LOCUS       sequence-211580-        4235 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Analogous to pKO but without the negative selection gene galK..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4235)
  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 4235)
  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: pKOdeltagalK (Kan)
FEATURES             Location/Qualifiers
     source          1..4235
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(123..938)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPHAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHNLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     846..865
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     930..949
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     primer_bind     1199..1221
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1213..1230
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1214..1230
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      complement(1474..2062)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1554..1573)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     promoter        2207..2652
                     /gene="sacR"
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             join(2653..2739,2740..4074)
                     /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"
ORIGIN
        1 gtcgggaaga tgcgtgatct gatccttcaa ctcagcaaaa gttcgattta ttcaacaaag
       61 ccgccgtccc gtcaagtcag cgtaatgctc tgccagtgtt acaaccaatt aaccaattct
      121 gattagaaaa actcatcgag catcaaatga aactgcaatt tattcatatc aggattatca
      181 ataccatatt tttgaaaaag ccgtttctgt aatgaaggag aaaactcacc gaggcagttc
      241 cataggatgg caagatcctg gtatcggtct gcgattccga ctcgtccaac atcaatacaa
      301 cctattaatt tcccctcgtc aaaaataagg ttatcaagtg agaaatcacc atgagtgacg
      361 actgaatccg gtgagaatgg caaaagatta tgcatttctt tccagacttg ttcaacaggc
      421 cagccattac gctcgtcatc aaaatcactc gcatcaacca aaccgttatt cattcgtgat
      481 tgcgcctgag cgagacgaaa tacgcgatcg ctgttaaaag gacaattaca aacaggaatc
      541 gaatgcaacc ggcgcaggaa cactgccagc gcatcaacaa tattttcacc tgaatcagga
      601 tattcttcta atacctggaa tgctgttttc ccggggatcg cagtggtgag taaccatgca
      661 tcatcaggag tacggataaa atgcttgatg gtcggaagag gcataaattc cgtcagccag
      721 tttagtctga ccatctcatc tgtaacatca ttggcaacgc tacctttgcc atgtttcaga
      781 aacaactctg gcgcatgggg cttcccatac aagcgataga ttgtcgcacc tgattgcccg
      841 acattatcgc gagcccattt atacccatat aaatcagcat ccatgttgga atttaatcgc
      901 ggcctcgagc aagacgtttc ccgttgaata tggctcataa caccccttgt attactgttt
      961 atgtaagcag acagttttat tgttcatgat gatatatttt tatcttgtgc aatgtaacat
     1021 cagagatttt gagacacaac gtggctttcc ccccccccga cgtcaggtgg ccccattcgc
     1081 cattcaggct gcgcaactgt tgggaagggc gtttcggtgc gggcctcttc gctattacgc
     1141 cagctggcga aagggggatg tgctgcaagg cgattaagtt gggtaacgcc agggttttcc
     1201 cagtcacgac gttgtaaaac gacggccagt gagcgcgacg taataagttg aagagcatct
     1261 gctcttcagc atcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaac
     1321 atggtcatag ctgtttcctt gcgtattggg cgctctccgc ttcctcgctc actgactcgc
     1381 tgcgctcggt cgttcgggta aagcctgggg tgcctaatga gcaaaaggcc agcaaaaggc
     1441 caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc cccctgacga
     1501 gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac tataaagata
