LOCUS       sequence-475922-        5425 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive expression Promoter (RpoD sigma factor, BT_RS06635), 
            integrates via IntN1; 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 5425)
  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 5425)
  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: pMS02
FEATURES             Location/Qualifiers
     source          1..5425
                     /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             1025..1090
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             join(4155..4223,4224..5015)
                     /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(4373..4392)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     oriT            5146..5255
                     /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 gggaattccc ctccacctta ttcttcccga
      421 ccgaacgagc ggtattcaag cggagtaagc ccggtacgtt ttttgaaaaa ggagaaaaaa
      481 tattcggtag actggaagcc cagaaaaaaa gaaatttctt ttactgactg ggaagttccc
      541 accaacattt gcttggcttt acgaagtttc agttgatatc agtgatctgg aagaagcaat
      601 gaaagctgct gttaagtctc cgaatcaggt attgttcctg acaggtgtat tcccatccgg
      661 taaacgcgga tactttgcag ttgatctgac tcaggaataa attataaatt aaggtaagaa
      721 gattgtagga taagctaatg aaatagaaaa aggatgccgt cacacaactt gtcggcattc
      781 ttttttgttt tattagttga aaatatagtg aaaaagttgc ctaaatatgt atgttaacaa
      841 attatttgtc gtaactttgc actccaaatc tgtttttaaa gacccgggca tgccaggatc
      901 cgaattcgag ctcggcgcgc ctgcaggtcg acaagcttgc ggccgcataa tgcttaagtc
      961 catatggcag atctcaattg gatatcggcc ggccacgcga tcgctgacgt cggtaccctc
     1021 gagtgactac aaggaccacg acggcgacta taaggaccac gacatcgact acaaggatga
     1081 cgacgacaag tgagggccct ataatttggt gaattggagt tattttcata tttttgcatc
     1141 ccgaagagtt tctcttaaag agagaaacat cttttgcata ccttttccga ccgaattttt
     1201 atgtcgtaaa gaggggcttt gcagggggtg gactcagaaa gatgagaata gatgactatt
     1261 gtagttgaaa cacatagaaa gttgctgata tacagaccga tacgcatatc gggatgaacc
     1321 atgagtacgt tcttttctca aaaaacataa atattcgaaa agagatgcaa taaattaagg
     1381 agaggttata gcttagcatg acaaaaaaga aattgcccgt tcgttttacg ggtcagcact
     1441 ttactattga taaagtgcta ataaaagatg caataagaca agcaaatata agtaatcagg
     1501 atacggtttt agatattggg gcaggcaagg ggtttcttac tgttcattta ttaaaaatcg
     1561 ccaacaatgt tgttgctatt gaaaacgaca cagctttggt tgaacattta cgaaaattat
     1621 tttctgatgc ccgaaatgtt caagttgtcg gttgtgattt taggaatttt gcagttccga
     1681 aatttccttt caaagtggtg tcaaatattc cttatggcat tacttccgat attttcaaaa
     1741 tcctgatgtt tgagagtctt ggaaattttc tgggaggttc cattgtcctt caattagaac
     1801 ctacacaaaa gttattttcg aggaagcttt acaatccata taccgttttc tatcatactt
     1861 tttttgattt gaaacttgtc tatgaggtag gtcctgaaag tttcttgcca ccgccaactg
     1921 tcaaatcagc cctgttaaac attaaaagaa aacacttatt ttttgatttt aagtttaaag
     1981 ccaaatactt agcatttatt tcctatctgt tagagaaacc tgatttatct gtaaaaacag
     2041 ctttaaagtc gattttcagg aaaagtcagg tcaggtcaat ttcggaaaaa ttcggtttaa
     2101 accttaatgc tcaaattgtt tgtttgtctc caagtcaatg gttaaactgt tttttggaaa
     2161 tgctggaagt tgtccctgaa aaatttcatc cttcgtaggc atgcctcatc ggattctttt
     2221 aatatcgcta ctctctacgt tcaaccaaaa gaaataatga cttactgcta tattttttgc
     2281 acgtgtgggg aaaatgtggg gaaaattcaa gcaaaagaaa aagctaagta ttgaactatc
     2341 aaatacttag ctttctttct tgtacccaga ccccgcattt gaaataatta aagtggggaa
     2401 aatgtgggta aaaagaaaaa tgcggaaaaa cgccacaatt acactgtatt tcaatatgtt
     2461 ataatcctat taaattttaa tccaagttta atcgaattgc aaaatattta gcagatgtgg
