LOCUS       sequence-274091-        4650 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  E. coli-Lactobacilli shuttle vector containing SpCas9, tracrRNA, and
            a repeat-spacer-repeat array.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 4650)
  AUTHORS   Leenay RT, Vento JM, Shah M, Martino ME, Leulier F, Beisel CL
  TITLE     Genome Editing with CRISPR-Cas9 in Lactobacillus plantarum Revealed 
            That Editing Outcomes Can Vary Across Strains and Between Methods.
  JOURNAL   Biotechnol J. 2019 Mar;14(3):e1700583. doi: 10.1002/biot.201700583. 
            Epub 2018 Sep 20.
  PUBMED    30156038
REFERENCE   2  (bases 1 to 4650)
  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: "Biotechnol 
            J."; date: "2019-03"; volume: "14(3):e1700583. doi"; pages: " 
            10.1002/biot.201700583. Epub 2018 Sep 20"
COMMENT     Sequence Label: pCB578
FEATURES             Location/Qualifiers
     source          1..4650
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     misc_RNA        complement(170..248)
                     /label=tracrRNA
                     /note="trans-activating CRISPR RNA for the Streptococcus 
                     pyogenes CRISPR/Cas9 system"
     CDS             544..4650
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
     CDS             544..4647
                     /codon_start=1
                     /product="Cas9 (Csn1) endonuclease from the Streptococcus 
                     pyogenes Type II CRISPR/Cas system"
                     /label=Cas9
                     /note="generates RNA-guided double strand breaks in DNA"
                     /translation="MDKKYSIGLDIGTNSVGWAVITDEYKVPSKKFKVLGNTDRHSIKK
                     NLIGALLFDSGETAEATRLKRTARRRYTRRKNRICYLQEIFSNEMAKVDDSFFHRLEES
                     FLVEEDKKHERHPIFGNIVDEVAYHEKYPTIYHLRKKLVDSTDKADLRLIYLALAHMIK
                     FRGHFLIEGDLNPDNSDVDKLFIQLVQTYNQLFEENPINASGVDAKAILSARLSKSRRL
                     ENLIAQLPGEKKNGLFGNLIALSLGLTPNFKSNFDLAEDAKLQLSKDTYDDDLDNLLAQ
                     IGDQYADLFLAAKNLSDAILLSDILRVNTEITKAPLSASMIKRYDEHHQDLTLLKALVR
                     QQLPEKYKEIFFDQSKNGYAGYIDGGASQEEFYKFIKPILEKMDGTEELLVKLNREDLL
                     RKQRTFDNGSIPHQIHLGELHAILRRQEDFYPFLKDNREKIEKILTFRIPYYVGPLARG
                     NSRFAWMTRKSEETITPWNFEEVVDKGASAQSFIERMTNFDKNLPNEKVLPKHSLLYEY
                     FTVYNELTKVKYVTEGMRKPAFLSGEQKKAIVDLLFKTNRKVTVKQLKEDYFKKIECFD
                     SVEISGVEDRFNASLGTYHDLLKIIKDKDFLDNEENEDILEDIVLTLTLFEDREMIEER
                     LKTYAHLFDDKVMKQLKRRRYTGWGRLSRKLINGIRDKQSGKTILDFLKSDGFANRNFM
                     QLIHDDSLTFKEDIQKAQVSGQGDSLHEHIANLAGSPAIKKGILQTVKVVDELVKVMGR
                     HKPENIVIEMARENQTTQKGQKNSRERMKRIEEGIKELGSQILKEHPVENTQLQNEKLY
                     LYYLQNGRDMYVDQELDINRLSDYDVDHIVPQSFLKDDSIDNKVLTRSDKNRGKSDNVP
                     SEEVVKKMKNYWRQLLNAKLITQRKFDNLTKAERGGLSELDKAGFIKRQLVETRQITKH
                     VAQILDSRMNTKYDENDKLIREVKVITLKSKLVSDFRKDFQFYKVREINNYHHAHDAYL
                     NAVVGTALIKKYPKLESEFVYGDYKVYDVRKMIAKSEQEIGKATAKYFFYSNIMNFFKT
                     EITLANGEIRKRPLIETNGETGEIVWDKGRDFATVRKVLSMPQVNIVKKTEVQTGGFSK