     1561 ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc tgccgcttac
     1621 cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata gctcacgctg
     1681 taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc acgaaccccc
     1741 cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca acccggtaag
     1801 acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag cgaggtatgt
     1861 aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta gaagaacagt
     1921 atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg gtagctcttg
     1981 atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc agcagattac
     2041 gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt ctgacgctca
     2101 gtggaacgaa ggcgcgcctg agtgagctga taccgctcgc cgcagccgaa cgaccgagcg
     2161 cagcgagtca gtgagcgagg aagcggaaga tcgggcccag ccggcccaca tatacctgcc
     2221 gttcactatt atttagtgaa atgagatatt atgatatttt ctgaattgtg attaaaaagg
     2281 caactttatg cccatgcaac agaaactata aaaaatacag agaatgaaaa gaaacagata
     2341 gattttttag ttctttaggc ccgtagtctg caaatccttt tatgattttc tatcaaacaa
     2401 aagaggaaaa tagaccagtt gcaatccaaa cgagagtcta atagaatgag gtcgaaaagt
     2461 aaatcgcgcg ggtttgttac tgataaagca ggcaagacct aaaatgtgta aagggcaaag
     2521 tgtatacttt ggcgtcaccc cttacatatt ttaggtcttt ttttattgtg cgtaactaac
     2581 ttgccatctt caaacaggag ggctggaaga agcagaccgc taacacagta cataaaaaag
     2641 gagacatgaa cgatgaacat caaaaagttt gcaaaacaag caacagtatt aacctttact
     2701 accgcactgc tggcaggagg cgcaactcaa gcgtttgcga aagaaacgaa ccaaaagcca
     2761 tataaggaaa catacggcat ttcccatatt acacgccatg atatgctgca aatccctgaa
     2821 cagcaaaaaa atgaaaaata taaagttcct gagttcgatt cgtccacaat taaaaatatc
     2881 tcttctgcaa aaggcctgga cgtttgggac agctggccat tacaaaacac tgacggcact
     2941 gtcgcaaact atcacggcta ccacatcgtc tttgcattag ccggagatcc taaaaatgcg
     3001 gatgacacat cgatttacat gttctatcaa aaagtcggcg aaacttctat tgacagctgg
     3061 aaaaacgctg gccgcgtctt taaagacagc gacaaattcg atgcaaatga ttctatccta
     3121 aaagaccaaa cacaagaatg gtcaggttca gccacattta catctgacgg aaaaatccgt
     3181 ttattctaca ctgatttctc cggtaaacat tacggcaaac aaacactgac aactgcacaa
     3241 gttaacgtat cagcatcaga cagctctttg aacatcaacg gtgtagagga ttataaatca
     3301 atctttgacg gtgacggaaa aacgtatcaa aatgtacagc agttcatcga tgaaggcaac
     3361 tacagctcag gcgacaacca tacgctgaga gatcctcact acgtagaaga taaaggccac
     3421 aaatacttag tatttgaagc aaacactgga actgaagatg gctaccaagg cgaagaatct
     3481 ttatttaaca aagcatacta tggcaaaagc acatcattct tccgtcaaga aagtcaaaaa
     3541 cttctgcaaa gcgataaaaa acgcacggct gagttagcaa acggcgctct cggtatgatt
     3601 gagctaaacg atgattacac actgaaaaaa gtgatgaaac cgctgattgc atctaacaca
     3661 gtaacagatg aaattgaacg cgcgaacgtc tttaaaatga acggcaaatg gtacctgttc
     3721 actgactccc gcggatcaaa aatgacgatt gacggcatta cgtctaacga tatttacatg
     3781 cttggttatg tttctaattc tttaactggc ccatacaagc cgctgaacaa aactggcctt
     3841 gtgttaaaaa tggatcttga tcctaacgat gtaaccttta cttactcaca cttcgctgta
     3901 cctcaagcga aaggaaacaa tgtcgtgatt acaagctata tgacaaacag aggattctac
     3961 gcagacaaac aatcaacgtt tgcgcctagc ttcctgctga acatcaaagg caagaaaaca
     4021 tctgttgtca aagacagcat ccttgaacaa ggacaattaa cagttaacaa ataaaaacgc
     4081 aaaagaaaat gccgatatcc tattggcatt gcggccgcgg tctcggcggt gggtcgtcta
     4141 gagaaagtga gggagccacg gttggtgaga gctttgttgt aggtggacca gttggtgatt
     4201 ttgaactttt gctttgccac ggaatggtct gcgtt
//
