcc tcttcagggc gataaaaaac
     3301 cacttccttc cccgcgaaac tcttcaatgc ctgccgtata tccttactgg cttccgcaga
     3361 ggtcaatccg aatatttcag catatttagc aacatggatc tcgcagatac cgtcatgttc
     3421 ctgtagggtg ccatcagatt ttctgatctg gtcaacgaac agatacagca tacgtttttg
     3481 atcccgggag agactatatg ccgcctcagt gaggtcgttt gactggacga ttcgcgggct
     3541 atttttacgt ttcttgtgat tgataaccgc tgtttccgcc atgacagatc catgtgaagt
     3601 gtgacaagtt tttagattgt cacactaaat aaaaaagagt caataagcag ggataacttt
     3661 gtgaaaaaac agcttcttct gagggcaatt tgtcacaggg ttaagggcaa tttgtcacag
     3721 acaggactgt catttgaggg tgatttgtca cactgaaagg gcaatttgtc acaacacctt
     3781 ctctagaacc agcatggata aaggcctaca aggcgctcta aaaaagaaga tctaaaaact
     3841 ataaaaaaaa taattataaa aatatccccg tggataagtg gataacccca agggaagttt
     3901 tttcaggcat cgtgtgtaag cagaatatat aagtgctgtt ccctggtgct tcctcgctca
     3961 ctcgaccggg agggttcgag aagggggggc accccccttc ggcgtgcgcg gtcacgcgca
     4021 cagggcgcag ccctggttaa aaacaaggtt tataaatatt ggtttaaaag caggttaaaa
     4081 gacaggttag cggtggccga aaaacgggcg gaaacccttg caaatgctgg attttctgcc
     4141 tgtggacagc ccctcaaatg tcaataggtg cgcccctcat ctgtcagcac tctgcccctc
     4201 aagtgtcaag gatcgcgccc ctcatctgtc agtagtcgcg cccctcaagt gtcaataccg
     4261 cagggcactt atccccaggc ttgtccacat catctgtggg aaactcgcgt aaaatcaggc
     4321 gttttcgccg atttgcgagg ctggccagct ccacgtcgcc ggccgaaatc gagcctgccc
     4381 ctcatctgtc aacgccgcgc cgggtgagtc ggcccctcaa gtgtcaacgt ccgcccctca
     4441 tctgtcagtg agggccaagt tttccgcgag gtatccacaa cgccggcggc cggccgcggt
     4501 gtctcgcaca cggcttcgac ggcgtttctg gcgcgtttgc agggccatag acggccgcca
     4561 gcccagcggc gagggcaacc agccgagggc ttcgccctgt cgctcgactg cggcgagcac
     4621 tactggctgt aaaaggacag accacatcat ggttctgtgt tcattaggtt gttctgtcca
     4681 ttgctgacat aatccgctcc acttcaacgt aacaccgcac gaagatttct attgttcctg
     4741 aaggcatatt caaatcgttt tcgttaccgc ttgcaggcat catgacagaa cactacttcc
     4801 tataaacgct acacaggctc ctgagattaa taatgcggat ctctacgata atgggagatt
     4861 ttcccgactg tttcgttcgc ttctcagtgg ataacagcca gcttctctgt ttaacagaca
     4921 aaaacagcat atccactcag ttccacattt ccatataaag gccaaggcat ttattctcag
     4981 gataattgtt tcagcatcgc aaccgcatca gactccggca tcgcaaactg cacccggtgc
     5041 cgggcagcca catccagcgc aaaaaccttc gtgtagactt ccgttgaact gatggactta
     5101 tgtcccatca ggctttgcag aactttcagc ggtataccgg catacagcat gtgcatcgca
     5161 taggaatggc ggaacgtatg tggtgtgacc ggaacagaga acgtcacacc gtcagcagca
     5221 gcggcggcaa ccgcctcccc aatccaggtc ctgaccgttc tgtccgtcac ttcccagatc
     5281 cgcgctttct ctgtccttcc tgtgcgacgg ttacgccgct ccatgagctt atcgcgaata
     5341 aatacctgtg acggaagatc acttcgcaga ataaataaat cctggtgtcc ctgttgatac
     5401 cgggaagccc tgggccaact tttggcgaaa atgagacgtt gacagaacag tagtacttgc
     5461 acagacgtcg gaattgccag ttgggccgcc ctctggtaag gttgggaagc cctgcaaagt
     5521 aaactggatg gctttcttgc cgccaaggat ctgatggcgc aggggatcaa gatctgatca
     5581 agagacagga tgaggatcgt ttcgcatgat tgaacaagat ggattgcacg caggttctcc
     5641 ggcggcttgg gtggagaggc tattcggcta tgactgggca caacagacaa tcggctgctc
     5701 tgatgccgcc gtgttccggc tgtcagcgca gggtcgcccg gttctttttg tcaagaccga