                     ESILPKRNSDKLIARKKDWDPKKYGGFDSPTVAYSVLVVAKVEKGKSKKLKSVKELLGI
                     TIMERSSFEKNPIDFLEAKGYKEVKKDLIIKLPKYSLFELENGRKRMLASAGELQKGNE
                     LALPSKYVNFLYLASHYEKLKGSPEDNEQKQLFVEQHKHYLDEIIEQISEFSKRVILAD
                     ANLDKVLSAYNKHRDKPIREQAENIIHLFTLTNLGAPAAFKYFDTTIDRKRYTSTKEVL
                     DATLIHQSITGLYETRIDLSQLGGD"
ORIGIN
        1 ttacgaaatc atcctgtgga gcttagtagg tttagcaaga tggcagcgcc taaatgtaga
       61 atgataaaag gattaagaga ttaatttccc taaaaatgat aaaacaagcg ttttgaaagc
      121 gcttgttttt ttggtttgca gtcagagtag aatagaagta tcaaaaaaag caccgactcg
      181 gtgccacttt ttcaagttga taacggacta gccttatttt aacttgctat gctgttttga
      241 atggttccaa caagattatt ttataacttt tataacaaat aatcaaggag aaattcaaag
      301 aaatttatca gccataaaac aatacttaat actatagaat gataacaaaa taaactactt
      361 tttaaaagaa ttttgtgtta taatctattt attattaagt attgggtaat attttttgaa
      421 gagatatttt gaaaaagaaa aattaaagca tattaaacta atttcggagg tcattaaaac
      481 tattattgaa atcatcaaac tcattatgga tttaatttaa actttttatt ttaggaggca
      541 aaaatggata agaaatactc aataggctta gatatcggca caaatagcgt cggatgggcg
      601 gtgatcactg atgaatataa ggttccgtct aaaaagttca aggttctggg aaatacagac
      661 cgccacagta tcaaaaaaaa tcttataggg gctcttttat ttgacagtgg agagacagcg
      721 gaagcgactc gtctcaaacg gacagctcgt agaaggtata cacgtcggaa gaatcgtatt
      781 tgttatctac aggagatttt ttcaaatgag atggcgaaag tagatgatag tttctttcat
      841 cgacttgaag agtctttttt ggtggaagaa gacaagaagc atgaacgtca tcctattttt
      901 ggaaatatag tagatgaagt tgcttatcat gagaaatatc caactatcta tcatctgcga
      961 aaaaaattgg tagattctac tgataaagcg gatttgcgct taatctattt ggccttagcg
     1021 catatgatta agtttcgtgg tcattttttg attgagggag atttaaatcc tgataatagt
     1081 gatgtggaca aactatttat ccagttggta caaacctaca atcaattatt tgaagaaaac
     1141 cctattaacg caagtggagt agatgctaaa gcgattcttt ctgcacgatt gagtaaatca
     1201 agacgattag aaaatctcat tgctcagctc cccggtgaga agaaaaatgg cttatttggg
     1261 aatctcattg ctttgtcatt gggtttgacc cctaatttta aatcaaattt tgatttggca
     1321 gaagatgcta aattacagct ttcaaaagat acttacgatg atgatttaga taatttattg
     1381 gcgcaaattg gagatcaata tgctgatttg tttttggcag ctaagaattt atcagatgct
     1441 attttacttt cagatatcct aagagtaaat actgaaataa ctaaggctcc cctatcagct
     1501 tcaatgatta aacgctacga tgaacatcat caagacttga ctcttttaaa agctttagtt
     1561 cgacaacaac ttccagaaaa gtataaagaa atcttttttg atcaatcaaa aaacggatat
     1621 gcaggttata ttgatggggg agctagccaa gaagaatttt ataaatttat caaaccaatt
     1681 ttagaaaaaa tggatggtac tgaggaatta ttggtgaaac taaatcgtga agatttgctg
     1741 cgcaagcaac ggacctttga caacggctct attccccatc aaattcactt gggtgagctg
     1801 catgctattt tgagaagaca agaagacttt tatccatttt taaaagacaa tcgtgagaag
     1861 attgaaaaaa tcttgacttt tcgaattcct tattatgttg gtccattggc gcgtggcaat
     1921 agtcgttttg catggatgac tcggaagtct gaagaaacaa ttaccccatg gaattttgaa
     1981 gaagttgtcg ataaaggtgc ttcagctcaa tcatttattg aacgcatgac aaactttgat
     2041 aaaaatcttc caaatgaaaa agtactacca aaacatagtt tgctttatga gtattttacg
     2101 gtttataacg aattgacaaa ggtcaaatat gttactgaag gaatgcgaaa accagcattt
