LOCUS       sequence-475946-        5285 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For allelic exchange into Bacteroides fragilis (pLGB13-based); ermC,
            aTC-inducible counterselection with ssbfe1; R6K ori, RP4 oriT.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5285)
  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 5285)
  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: pMS01
FEATURES             Location/Qualifiers
     source          1..5285
                     /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(416..1039)
                     /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     947..966
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     protein_bind    1310..1328
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1372..1390
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             join(4015..4083,4084..4875)
                     /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(4233..4252)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     oriT            5006..5115
                     /note="incP origin of transfer"
ORIGIN
        1 gatctgaaga tcagcagttc aacctgttga tagtacgtac taagctctca tgtttcacgt
       61 actaagctct catgtttcac 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 gggaattccc ctccaccgcg gtggcttaag
      421 acccactttc acatttaagt tgtttttcta atccgcatat gatcaattca aggccgaata
      481 agaaggctgg ctctgcacct tggtgatcaa ataattcgat agcttgtcgt aataatggcg
      541 gcatactatc agtagtaggt gtttcccttt cttctttagc gacttgatgc tcttgatctt
      601 ccaatacgca acctaaagta aaatgcccca cagcgctgag tgcatataat gcattctcta
      661 gtgaaaaacc ttgttggcat aaaaaggcta attgattttc gagagtttca tactgttttt
      721 ctgtaggccg tgtacctaaa tgtacttttg ctccatcgcg atgacttagt aaagcacatc
      781 taaaactttt agcgttatta cgtaaaaaat cttgccagct ttccccttct aaagggcaaa
      841 agtgagtatg gtgcctatct aacatctcaa tggctaaggc gtcgagcaaa gcccgcttat
      901 tttttacatg ccaatacaat gtaggctgct ctacacctag cttctgggcg agtttacggg
      961 ttgttaaacc ttcgattccg acctcattaa gcagctctaa tgcgctgtta atcactttac
     1021 ttttatctaa tctagacata ttcgtttaat atcataaata atttatttta ttttaaaatg
     1081 cgcgggtgca aaggtaagag gttttatttt aactaccaaa tgttttcgga agttttttcg
     1141 cttttctttt tctatcgttt ctcagactct cttagcgaaa gggaaagaag gtaaagaaga
     1201 aaaacaaaac gccttttctt tttttgcacc cgctttccaa gagaagaaag ccttgttaaa
     1261 ttgacttagt gtaaaagcgc agtactgctt gaccataaga acaaaaaaat ctctatcact
     1321 gatagggata aagtttggaa gataaagcta aaagttctta tctttgcagt ctccctatca
     1381 gtgatagaga cgaaataaag acatataaaa gaaaagacac atgaagaaaa ttttatcttt
     1441 gcttgtgatg gctattgtag ctatacagtt ctcatttgcc ggcgcttata gaaatgaata
     1501 taaaacgcaa agtgaattta atgtcatctt ggaacggggg gatgactatg aaggatttgt
     1561 agttggcttg ggatatactt ggatgagtag caaggtaatt ctaccggtaa atcaaaacgg
     1621 atggagtcca atatctcgga atgtttcggt ggacgagagt tttcatacga tagtctcaga
     1681 aaggaaatac gatacctccc aatatgccta tgaaaaaagt ctgatgcaag atccaacaaa
     1741 agtttccgaa aaggtccgtg acttaatagt taaaaacaaa ggtaataata tcactgagat
     1801 aaatttaggc caagaaaagc aatatttgcc cacggataat agtcagataa gtattgtaat
     1861 tactgacact ggcagtcggt acgaaattgt gattagtgcc acggataact caaatggaaa
     1921 gacttatgag gcaaagtatg agagtttgac ggacttggtg tcagcggtac gcgattctgg
     1981 tagcctgcct gctgtaaata aggaaggacc caatctggaa ggactggcag ggttaggatt
     2041 cggaattgct gaaacagccg gaaattgggc tgagaagatt atggataatc gaggtgcgta
     2101 cttacctaag cagatgcgtt tctcgcccaa aacgcttccg ccgattataa gattgccttt
     2161 agggaactat caagtccccg ctaaaggtat gagtagagtg cgtggagtag gtaaagcttt
     2221 gggatgggca ggaatggtgc ttactggcta tcaagttgtg cgtgatgtgc aaaatggtcg
