LOCUS       sequence-211180-        5685 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Suicide vector for generation of gene deletions in mycobacteria. 
            Contains the two genes sacB and galK for negative selection. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5685)
  AUTHORS   Arnold FM, Hohl M, Remm S, Koliwer-Brandl H, Adenau S, Chusri S, 
            Sander P, Hilbi H, Seeger MA
  TITLE     A uniform cloning platform for mycobacterial genetics and protein 
            production.
  JOURNAL   Sci Rep. 2018 Jun 22;8(1):9539. doi: 10.1038/s41598-018-27687-5.
  PUBMED    29934571
REFERENCE   2  (bases 1 to 5685)
  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: "Sci Rep."; 
            date: "2018-06-22"; volume: "8(1):9539. doi"; pages: " 
            10.1038/s41598-018-27687-5"
COMMENT     Sequence Label: pKO (Apr)
FEATURES             Location/Qualifiers
     source          1..5685
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(105..110,111..128,129..135)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    141..160
                     /label=lac operator (symmetric)
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG). The 
                     symmetric lac operator was optimized for tight binding of 
                     lac repressor."
     primer_bind     1736..1758
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1750..1767
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1751..1767
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      complement(2011..2599)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2091..2110)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2744..3189
                     /gene="sacR"
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             join(3190..3276,3277..4611)
                     /codon_start=1
                     /gene="Bacillus subtilis sacB"
                     /product="secreted levansucrase that renders bacterial 
                     growth sensitive to sucrose"
                     /label=SacB
                     /note="negative selection marker"
                     /translation="MNIKKFAKQATVLTFTTALLAGGATQAFAKETNQKPYKETYGISH
                     ITRHDMLQIPEQQKNEKYKVPEFDSSTIKNISSAKGLDVWDSWPLQNTDGTVANYHGYH
                     IVFALAGDPKNADDTSIYMFYQKVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEW
                     SGSATFTSDGKIRLFYTDFSGKHYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDG
                     KTYQNVQQFIDEGNYSSGDNHTLRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKA
                     YYGKSTSFFRQESQKLLQSDKKRTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDE
                     IERANVFKMNGKWYLFTDSRGSKMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLK
                     MDLDPNDVTFTYSHFAVPQAKGNNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSV
                     VKDSILEQGQLTVNK"
     CDS             4796..5596
                     /codon_start=1
                     /gene="aac(3)IV"
                     /product="aminoglycoside 3-N-acetyltransferase type IV     
                               "
                     /label=ApmR
                     /note="confers resistance to apramycin (Paget and Davies, 
                     1996)"
                     /translation="MSSAVECNVVQYEWRKAELIGQLLNLGVTPGGVLLVHSSFRSVRP
                     LEDGPLGLIEALRAALGPGGTLVMPSWSGLDDEPFDPATSPVTPDLGVVSDTFWRLPNV
                     KRSAHPFAFAAAGPQAEQIISDPLPLPPHSPASPVARVHELDGQVLLLGVGHDANTTLH
                     LAELMAKVPYGVPRHCTILQDGKLVRVDYLENDHCCERFALADRWLKEKSLQKEGPVGH
                     AFARLIRSRDIVATALGQLGRDPLIFLHPPEAGCEECDAARQSIG"
