LOCUS       sequence-313926-        7134 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bile Acid inducible sgRNA-4 with PM4 driving Nanoluc expression (NOT
            Gate 4), pNBU1 backbone, AmpR, ErmR.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7134)
  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 7134)
  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: pMT451
FEATURES             Location/Qualifiers
     source          1..7134
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     oriT            360..469
                     /note="incP origin of transfer"
     rep_origin      640..1028
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     misc_RNA        1261..1336
                     /label=gRNA scaffold
                     /note="guide RNA scaffold for the Streptococcus pyogenes 
                     CRISPR/Cas9 system"
     misc_RNA        1521..1570
                     /label=sTRSV HHRz
                     /note="hammerhead ribozyme from the tobacco ringspot virus 
                     satellite RNA (Khvorova et al., 2003)"
     CDS             join(6503..6571,6572..7134,1..229)
                     /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(6721..6740)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
       61 taaatctgga gccggtgagc gtgggtcacg cggtatcatt gcagcactgg ggccagatgg
      121 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
      181 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
      241 agtttactca taacgcgtca attcgagggg gatcaattcc gtgataggtg ggctgccctt
      301 cctggttggc ttggtttcat cagccatccg cttgccctca tctgttacgc cggcggtagc
      361 cggccagcct cgcagagcag gattcccgtt gagcaccgcc aggtgcgaat aagggacagt
      421 gaagaaggaa cacccgctcg cgggtgggcc tacttcacct atcctgcccg gctgacgccg
      481 ttggatacac caaggaaagt ctacacgaac cctttggcaa aatcctgtat atcgtgcgaa
      541 aaaggatgga tataccgaaa aaatcgctat aatgaccccg aagcagggtt atgcagcgga
      601 aaacggaatt gatccggcca cgatgcgtcc ggcgtagagg atctgaagat cagcagttca
      661 acctgttgat agtacgtact aagctctcat gtttcacgta ctaagctctc atgtttaacg
      721 tactaagctc tcatgtttaa cgaactaaac cctcatggct aacgtactaa gctctcatgg
      781 ctaacgtact aagctctcat gtttcacgta ctaagctctc atgtttgaac aataaaatta
      841 atataaatca gcaacttaaa tagcctctaa ggttttaagt tttataagaa aaaaaagaat
      901 atataaggct tttaaagctt ttaaggttta acggttgtgg acaacaagcc agggatgtaa
      961 cgcactgaga agcccttaga gcctctcaaa gcaattttga gtgacacagg aacacttaac
     1021 ggctgacatg ggaattcccc tccaccgcgg tgtattaatg cggctgcaat ttttttgggc
     1081 ggggccgccc aaaaaaatcc tagcaccctg cagggtctcc cgctttacaa agaaaattcg
     1141 acaaactgtt atttttctat ctatttaaat gtacacccgt ttgtttactt tactttagtt
     1201 atgtaccttt gtcggcaatg tacacccgaa agtttacttt caaagatact tatgttagaa
     1261 gttttagagc tagaaatagc aagttaaaat aaggctagtc cgttatcaac ttgaaaaagt
     1321 ggcaccgagt cggtgctttt tttggatcca acgcatgaga aagcccccgg aagatcacct
     1381 tccgggggct tttttattgc gcgcggccgc tgctgtcata ggccggttgt ccggtagtcc
     1441 ggatgattta aggaagattt ctcttctttt ttcactttct ccgaacaacc tccattctaa
     1501 cataagtatc tttgcagtag ctgtcaccgg atgtgctttc cggtctgatg agtccgtgag
     1561 gacgaaacag cctctacaaa taattttgtt taacgcattt taaaataaaa taaattattt
     1621 atttaattaa acgaatatgg tttttactct ggaagatttt gttggcgatt ggcgtcagac
     1681 cgcgggttat aatttggatc aagtcctgga acagggtggc gtaagctctc tgttccagaa
