LOCUS       sequence-344706-        7225 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  SIBR Intron 2 (Int2) interrupts the CDS of FnCas12a.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7225)
  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 7225)
  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: pSIBR002
FEATURES             Location/Qualifiers
     source          1..7225
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(2283..2288,2289..2306,2307..2313)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     primer_bind     complement(2567..2586)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             2578..3393
                     /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(3464..4126)
                     /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      4127..4898
                     /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(5013..5099)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     5114..5136
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     promoter        join(5143..5148,5149..5166,5167..5173)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
ORIGIN
        1 ggagtgctac tttgagctag cgaatatagt gcctctttat aacaaaatta gaaactatat
       61 aactcaaaag ccatatagtg atgagaaatt taagctcaat tttgagaact cgactttggc
      121 taatggttgg gataaaaata aagagcctga caatacggca attttattta tcaaagatga
      181 taaatattat ctgggtgtga tgaataagaa aaataacaaa atatttgatg ataaagctat
      241 caaagaaaat aaaggcgagg gttataaaaa aattgtttat aaacttttac ctggcgcaaa
      301 taaaatgtta cctaaggttt tcttttctgc taaatctata aaattttata atcctagtga
      361 agatatactt agaataagaa atcattccac acatacaaaa aatggtagtc ctcaaaaagg
      421 atatgaaaaa tttgagttta atattgaaga ttgccgaaaa tttatagatt tttataaaca
      481 gtctataagt aagcatccgg agtggaaaga ttttggattt agattttctg atactcaaag
      541 atataattct atagatgaat tttatagaga agttgaaaat caaggctaca aactaacttt
      601 tgaaaatata tcagagagct atattgatag cgtagttaat cagggtaaat tgtacctatt
      661 ccaaatctat aataaagatt tttcagctta tagcaaaggg cgaccaaatc tacatacttt
      721 atattggaaa gcgctgtttg atgagagaaa tcttcaagat gtggtttata agctaaatgg
      781 tgaggcagag cttttttatc gtaaacaatc aatacctaaa aaaatcactc acccagctaa
      841 agaggcaata gctaataaaa acaaagataa tcctaaaaaa gagagtgttt ttgaatatga
      901 tttaatcaaa gataaacgct ttactgaaga taagtttttc tttcactgtc ctattacaat
      961 caattttaaa tctagtggag ctaataagtt taatgatgaa atcaatttat tgctaaaaga
     1021 aaaagcaaat gatgttcata tattaagtat agatagaggt gaaagacatt tagcttacta
     1081 tactttggta gatggtaaag gcaatatcat caaacaagat actttcaaca tcattggtaa
     1141 tgatagaatg aaaacaaact accatgataa gcttgctgca atagagaaag atagggattc
     1201 agctaggaaa gactggaaaa agataaataa catcaaagag atgaaagagg gctatctatc
     1261 tcaggtagtt catgaaatag ctaagctagt tatagagtat aatgctattg tggtttttga
     1321 ggatttaaat tttggattta aaagagggcg tttcaaggta gagaagcagg tctatcaaaa
     1381 gttagaaaaa atgctaattg agaaactaaa ctatctagtt ttcaaagata atgagtttga
     1441 taaaactggg ggagtgctta gagcttatca gctaacagca ccttttgaga cttttaaaaa
     1501 gatgggtaaa caaacaggta ttatctacta tgtaccagct ggttttactt caaaaatttg
     1561 tcctgtaact ggttttgtaa atcagttata tcctaagtat gaaagtgtca gcaaatctca
     1621 agagttcttt agtaagtttg acaagatttg ttataacctt gataagggct attttgagtt
     1681 tagttttgat tataaaaact ttggtgacaa ggctgccaaa ggcaagtgga ctatagctag
     1741 ctttgggagt agattgatta actttagaaa ttcagataaa aatcataatt gggatactcg
