LOCUS       sequence-263619-        3565 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ORBIT integrating plasmid for C-terminal tagging with FLAG-His tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3565)
  AUTHORS   Murphy KC, Nelson SJ, Nambi S, Papavinasasundaram K, Baer CE, 
            Sassetti CM
  TITLE     ORBIT: a New Paradigm for Genetic Engineering of Mycobacterial 
            Chromosomes.
  JOURNAL   MBio. 2018 Dec 11;9(6). pii: mBio.01467-18. doi: 
            10.1128/mBio.01467-18.
  PUBMED    30538179
REFERENCE   2  (bases 1 to 3565)
  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: "MBio."; 
            date: "2018-12-11"; volume: "9(6). pii: mBio.01467-18. doi"; pages: 
            " 10.1128/mBio.01467-18"
COMMENT     Sequence Label: pKM491-Flag-His
FEATURES             Location/Qualifiers
     source          1..3565
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    440..477
                     /label=Bxb1 attB
                     /bound_moiety="Mycobacteriophage Bxb1 integrase"
                     /note="Minimal attB site for the Mycopbacteriophage Bxb1 
                     integrase (Voutev and Mann, 2017)"
     CDS             510..533
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             534..551
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     terminator      594..621
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     rep_origin      complement(788..1376)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(868..887)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     complement(1641..1660)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     terminator      complement(3513..3559)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
ORIGIN
        1 ggcagtcgat cgagaggtga ccacaacgac gcgcccgctt tgatcgggga cgtctgcggc
       61 cgaccattta cgggtcttgt tgtcgttggc ggtcatgggc cgaacatact cacccggatc
      121 ggagggccga ggacaaggtc gaacgagggg catgacccgg tgcggggctt cttgcactcg
      181 gcataggcga gtgctaagaa taacgttggc actcgcgacc ggtgagtgct aggtcgggac
      241 ggtgaggcca ggcccgtcgt cgcagcgagt ggcagcgagg acaacttgag ccgtccgtcg
      301 cgggcactgc gcccggccag cgtaagtagc ggggttgccg tcacccggtg acccccgttt
      361 catccccgat ccggaggaat cacttcgcaa tggccaagac aattgcggat ccatcgatag
      421 aaaggaggaa ggaaagcttg gcttgtcgac gacggcggtc tccgtcgtca ggatcatcgg
      481 gcggcggcgg cgagaacctg tacttccagg actacaagga cgacgacgac aagcaccacc
      541 accaccacca ctgatgactc gagtctagag catgcactag tgcggccgct agcagaaggc
      601 catcctgacg gatggccttt tcttaagtcg cgttgcgctc ggtcgttcgg ctgcggcgag
      661 cggtatcagc tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag
      721 gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc
      781 tggcgttttt ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc
      841 agaggtggcg aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc
      901 tcgtgcgctc tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctcccct
      961 cgggaagcgt ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg
     1021 ttcgctccaa gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat
     1081 ccggtaacta tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag
     1141 ccactggtaa caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt
     1201 ggtggcctaa ctacggctac actagaagga cagtatttgg tatctgcgct ctgctgaagc
     1261 cagttacctt cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta
     1321 gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag
     1381 atcctttgat cttttctacg gggtctgacg ctcatggaac tagagcatga gattatcaaa
     1441 aaggatcttc acctagatcc ttttaaatta aaaatcggca cgtaagaggt tccaactttc
     1501 accataatga aataagatca ctaccgggcg tattttttga gttatcgaga ttttcaggag
     1561 ctaaggaagc taaaatggag aaaaaaatca ctggatatac caccgttgat atatcccaat
     1621 ggcatcgtaa agaacatttt gaggcatttc agtcagttgc tcaatgtacc tataaccaga
     1681 ccgttcagct ggatattacg gcctttttaa agaccgtaaa gaaaaataag cacaagtttt
     1741 atccggcctt tattcacatt cttgcccgcc tgatgaatgc tcatccggaa ttccgtatgg
     1801 caatgaaaga cggtgagctg gtgatatggg atagtgttca cccttgttac accgttttcc
     1861 atgagcaaac tgaaacgttt tcatcgctct ggagtgaata ccacgacgat ttccggcagt
     1921 ttctacacat atattcgcaa gatgtggcgt gttacggtga aaacctggcc tatttcccta
     1981 aagggtttat tgagaatatg tttttcgtct cagccaatcc ctgggtgagt ttcaccagtt
     2041 ttgatttaaa cgtggccaat atggacaact tcttcgcccc cgttttcacc atgggcaaat
     2101 attatacgca aggcgacaag gtgctgatgc cgctggcgat tcaggttcat catgccgttt
     2161 gtgatggctt ccatgtcggc agaatgctta atgaattaca acagtactgc gatgagtggc
     2221 tggcggaaac ggcaatgctg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     2281 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     2341 tcatccatag ttgcctgact ccccgacgtg gccgaccagc ccgtcatcgt caacgcctga
     2401 tccgcggtgc ggacaggccg tgtcgcgacc ggccgtgcgg aattaagccg gcccgtaccc
     2461 tgtgaataga ggtccgctgt gacacaagaa tccctgttac ttctcgaccg tattgattcg
     2521 gatgattcct acgcgagcct gcggaacgac caggaattct gggagccgct ggcccgccga
     2581 gccctggagg agctcgggct gccggtgccg ccggtgctgc gggtgcccgg cgagagcacc
     2641 aaccccgtac tggtcggcga gcccggcccg gtgatcaagc tgttcggcga gcactggtgc
     2701 ggtccggaga gcctcgcgtc ggagtcggag gcgtacgcgg tcctggcgga cgccccggtg
     2761 ccggtgcccc gcctcctcgg ccgcggcgag ctgcggcccg gcaccggagc ctggccgtgg
     2821 ccctacctgg tgatgagccg gatgaccggc accacctggc ggtccgcgat ggacggcacg
     2881 accgaccgga acgcgctgct cgccctggcc cgcgaactcg gccgggtgct cggccggctg
     2941 cacagggtgc cgctgaccgg gaacaccgtg ctcacccccc attccgaggt cttcccggaa
     3001 ctgctgcggg aacgccgcgc ggcgaccgtc gaggaccacc gcgggtgggg ctacctctcg
     3061 ccccggctgc tggaccgcct ggaggactgg ctgccggacg tggacacgct gctggccggc
     3121 cgcgaacccc ggttcgtcca cggcgacctg cacgggacca acatcttcgt ggacctggcc
     3181 gcgaccgagg tcaccgggat cgtcgacttc accgacgtct atgcgggaga ctcccgctac
     3241 agcctggtgc aactgcatct caacgccttc cggggcgacc gcgagatcct ggccgcgctg
     3301 ctcgacgggg cgcagtggaa gcggaccgag gacttcgccc gcgaactgct cgccttcacc
     3361 ttcctgcacg acttcgaggt gttcgaggag accccgctgg atctctccgg cttcaccgat
     3421 ccggaggaac tggcgcagtt cctctggggg ccgccggaca ccgcccccgg cgcctgacgc
     3481 cccctctagc tgatcaccgc ggccatgatg gcataaaacg aaaggcccag tctttcgact
     3541 gagcctttcg ttttatttga tgcct
//