     1741 cctgggtgtg agcgtgacgc cgattcagcg catcgttctg tccggcgaga acggtctgaa
     1801 aattgatatt catgtgatca tcccgtacga aggcctgagc ggtgaccaaa tgggtcaaat
     1861 cgagaaaatc tttaaagtcg tctacccagt tgacgatcac cacttcaagg ttatcttgca
     1921 ttacggtacg ctggtgattg atggtgtgac cccgaatatg attgactatt tcggccgtcc
     1981 gtatgaaggc attgccgttt ttgacggtaa aaagatcacc gtcaccggta ccctgtggaa
     2041 tggcaataag attattgacg agcgtctgat taacccggac ggcagcctgc tgttccgcgt
     2101 gaccatcaac ggtgtcacgg gttggcgtct gtgcgagcgc atcctggcat aatgaactgg
     2161 gatccctcgg taccaaattt tcgaaaaaag acgctgaaaa gcgtcttttt tcgttttggt
     2221 ccgcggccgc cggccgctga tctggaagaa gcaatgaaag ctgctgttaa gtctccgaat
     2281 caggtattgt tcctgacagg tgtattccca tccggtaaac gcggatactt tgcagttgat
     2341 ctgactcagg aataaattat aaattaaggt aagaagattg taggataagc taatgaaata
     2401 gaaaaaggat gccgtcacac aacttgtcgg cattcttttt tgttttatta gttgaaaata
     2461 tagtgaaaaa gttgcctaaa tatgtatgtt aacaaattat ttgtcgtaac tttgcactcc
     2521 aaatctgttt ttaacatatg gcactaatgc agaaaaaatt gactcgttct cagcagaaac
     2581 acttggacat catcaacgct gctaaagaag aattcatcga attcggtttc ttggctgcta
     2641 acatggaccg tatcacttct tctgctgaag tatctaaacg tactttgtac cgtcacttcg
     2701 aatctaaaga agtattgttc gaatctgtat tgactatcat caacgactct gtaaacgaat
     2761 ctatctctta ccacttcgac ccgaacaaat ctactgaaga acagttgact gaaatcgctt
     2821 acaaagaaat cgacgtattg tacaaaactt acggtatcgc tttggctcgt actatcgtaa
     2881 tggaattctt gcgtcagccg gaaatggcta aaactttgat ccagaacatc tactctatcc
     2941 gtgctatcac tcagtggttc cgttctgcta tcgaagctaa acgtttgaaa gacgctgacc
     3001 cgaaattgat gactgacgta tacgtatctt tgttccaggg tttgttcttc tggccgcagg
     3061 taatgcactt ggacttggaa ccgcacggtg aagaattgtc tcagaaaatc gaaactttga
     3121 ctactatctt cttgcagtct tacggtgtag ctgaataaca atgaatgggc tttcctcggt
     3181 accaaattcc agaaaagagg cctcccgaaa ggggggcctt ttttcgtttt ggtccctaat
     3241 tgcctatctt ccagtgatgg aacagcattt gtgcattggc tgcaacaatc agccttactt
     3301 gtgcctgttc tatttccgaa ccgaccgctt gtatgaatgc atcaaaattc gttttctcta
     3361 cgttggattc cttgttgctc atattgtgat gataatttct acaaatatag tcattggtaa
     3421 ctatctatga aactgtttga tacttttatc agtccagtag ttttacaagg tctttcttca
     3481 tttcttcgtc aatatccctg tatcgtctga aagctttgct tccctccttg tgtcccgaca
     3541 gtgcggaaac aaggttcggg tctttcactt ttttatagat attgccgata aacgtacgtc
     3601 ttgccagatg gctgcttgcc acttcataaa taggtcgttt gatttcgttg tgcgtcaacg
     3661 ggtctaagat tgttacgatg cggtcaactc cagctaattt gaatatcttt tttatggcat
     3721 cattgtactt ttgctcggat atgaacggca acagttttcc ctcatattct ttgtagcgtt
     3781 caaggatttc tttcgctttg tcgttaagtg gaacacgtac cgtaaccgga ttcccctctt
     3841 tggttttctt gggaatatat tctatggctt cattgaccac atttagtttg gtcattcggt
     3901 acaggtcgct caccctgcat cctatcagtg tctgaaatat gaatatatcc ctctgtattg
     3961 ccagttgtgg ggtggcagaa aggtctgcat taaaaatcct gtccctttct tcgagtgtta
     4021 tataataagg tgtaccatat gtacactcct ctatcggaaa cttgtcgaaa ggtctgtttg
     4081 tggtgcgttt gttatcgaag caccacagga agaatgtgcg tattcttgaa aaacagtcta
     4141 tcagcgtgtt tttgcttctg ggctgtggtg tcctcttttc gggaatggct tcataaatgc
     4201 tcgggtaaag ttcataatac tggtattcgt tctgaaagaa atcccacata tcccgaagcg
     4261 tgtcaggtgt taccaaatcc acatcaagga taaagccctt ttgtcctctc tttgtagccc
     4321 ttacatatag ttcataacgc agtaaggctc ttttgacaac ccggaaattt ttctttcgta