     2521 ggaaaatgct ggggaaaata tttatatttg cagcagagta aaattcaacc tgaaagtcta
     2581 tgaaagtaac ctttatcatt aaaaaagcag ccaaacgata tgatacagaa tccatggcta
     2641 caatctatgt ccgttttaga aacggaaggc agttagactc cgttgctcct actcagttag
     2701 ccatcaatcc caatctatgg gatgataaag acgaatgtgt aaaaacgaaa gctgtctgca
     2761 atgaagaaat gcgtacccat ataaatgaag agatacgcca gttgaaaacc tatatcgaga
     2821 aggtatatca acaagaaaag gaagcaatag acaaagaatg gctaaaaaca acacttgata
     2881 aattttacca tcctgaaaaa tattttttgc cggaggaagt ggttatcaag cctaccattg
     2941 gagaactatt cgatgaattt ctaaacaagc accctttgtc ggaagtacga aagaaaaatt
     3001 tccgggttgt caaaagagcc ttactgcgtt atgaactata tgtaagggct acaaagagag
     3061 gacaaaaggg ctttatcctt gatgtggatt tggtaacacc tgacacgctt cgggatatgt
     3121 gggatttctt tcagaacgaa taccggtatt atgaacttta cccgagcatt tatgaagcca
     3181 ttcccgaaaa gaggacacca cagcccagaa gcaaaaacac gctgatagac tgtttttcaa
     3241 gaatacgcac attcttcctg tggtgcttcg ataacaaacg caccacaaac agacctttcg
     3301 acaagtttcc gatagaggag tgtacatatg gtacacctta ttatataaca ctcgaagaaa
     3361 gggacaggat ttttaatgca gacctttctg ccaccccaca actggcaata cagagggata
     3421 tattcatatt tcagacactg ataggatgca gggtgagcga cctgtaccga atgaccaaac
     3481 taaatgtggt caatgaagcc atagaatata ttcccaagaa aaccaaagag gggaatccgg
     3541 ttacggtacg tgttccactt aacgacaaag cgaaagaaat ccttgaacgc tacaaagaat
     3601 atgagggaaa actgttgccg ttcatatccg agcaaaagta caatgatgcc ataaaaaaga
     3661 tattcaaatt agctggagtt gaccgcatcg taacaatctt agacccgttg acgcacaacg
     3721 aaatcaaacg acctatttat gaagtggcaa gcagccatct ggcaagacgt acgtttatcg
     3781 gcaatatcta taaaaaagtg aaagacccga accttgtttc cgcactgtcg ggacacaagg
     3841 agggaagcaa agctttcaga cgatacaggg atattgacga agaaatgaag aaagaccttg
     3901 taaaactact ggactgataa aagtatcaaa cagtttcata gatagttagg ctagcatata
     3961 gcgacttttt tacaaaatgc taaaatttgt tgatttccgt caagcaattg ttgagcaaaa
     4021 atgtctttta cgataaaatg atacctcaat atcaactgtt tagcaaaacg atatttctct
     4081 taaagagaga aacacctttt tgttcaccaa tccccgactt ttaatcccgc ggccatgatt
     4141 gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg
     4201 cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag
     4261 atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg
     4321 agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg
     4381 gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt
     4441 ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga
     4501 cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac
     4561 ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc
     4621 atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc
     4681 gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac
     4741 tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag
     4801 gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg
     4861 gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta
     4921 tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg
     4981 ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcataac
     5041 gcgtcaattc gagggggatc aattccgtga taggtgggct gcccttcctg gttggcttgg
     5101 tttcatcagc catccgcttg ccctcatctg ttacgccggc ggtagccggc cagcctcgca
     5161 gagcaggatt cccgttgagc accgccaggt gcgaataagg gacagtgaag aaggaacacc
     5221 cgctcgcggg tgggcctact tcacctatcc tgcccggctg acgccgttgg atacaccaag
     5281 gaaagtctac acgaaccctt tggcaaaatc ctgtatatcg tgcgaaaaag gatggatata
     5341 ccgaaaaaat cgctataatg accccgaagc agggttatgc agcggaaaac ggaattgatc
     5401 cggccacgat gcgtccggcg tagag
//
