LOCUS       sequence-226944-        4009 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Spectinomycin resistance cassette.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4009)
  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 4009)
  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 specR cassette
FEATURES             Location/Qualifiers
     source          1..4009
                     /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(1841..1847,1848..1865,1866..1871))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    1886..1907
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2027..2044)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2198..2786)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2278..2297)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2957..3748,3749..3817))
                     /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     3580..3599
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3818..3922)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3990..4008
                     /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 ctcgaatatc ttcttacttc tgtataataa agaataatta
      421 ttaatctgta gacaaattgt gaaaggatgt acttaaacgc taacggtcag ctttattgaa
      481 cagtaattta agtatatgtc caatctaggg taagtaaatt gagtatcaat ataaacttta
      541 tatgaacata atcaacgagg tgaaatcatg agcaatttga ttaacggaaa aataccaaat
      601 caagcgattc aaacattaaa aatcgtaaaa gatttatttg gaagttcaat agttggagta
      661 tatctatttg gttcagcagt aaatggtggt ttacgcatta acagcgatgt agatgttcta
      721 gtcgtcgtga atcatagttt acctcaatta actcgaaaaa aactaacaga aagactaatg
      781 actatatcag gaaagattgg aaatacggat tctgttagac cacttgaagt tacggttata
      841 aataggagtg aagttgtccc ttggcaatat cctccaaaaa gagaatttat atacggtgag
      901 tggctcaggg gtgaatttga gaatggacaa attcaggaac caagctatga tcctgatttg
      961 gctattgttt tagcacaagc aagaaagaat agtatttctc tatttggtcc tgattcttca
     1021 agtatacttg tctccgtacc tttgacagat attcgaagag caattaagga ttctttgcca
     1081 gaactaattg aggggataaa aggtgatgag cgtaatgtaa ttttaaccct agctcgaatg
     1141 tggcaaacag tgactactgg tgaaattacc tcgaaagatg tcgctgcaga atgggctata
     1201 cctcttttac ctaaagagca tgtaacttta ctggatatag ctagaaaagg ctatcgggga
     1261 gagtgtgatg ataagtggga aggactatat tcaaaggtga aagcactcgt taagtatatg
     1321 aaaaattcta tagaaacttc tctcaattaa gaaaaaggcc ccaacatctt ggggcctttt
     1381 tcttttttat ctacgagacc caatgaatct ttctagaaga tctcctacaa tattctcagc
     1441 tgccatggaa aatcgatgtt cttcttttat tctctcaaga ttttcaggct gtatattaaa
     1501 acttatatta agaactatgc taaccacctc atcaggaacc gttgtaggtg gcgtgggttt
     1561 tcttggcaat cgactctcat gaaaactacg agctaaatat tcaatatgtt cctcttgacc
     1621 aactttattc tgcatttttt ttgaacgagg tttagagcaa gcttcaggaa actgagacag
     1681 gaattttatt aaaaatttaa attttgaaga aagttcaggg ttaatagcat ccattttttg
     1741 ctttgcaagt tcctcagcat tcttaacaaa agacgtctct tttgacatgt ttaaagttta
     1801 aacctcctgt gtgaaattat tatccgctca taattccaca cattatacga gccggaagca
     1861 taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct
     1921 cactgccaat tgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     1981 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     2041 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     2101 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     2161 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     2221 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     2281 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     2341 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     2401 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     2461 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     2521 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     2581 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     2641 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     2701 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     2761 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     2821 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     2881 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     2941 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     3001 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     3061 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     3121 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     3181 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     3241 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     3301 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     3361 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     3421 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     3481 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     3541 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     3601 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     3661 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     3721 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     3781 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     3841 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     3901 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtctaagaaa
     3961 ccattattat catgacatta acctataaaa ataggcgtat cacgaggcc
//