     4381 cttccgacaa agggtgcttg tttagaaatt catcgaatag ttctccaatg gtaggcttga
     4441 taaccacttc ctccggcaaa aaatattttt caggatggta aaatttatca agtgttgttt
     4501 ttagccattc tttgtctatt gcttcctttt cttgttgata taccttctcg atataggttt
     4561 tcaactggcg tatctcttca tttatatggg tacgcatttc ttcattgcag acagctttcg
     4621 tttttacaca ttcgtcttta tcatcccata gattgggatt gatggctaac tgagtaggag
     4681 caacggagtc taactgcctt ccgtttctaa aacggacata gattgtagcc atggattctg
     4741 tatcatatcg tttggctgct tttttaatga taaaggttac tttcatagac tttcaggttg
     4801 aattttactc tgctgcaaat ataaatattt tccccagcat tttccccaca tctgctaaat
     4861 attttgcaat tcgattaaac ttggattaaa atttaatagg attataacat attgaaatac
     4921 agtgtaattg tggcgttttt ccgcattttt ctttttaccc acattttccc cactttaatt
     4981 atttcaaatg cggggtctgg gtacaagaaa gaaagctaag tatttgatag ttcaatactt
     5041 agctttttct tttgcttgaa ttttccccac attttcccca cacgtgcaaa aaatatagca
     5101 gtaagtcatt atttcttttg gttgaacgta gagagtagcg atattaaaag aatccgatga
     5161 gaaaagacta atatttatct atccattcag tttgatttct caggacttta catcgtcctg
     5221 aaagtatttg ttggtaccga ggacgcgtaa acatttacag ttgcatgtgg cctattgttt
     5281 ttagccgtta aatattttat aactattaaa tagcgataca aattgttcga aactaatatt
     5341 gtttatatca tatattctcg catgttttaa agctttatta aattgatttt ttgtaaacag
     5401 tttttcgtac tctttgttaa cccatttcat tacaaaagtt tcatattttt ttctctcttt
     5461 aaatgccatt tttgctggct ttctttttaa tacaattaat gtgctatcca ctttaggttt
     5521 tggatggaaa taatacctag gaatttttgc taatatagaa atatctacct ctgccattaa
     5581 cagcaatgct agtgatctgt ttgtatctaa taacatttta gcaaaaccat attccactat
     5641 taaataactt attgtggctg aactttcaaa aacaattttt cgaattatat ttgtgcttat
     5701 gttgtaaggt atgctgccaa atattttata tggattgtgg ctaggaaatg taaatttcag
     5761 tatatcatca tttactattt gatagttagg ataatttaag agcttattac gagttacctc
     5821 acataattta gaatcaattt ctatcgccgt tacaaaatta catctcttta ccaatccagc
     5881 agtaaaatga cctttccctg cacctatttc aaagatgtta tctttttcat ctaaacttat
     5941 gcaattcatt attttttcta tgtgatattt tgaagtaata aaattttgac tatcttttat
     6001 atttactttg ttcattataa cctctcctta atttattgca tctcttttcg aatatttatg
     6061 ttttttgaga aaagaacgta ctcatggttc atcccgatat gcgtatcggt ctgtatatca
     6121 gcaactttct atgtgtttca actacaatag tcatctattc tcatctttct gagtccaccc
     6181 cctgcaaagc ccctctttac gacataaaaa ttcggtcgga aaaggtatgc aaaagatgtt
     6241 tctctcttta agagaaactc ttcgggatgc aaaaatatga aaataactcc aattcaccaa
     6301 attatatagc gactttttta caaaatgcta aaatttgttg atttccgtca agcaattgtt
     6361 gagcaaaaat gtcttttacg ataaaatgat acctcaatat caactgttta gcaaaacgat
     6421 atttctctta aagagagaaa cacctttttg ttcaccaatc cccgactttt aatcccgcgg
     6481 ccatgattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt
     6541 ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga
     6601 tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa
     6661 gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct
     6721 gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat
     6781 acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga
     6841 tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc
     6901 caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat
     6961 gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa
     7021 cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac
     7081 tggcgaacta cttactctag cttcccggca acaattaata gactggatgg aggc
//