ORIGIN
        1 tttcttcagc tcgctgatga tatgctgacg ctcaatgccg tttggcctcc gactaacgaa
       61 aatcccgcat ttggacggct gatccatgga tccagacccc aggcttgaca ctttatgctt
      121 ccggctcgta taatgtgtgg aattgtgagc gctcacaatt cgcatgcggt accaagctga
      181 tccgataaca caggaacaga tcttccggag tgtaagaaat gagtctgaaa gaaaaaacac
      241 aatctctgtt tgccaacgca tttggctacc ctgccactca caccattcag gcgcctggcc
      301 gcgtgaattt gattggtgaa cacaccgact acaacgacgg tttcgttctg ccctgcgcga
      361 ttgattatca aaccgtgatc agttgtgcac cacgcgatga ccgtaaagtt cgcgtgatgg
      421 cagccgatta tgaaaatcag ctcgacgagt tttccctcga tgcgcccatt gtcgcacatg
      481 aaaactatca atgggctaac tacgttcgtg gcgtggtgaa acatctgcaa ctgcgtaaca
      541 acagcttcgg cggcgtggac atggtgatca gcggcaatgt gccgcagggt gccgggttaa
      601 gttcttccgc ttcactggaa gtcgcggtcg gaaccgtatt gcagcagctt tatcatctgc
      661 cgctggacgg cgcacaaatc gcgcttaacg gtcaggaagc agaaaaccag tttgtaggct
      721 gtaactgcgg gatcatggat cagctaattt ccgcgctcgg caagaaagat catgccttgc
      781 tgatcgattg ccgctcactg gggaccaaag cagtttccat gcccaaaggt gtggctgtcg
      841 tcatcatcaa cagtaacttc aaacgtaccc tggttggcag cgaatacaac acccgtcgtg
      901 aacagtgcga aaccggtgcg cgtttcttcc agcagccagc cctgcgtgat gtcaccattg
      961 aagagttcaa cgctgttgcg catgaactgg acccgatcgt ggcaaaacgc gtgcgtcata
     1021 tactgactga aaacgcccgc accgttgaag ctgccagcgc gctggagcaa ggcgacctga
     1081 aacgtatggg cgagttgatg gcggagtctc atgcctctat gcgcgatgat ttcgaaatca
     1141 ccgtgccgca aattgacact ctggtagaaa tcgtcaaagc tgtgattggc gacaaaggtg
     1201 gcgtacgcat gaccggcggc ggatttggcg gctgtatcgt cgcgctgatc ccggaagaac
     1261 tggtgcctgc cgtacagcaa gctgtcgctg aacaatatga agcaaaaaca ggtattaaag
     1321 agacttttta cgtttgtaaa ccatcacaag gagcaggaca gtgctgaacg aaactcctgc
     1381 agcgcgtagt acctcacacc gtcctgcggg ttgacgtatg aggcttggat cctcccgctg
     1441 cgctgagtct tcagcgatcc ccatccgcga cgtgccaact aggtctcctc tcgtcgtgaa
     1501 caaggctacc gggttgcaac tcctgtgcaa ctctcaggct tcaacgcgct tctacgacct
     1561 gcaatttctt tccacttaga ggatgcagcc gagagggggt aaaaacctcc cattcgccat
     1621 tcaggctgcg caactgttgg gaagggcgtt tcggtgcggg cctcttcgct attacgccag
     1681 ctggcgaaag ggggatgtgc tgcaaggcga ttaagttggg taacgccagg gttttcccag
     1741 tcacgacgtt gtaaaacgac ggccagtgag cgcgacgtaa taagttgaag agcatctgct
     1801 cttcagcatc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaacatg
     1861 gtcatagctg tttccttgcg tattgggcgc tctccgcttc ctcgctcact gactcgctgc
     1921 gctcggtcgt tcgggtaaag cctggggtgc ctaatgagca aaaggccagc aaaaggccag
     1981 gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg ctccgccccc ctgacgagca
     2041 tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg acaggactat aaagatacca
     2101 ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt ccgaccctgc cgcttaccgg
     2161 atacctgtcc gcctttctcc cttcgggaag cgtggcgctt tctcatagct cacgctgtag
     2221 gtatctcagt tcggtgtagg tcgttcgctc caagctgggc tgtgtgcacg aaccccccgt
     2281 tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt gagtccaacc cggtaagaca
     2341 cgacttatcg ccactggcag cagccactgg taacaggatt agcagagcga ggtatgtagg
     2401 cggtgctaca gagttcttga agtggtggcc taactacggc tacactagaa gaacagtatt
     2461 tggtatctgc gctctgctga agccagttac cttcggaaaa agagttggta gctcttgatc
     2521 cggcaaacaa accaccgctg gtagcggtgg tttttttgtt tgcaagcagc agattacgcg
     2581 cagaaaaaaa ggatctcaag aagatccttt gatcttttct acggggtctg acgctcagtg
     2641 gaacgaaggc gcgcctgagt gagctgatac cgctcgccgc agccgaacga ccgagcgcag
     2701 cgagtcagtg agcgaggaag cggaagatcg ggcccagccg gcccacatat acctgccgtt
     2761 cactattatt tagtgaaatg agatattatg atattttctg aattgtgatt aaaaaggcaa
