LOCUS       sequence-476753-        5855 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible Promoter (anhydrotetracycline-induced, tetO/tetR system), 
            replicates via RepA; ermF, C-terminal 3x-FLAG, R6K ori, RP4 oriT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5855)
  AUTHORS   Schnizlein MK, Dubey AA, Fiebig A, Crosson S
  TITLE     Genetic- and culture-based tools for studying Bacteroides fragilis.
  JOURNAL   Microbiol Resour Announc. 2025 May 8;14(5):e0000625. doi: 
            10.1128/mra.00006-25. Epub 2025 Mar 25.
  PUBMED    40130927
REFERENCE   2  (bases 1 to 5855)
  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: "Microbiol 
            Resour Announc."; date: "2025-05-8"; volume: "14(5):e0000625. doi"; 
            pages: " 10.1128/mra.00006-25. Epub 2025 Mar 25"
COMMENT     Sequence Label: pMS05
FEATURES             Location/Qualifiers
     source          1..5855
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..389
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     CDS             complement(419..1042)
                     /codon_start=1
                     /gene="tetR from transposon Tn10"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     950..969
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     protein_bind    1313..1331
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1375..1393
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             1566..1631
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             join(4585..4653,4654..5445)
                     /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(4803..4822)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     oriT            5576..5685
                     /note="incP origin of transfer"
ORIGIN
        1 gatctgaaga tcagcagttc aacctgttga tagtacgtac taagctctca tgtttcacgt
       61 actaagctct catgtttaac gtactaagct ctcatgttta acgaactaaa ccctcatggc
      121 taacgtacta agctctcatg gctaacgtac taagctctca tgtttcacgt actaagctct
      181 catgtttgaa caataaaatt aatataaatc agcaacttaa atagcctcta aggttttaag
      241 ttttataaga aaaaaaagaa tatataaggc ttttaaagct tttaaggttt aacggttgtg
      301 gacaacaagc cagggatgta acgcactgag aagcccttag agcctctcaa agcaattttg
      361 agtgacacag gaacacttaa cggctgacat gggaattctt cccctccacc gcggtggctt
      421 aagacccact ttcacattta agttgttttt ctaatccgca tatgatcaat tcaaggccga
      481 ataagaaggc tggctctgca ccttggtgat caaataattc gatagcttgt cgtaataatg
      541 gcggcatact atcagtagta ggtgtttccc tttcttcttt agcgacttga tgctcttgat
      601 cttccaatac gcaacctaaa gtaaaatgcc ccacagcgct gagtgcatat aatgcattct
      661 ctagtgaaaa accttgttgg cataaaaagg ctaattgatt ttcgagagtt tcatactgtt
      721 tttctgtagg ccgtgtacct aaatgtactt ttgctccatc gcgatgactt agtaaagcac
      781 atctaaaact tttagcgtta ttacgtaaaa aatcttgcca gctttcccct tctaaagggc
      841 aaaagtgagt atggtgccta tctaacatct caatggctaa ggcgtcgagc aaagcccgct
      901 tattttttac atgccaatac aatgtaggct gctctacacc tagcttctgg gcgagtttac
      961 gggttgttaa accttcgatt ccgacctcat taagcagctc taatgcgctg ttaatcactt
     1021 tacttttatc taatctagac atattcgttt aatatcataa ataatttatt ttattttaaa
     1081 atgcgcgggt gcaaaggtaa gaggttttat tttaactacc aaatgttttc ggaagttttt
     1141 tcgcttttct ttttctatcg tttctcagac tctcttagcg aaagggaaag aaggtaaaga
     1201 agaaaaacaa aacgcctttt ctttttttgc acccgctttc caagagaaga aagccttgtt
     1261 aaattgactt agtgtaaaag cgcagtactg cttgaccata agaacaaaaa aatctctatc
     1321 actgataggg ataaagtttg gaagataaag ctaaaagttc ttatctttgc agtctcccta
     1381 tcagtgatag agacgaaata aagacatata aaagaaaaga caccccgggc atgccaggat
     1441 ccgaattcga gctcggcgcg cctgcaggtc gacaagcttg cggccgcata atgcttaagt
     1501 ccatatggca gatctcaatt ggatatcggc cggccacgcg atcgctgacg tcggtaccct
     1561 cgagtgacta caaggaccac gacggcgact ataaggacca cgacatcgac tacaaggatg
     1621 acgacgacaa gtgagggccc tataatttgg tgaattggag ttattttcat atttttgcat
     1681 cccgaagagt ttctcttaaa gagagaaaca tcttttgcat accttttccg accgaatttt
     1741 tatgtcgtaa agaggggctt tgcagggggt ggactcagaa agatgagaat agatgactat
     1801 tgtagttgaa acacatagaa agttgctgat atacagaccg atacgcatat cgggatgaac
     1861 catgagtacg ttcttttctc aaaaaacata aatattcgaa aagagatgca ataaattaag
     1921 gagaggttat agcttagcat gacaaaaaag aaattgcccg ttcgttttac gggtcagcac
     1981 tttactattg ataaagtgct aataaaagat gcaataagac aagcaaatat aagtaatcag
     2041 gatacggttt tagatattgg ggcaggcaag gggtttctta ctgttcattt attaaaaatc
     2101 gccaacaatg ttgttgctat tgaaaacgac acagctttgg ttgaacattt acgaaaatta
