LOCUS       sequence-148756-        9342 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Output plasmid: contains 5-state register that regulates expression 
            of GFP..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9342)
  AUTHORS   Roquet N, Soleimany AP, Ferris AC, Aaronson S, Lu TK
  TITLE     Synthetic recombinase-based state machines in living cells.
  JOURNAL   Science. 2016 Jul 22;353(6297):aad8559. doi: 
            10.1126/science.aad8559. Epub 2016 Jul 21.
  PUBMED    27463678
REFERENCE   2  (bases 1 to 9342)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Science."; 
            date: "2016-07-22"; volume: "353(6297):aad8559. doi"; pages: " 
            10.1126/science.aad8559. Epub 2016 Jul 21"
COMMENT     Sequence Label: pNR163
FEATURES             Location/Qualifiers
     source          1..9342
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      804..1023
                     /label=ori2
                     /note="secondary origin of replication for the bacterial F 
                     plasmid; also known as oriS"
     rep_origin      1091..1622
                     /label=oriV
                     /note="incP origin of replication"
     rep_origin      1099..1713
                     /label=oriV
                     /note="origin of replication for the bacterial F plasmid"
     promoter        2571..2673
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             2674..3333
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(2740..2759)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     primer_bind     complement(3928..3948)
                     /label=GFP-F
                     /note="GFP, forward primer. Does NOT anneal to EGFP"
     primer_bind     4543..4571
                     /label=GFP-R
                     /note="GFP, reverse primer. Does NOT anneal to EGFP"
     RBS             4607..4615
                     /label=Shine-Dalgarno sequence
                     /note="full consensus sequence for ribosome-binding sites 
                     upstream of start codons in E. coli; complementary to a 
                     region in the 3' end of the 16S rRNA (Chen et al., 1994)"
     misc_RNA        complement(4658..4707)
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     protein_bind    4845..4893
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="attB site for the Mycopbacteriophage Bxb1 integrase 
                     (Xu et al., 2013)"
     terminator      5199..5270
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     protein_bind    5495..5552
                     /label=Bxb1 attP
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="attP site for the Mycopbacteriophage Bxb1 integrase 
                     (Xu et al., 2013)"
     misc_feature    6029..6502
                     /gene="sopC"
                     /label=sopC