     2161 ctttcaggtg aacagaagaa agccattgtt gatttactct tcaaaacaaa tcgaaaagta
     2221 accgttaagc aattaaaaga agattatttc aaaaaaatag aatgttttga tagtgttgaa
     2281 atttcaggag ttgaagatag atttaatgct tcattaggta cctaccatga tttgctaaaa
     2341 attattaaag ataaagattt tttggataat gaagaaaatg aagatatctt agaggatatt
     2401 gttttaacat tgaccttatt tgaagatagg gagatgattg aggaaagact taaaacatat
     2461 gctcacctct ttgatgataa ggtgatgaaa cagcttaaac gtcgccgtta tactggttgg
     2521 ggacgtttgt ctcgaaaatt gattaatggt attagggata agcaatctgg caaaacaata
     2581 ttagattttt tgaaatcaga tggttttgcc aatcgcaatt ttatgcagct gatccatgat
     2641 gatagtttga catttaaaga agacattcaa aaagcacaag tgtctggaca aggcgatagt
     2701 ttacatgaac atattgcaaa tttagctggt agccctgcta ttaaaaaagg tattttacag
     2761 actgtaaaag ttgttgatga attggtcaaa gtaatggggc ggcataagcc agaaaatatc
     2821 gttattgaaa tggcacgtga aaatcagaca actcaaaagg gccagaaaaa ttcgcgagag
     2881 cgtatgaaac gaatcgaaga aggtatcaaa gaattaggaa gtcagattct taaagagcat
     2941 cctgttgaaa atactcaatt gcaaaatgaa aagctctatc tctattatct ccaaaatgga
     3001 agagacatgt atgtggacca agaattagat attaatcgtt taagtgatta tgatgtcgat
     3061 cacattgttc cacaaagttt ccttaaagac gattcaatag acaataaggt cttaacgcgt
     3121 tctgataaaa atcgtggtaa atcggataac gttccaagtg aagaagtagt caaaaagatg
     3181 aaaaactatt ggagacaact tctaaacgcc aagttaatca ctcaacgtaa gtttgataat
     3241 ttaacgaaag ctgaacgtgg aggtttgagt gaacttgata aagctggttt tatcaaacgc
     3301 caattggttg aaactcgcca aatcactaag catgtggcac aaattttgga tagtcgcatg
     3361 aatactaaat acgatgaaaa tgataaactt attcgagagg ttaaagtgat taccttaaaa
     3421 tctaaattag tttctgactt ccgaaaagat ttccaattct ataaagtacg tgagattaac
     3481 aattaccatc atgcccatga tgcgtatcta aatgccgtcg ttggaactgc tttgattaag
     3541 aaatatccaa aacttgaatc ggagtttgtc tatggtgatt ataaagttta tgatgttcgt
     3601 aaaatgattg ctaagtctga gcaagaaata ggcaaagcaa ccgcaaaata tttcttttac
     3661 tctaatatca tgaacttctt caaaacagaa attacacttg caaatggaga gattcgcaaa
     3721 cgccctctaa tcgaaactaa tggggaaact ggagaaattg tctgggataa agggcgagat
     3781 tttgccacag tgcgcaaagt attgtccatg ccccaagtca atattgtcaa gaaaacagaa
     3841 gtacagacag gcggattctc caaggagtca attttaccaa aaagaaattc ggacaagctt
     3901 attgctcgta aaaaagactg ggatccaaaa aaatatggtg gttttgatag tccaacggta
     3961 gcttattcag tcctagtggt tgctaaggtg gaaaaaggga aatcgaagaa gttaaaatcc
     4021 gttaaagagt tactagggat cacaattatg gaaagaagtt cctttgaaaa aaatccgatt
     4081 gactttttag aagctaaagg atataaggaa gttaaaaaag acttaatcat taaactacct
     4141 aaatatagtc tttttgagtt agaaaacggt cgtaaacgga tgctggctag tgccggagaa
     4201 ttacaaaaag gaaatgagct ggctctgcca agcaaatatg tgaatttttt atatttagct
     4261 agtcattatg aaaagttgaa gggtagtcca gaagataacg aacaaaaaca attgtttgtg
     4321 gagcagcata agcattattt agatgagatt attgagcaaa tcagtgaatt ttctaagcgt
     4381 gttattttag cagatgccaa tttagataaa gttcttagtg catataacaa acatagagac
     4441 aaaccaatac gtgaacaagc agaaaatatt attcatttat ttacgttgac gaatcttgga
     4501 gctcccgctg cttttaaata ttttgataca acaattgatc gtaaacgata tacgtctaca
     4561 aaagaagttt tagatgccac tcttatccat caatccatca ctggtcttta tgaaacacgc
     4621 attgatttga gtcagctagg aggtgactga
//
