LOCUS       sequence-181787-        7897 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Module A, Promoter: OsAct1, Gene: TaCas9, Terminator: HSP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7897)
  AUTHORS   Cermak T, Curtin SJ, Gil-Humanes J, Cegan R, Kono TJY, Konecna E, 
            Belanto JJ, Starker CG, Mathre JW, Greenstein RL, Voytas DF
  TITLE     A multi-purpose toolkit to enable advanced genome engineering in 
            plants.
  JOURNAL   Plant Cell. 2017 May 18. pii: tpc.00922.2016. doi: 
            10.1105/tpc.16.00922.
  PUBMED    28522548
REFERENCE   2  (bases 1 to 7897)
  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: "Plant Cell.
            2017 May 18. pii: tpc.00922.2016. doi: 10.1105/tpc.16.00922."
COMMENT     Sequence Label: pMOD_A1111
FEATURES             Location/Qualifiers
     source          1..7897
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      80..668
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     569..588
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     822..839
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        884..1725
                     /label=Act1 promoter
                     /note="rice actin promoter"
     CDS             1728..1745
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2314..6417
                     /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             2314..6417
                     /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             6418..6465
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
     terminator      6477..6725
                     /label=HSP terminator
                     /note="efficient transcription terminator from the 
                     Arabidopsis thaliana heat shock protein 18.2 gene (Nagaya 
                     et al., 2010)"
     promoter        6805..6909
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gccttgacgg caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
       61 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
      121 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ctaactcttt
      181 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
      241 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
      301 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
      361 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgctcacagc
      421 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
      481 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
      541 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
      601 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
      661 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
      721 ctcacatgtt ctttcctgcg ttatcccctg attctgtgga taaccgtatt accgcctttg
      781 agtgagctga taccgctcgc cgcagccgaa cgaccgagcg cagcgagtca gtgagcgagg
      841 aagcggaaga gcgcccacct gctcatgatc tgcaggcgcg ccgtcgaggt cattcatatg
      901 cttgagaaga gagtcgggat agtccaaaat aaaacaaagg taagattacc tggtcaaaag
      961 tgaaaacatc agttaaaagg tggtataaag taaaatatcg gtaataaaag gtggcccaaa
     1021 gtgaaattta ctcttttcta ctattataaa aattgaggat gtttttgtcg gtactttgat
     1081 acgtcatttt tgtatgaatt ggtttttaag tttattcgct tttggaaatg catatctgta
     1141 tttgagtcgg gttttaagtt cgtttgcttt tgtaaataca gagggatttg tataagaaat
     1201 atctttaaaa aaacccatat gctaatttga cataattttt gagaaaaata tatattcagg
     1261 cgaattctca caatgaacaa taataagatt aaaatagctt tcccccgttg cagcgcatgg
     1321 gtattttttc tagtaaaaat aaaagataaa cttagactca aaacatttac aaaaacaacc
     1381 cctaaagttc ctaaagccca aagtgctatc cacgatccat agcaagccca gcccaaccca
     1441 acccaaccca acccacccca gtccagccaa ctggacaata gtctccacac ccccccacta
     1501 tcaccgtgag ttgtccgcac gcaccgcacg tctcgcagcc aaaaaaaaaa aaagaaagaa
     1561 aaaaaagaaa aagaaaaaac agcagggggt ccgggtcgtg ggggccggaa acgcgaggag
     1621 gatcgcgagc cagcgacgag gccggccctc cctccgcttc caaagaaacg ccccccatcg