                     /note="centromere-like partitioning region of the bacterial
                     F plasmid"
     CDS             complement(6575..7546)
                     /codon_start=1
                     /gene="sopB"
                     /product="partitioning protein for the bacterial F plasmid"
                     /label=sopB
                     /translation="MKRAPVIPKHTLNTQPVEDTSLSTPAAPMVDSLIARVGVMARGNA
                     ITLPVCGRDVKFTLEVLRGDSVEKTSRVWSGNERDQELLTEDALDDLIPSFLLTGQQTP
                     AFGRRVSGVIEIADGSRRRKAAALTESDYRVLVGELDDEQMAALSRLGNDYRPTSAYER
                     GQRYASRLQNEFAGNISALADAENISRKIITRCINTAKLPKSVVALFSHPGELSARSGD
                     ALQKAFTDKEELLKQQASNLHEQKKAGVIFEAEEVITLLTSVLKTSSASRTSLSSRHQF
                     APGATVLYKGDKMVLNLDRSRVPTECIEKIEAILKELEKPAP"
     CDS             complement(7546..8721)
                     /codon_start=1
                     /gene="sopA"
                     /product="partitioning protein for the bacterial F plasmid"
                     /label=sopA
                     /translation="MFRMKLMETLNQCINAGHEMTKAIAIAQFNDDSPEARKITRRWRI
                     GEAADLVGVSSQAIRDAEKAGRLPHPDMEIRGRVEQRVGYTIEQINHMRDVFGTRLRRA
                     EDVFPPVIGVAAHKGGVYKTSVSVHLAQDLALKGLRVLLVEGNDPQGTASMYHGWVPDL
                     HIHAEDTLLPFYLGEKDDVTYAIKPTCWPGLDIIPSCLALHRIETELMGKFDEGKLPTD
                     PHLMLRLAIETVAHDYDVIVIDSAPNLGIGTINVVCAADVLIVPTPAELFDYTSALQFF
                     DMLRDLLKNVDLKGFEPDVRILLTKYSNSNGSQSPWMEEQIRDAWGSMVLKNVVRETDE
                     VGKGQIRMRTVFEQAIDQRSSTGAWRNALSIWEPVCNEIFDRLIKPRWEIR"
     misc_feature    9047..9297
                     /gene="incC"
                     /label=incC
                     /note="incompatibility region of the bacterial F plasmid"
     CDS             complement(join(9300..9342,1..713))
                     /codon_start=1
                     /gene="repE"
                     /product="replication initiation protein for the bacterial 
                     F plasmid"
                     /label=repE
                     /translation="MAETAVINHKKRKNSPRIVQSNDLTEAAYSLSRDQKRMLYLFVDQ
                     IRKSDGTLQEHDGICEIHVAKYAEIFGLTSAEASKDIRQALKSFAGKEVVFYRPEEDAG
                     DEKGYESFPWFIKRAHSPSRGLYSVHINPYLIPFFIGLQNRFTQFRLSETKEITNPYAM
                     RLYESLCQYRKPDGSGIVSLKIDWIIERYQLPQSYQRMPDFRRRFLQVCVNEINSRTPM
                     RLSYIEKKKGRQTTHIVFSFRDITSMTTG"
ORIGIN
        1 atatgagtcg tctggcggcc tttctttttc tcaatgtatg agaggcgcat tggagttctg
       61 ctgttgatct cattaacaca gacctgcagg aagcggcggc ggaagtcagg catacgctgg
      121 taactttgag gcagctggta acgctctatg atccagtcga ttttcagaga gacgatgcct
      181 gagccatccg gcttacgata ctgacacagg gattcgtata aacgcatggc atacggattg
      241 gtgatttctt ttgtttcact aagccgaaac tgcgtaaacc ggttctgtaa cccgataaag