     2161 ttttctgatg cccgaaatgt tcaagttgtc ggttgtgatt ttaggaattt tgcagttccg
     2221 aaatttcctt tcaaagtggt gtcaaatatt ccttatggca ttacttccga tattttcaaa
     2281 atcctgatgt ttgagagtct tggaaatttt ctgggaggtt ccattgtcct tcaattagaa
     2341 cctacacaaa agttattttc gaggaagctt tacaatccat ataccgtttt ctatcatact
     2401 ttttttgatt tgaaacttgt ctatgaggta ggtcctgaaa gtttcttgcc accgccaact
     2461 gtcaaatcag ccctgttaaa cattaaaaga aaacacttat tttttgattt taagtttaaa
     2521 gccaaatact tagcatttat ttcctatctg ttagagaaac ctgatttatc tgtaaaaaca
     2581 gctttaaagt cgattttcag gaaaagtcag gtcaggtcaa tttcggaaaa attcggttta
     2641 aaccttaatg ctcaaattgt ttgtttgtct ccaagtcaat ggttaaactg ttttttggaa
     2701 atgctggaag ttgtccctga aaaatttcat ccttcgtagg catgcggcgc tccccgaccg
     2761 atgattttta agtgactgat tttgtgctgt tttgggggta tattaagaat aaaagaaata
     2821 gaataagtta agtacttgat acacaatata gggcattttc catattggaa attctcattt
     2881 tccaatctag aaaataccga tttcctaata tgatactaaa taggaaaata atatttccct
     2941 taatattgtt tttatggaaa ataatagttt actttgtgga gaataatatt tcccaaaaac
     3001 atatcaaaat ggaaaataaa aaagcagtta agttaaccga ttttcaaaag aacgaagaaa
     3061 atccttttat gaaacaagct atagaaggta ttgaaaatca tgttgttaaa aagtataaga
     3121 gtaatagtgg tggcgataag agagctgtag tagctttagc cgacactgaa actggagaag
     3181 tgtttaagac ttcgtttatc cgtcaaatag aagtagatga agaacaattc actaaattgt
     3241 atctttctaa ctttgctgca ttctttgacc tatcacaagc agctattcgg gtttttggtt
     3301 actttatgac ctgcatgaaa cccaaaaatg atttaatcat tttcaataga aaaaaatgcc
     3361 tagaatatac caaatacaaa acagacaaag ccgtttataa aggacttgca gaacttgtaa
     3421 aagctgaaat catagcccga ggaccagccg ataatctttg gtttattaat cctctgatag
     3481 tattcaatgg tgaccgagtg acatttgcta aaacatacgt tcggaaaaag actttagctg
     3541 cccaaaagaa agaagaagca gagaaacgac aattatcact tggctttgat gaacagtaac
     3601 actccattga gtgaagctgc cgtttggtcg ctcccccttt gggggcgggg ggggatagat
     3661 aaagttcctc tatgtaaagt tataatgggg gatgaaaggc aaggttcgct aaccttaccc
     3721 cgattgttta tgcttcccag cattcaatct atccccgttt atgggtttct ctttcaccct
     3781 acaaagataa cctcatgggg gaaaaatgtc caagaatatg gggtaaaaac tatcaagtcg
     3841 gtagaaaata agtatctttg aagtacattt ttagtagagg tactggtgtg cctaggcgaa
     3901 gcagataggc aaaaataaag ccgctggaga tgtcggatgt ccgtcaatta actccagcgg
     3961 cttgtatttt ggtaactctc ccgtatcggt cagctaccgt ttggtcagca acaaaggtag
     4021 tactttatta tcgcaaatcc aaatgttttt ccctttattt gtgcatttac aaaacattca
     4081 ctaacttcgc acctatgcag tatggtgata gcacactagc accctaaaga gtgcgtgtgt
     4141 aaggggacaa gcccccttaa ccccctgtca gccggctgtc accggctgtt atcgattgtc
     4201 acaaaaatgt taacttatgg gagcaacaag tattcatgta caagcagtga agccggggag
     4261 cgagattcac aactttaggg aaaaagagtt ggactatgtt cgtcccgaac ttagtcattt
     4321 gaatgaaagc tgggttggag atagcatttc ccatcggctg gagagtgcaa aacaaagatg
     4381 ctagcatata gcgacttttt tacaaaatgc taaaatttgt tgatttccgt caagcaattg
     4441 ttgagcaaaa atgtctttta cgataaaatg atacctcaat atcaactgtt tagcaaaacg
     4501 atatttctct taaagagaga aacacctttt tgttcaccaa tccccgactt ttaatcccgc
     4561 ggccatgatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     4621 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     4681 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     4741 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     4801 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     4861 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     4921 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     4981 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     5041 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     5101 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     5161 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     5221 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     5281 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     5341 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     5401 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     5461 tactcataac gcgtcaattc gagggggatc aattccgtga taggtgggct gcccttcctg
     5521 gttggcttgg tttcatcagc catccgcttg ccctcatctg ttacgccggc ggtagccggc
     5581 cagcctcgca gagcaggatt cccgttgagc accgccaggt gcgaataagg gacagtgaag
     5641 aaggaacacc cgctcgcggg tgggcctact tcacctatcc tgcccggctg acgccgttgg
     5701 atacaccaag gaaagtctac acgaaccctt tggcaaaatc ctgtatatcg tgcgaaaaag
     5761 gatggatata ccgaaaaaat cgctataatg accccgaagc agggttatgc agcggaaaac
     5821 ggaattgatc cggccacgat gcgtccggcg tagag
//