     2281 atttgccgtg gcaggtacaa gaattgccgt agcaggttta gcttatggcg ttacttttat
     2341 tccctatgtc ggttgggtct tagctatcgg tatcggagtg gctgattata cttggggtga
     2401 cgagttctat gactggatag acaatagagc ttctgaattg gaaatgtggt gggacggtgt
     2461 aagattagca ttatgagtgg atccgaattc gagctcggcg cgcctgcagg tcgacaagct
     2521 tgcggccgca taatgcttaa gtccatatgg cagatctcaa ttggatatcg gccggccacg
     2581 cgatcgctga cgtcggtacc ctcgagtctg gtaaagaacc cgggctgcag gaattcgata
     2641 tcaagcttat cgataccgtc gactaattgc ctatcttcca gtgatggaac agcatttgtg
     2701 cattggctgc aacaatcagc cttacttgtg cctgttctat ttccgaaccg accgcttgta
     2761 tgaatccaac agttgcatgt ggcctattgt ttttagccgt taaatatttt ataactatta
     2821 aatagcgata caaattgttc gaaactaata ttgtttatat catatattct cgcatgtttt
     2881 aaagctttat taaattgatt ttttgtaaac agtttttcgt actctttgtt aacccatttc
     2941 attacaaaag tttcatattt ttttctctct ttaaatgcca tttttgctgg ctttcttttt
     3001 aatacaatta atgtgctatc cactttaggt tttggatgga aataatacct aggaattttt
     3061 gctaatatag aaatatctac ctctgccatt aacagcaatg ctagtgatct gtttgtatct
     3121 aataacattt tagcaaaacc atattccact attaaataac ttattgtggc tgaactttca
     3181 aaaacaattt ttcgaattat atttgtgctt atgttgtaag gtatgctgcc aaatatttta
     3241 tatggattgt ggctaggaaa tgtaaatttc agtatatcat catttactat ttgatagtta
     3301 ggataattta agagcttatt acgagttacc tcacataatt tagaatcaat ttctatcgcc
     3361 gttacaaaat tacatctctt taccaatcca gcagtaaaat gacctttccc tgcacctatt
     3421 tcaaagatgt tatctttttc atctaaactt atgcaattca ttattttttc tatgtgatat
     3481 tttgaagtaa taaaattttg actatctttt atatttactt tgttcattat aacctctcct
     3541 taatttattg catctctttt cgaatattta tgttttttga gaaaagaacg tactcatggt
     3601 tcatcccgat atgcgtatcg gtctgtatat cagcaacttt ctatgtgttt caactacaat
     3661 agtcatctat tctcatcttt ctgagtccac cccctgcaaa gcccctcttt acgacataaa
     3721 aattcggtcg gaaaaggtat gcaaaagatg tttctctctt taagagaaac tcttcgggat
     3781 gcaaaaatat gaaaataact ccaattcacc aaattatata gcgacttttt tacaaaatgc
     3841 taaaatttgt tgatttccgt caagcaattg ttgagcaaaa atgtctttta cgataaaatg
     3901 atacctcaat atcaactgtt tagcaaaacg atatttctct taaagagaga aacacctttt
     3961 tgttcaccaa tccccgactt ttaatcccgc ggccatgatt gaaaaaggaa gagtatgagt
     4021 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     4081 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     4141 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     4201 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
     4261 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     4321 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     4381 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     4441 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     4501 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     4561 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     4621 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     4681 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     4741 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     4801 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     4861 attaagcatt ggtaactgtc agaccaagtt tactcataac gcgtcaattc gagggggatc
     4921 aattccgtga taggtgggct gcccttcctg gttggcttgg tttcatcagc catccgcttg
     4981 ccctcatctg ttacgccggc ggtagccggc cagcctcgca gagcaggatt cccgttgagc
     5041 accgccaggt gcgaataagg gacagtgaag aaggaacacc cgctcgcggg tgggcctact
     5101 tcacctatcc tgcccggctg acgccgttgg atacaccaag gaaagtctac acgaaccctt
     5161 tggcaaaatc ctgtatatcg tgcgaaaaag gatggatata ccgaaaaaat cgctataatg
     5221 accccgaagc agggttatgc agcggaaaac ggaattgatc cggccacgat gcgtccggcg
     5281 tagag
//