      301 aagggaatga gatatgggtt gatatgtaca ctgtaaagcc ctctggatgg actgtgcgca
      361 cgtttgataa accaaggaaa agattcatag cctttttcat cgccggcatc ctcttcaggg
      421 cgataaaaaa ccacttcctt ccccgcgaaa ctcttcaatg cctgccgtat atccttactg
      481 gcttccgcag aggtcaatcc gaatatttca gcatatttag caacatggat ctcgcagata
      541 ccgtcatgtt cctgtagggt gccatcagat tttctgatct ggtcaacgaa cagatacagc
      601 atacgttttt gatcccggga gagactatat gccgcctcag tgaggtcgtt tgactggacg
      661 attcgcgggc tatttttacg tttcttgtga ttgataaccg ctgtttccgc catgacagat
      721 ccatgtgaag tgtgacaagt ttttagattg tcacactaaa taaaaaagag tcaataagca
      781 gggataactt tgtgaaaaaa cagcttcttc tgagggcaat ttgtcacagg gttaagggca
      841 atttgtcaca gacaggactg tcatttgagg gtgatttgtc acactgaaag ggcaatttgt
      901 cacaacacct tctctagaac cagcatggat aaaggcctac aaggcgctct aaaaaagaag
      961 atctaaaaac tataaaaaaa ataattataa aaatatcccc gtggataagt ggataacccc
     1021 aagggaagtt ttttcaggca tcgtgtgtaa gcagaatata taagtgctgt tccctggtgc
     1081 ttcctcgctc actcgaccgg gagggttcga gaaggggggg cacccccctt cggcgtgcgc
     1141 ggtcacgcgc acagggcgca gccctggtta aaaacaaggt ttataaatat tggtttaaaa
     1201 gcaggttaaa agacaggtta gcggtggccg aaaaacgggc ggaaaccctt gcaaatgctg
     1261 gattttctgc ctgtggacag cccctcaaat gtcaataggt gcgcccctca tctgtcagca
     1321 ctctgcccct caagtgtcaa ggatcgcgcc cctcatctgt cagtagtcgc gcccctcaag
     1381 tgtcaatacc gcagggcact tatccccagg cttgtccaca tcatctgtgg gaaactcgcg
     1441 taaaatcagg cgttttcgcc gatttgcgag gctggccagc tccacgtcgc cggccgaaat
     1501 cgagcctgcc cctcatctgt caacgccgcg ccgggtgagt cggcccctca agtgtcaacg
     1561 tccgcccctc atctgtcagt gagggccaag ttttccgcga ggtatccaca acgccggcgg
     1621 ccggccgcgg tgtctcgcac acggcttcga cggcgtttct ggcgcgtttg cagggccata
     1681 gacggccgcc agcccagcgg cgagggcaac cagccgaggg cttcgccctg tcgctcgact
     1741 gcggcgagca ctactggctg taaaaggaca gaccacatca tggttctgtg ttcattaggt
     1801 tgttctgtcc attgctgaca taatccgctc cacttcaacg taacaccgca cgaagatttc
     1861 tattgttcct gaaggcatat tcaaatcgtt ttcgttaccg cttgcaggca tcatgacaga
     1921 acactacttc ctataaacgc tacacaggct cctgagatta ataatgcgga tctctacgat
     1981 aatgggagat tttcccgact gtttcgttcg cttctcagtg gataacagcc agcttctctg
     2041 tttaacagac aaaaacagca tatccactca gttccacatt tccatataaa ggccaaggca
     2101 tttattctca ggataattgt ttcagcatcg caaccgcatc agactccggc atcgcaaact
     2161 gcacccggtg ccgggcagcc acatccagcg caaaaacctt cgtgtagact tccgttgaac
     2221 tgatggactt atgtcccatc aggctttgca gaactttcag cggtataccg gcatacagca
     2281 tgtgcatcgc ataggaatgg cggaacgtat gtggtgtgac cggaacagag aacgtcacac
     2341 cgtcagcagc agcggcggca accgcctccc caatccaggt cctgaccgtt ctgtccgtca
     2401 cttcccagat ccgcgctttc tctgtccttc ctgtgcgacg gttacgccgc tccatgagct
     2461 tatcgcgaat aaatacctgt gacggaagat cacttcgcag aataaataaa tcctggtgtc
     2521 cctgttgata ccgggaagcc ctgggccaac ttttggcgaa aatgagacgt tgatcggcac