     1681 ccactatata catacccccc cctctcctcc catcccccca accctaccac caccaccacc
     1741 accacctcca cctcctcccc cctcgctgcc ggacgacgag ctcctccccc ctccccctcc
     1801 gccgccgccg cgccggtaac caccccgccc ctctcctctt tctttctccg tttttttttt
     1861 ccgtctcggt ctcgatcttt ggccttggta gtttgggtgg gcgagaggcg gcttcgtgcg
     1921 cgcccagatc ggtgcgcggg aggggcggga tctcgcggct ggggctctcg ccggcgtgga
     1981 tccggcccgg atctcgcggg gaatggggct ctcggatgta gatctgcgat ccgccgttgt
     2041 tgggggagat gatggggggt ttaaaatttc cgccatgcta aacaagatca ggaagagggg
     2101 aaaagggcac tatggtttat atttttatat atttctgctg cttcgtcagg cttagatgtg
     2161 ctagatcttt ctttcttctt tttgtgggta gaatttgaat ccctcagcat tgttcatcgg
     2221 tagtttttct tttcatgatt tgtgacaaat gcagcctcgt gcggagcttt tttgtaggta
     2281 gacgataagc ttcctgcagg gcgatctatt cgaatggaca agaagtactc gatcggcctc
     2341 gacatcggga cgaactcagt tggctgggcc gtgatcaccg acgagtacaa ggtgccctct
     2401 aagaagttca aggtcctggg gaacaccgac cgccattcca tcaagaagaa cctcatcggc
     2461 gctctcctgt tcgacagcgg ggagaccgct gaggctacga ggctcaagag aaccgctagg
     2521 cgccggtaca cgagaaggaa gaacaggatc tgctacctcc aagagatttt ctccaacgag
     2581 atggccaagg ttgacgattc attcttccac cgcctggagg agtctttcct cgtggaggag
     2641 gataagaagc acgagcggca tcccatcttc ggcaacatcg tggacgaggt tgcctaccac
     2701 gagaagtacc ctacgatcta ccatctgcgg aagaagctcg tggactccac cgataaggcg
     2761 gacctcagac tgatctacct cgctctggcc cacatgatca agttccgcgg ccatttcctg
     2821 atcgaggggg atctcaaccc agacaacagc gatgttgaca agctgttcat ccaactcgtg
     2881 cagacctaca accaactctt cgaggagaac ccgatcaacg cctctggcgt ggacgcgaag
     2941 gctatcctgt ccgcgaggct ctcgaagtcc aggaggctgg agaacctgat cgctcagctc
     3001 ccaggcgaga agaagaacgg cctgttcggg aacctcatcg ctctcagcct ggggctcacc
     3061 ccgaacttca agtcgaactt cgatctcgct gaggacgcca agctgcaact ctccaaggac
     3121 acctacgacg atgacctcga taacctcctg gcccagatcg gcgatcaata cgcggacctg
     3181 ttcctcgctg ccaagaacct gtcggacgcc atcctcctgt cagatatcct ccgcgtgaac
     3241 accgagatca cgaaggctcc actctctgcc tccatgatca agcgctacga cgagcaccat
     3301 caggatctga ccctcctgaa ggcgctggtc cgccaacagc tcccggagaa gtacaaggag
     3361 attttcttcg atcagtcgaa gaacggctac gctgggtaca tcgacggcgg ggcctcacaa
     3421 gaggagttct acaagttcat caagccaatc ctggagaaga tggacggcac ggaggagctc
     3481 ctggtgaagc tcaacaggga ggacctcctg cggaagcaga gaaccttcga taacggcagc
     3541 atcccccacc aaatccatct cggggagctg cacgccatcc tgagaaggca agaggacttc
     3601 taccctttcc tcaaggataa ccgggagaag atcgagaaga tcctgacctt cagaatccca
     3661 tactacgtcg gccctctcgc gcgggggaac tcaagattcg cttggatgac ccgcaagtct
     3721 gaggagacca tcacgccgtg gaacttcgag gaggtggtgg acaagggcgc tagcgctcag
     3781 tcgttcatcg agaggatgac caacttcgac aagaacctgc ccaacgagaa ggtgctccct
     3841 aagcactcgc tcctgtacga gtacttcacc gtctacaacg agctcacgaa ggtgaagtac
     3901 gtcaccgagg gcatgcgcaa gccagcgttc ctgtccgggg agcagaagaa ggctatcgtg
     3961 gacctcctgt tcaagaccaa ccggaaggtc acggttaagc aactcaagga ggactacttc
     4021 aagaagatcg agtgcttcga ttcggtcgag atcagcggcg ttgaggaccg cttcaacgcc
     4081 agcctcggga cctaccacga tctcctgaag atcatcaagg ataaggactt cctggacaac
     4141 gaggagaacg aggatatcct ggaggacatc gtgctgaccc tcacgctgtt cgaggacagg
     4201 gagatgatcg aggagcgcct gaagacgtac gcccatctct tcgatgacaa ggtcatgaag
     4261 caactcaagc gccggagata caccggctgg gggaggctgt cccgcaagct catcaacggc
     4321 atccgggaca agcagtccgg gaagaccatc ctcgacttcc tcaagagcga tggcttcgcc
     4381 aacaggaact tcatgcaact gatccacgat gacagcctca ccttcaagga ggatatccaa
     4441 aaggctcaag tgagcggcca gggggactcg ctgcacgagc atatcgcgaa cctcgctggc
     4501 tcccccgcga tcaagaaggg catcctccag accgtgaagg ttgtggacga gctcgtgaag
     4561 gtcatgggcc ggcacaagcc tgagaacatc gtcatcgaga tggccagaga gaaccaaacc
     4621 acgcagaagg ggcaaaagaa ctctagggag cgcatgaagc gcatcgagga gggcatcaag
     4681 gagctggggt cccaaatcct caaggagcac ccagtggaga acacccaact gcagaacgag
     4741 aagctctacc tgtactacct ccagaacggc agggatatgt acgtggacca agagctggat
