LOCUS       sequence-353367-        6937 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive expression of FnCas12a.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6937)
  AUTHORS   Patinios C, Creutzburg SCA, Arifah AQ, Adiego-Perez B, Gyimah EA, 
            Ingham CJ, Kengen SWM, van der Oost J, Staals RHJ
  TITLE     Streamlined CRISPR genome engineering in wild-type bacteria using 
            SIBR-Cas.
  JOURNAL   Nucleic Acids Res. 2021 Oct 6. pii: 6382381. doi: 
            10.1093/nar/gkab893.
  PUBMED    34614191
REFERENCE   2  (bases 1 to 6937)
  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: "Nucleic 
            Acids Res. 2021 Oct 6. pii: 6382381. doi: 10.1093/nar/gkab893."
COMMENT     Sequence Label: pSIBR005
FEATURES             Location/Qualifiers
     source          1..6937
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(137..142,143..160,161..167)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     primer_bind     complement(421..440)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             432..1247
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(1318..1980)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     rep_origin      1981..2752
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     terminator      complement(2867..2953)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     2968..2990
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     promoter        join(2997..3002,3003..3020,3021..3027)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
ORIGIN
        1 ctcaatttgg ttatcaaaaa tgaagagtat tttgagttcg tgcagaatag gaataactaa
       61 gcggccgcat aatgatgtgt tatcattgat gcgaggtcgc ctatacctcc ccgcttcggc
      121 ggggtttttt cccgggttta cactttatgc ttccggctcg tataatgtgt ggtctaagaa
      181 ctttaaataa tttctactgt tgtagatatg tcttcctgag tgacgagcaa cgagactgga
      241 agacatgtct aagaacttta aataatttct actgttgtag atagataccg gacaacgtgt
      301 ctccccgctt cggcggggtt ttttcctagg ttgtgtctca aaatctctga tgttacattg
      361 cacaagataa aaatatatca tcatgaacaa taaaactgtc tgcttacata aacagtaata
      421 caaggggtgt tatgagccat attcagcgtg aaacgagctg tagccgtccg cgtctgaaca
      481 gcaacatgga tgcggatctg tatggctata aatgggcgcg tgataacgtg ggtcagagcg
      541 gcgcgaccat ttatcgtctg tatggcaaac cggatgcgcc ggaactgttt ctgaaacatg
      601 gcaaaggcag cgtggcgaac gatgtgaccg atgaaatggt gcgtctgaac tggctgaccg
      661 aatttatgcc gctgccgacc attaaacatt ttattcgcac cccggatgat gcgtggctgc
      721 tgaccaccgc gattccgggc aaaaccgcgt ttcaggtgct ggaagaatat ccggatagcg
      781 gcgaaaacat tgtggatgcg ctggccgtgt ttctgcgtcg tctgcatagc attccggtgt
      841 gcaactgccc gtttaacagc gatcgtgtgt ttcgtctggc ccaggcgcag agccgtatga
      901 acaacggcct ggtggatgcg agcgattttg atgatgaacg taacggctgg ccggtggaac
      961 aggtgtggaa agaaatgcat aaactgctgc cgtttagccc ggatagcgtg gtgacccacg
     1021 gcgattttag cctggataac ctgattttcg atgaaggcaa actgattggc tgcattgatg
     1081 tgggccgtgt gggcattgcg gatcgttatc aggatctggc cattctgtgg aactgcctgg
     1141 gcgaatttag cccgagcctg caaaaacgtc tgtttcagaa atatggcatt gataatccgg
     1201 atatgaacaa actgcaattt catctgatgc tggatgaatt tttctaataa ttaattggac
     1261 cgcggtccga gagacgcatg gcagtatctc agactgtgta tcggcacgtc tcactactta
     1321 ccggcgcggc agcgttaccc gtgtcggcgg ctccaacggc tcgccatcgt ccagaaaaca
     1381 cggctcatcg ggcatcggca ggcgctgctg cccgcgccgt tcccattcct ccgtttcggt
     1441 caaggctggc aggtctggtt ccatgcccgg aatgccgggc tggctgggcg gctcctcgcc