     2581 gtaagaggtt ccaactttca ccataatgaa ataagatcac taccgggcgt attttttgag
     2641 ttatcgagat tttcaggagc taaggaagct aaaatggaga aaaaaatcac tggatatacc
     2701 accgttgata tatcccaatg gcatcgtaaa gaacattttg aggcatttca gtcagttgct
     2761 caatgtacct ataaccagac cgttcagctg gatattacgg cctttttaaa gaccgtaaag
     2821 aaaaataagc acaagtttta tccggccttt attcacattc ttgcccgcct gatgaatgct
     2881 catccggaat ttcgtatggc aatgaaagac ggtgagctgg tgatatggga tagtgttcac
     2941 ccttgttaca ccgttttcca tgagcaaact gaaacgtttt catcgctctg gagtgaatac
     3001 cacgacgatt tccggcagtt tctacacata tattcgcaag atgtggcgtg ttacggtgaa
     3061 aacctggcct atttccctaa agggtttatt gagaatatgt ttttcgtctc agccaatccc
     3121 tgggtgagtt tcaccagttt tgatttaaac gtggccaata tggacaactt cttcgccccc
     3181 gttttcacca tgggcaaata ttatacgcaa ggcgacaagg tgctgatgcc gctggcgatt
     3241 caggttcatc atgccgtttg tgatggcttc catgtcggca gaatgcttaa tgaattacaa
     3301 cagtactgcg atgagtggca gggcggggcg taagacgtct aagtaagtgc ccattcagcc
     3361 agtataacca gataaaaaaa atccttagct ttcgctaagg atgatttctg tcagccagga
     3421 aggtataaca ggaaaaatta aaggggagat aaaatccccc ctttttggtt aactcgagga
     3481 ttcgaaaagg tgaaatctcc tttcacggcc cattcctcat ggatgggccg tttatttccc
     3541 cgagccagac gtccaccata ccttcgatat tcgcgcccac tctcccatta gtcggagtgg
     3601 atgtgttgcg attgcccgct agatcaatgt gcacgtcgct aacaccgtgc gtttgactat
     3661 tttacctctg gcggtgataa tggttgcata tagtgaacaa ggattaacgc cgagtgcggt
     3721 attcctctgg gcatgccaac acaattaaca tctcaatcaa ggtaaatgct ttttgctttt
     3781 tttgccgtgg tgcggatcta gctaccacct tccccttgat aaagcaccct ttggtcgaaa
     3841 aaaaaagccc gcactgtcag gtgcgggctt ttttctgtgt ttccttattt gtagagatca
     3901 tccatgccat gtgtaatccc agcagctgtt acaaactcaa gaaggaccat gtggtctctc
     3961 ttttcgttgg gatctttcga aagggcagat tgtgtggaca ggtaatggtt gtctggtaaa
     4021 aggacagggc catcgccaat tggagtattt tgttgataat ggtctgctag ttgaacgctt
     4081 ccatcttcaa tgttgtgtct aattttgaag ttaactttga ttccattctt ttgtttgtct
     4141 gccatgatgt atacattgtg tgagttatag ttgtattcca atttgtgtcc aagaatgttt
     4201 ccatcttctt taaaatcaat accttttaac tcgattctat taacaagggt atcaccttca
     4261 aacttgactt cagcacgtgt cttgtagttc ccgtcatctt tgaaaaatat agttctttcc
     4321 tgtacataac cttcgggcat ggcactcttg aaaaagtcat gctgtttcat atgatctggg
     4381 tatctcgcaa agcattgaac accataaccg aaagtagtga caagtgttgg ccatggaaca
     4441 ggtagttttc cagtagtgca aataaattta agggtaagtt ttccgtatgt tgcatcacct
     4501 tcaccctctc cactgacaga aaatttgtgc ccattaacat caccatctaa ttcaacaaga
     4561 attgggacaa ctccagtgaa aagttcttct cctttactca ttagtgacct ccttaatata
     4621 ggtcggcttc gaccttaaac aaaattattt gtagaggctg tttcgtcctc acggactcat
     4681 cagaccggaa agcacatccg gtgacagctg tgccatccct cgtcctcacc cgcacttgag
     4741 tccacggctg atgccaacac aattaacatc agaatcaagg taaatgcttt ttgctttttt
     4801 tgcgggctca ctagggaatg gcgtcgtccg tgacattctg tgcgcggccg gcttgtcgac