     2821 ctttatgccc atgcaacaga aactataaaa aatacagaga atgaaaagaa acagatagat
     2881 tttttagttc tttaggcccg tagtctgcaa atccttttat gattttctat caaacaaaag
     2941 aggaaaatag accagttgca atccaaacga gagtctaata gaatgaggtc gaaaagtaaa
     3001 tcgcgcgggt ttgttactga taaagcaggc aagacctaaa atgtgtaaag ggcaaagtgt
     3061 atactttggc gtcacccctt acatatttta ggtctttttt tattgtgcgt aactaacttg
     3121 ccatcttcaa acaggagggc tggaagaagc agaccgctaa cacagtacat aaaaaaggag
     3181 acatgaacga tgaacatcaa aaagtttgca aaacaagcaa cagtattaac ctttactacc
     3241 gcactgctgg caggaggcgc aactcaagcg tttgcgaaag aaacgaacca aaagccatat
     3301 aaggaaacat acggcatttc ccatattaca cgccatgata tgctgcaaat ccctgaacag
     3361 caaaaaaatg aaaaatataa agttcctgag ttcgattcgt ccacaattaa aaatatctct
     3421 tctgcaaaag gcctggacgt ttgggacagc tggccattac aaaacactga cggcactgtc
     3481 gcaaactatc acggctacca catcgtcttt gcattagccg gagatcctaa aaatgcggat
     3541 gacacatcga tttacatgtt ctatcaaaaa gtcggcgaaa cttctattga cagctggaaa
     3601 aacgctggcc gcgtctttaa agacagcgac aaattcgatg caaatgattc tatcctaaaa
     3661 gaccaaacac aagaatggtc aggttcagcc acatttacat ctgacggaaa aatccgttta
     3721 ttctacactg atttctccgg taaacattac ggcaaacaaa cactgacaac tgcacaagtt
     3781 aacgtatcag catcagacag ctctttgaac atcaacggtg tagaggatta taaatcaatc
     3841 tttgacggtg acggaaaaac gtatcaaaat gtacagcagt tcatcgatga aggcaactac
     3901 agctcaggcg acaaccatac gctgagagat cctcactacg tagaagataa aggccacaaa
     3961 tacttagtat ttgaagcaaa cactggaact gaagatggct accaaggcga agaatcttta
     4021 tttaacaaag catactatgg caaaagcaca tcattcttcc gtcaagaaag tcaaaaactt
     4081 ctgcaaagcg ataaaaaacg cacggctgag ttagcaaacg gcgctctcgg tatgattgag
     4141 ctaaacgatg attacacact gaaaaaagtg atgaaaccgc tgattgcatc taacacagta
     4201 acagatgaaa ttgaacgcgc gaacgtcttt aaaatgaacg gcaaatggta cctgttcact
     4261 gactcccgcg gatcaaaaat gacgattgac ggcattacgt ctaacgatat ttacatgctt
     4321 ggttatgttt ctaattcttt aactggccca tacaagccgc tgaacaaaac tggccttgtg
     4381 ttaaaaatgg atcttgatcc taacgatgta acctttactt actcacactt cgctgtacct
     4441 caagcgaaag gaaacaatgt cgtgattaca agctatatga caaacagagg attctacgca
     4501 gacaaacaat caacgtttgc gcctagcttc ctgctgaaca tcaaaggcaa gaaaacatct
     4561 gttgtcaaag acagcatcct tgaacaagga caattaacag ttaacaaata aaaacgcaaa
     4621 agaaaatgcc gatatcctat tggcattgcg gccgcggtct cggcggtggg tcgtctagaa
     4681 ggatcttcac ctagatcctt ttggttcatg tgcagctcca tcagcaaaag gggatgataa
     4741 gtttatcacc accgactatt tgcaacagtg ccgttgatcg tgctatgatc gactgatgtc
     4801 atcagcggtg gagtgcaatg tcgtgcaata cgaatggcga aaagccgagc tcatcggtca
     4861 gcttctcaac cttggggtta cccccggcgg tgtgctgctg gtccacagct ccttccgtag
     4921 cgtccggccc ctcgaagatg ggccacttgg actgatcgag gccctgcgtg ctgcgctggg
     4981 tccgggaggg acgctcgtca tgccctcgtg gtcaggtctg gacgacgagc cgttcgatcc
     5041 tgccacgtcg cccgttacac cggaccttgg agttgtctct gacacattct ggcgcctgcc
     5101 aaatgtaaag cgcagcgccc atccatttgc ctttgcggca gcggggccac aggcagagca
     5161 gatcatctct gatccattgc ccctgccacc tcactcgcct gcaagcccgg tcgcccgtgt
     5221 ccatgaactc gatgggcagg tacttctcct cggcgtggga cacgatgcca acacgacgct
     5281 gcatcttgcc gagttgatgg caaaggttcc ctatggggtg ccgagacact gcaccattct
     5341 tcaggatggc aagttggtac gcgtcgatta tctcgagaat gaccactgct gtgagcgctt
     5401 tgccttggcg gacaggtggc tcaaggaaaa gagccttcag aaggaaggtc cagtcggtca
     5461 tgcctttgct cggttgatcc gctcccgcga cattgtggcg acagccctgg gtcaactggg
     5521 ccgagatccg ttgatcttcc tgcatccgcc agaggcggga tgcgaagaat gcgatgccgc
     5581 tcgccagtcg attggctgag ctcatgagcg gagaacgaga tgacgttgga ggggcaaggt
     5641 cgcgctgatt gctggggcaa cacgtggagc ggatcgggga ttgtc
//