     1801 agaagtttat ccaactaaag agttggagaa attgctaaaa gattattcta tcgaatatgg
     1861 gcatggcgaa tgtatcaaag cagctatttg cggtgagagc gacaaaaagt tttttgctaa
     1921 gctaactagt gtcctaaata ctatcttaca aatgcgtaac tcaaaaacag gtactgagtt
     1981 agattatcta atttcaccag tagcagatgt aaatggcaat ttctttgatt cgcgacaggc
     2041 gccaaaaaat atgcctcaag atgctgatgc caatggtgct tatcatattg ggctaaaagg
     2101 tctgatgcta ctaggtagga tcaaaaataa tcaagagggc aaaaaactca atttggttat
     2161 caaaaatgaa gagtattttg agttcgtgca gaataggaat aactaagcgg ccgcataatg
     2221 atgtgttatc attgatgcga ggtcgcctat acctccccgc ttcggcgggg ttttttcccg
     2281 ggtttacact ttatgcttcc ggctcgtata atgtgtggtc taagaacttt aaataatttc
     2341 tactgttgta gatatgtctt cctgagtgac gagcaacgag actggaagac atgtctaaga
     2401 actttaaata atttctactg ttgtagatag ataccggaca acgtgtctcc ccgcttcggc
     2461 ggggtttttt cctaggttgt gtctcaaaat ctctgatgtt acattgcaca agataaaaat
     2521 atatcatcat gaacaataaa actgtctgct tacataaaca gtaatacaag gggtgttatg
     2581 agccatattc agcgtgaaac gagctgtagc cgtccgcgtc tgaacagcaa catggatgcg
     2641 gatctgtatg gctataaatg ggcgcgtgat aacgtgggtc agagcggcgc gaccatttat
     2701 cgtctgtatg gcaaaccgga tgcgccggaa ctgtttctga aacatggcaa aggcagcgtg
     2761 gcgaacgatg tgaccgatga aatggtgcgt ctgaactggc tgaccgaatt tatgccgctg
     2821 ccgaccatta aacattttat tcgcaccccg gatgatgcgt ggctgctgac caccgcgatt
     2881 ccgggcaaaa ccgcgtttca ggtgctggaa gaatatccgg atagcggcga aaacattgtg
     2941 gatgcgctgg ccgtgtttct gcgtcgtctg catagcattc cggtgtgcaa ctgcccgttt
     3001 aacagcgatc gtgtgtttcg tctggcccag gcgcagagcc gtatgaacaa cggcctggtg
     3061 gatgcgagcg attttgatga tgaacgtaac ggctggccgg tggaacaggt gtggaaagaa
     3121 atgcataaac tgctgccgtt tagcccggat agcgtggtga cccacggcga ttttagcctg
     3181 gataacctga ttttcgatga aggcaaactg attggctgca ttgatgtggg ccgtgtgggc
     3241 attgcggatc gttatcagga tctggccatt ctgtggaact gcctgggcga atttagcccg
     3301 agcctgcaaa aacgtctgtt tcagaaatat ggcattgata atccggatat gaacaaactg
     3361 caatttcatc tgatgctgga tgaatttttc taataattaa ttggaccgcg gtccgagaga
     3421 cgcatggcag tatctcagac tgtgtatcgg cacgtctcac tacttaccgg cgcggcagcg
     3481 ttacccgtgt cggcggctcc aacggctcgc catcgtccag aaaacacggc tcatcgggca
     3541 tcggcaggcg ctgctgcccg cgccgttccc attcctccgt ttcggtcaag gctggcaggt
     3601 ctggttccat gcccggaatg ccgggctggc tgggcggctc ctcgccgggg ccggtcggta
     3661 gttgctgctc gcccggatac agggtcggga tgcggcgcag gtcgccatgc cccaacagcg
     3721 attcgtcctg gtcgtcgtga tcaaccacca cggcggcact gaacaccgac aggcgcaact
     3781 ggtcgcgggg ctggccccac gccacgcggt cattgaccac gtaggccgac acggtgccgg
     3841 ggccgttgag cttcacgacg gagatccagc gctcggccac caagtccttg actgcgtatt
     3901 ggaccgtccg caaagaacgt ccgatgagct tggaaagtgt tttctggctg accaccacgg
     3961 cgttctggtg gcccatctgc gccacgaggt gatgcagcag cattgccgcc gtgggtttcc
     4021 tcgcaataag cccggcccac gcctcatgcg ctttgcgttc cgtttgcacc cagtgaccgg
     4081 gcttgttctt ggcttgaatg ccgatttctc tggactgcgt ggccatgctt atctccatgc
     4141 ggtaggggtg ccgcacggtt gcggcaccat gcgcaatcag ctgcaacttt tcggcagcgc
     4201 gacaacaatt atgcgttgcg taaaagtggc agtcaattac agattttctt taacctacgc
     4261 aatgagctat tgcggggggt gccgcaatga gctgttgcgt accccccttt tttaagttgt
     4321 tgatttttaa gtctttcgca tttcgcccta tatctagttc tttggtgccc aaagaagggc
     4381 acccctgcgg ggttccccca cgccttcggc gcggctcccc ctccggcaaa aagtggcccc
     4441 tccggggctt gttgatcgac tgcgcggcct tcggccttgc ccaaggtggc gctgccccct