     4861 gacggcggtc tccgtcgtca ggatcatccg ggccagcggc tgtctatgcc agaagtcggt
     4921 caggtcggag tcctagaaag gagtttttta gttaccttaa ttctaataaa cgaaataaaa
     4981 actcgccgtt cgcgcgacat gacatgcgcc gccgattgat tcagaaagtt aaacaaaatt
     5041 atttgtagag ggaaaccgtt gtggtctccc tgaatatatt atacgagcct tatgcatgcc
     5101 cgtaaagtta tccagcaacc actcatagac ctagggcagc agatagggac gacgtggtgt
     5161 tagctgtgac agacctccca ctcacactcc ccaggcatca aataaaacga aaggctcagt
     5221 cgaaagactg ggcctttcgt tttatctgtt gtttgtcggt gaacgctctc gggcccactc
     5281 gcagcttaag ttcagccaaa aaacttaaga ccgccggtct tgtccactac cttgcagtaa
     5341 tgcggtggac aggatcggcg gttttctttt ctcttctcaa cccgaatggg agtgaacacc
     5401 aacgcatgag aaagcccccg gaagatcacc ttccgggggc ttttttattg cgcgcattga
     5461 gctacccggt ggccgacaca atctccctgc tcaggtcgtg gtttgtctgg tcaaccaccg
     5521 cggtctcagt ggtgtacggt acaaaccccg acgcaatagc gccaaccatg caccatagcc
     5581 agcctgacag tagccaaagg agttttttag ttaccttaat tgaaataaac gaaataaaaa
     5641 ctcgcgcagc tgagagcgac acagttgcct cactcctagg gtgtaatagg cactaaaaga
     5701 gtgatatggc acgcgcatgt ggaacgccgt tagagggttg ctgagccatc gtggtcctta
     5761 gactcgggaa aaacaaaaac ccgccgaagc gggttaagtg cggggcggct agcccgccac
     5821 ctaagttatg agtcaggaaa aaaggcgaca gagtaatctg tcgccttttt tctttgcttg
     5881 ctttgtaatc actcgatgac caaaagtaaa aacccgccga agcgggtttt tacgtaaaac
     5941 aggtgaaact gacgctagcc ggtcacatgt caatacttgc ccttgacagg cattgatgga
     6001 atcgtagtct cacgctgata gtctgatcga caatacaagt gggaccgtgg tcccagaccg
     6061 ataatcagac cgacaacacg agtgggatcg tggtcccaga ctaataatca gaccgacgat
     6121 acgagtggga ccgtggtccc agactaataa tcagaccgac gatacgagtg ggaccgtggt
     6181 tccagactaa taatcagacc gacgatacga gtgggaccgt ggtcccagac taataatcag
     6241 accgacgata cgagtgggac catggtccca gactaataat cagaccgacg atacgagtgg
     6301 gaccgtggtc ccagtctgat tatcagaccg acgatacgag tgggaccgtg gtcccagact
     6361 aataatcaga ccgacgatac gagtgggacc gtggtcccag actaataatc agaccgacga
     6421 tacgagtggg accgtggtcc cagtctgatt atcagaccga cgatacaagt ggaacagtgg
     6481 gcccagagag aatattcagg ccagttatgc tttctggcct gtaacaaagg acattaagta
     6541 aagacagata aacgtagact aaaacgtggt cgcatcaggg tgctggcttt tcaagttcct
     6601 taagaatggc ctcaattttc tctatacact cagttggaac acgggacctg tccaggttaa
     6661 gcaccatttt atcgccctta tacaatactg tcgctccagg agcaaactga tgtcgtgagc
     6721 ttaaactagt tcttgatgca gatgacgttt taagcacaga agttaaaaga gtgataactt
     6781 cttcagcttc aaatatcacc ccagcttttt tctgctcatg aaggttagat gcctgctgct
     6841 taagtaattc ctctttatct gtaaaggctt tttgaagtgc atcacctgac cgggcagata
     6901 gttcaccggg gtgagaaaaa agagcaacaa ctgatttagg caatttggcg gtgttgatac
     6961 agcgggtaat aatcttacgt gaaatatttt ccgcatcagc cagcgcagaa atatttccag
     7021 caaattcatt ctgcaatcgg cttgcataac gctgaccacg ttcataagca cttgttgggc
