LOCUS       sequence-313930-        7477 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  aTC and Bile Acid inducible sgRNA-4 with PM4 driving Nanoluc 
            expression (NOR Gate), AmpR, ErmR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7477)
  AUTHORS   Taketani M, Zhang J, Zhang S, Triassi AJ, Huang YJ, Griffith LG, 
            Voigt CA
  TITLE     Genetic circuit design automation for the gut resident species 
            Bacteroides thetaiotaomicron.
  JOURNAL   Nat Biotechnol. 2020 Aug;38(8):962-969. doi: 
            10.1038/s41587-020-0468-5. Epub 2020 Mar 30.
  PUBMED    32231334
REFERENCE   2  (bases 1 to 7477)
  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"; doi: 
            "10.1038/s41587-020-0468-5"; journalName: "Nat Biotechnol"; date: 
            "2020-08"; volume: "38"; issue: "8"; pages: "962-969"
COMMENT     Sequence Label: pMT455
FEATURES             Location/Qualifiers
     source          1..7477
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    485..503
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    547..565
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_RNA        590..665
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_RNA        876..951
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_RNA        1136..1185
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             join(6118..6186,6187..6978)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(6336..6355)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     oriT            7109..7218
                     /note="incP origin of transfer"
     rep_origin      join(7389..7477,1..300)
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
ORIGIN
        1 tctcatgttt aacgaactaa accctcatgg ctaacgtact aagctctcat ggctaacgta
       61 ctaagctctc atgtttcacg tactaagctc tcatgtttga acaataaaat taatataaat
      121 cagcaactta aatagcctct aaggttttaa gttttataag aaaaaaaaga atatataagg
      181 cttttaaagc ttttaaggtt taacggttgt ggacaacaag ccagggatgt aacgcactga
      241 gaagccctta gagcctctca aagcaatttt gagtgacaca ggaacactta acggctgaca
      301 tgggaattcc cctccaccgc ggtgtattaa tgcggctgca atttttttgg gcggggccgc
      361 ccaaaaaaat cctagcaccc tgcagggtct cccgcttttt gcacccgctt tccaagagaa
      421 gaaagccttg ttaaattgac ttagtgtaaa agcgcagtac tgcttgacca taagaacaaa
      481 aaaatctcta tcactgatag ggataaagtt tggaagataa agctaaaagt tcttatcttt
      541 gcagtctccc tatcagtgat agagacgcac aaagatactt atgttagaag ttttagagct
      601 agaaatagca agttaaaata aggctagtcc gttatcaact tgaaaaagtg gcaccgagtc
      661 ggtgcttttt ttggatccaa cgcatgagaa agcccccgga agatcacctt ccgggggctt
      721 ttttattgcg cgcggccgct tacaaagaaa attcgacaaa ctgttatttt tctatctatt
      781 taaatgtaca cccgtttgtt tactttactt tagttatgta cctttgtcgg caatgtacac
      841 ccgaaagttt actttcaaag atacttatgt tagaagtttt agagctagaa atagcaagtt
      901 aaaataaggc tagtccgtta tcaacttgaa aaagtggcac cgagtcggtg ctttttttgg
      961 atccaacgca tgagaaagcc cccggaagat caccttccgg gggctttttt attgcgcgcg
     1021 gccgctgctg tcataggccg gttgtccggt agtccggatg atttaaggaa gatttctctt
     1081 cttttttcac tttctccgaa caacctccat tctaacataa gtatctttgc agtagctgtc
     1141 accggatgtg ctttccggtc tgatgagtcc gtgaggacga aacagcctct acaaataatt
     1201 ttgtttaacg cattttaaaa taaaataaat tatttattta attaaacgaa tatggttttt
     1261 actctggaag attttgttgg cgattggcgt cagaccgcgg gttataattt ggatcaagtc
     1321 ctggaacagg gtggcgtaag ctctctgttc cagaacctgg gtgtgagcgt gacgccgatt
     1381 cagcgcatcg ttctgtccgg cgagaacggt ctgaaaattg atattcatgt gatcatcccg
     1441 tacgaaggcc tgagcggtga ccaaatgggt caaatcgaga aaatctttaa agtcgtctac
     1501 ccagttgacg atcaccactt caaggttatc ttgcattacg gtacgctggt gattgatggt