     4501 tggaaccccc gcactcgccg ccgtgaggct cggggggcag gcgggcgggc ttcgcccttc
     4561 gactgccccc actcgcatag gcttgggtcg ttccaggcgc gtcaaggcca agccgctgcg
     4621 cggtcgctgc gcgagccttg acccgccttc cacttggtgt ccaaccggca agcgaagcgc
     4681 gcaggccgca ggccggaggc ttttccccag agaaaattaa aaaaattgat ggggcaaggc
     4741 cgcaggccgc gcagttggag ccggtgggta tgtggtcgaa ggctgggtag ccggtgggca
     4801 atccctgtgg tcaagctcgt gggcaggcgc agcctgtcca tcagcttgtc cagcagggtt
     4861 gtccacgggc cgagcgaagc gagccagccg gtggccgctc gcggccatcg tccacatatc
     4921 cacgggctgg caagggagcg cagcgaccgc gcagggcgaa gcccggagag caagcccgta
     4981 gggggggcgc gcccagctgt ctagggcggc ggatttgtcc tactcaggag agcgttcacc
     5041 gacaaacaac agataaaacg aaaggcccag tctttcgact gagcctttcg ttttatttga
     5101 tgcctttaat taaagcggat aacaatttca cacaggaggc cgtttacact ttatgcttcc
     5161 ggctcgtata atgtgtgggg aaaccacaac ggtttccctc tagaaataat tttgtttaac
     5221 tataagaagg agatatacat atgtcctcgg gttaattgag gcctgagtat aaggtgactt
     5281 atacttgtaa tctatctaaa cggggaacct ctctagtaga caatcccgtg ctaaattgat
     5341 accagcatcg tcttgatgcc cttggcagca taaatgccta acgactatcc ctttggggag
     5401 tagggtcaag tgactcgaaa cgatagacaa cttgctttaa caagttggag atatagtctg
     5461 ctctgcatgg tgacatgcag ctggatataa ttccggggta agattaacga ccttatctga
     5521 acataatgct atcaatttat caagaatttg ttaataaata tagtttaagt aaaactctaa
     5581 gatttgagtt aatcccacag ggtaaaacac ttgaaaacat aaaagcaaga ggtttgattt
     5641 tagatgatga gaaaagagct aaagactaca aaaaggctaa acaaataatt gataaatatc
     5701 atcagttttt tatagaggag atattaagtt cggtttgtat tagcgaagat ttattacaaa
     5761 actattctga tgtttatttt aaacttaaaa agagtgatga tgataatcta caaaaagatt
     5821 ttaaaagtgc aaaagatacg ataaagaaac aaatatctga atatataaag gactcagaga
     5881 aatttaagaa tttgtttaat caaaacctta tcgatgctaa aaaagggcaa gagtcagatt
     5941 taattctatg gctaaagcaa tctaaggata atggtataga actatttaaa gccaatagtg
     6001 atatcacaga tatagatgag gcgttagaaa taatcaaatc ttttaaaggt tggacaactt
     6061 attttaaggg ttttcatgaa aatagaaaaa atgtttatag tagcaatgat attcctacat
     6121 ctattattta taggatagta gatgataatt tgcctaaatt tctagaaaat aaagctaagt
     6181 atgagagttt aaaagacaaa gctccagaag ctataaacta tgaacaaatt aaaaaagatt
     6241 tggcagaaga gctaaccttt gatattgact acaaaacatc tgaagttaat caaagagttt
     6301 tttcacttga tgaagttttt gagatagcaa actttaataa ttatctaaat caaagtggta
     6361 ttactaaatt taatactatt attggtggta aatttgtaaa tggtgaaaat acaaagagaa
     6421 aaggtataaa tgaatatata aatctatact cacagcaaat aaatgataaa acactcaaaa
     6481 aatataaaat gagtgtttta tttaagcaaa ttttaagtga tacagaatct aaatcttttg
     6541 taattgataa gttagaagat gatagtgatg tagttacaac gatgcaaagt ttttatgagc
     6601 aaatagcagc ttttaaaaca gtagaagaaa aatctattaa agaaacacta tctttattat
     6661 ttgatgattt aaaagctcaa aaacttgatt tgagtaaaat ttattttaaa aatgataaat
     6721 ctcttactga tctatcacaa caagtttttg atgattatag tgttattggt acagcggtac
     6781 tagaatatat aactcaacaa atagcaccta aaaatcttga taaccctagt aagaaagagc
     6841 aagaattaat agccaaaaaa actgaaaaag caaaatactt atctctagaa actataaagc
     6901 ttgccttaga agaatttaat aagcatagag atatagataa acagtgtagg tttgaagaaa
     6961 tacttgcaaa ctttgcggct attccgatga tatttgatga aatagctcaa aacaaagaca
     7021 atttggcaca gatatctatc aaatatcaaa atcaaggtaa aaaagaccta cttcaagcta
     7081 gtgcggaaga tgatgttaaa gctatcaagg atcttttaga tcaaactaat aatctcttac
     7141 ataaactaaa aatatttcat attagtcagt cagaagataa ggcaaatatt ttagacaagg
     7201 atgagcattt ttatctagta tttga
//