     1501 ggggccggtc ggtagttgct gctcgcccgg atacagggtc gggatgcggc gcaggtcgcc
     1561 atgccccaac agcgattcgt cctggtcgtc gtgatcaacc accacggcgg cactgaacac
     1621 cgacaggcgc aactggtcgc ggggctggcc ccacgccacg cggtcattga ccacgtaggc
     1681 cgacacggtg ccggggccgt tgagcttcac gacggagatc cagcgctcgg ccaccaagtc
     1741 cttgactgcg tattggaccg tccgcaaaga acgtccgatg agcttggaaa gtgttttctg
     1801 gctgaccacc acggcgttct ggtggcccat ctgcgccacg aggtgatgca gcagcattgc
     1861 cgccgtgggt ttcctcgcaa taagcccggc ccacgcctca tgcgctttgc gttccgtttg
     1921 cacccagtga ccgggcttgt tcttggcttg aatgccgatt tctctggact gcgtggccat
     1981 gcttatctcc atgcggtagg ggtgccgcac ggttgcggca ccatgcgcaa tcagctgcaa
     2041 cttttcggca gcgcgacaac aattatgcgt tgcgtaaaag tggcagtcaa ttacagattt
     2101 tctttaacct acgcaatgag ctattgcggg gggtgccgca atgagctgtt gcgtaccccc
     2161 cttttttaag ttgttgattt ttaagtcttt cgcatttcgc cctatatcta gttctttggt
     2221 gcccaaagaa gggcacccct gcggggttcc cccacgcctt cggcgcggct ccccctccgg
     2281 caaaaagtgg cccctccggg gcttgttgat cgactgcgcg gccttcggcc ttgcccaagg
     2341 tggcgctgcc cccttggaac ccccgcactc gccgccgtga ggctcggggg gcaggcgggc
     2401 gggcttcgcc cttcgactgc ccccactcgc ataggcttgg gtcgttccag gcgcgtcaag
     2461 gccaagccgc tgcgcggtcg ctgcgcgagc cttgacccgc cttccacttg gtgtccaacc
     2521 ggcaagcgaa gcgcgcaggc cgcaggccgg aggcttttcc ccagagaaaa ttaaaaaaat
     2581 tgatggggca aggccgcagg ccgcgcagtt ggagccggtg ggtatgtggt cgaaggctgg
     2641 gtagccggtg ggcaatccct gtggtcaagc tcgtgggcag gcgcagcctg tccatcagct
     2701 tgtccagcag ggttgtccac gggccgagcg aagcgagcca gccggtggcc gctcgcggcc
     2761 atcgtccaca tatccacggg ctggcaaggg agcgcagcga ccgcgcaggg cgaagcccgg
     2821 agagcaagcc cgtagggggg gcgcgcccag ctgtctaggg cggcggattt gtcctactca
     2881 ggagagcgtt caccgacaaa caacagataa aacgaaaggc ccagtctttc gactgagcct
     2941 ttcgttttat ttgatgcctt taattaaagc ggataacaat ttcacacagg aggccgttta
     3001 cactttatgc ttccggctcg tataatgtgt ggggaaacca caacggtttc cctctagaaa
     3061 taattttgtt taactataag aaggagatat acatatgtca atttatcaag aatttgttaa
     3121 taaatatagt ttaagtaaaa ctctaagatt tgagttaatc ccacagggta aaacacttga
     3181 aaacataaaa gcaagaggtt tgattttaga tgatgagaaa agagctaaag actacaaaaa
     3241 ggctaaacaa ataattgata aatatcatca gttttttata gaggagatat taagttcggt
     3301 ttgtattagc gaagatttat tacaaaacta ttctgatgtt tattttaaac ttaaaaagag
     3361 tgatgatgat aatctacaaa aagattttaa aagtgcaaaa gatacgataa agaaacaaat
     3421 atctgaatat ataaaggact cagagaaatt taagaatttg tttaatcaaa accttatcga
     3481 tgctaaaaaa gggcaagagt cagatttaat tctatggcta aagcaatcta aggataatgg
     3541 tatagaacta tttaaagcca atagtgatat cacagatata gatgaggcgt tagaaataat
     3601 caaatctttt aaaggttgga caacttattt taagggtttt catgaaaata gaaaaaatgt
     3661 ttatagtagc aatgatattc ctacatctat tatttatagg atagtagatg ataatttgcc
     3721 taaatttcta gaaaataaag ctaagtatga gagtttaaaa gacaaagctc cagaagctat
     3781 aaactatgaa caaattaaaa aagatttggc agaagagcta acctttgata ttgactacaa
     3841 aacatctgaa gttaatcaaa gagttttttc acttgatgaa gtttttgaga tagcaaactt
     3901 taataattat ctaaatcaaa gtggtattac taaatttaat actattattg gtggtaaatt
     3961 tgtaaatggt gaaaatacaa agagaaaagg tataaatgaa tatataaatc tatactcaca
     4021 gcaaataaat gataaaacac tcaaaaaata taaaatgagt gttttattta agcaaatttt
     4081 aagtgataca gaatctaaat cttttgtaat tgataagtta gaagatgata gtgatgtagt
     4141 tacaacgatg caaagttttt atgagcaaat agcagctttt aaaacagtag aagaaaaatc