     1561 gtgaccccga atatgattga ctatttcggc cgtccgtatg aaggcattgc cgtttttgac
     1621 ggtaaaaaga tcaccgtcac cggtaccctg tggaatggca ataagattat tgacgagcgt
     1681 ctgattaacc cggacggcag cctgctgttc cgcgtgacca tcaacggtgt cacgggttgg
     1741 cgtctgtgcg agcgcatcct ggcataatga actgggatcc ctcggtacca aattttcgaa
     1801 aaaagacgct gaaaagcgtc ttttttcgtt ttggtccgcg gccgccggcc gctgatctgg
     1861 aagaagcaat gaaagctgct gttaagtctc cgaatcaggt attgttcctg acaggtgtat
     1921 tcccatccgg taaacgcgga tactttgcag ttgatctgac tcaggaataa attataaatt
     1981 aaggtaagaa gattgtagga taagctaatg aaatagaaaa aggatgccgt cacacaactt
     2041 gtcggcattc ttttttgttt tattagttga aaatatagtg aaaaagttgc ctaaatatgt
     2101 atgttaacaa attatttgtc gtaactttgc actccaaatc tgtttttaac atatggcact
     2161 aatgcagaaa aaattgactc gttctcagca gaaacacttg gacatcatca acgctgctaa
     2221 agaagaattc atcgaattcg gtttcttggc tgctaacatg gaccgtatca cttcttctgc
     2281 tgaagtatct aaacgtactt tgtaccgtca cttcgaatct aaagaagtat tgttcgaatc
     2341 tgtattgact atcatcaacg actctgtaaa cgaatctatc tcttaccact tcgacccgaa
     2401 caaatctact gaagaacagt tgactgaaat cgcttacaaa gaaatcgacg tattgtacaa
     2461 aacttacggt atcgctttgg ctcgtactat cgtaatggaa ttcttgcgtc agccggaaat
     2521 ggctaaaact ttgatccaga acatctactc tatccgtgct atcactcagt ggttccgttc
     2581 tgctatcgaa gctaaacgtt tgaaagacgc tgacccgaaa ttgatgactg acgtatacgt
     2641 atctttgttc cagggtttgt tcttctggcc gcaggtaatg cacttggact tggaaccgca
     2701 cggtgaagaa ttgtctcaga aaatcgaaac tttgactact atcttcttgc agtcttacgg
     2761 tgtagctgaa taacaatgaa tgggctttcc tcggtaccaa attccagaaa agaggcctcc
     2821 cgaaaggggg gccttttttc gttttggtcc ctaattgcct atcttccagt gatggaacag
     2881 catttgtgca ttggctgcaa caatcagcct tacttgtgcc tgttctattt ccgaaccgac
     2941 cgcttgtatg aatgcatcaa aattcgtttt ctctacgttg gattccttgt tgctcatatt
     3001 gtgatgataa tttctacaaa tatagtcatt ggtaactatc tatgaaactg tttgatactt
     3061 ttatcagtcc agtagtttta caaggtcttt cttcatttct tcgtcaatat ccctgtatcg
     3121 tctgaaagct ttgcttccct ccttgtgtcc cgacagtgcg gaaacaaggt tcgggtcttt
     3181 cactttttta tagatattgc cgataaacgt acgtcttgcc agatggctgc ttgccacttc
     3241 ataaataggt cgtttgattt cgttgtgcgt caacgggtct aagattgtta cgatgcggtc
     3301 aactccagct aatttgaata tcttttttat ggcatcattg tacttttgct cggatatgaa
     3361 cggcaacagt tttccctcat attctttgta gcgttcaagg atttctttcg ctttgtcgtt
     3421 aagtggaaca cgtaccgtaa ccggattccc ctctttggtt ttcttgggaa tatattctat
     3481 ggcttcattg accacattta gtttggtcat tcggtacagg tcgctcaccc tgcatcctat
     3541 cagtgtctga aatatgaata tatccctctg tattgccagt tgtggggtgg cagaaaggtc
     3601 tgcattaaaa atcctgtccc tttcttcgag tgttatataa taaggtgtac catatgtaca
     3661 ctcctctatc ggaaacttgt cgaaaggtct gtttgtggtg cgtttgttat cgaagcacca
     3721 caggaagaat gtgcgtattc ttgaaaaaca gtctatcagc gtgtttttgc ttctgggctg
     3781 tggtgtcctc ttttcgggaa tggcttcata aatgctcggg taaagttcat aatactggta
     3841 ttcgttctga aagaaatccc acatatcccg aagcgtgtca ggtgttacca aatccacatc
     3901 aaggataaag cccttttgtc ctctctttgt agcccttaca tatagttcat aacgcagtaa
     3961 ggctcttttg acaacccgga aatttttctt tcgtacttcc gacaaagggt gcttgtttag
     4021 aaattcatcg aatagttctc caatggtagg cttgataacc acttcctccg gcaaaaaata
     4081 tttttcagga tggtaaaatt tatcaagtgt tgtttttagc cattctttgt ctattgcttc
     4141 cttttcttgt tgatatacct tctcgatata ggttttcaac tggcgtatct cttcatttat
     4201 atgggtacgc atttcttcat tgcagacagc tttcgttttt acacattcgt ctttatcatc
     4261 ccatagattg ggattgatgg ctaactgagt aggagcaacg gagtctaact gccttccgtt
     4321 tctaaaacgg acatagattg tagccatgga ttctgtatca tatcgtttgg ctgctttttt
     4381 aatgataaag gttactttca tagactttca ggttgaattt tactctgctg caaatataaa
     4441 tattttcccc agcattttcc ccacatctgc taaatatttt gcaattcgat taaacttgga