     4801 atcaaccgcc tcagcgatta cgacgtcgat catatcgttc cccagtcttt cctgaaggat
     4861 gactccatcg acaacaaggt cctcaccagg tcggacaaga accgcggcaa gtcagataac
     4921 gttccatctg aggaggtcgt taagaagatg aagaactact ggaggcagct cctgaacgcc
     4981 aagctgatca cgcaaaggaa gttcgacaac ctcaccaagg ctgagagagg cgggctctca
     5041 gagctggaca aggccggctt catcaagcgg cagctggtcg agaccagaca aatcacgaag
     5101 cacgttgcgc aaatcctcga ctctcggatg aacacgaagt acgatgagaa cgacaagctg
     5161 atcagggagg ttaaggtgat caccctgaag tctaagctcg tctccgactt caggaaggat
     5221 ttccagttct acaaggttcg cgagatcaac aactaccacc atgcccatga cgcttacctc
     5281 aacgctgtgg tcggcaccgc tctgatcaag aagtacccaa agctggagtc cgagttcgtg
     5341 tacggggact acaaggttta cgatgtgcgc aagatgatcg ccaagtcgga gcaagagatc
     5401 ggcaaggcta ccgccaagta cttcttctac tcaaacatca tgaacttctt caagaccgag
     5461 atcacgctgg ccaacggcga gatccggaag agaccgctca tcgagaccaa cggcgagacg
     5521 ggggagatcg tgtgggacaa gggcagggat ttcgcgaccg tccgcaaggt tctctccatg
     5581 ccccaggtga acatcgtcaa gaagaccgag gtccaaacgg gcgggttctc aaaggagtct
     5641 atcctgccta agcggaacag cgacaagctc atcgccagaa agaaggactg ggacccaaag
     5701 aagtacggcg ggttcgacag ccctaccgtg gcctactcgg tcctggttgt ggcgaaggtt
     5761 gagaagggca agtccaagaa gctcaagagc gtgaaggagc tcctggggat caccatcatg
     5821 gagaggtcca gcttcgagaa gaacccaatc gacttcctgg aggccaaggg ctacaaggag
     5881 gtgaagaagg acctgatcat caagctcccg aagtactctc tcttcgagct ggagaacggc
     5941 aggaagagaa tgctggcttc cgctggcgag ctccagaagg ggaacgagct cgcgctgcca
     6001 agcaagtacg tgaacttcct ctacctggct tcccactacg agaagctcaa gggcagcccg
     6061 gaggacaacg agcaaaagca gctgttcgtc gagcagcaca agcattacct cgacgagatc
     6121 atcgagcaaa tctccgagtt cagcaagcgc gtgatcctcg ccgacgcgaa cctggataag
     6181 gtcctctccg cctacaacaa gcaccgggac aagcccatca gagagcaagc ggagaacatc
     6241 atccatctct tcaccctgac gaacctcggc gctcctgctg ctttcaagta cttcgacacc
     6301 acgatcgatc ggaagagata cacctccacg aaggaggtcc tggacgcgac cctcatccac
     6361 cagtcgatca ccggcctgta cgagacgagg atcgacctct cacaactcgg cggggataag
     6421 agacccgcag caaccaagaa ggcagggcaa gcaaagaaga agaagtgact cgagtatatg
     6481 aagatgaaga tgaaatattt ggtgtgtcaa ataaaaagct tgtgtgctta agtttgtgtt
     6541 tttttcttgg cttgttgtgt tatgaatttg tggctttttc taatattaaa tgaatgtaag
     6601 atcacattat aatgaataaa caaatgtttc tataatccat tgtgaatgtt ttgttggatc
     6661 tcttctgcag catataacta ctgtatgtgc tatggtatgg actatggaat atgattaaag
     6721 ataaggagct cgttaaccgg tgacggacgt gggcaggtgt gatgaaaagc cttagggtca
     6781 ggtggcactt ttcggggaaa tgtgcgatga acccctattt gtttattttt ctaaatacat
     6841 tcaaatatgt atccgctaat gagacaataa ccctgataaa tgcttcaatt atattgaaaa
     6901 aggaagagta tgagtatcca acatttccgt gtcgccctta ttcccttttt tgcggccttc
     6961 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     7021 ttgggtgccc gagtgggtta cattgaactg gatctcaaca gcggcaaaat cttagagagt
     7081 tttcgccccg aagaacgctt cccaatgatg agcactttca aagttctgct atgtggcgcg
     7141 gtgttatccc gtattgatgc tgggcaagag caactcggtc gccgcataca ctattctcag
     7201 aatgacttgg ttgagtattc accagtcaca gaaaagcatc ttacggatgg catgacagta
     7261 agagaattat gtagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg
     7321 acaacgattg gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta
     7381 actcgccttg atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac
     7441 accacgatgc ctgtagcaat ggcaacaact ctccgcaaac tattaactgg cgaactactt
     7501 actcttgctt cacgccagca actcattgac tggatggagg cggacaaagt tgcaggacca
     7561 cttctgcgct cggcacttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     7621 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     7681 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     7741 ataggtgcct cactgattaa gcattggtaa ctgtcagacg aagggaaata aatagtagcc
     7801 cgcctgatgt gcgggctttt ttttggtggt ggttaccagt gtcgcttagc taccgtaggt
     7861 cctcttaggc ggtccgatta gcgacatagt catcaat
//