     7081 gataatcgtt acccaatctg gataatgcag ccatctgctc atcatccagc tcgccaacca
     7141 gaacacgata atcactttcg gtaagtgcag cagctttacg acggcgactc ccatcggcaa
     7201 tttctatgac accagatact cttcgaccga acgccggtgt ctgttgacca gtcagtagaa
     7261 aagaagggat gagatcatcc agtgcgtcct cagtaagcag ctcctggtca cgttcattac
     7321 ctgaccatac ccgagaggtc ttctcaacac tatcaccccg gagcacttca agagtaaact
     7381 tcacatcccg accacataca ggcaaagtaa tggcattacc gcgagccatt actcctacgc
     7441 gcgcaattaa cgaatccacc atcggggcag ctggtgtcga taacgaagta tcttcaaccg
     7501 gttgagtatt gagcgtatgt tttggaataa caggcgcacg cttcattatc taatctccca
     7561 gcgtggttta atcagacgat cgaaaatttc attgcagaca ggttcccaaa tagaaagagc
     7621 atttctccag gcaccagttg aagagcgttg atcaatggcc tgttcaaaaa cagttctcat
     7681 ccggatctga cctttaccaa cttcatccgt ttcacgtaca acatttttta gaaccatgct
     7741 tccccaggca tcccgaattt gctcctccat ccacggggac tgagagccat tactattgct
     7801 gtatttggta agcaaaatac gtacatcagg ctcgaaccct ttaagatcaa cgttcttgag
     7861 cagatcacga agcatatcga aaaactgcag tgcggaggtg tagtcaaaca actcagcagg
     7921 cgtgggaaca atcagcacat cagcagcaca tacgacatta atcgtgccga tacccaggtt
     7981 aggcgcgctg tcaataacta tgacatcata gtcatgagca acagtttcaa tggccagtcg
     8041 gagcatcagg tgtggatcgg tgggcagttt accttcatca aatttgccca ttaactcagt
     8101 ttcaatacgg tgcagagcca gacaggaagg aataatgtca agccccggcc agcaagtggg
     8161 ctttattgca taagtgacat cgtccttttc cccaagatag aaaggcagga gagtgtcttc
     8221 tgcatgaata tgaagatctg gtacccatcc gtgatacatt gaggctgttc cctgggggtc
     8281 gttaccttcc acgagcaaaa cacgtagccc cttcagagcc agatcctgag caagatgaac
     8341 agaaactgag gttttgtaaa cgccaccttt atgggcagca accccgatca ccggtggaaa
     8401 tacgtcttca gcacgtcgca atcgcgtacc aaacacatca cgcatatgat taatttgttc
     8461 aattgtataa ccaacacgtt gctcaacccg tcctcgaatt tccatatccg ggtgcggtag
     8521 tcgccctgct ttctcggcat ctctgatagc ctgagaagaa accccaacta aatccgctgc
     8581 ttcacctatt ctccagcgcc gggttatttt cctcgcttcc gggctgtcat cattaaactg
     8641 tgcaatggcg atagccttcg tcatttcatg accagcgttt atgcactggt taagtgtttc
     8701 catgagtttc attctgaaca tcctttaatc attgctttgc gtttttttat taaatcttgc
     8761 aatttactgc aaagcaacaa caaaatcgca aagtcatcaa aaaaccgcaa agttgtttaa
     8821 aataagagca acactacaaa aggagataag aagagcacat acctcagtca cttattatca
     8881 ctagcgctcg ccgcagccgt gtaaccgagc atagcgagcg aactggcgag gaagcaaaga
     8941 agaactgttc tgtcagatag ctcttacgct cagcgcaaga agaaatatcc accgtgggaa
     9001 aaactccagg tagaggtaca cacgcggata gccaattcag agtaataaac tgtgataatc
     9061 aaccctcatc aatgatgacg aactaacccc cgatatcagg tcacatgacg aagggaaaga
     9121 gaaggaaatc aactgtgaca aactgccctc aaatttggct tccttaaaaa ttacagttca
     9181 aaaagtatga gaaaatccat gcaggctgaa ggaaacagca aaactgtgac aaattaccct
     9241 cagtaggtca gaacaaatgt gacgaaccac cctcaaatct gtgacagata accctcagac
     9301 tatcctgtcg tcatggaagt gatatcgcgg aaggaaaata cg
//