     4501 ttaaaattta ataggattat aacatattga aatacagtgt aattgtggcg tttttccgca
     4561 tttttctttt tacccacatt ttccccactt taattatttc aaatgcgggg tctgggtaca
     4621 agaaagaaag ctaagtattt gatagttcaa tacttagctt tttcttttgc ttgaattttc
     4681 cccacatttt ccccacacgt gcaaaaaata tagcagtaag tcattatttc ttttggttga
     4741 acgtagagag tagcgatatt aaaagaatcc gatgagaaaa gactaatatt tatctatcca
     4801 ttcagtttga tttctcagga ctttacatcg tcctgaaagt atttgttggt accgaggacg
     4861 cgtaaacatt tacagttgca tgtggcctat tgtttttagc cgttaaatat tttataacta
     4921 ttaaatagcg atacaaattg ttcgaaacta atattgttta tatcatatat tctcgcatgt
     4981 tttaaagctt tattaaattg attttttgta aacagttttt cgtactcttt gttaacccat
     5041 ttcattacaa aagtttcata tttttttctc tctttaaatg ccatttttgc tggctttctt
     5101 tttaatacaa ttaatgtgct atccacttta ggttttggat ggaaataata cctaggaatt
     5161 tttgctaata tagaaatatc tacctctgcc attaacagca atgctagtga tctgtttgta
     5221 tctaataaca ttttagcaaa accatattcc actattaaat aacttattgt ggctgaactt
     5281 tcaaaaacaa tttttcgaat tatatttgtg cttatgttgt aaggtatgct gccaaatatt
     5341 ttatatggat tgtggctagg aaatgtaaat ttcagtatat catcatttac tatttgatag
     5401 ttaggataat ttaagagctt attacgagtt acctcacata atttagaatc aatttctatc
     5461 gccgttacaa aattacatct ctttaccaat ccagcagtaa aatgaccttt ccctgcacct
     5521 atttcaaaga tgttatcttt ttcatctaaa cttatgcaat tcattatttt ttctatgtga
     5581 tattttgaag taataaaatt ttgactatct tttatattta ctttgttcat tataacctct
     5641 ccttaattta ttgcatctct tttcgaatat ttatgttttt tgagaaaaga acgtactcat
     5701 ggttcatccc gatatgcgta tcggtctgta tatcagcaac tttctatgtg tttcaactac
     5761 aatagtcatc tattctcatc tttctgagtc caccccctgc aaagcccctc tttacgacat
     5821 aaaaattcgg tcggaaaagg tatgcaaaag atgtttctct ctttaagaga aactcttcgg
     5881 gatgcaaaaa tatgaaaata actccaattc accaaattat atagcgactt ttttacaaaa
     5941 tgctaaaatt tgttgatttc cgtcaagcaa ttgttgagca aaaatgtctt ttacgataaa
     6001 atgatacctc aatatcaact gtttagcaaa acgatatttc tcttaaagag agaaacacct
     6061 ttttgttcac caatccccga cttttaatcc cgcggccatg attgaaaaag gaagagtatg
     6121 agtattcaac atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt
     6181 tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga
     6241 gtgggttaca tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa
     6301 gaacgttttc caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt
     6361 attgacgccg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt
     6421 gagtactcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc
     6481 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga
     6541 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat
     6601 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct
     6661 gtagcaatgg caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc
     6721 cggcaacaat taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg
     6781 gcccttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtcacgc
     6841 ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg
     6901 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca
     6961 ctgattaagc attggtaact gtcagaccaa gtttactcat aacgcgtcaa ttcgaggggg
     7021 atcaattccg tgataggtgg gctgcccttc ctggttggct tggtttcatc agccatccgc
     7081 ttgccctcat ctgttacgcc ggcggtagcc ggccagcctc gcagagcagg attcccgttg
     7141 agcaccgcca ggtgcgaata agggacagtg aagaaggaac acccgctcgc gggtgggcct
     7201 acttcaccta tcctgcccgg ctgacgccgt tggatacacc aaggaaagtc tacacgaacc
     7261 ctttggcaaa atcctgtata tcgtgcgaaa aaggatggat ataccgaaaa aatcgctata
     7321 atgaccccga agcagggtta tgcagcggaa aacggaattg atccggccac gatgcgtccg
     7381 gcgtagagga tctgaagatc agcagttcaa cctgttgata gtacgtacta agctctcatg
     7441 tttcacgtac taagctctca tgtttaacgt actaagc
//