     5761 cctgtccggt gccctgaatg aactgcaaga cgaggcagcg cggctatcgt ggctggccac
     5821 gacgggcgtt ccttgcgcgg ctgtgctcga cgttgtcact gaagcgggaa gggactggct
     5881 gctattgggc gaagtgccgg ggcaggatct cctgtcatct caccttgctc ctgccgagaa
     5941 agtatccatc atggctgatg caatgcggcg gctgcatacg cttgatccgg ctacctgccc
     6001 attcgaccac caagcgaaac atcgcatcga gcgagcacgt actcggatgg aagccggtct
     6061 tgtcgatcag gatgatctgg acgaagagca tcaggggctc gcgccagccg aactgttcgc
     6121 caggctcaag gcgcgtatgc ccgacggcga ggatctcgtc gtgacccacg gcgatgcctg
     6181 cttgccgaat atcatggtgg aaaatggccg cttttctgga ttcatcgact gtggccggct
     6241 gggtgtggcg gaccgctatc aggacatagc gttggctacc cgtgatattg ctgaagagct
     6301 tggcggcgaa tgggctgacc gcttcctcgt gctttacggt atcgccgctc ccgattcgca
     6361 gcgcatcgcc ttctatcgcc ttcttgacga gttcttctga tttttttaag gcagttattg
     6421 gtgcccttaa acgcctggtt gctacgcctg aataagtgat aataagcgga tgaatggcag
     6481 aaattcgatg ataagctgtc aaacatgaga attggtcgag acggtcccta gggcgataga
     6541 tatcaaccct ggcgccgggg gacaattgtg tagtcccggg aacgagcata tggagcgact
     6601 taagcagtcc ccatggggct gaaatgagct gttgacaatt aatcatccgg ctcgtataat
     6661 gtgtggaatt gtgagcggat aacaatttca cacgtcgacc gccttcggcg aagctaggga
     6721 cgagagctag catgtctcgt ttagataaaa gtaaagtgat taacagcgca ttagagctgc
     6781 ttaatgaggt cggaatcgaa ggtttaacaa cccgtaaact cgcccagaag ctaggtgtag
     6841 agcagcctac attgtattgg catgtaaaaa ataagcgggc tttgctcgac gccttagcca
     6901 ttgagatgtt agataggcac catactcact tttgcccttt agaaggggaa agctggcaag
     6961 attttttacg taataacgct aaaagtttta gatgtgcttt actaagtcat cgcgatggag
     7021 caaaagtaca tttaggtaca cggcctacag aaaaacagta tgaaactctc gaaaatcaat
     7081 tagccttttt atgccaacaa ggtttttcac tagagaatgc attatatgca ctcagcgctg
     7141 tggggcattt tactttaggt tgcgtattgg aagatcaaga gcatcaagtc gctaaagaag
     7201 aaagggaaac acctactact gatagtatgc cgccattatt acgacaagct atcgaattat
     7261 ttgatcacca aggtgcagag ccagccttct tattcggcct tgaattgatc atctgcggat
     7321 tagaaaaaca acttaaatgt gaaagtgggt ctatggtgag caagggcgag gaggataaca
     7381 tggccatcat caaggagttc atgcgcttca aggttcacat ggagggctcc gtgaacggcc
     7441 acgagttcga gatcgagggc gagggcgagg gccgccccta cgagggcacc cagaccgcca
     7501 agctgaaggt gaccaagggt ggccccctgc ccttcgcctg ggacatcctg tcccctcagt
     7561 tcatgtacgg ctccaaggcc tacgtgaagc accccgccga catccccgac tacttgaagc
     7621 tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg
     7681 tgaccgtgac ccaggactcc tccctgcaag acggcgagtt catctacaag gtgaagctgc
     7741 gcggcaccaa cttcccctcc gacggccccg taatgcagaa gaagactatg ggctgggagg
     7801 cctcctccga gcggatgtac cccgaggacg gcgcgctgaa gggcgagatc aagcagaggc
     7861 tgaagctgaa ggacggcggc cactacgacg ctgaggtcaa gaccacctac aaggccaaga
     7921 agcccgtgca actgcccggc gcgtacaacg tcaacatcaa gttggacatc acctcccaca
     7981 acgaggacta caccatcgtg gaacagtacg aacgcgccga gggccgccac tccaccggcg
     8041 gcatggacga gctgtataag taagatatct atcgccctag ggaccgtctc gagagaatca
     8101 atattaatcc aacgcgtggc atcaaataaa acgaaaggct cagtcgaaag actgggcctt
     8161 tcgttttatc tgttgtttgt cggtgaacgc tctcctgagt aggacaaatc cgccgcccta
     8221 gacttaggcg ttcggctgcg gcgagcggta tcagctcact caaag
//
