LOCUS       sequence-226875-        3226 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  C-terminal Pk6-tag.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 3226)
  AUTHORS   Diebold-Durand ML, Burmann F, Gruber S
  TITLE     High-Throughput Allelic Replacement Screening in Bacillus subtilis.
  JOURNAL   Methods Mol Biol. 2019;2004:49-61. doi: 10.1007/978-1-4939-9520-2_5.
  PUBMED    31147909
REFERENCE   2  (bases 1 to 3226)
  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: "Methods Mol
            Biol."; date: "2019"; volume: "2004:49-61. doi"; pages: " 
            10.1007/978-1-4939-9520-2_5"
COMMENT     Sequence Label: pJet1.2 Pk6-tag
FEATURES             Location/Qualifiers
     source          1..3226
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     305..324
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        305..323
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(join(1058..1064,1065..1082,1083..1088))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    1103..1124
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1244..1261)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1415..2003)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1495..1514)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2174..2965,2966..3034))
                     /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     2797..2816
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3035..3139)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3207..3225
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 gcccctgcag ccgaattata ttatttttgc caaataattt ttaacaaaag ctctgaagtc
       61 ttcttcattt aaattcttag atgatacttc atctggaaaa ttgtcccaat tagtagcatc
      121 acgctgtgag taagttctaa accatttttt tattgttgta ttatctctaa tcttactact
      181 cgatgagttt tcggtattat ctctattttt aacttggagc aggttccatt cattgttttt
      241 ttcatcatag tgaataaaat caactgcttt aacacttgtg cctgaacacc atatccatcc
      301 ggcgtaatac gactcactat agggagagcg gccgccagat cttccggatg gctcgagttt
      361 ttcagcaaga tgttacaggt ctcaagcggt tctgctgcta gaggtattcc taacccttta
      421 ttgggattag atggaatccc aaatcctttg ctaggtcttg acggcatacc caatccatta
      481 ttaggcttgg atggatcctc taggggtatt cctaaccctt tattgggatt agatggaatc
      541 ccaaatcctt tgctaggtct tgacggcata cccaatccat tattaggctt ggatggatcc
      601 tctagctaag tgagacccaa tgaatctttc tagaagatct cctacaatat tctcagctgc
      661 catggaaaat cgatgttctt cttttattct ctcaagattt tcaggctgta tattaaaact
      721 tatattaaga actatgctaa ccacctcatc aggaaccgtt gtaggtggcg tgggttttct
      781 tggcaatcga ctctcatgaa aactacgagc taaatattca atatgttcct cttgaccaac
      841 tttattctgc attttttttg aacgaggttt agagcaagct tcaggaaact gagacaggaa
      901 ttttattaaa aatttaaatt ttgaagaaag ttcagggtta atagcatcca ttttttgctt
      961 tgcaagttcc tcagcattct taacaaaaga cgtctctttt gacatgttta aagtttaaac
     1021 ctcctgtgtg aaattattat ccgctcataa ttccacacat tatacgagcc ggaagcataa
     1081 agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg ttgcgctcac
     1141 tgccaattgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac
     1201 gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc
     1261 tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt
     1321 tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg
     1381 ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg
     1441 agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat
     1501 accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta
     1561 ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct
     1621 gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc
     1681 ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa
     1741 gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg
     1801 taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag
     1861 tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt
     1921 gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta
     1981 cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc
     2041 agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca
     2101 cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa
     2161 cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat
     2221 ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct
     2281 taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt
     2341 tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat
     2401 ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta
     2461 atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg
     2521 gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt
     2581 tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg
     2641 cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg
     2701 taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc
     2761 ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa
     2821 ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac
     2881 cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt
     2941 ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg
     3001 gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa
     3061 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     3121 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc taagaaacca
     3181 ttattatcat gacattaacc tataaaaata ggcgtatcac gaggcc
//