     4201 tattaaagaa acactatctt tattatttga tgatttaaaa gctcaaaaac ttgatttgag
     4261 taaaatttat tttaaaaatg ataaatctct tactgatcta tcacaacaag tttttgatga
     4321 ttatagtgtt attggtacag cggtactaga atatataact caacaaatag cacctaaaaa
     4381 tcttgataac cctagtaaga aagagcaaga attaatagcc aaaaaaactg aaaaagcaaa
     4441 atacttatct ctagaaacta taaagcttgc cttagaagaa tttaataagc atagagatat
     4501 agataaacag tgtaggtttg aagaaatact tgcaaacttt gcggctattc cgatgatatt
     4561 tgatgaaata gctcaaaaca aagacaattt ggcacagata tctatcaaat atcaaaatca
     4621 aggtaaaaaa gacctacttc aagctagtgc ggaagatgat gttaaagcta tcaaggatct
     4681 tttagatcaa actaataatc tcttacataa actaaaaata tttcatatta gtcagtcaga
     4741 agataaggca aatattttag acaaggatga gcatttttat ctagtatttg aggagtgcta
     4801 ctttgagcta gcgaatatag tgcctcttta taacaaaatt agaaactata taactcaaaa
     4861 gccatatagt gatgagaaat ttaagctcaa ttttgagaac tcgactttgg ctaatggttg
     4921 ggataaaaat aaagagcctg acaatacggc aattttattt atcaaagatg ataaatatta
     4981 tctgggtgtg atgaataaga aaaataacaa aatatttgat gataaagcta tcaaagaaaa
     5041 taaaggcgag ggttataaaa aaattgttta taaactttta cctggcgcaa ataaaatgtt
     5101 acctaaggtt ttcttttctg ctaaatctat aaaattttat aatcctagtg aagatatact
     5161 tagaataaga aatcattcca cacatacaaa aaatggtagt cctcaaaaag gatatgaaaa
     5221 atttgagttt aatattgaag attgccgaaa atttatagat ttttataaac agtctataag
     5281 taagcatccg gagtggaaag attttggatt tagattttct gatactcaaa gatataattc
     5341 tatagatgaa ttttatagag aagttgaaaa tcaaggctac aaactaactt ttgaaaatat
     5401 atcagagagc tatattgata gcgtagttaa tcagggtaaa ttgtacctat tccaaatcta
     5461 taataaagat ttttcagctt atagcaaagg gcgaccaaat ctacatactt tatattggaa
     5521 agcgctgttt gatgagagaa atcttcaaga tgtggtttat aagctaaatg gtgaggcaga
     5581 gcttttttat cgtaaacaat caatacctaa aaaaatcact cacccagcta aagaggcaat
     5641 agctaataaa aacaaagata atcctaaaaa agagagtgtt tttgaatatg atttaatcaa
     5701 agataaacgc tttactgaag ataagttttt ctttcactgt cctattacaa tcaattttaa
     5761 atctagtgga gctaataagt ttaatgatga aatcaattta ttgctaaaag aaaaagcaaa
     5821 tgatgttcat atattaagta tagatagagg tgaaagacat ttagcttact atactttggt
     5881 agatggtaaa ggcaatatca tcaaacaaga tactttcaac atcattggta atgatagaat
     5941 gaaaacaaac taccatgata agcttgctgc aatagagaaa gatagggatt cagctaggaa
     6001 agactggaaa aagataaata acatcaaaga gatgaaagag ggctatctat ctcaggtagt
     6061 tcatgaaata gctaagctag ttatagagta taatgctatt gtggtttttg aggatttaaa
     6121 ttttggattt aaaagagggc gtttcaaggt agagaagcag gtctatcaaa agttagaaaa
     6181 aatgctaatt gagaaactaa actatctagt tttcaaagat aatgagtttg ataaaactgg
     6241 gggagtgctt agagcttatc agctaacagc accttttgag acttttaaaa agatgggtaa
     6301 acaaacaggt attatctact atgtaccagc tggttttact tcaaaaattt gtcctgtaac
     6361 tggttttgta aatcagttat atcctaagta tgaaagtgtc agcaaatctc aagagttctt
     6421 tagtaagttt gacaagattt gttataacct tgataagggc tattttgagt ttagttttga
     6481 ttataaaaac tttggtgaca aggctgccaa aggcaagtgg actatagcta gctttgggag
     6541 tagattgatt aactttagaa attcagataa aaatcataat tgggatactc gagaagttta
     6601 tccaactaaa gagttggaga aattgctaaa agattattct atcgaatatg ggcatggcga
     6661 atgtatcaaa gcagctattt gcggtgagag cgacaaaaag ttttttgcta agctaactag
     6721 tgtcctaaat actatcttac aaatgcgtaa ctcaaaaaca ggtactgagt tagattatct
     6781 aatttcacca gtagcagatg taaatggcaa tttctttgat tcgcgacagg cgccaaaaaa
     6841 tatgcctcaa gatgctgatg ccaatggtgc ttatcatatt gggctaaaag gtctgatgct
     6901 actaggtagg atcaaaaata atcaagaggg caaaaaa
//
