LOCUS       sequence-252671-        6033 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Level0 LbCas12a, rice codon optimized.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6033)
  AUTHORS   Hahn F, Korolev A, Sanjurjo Loures L, Nekrasov V
  TITLE     A modular cloning toolkit for genome editing in plants.
  JOURNAL   BMC Plant Biol. 2020 Apr 23;20(1):179. doi: 
            10.1186/s12870-020-02388-2.
  PUBMED    32326901
REFERENCE   2  (bases 1 to 6033)
  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: "BMC Plant 
            Biol."; date: "2020-04-23"; volume: "20(1):179. doi"; pages: " 
            10.1186/s12870-020-02388-2"
COMMENT     Sequence Label: pFH17
FEATURES             Location/Qualifiers
     source          1..6033
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(24..42)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        110..214
                     /gene="bla"
                     /label=AmpR promoter
     CDS             436..1227
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      1398..1986
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1887..1906
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2140..2157
                     /label=L4440
                     /note="L4440 vector, forward primer"
     CDS             2241..2261
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             2286..5966
                     /codon_start=1
                     /product="CRISPR-associated protein from Lachnospiraceae 
                     bacterium ND2006 (Zetsche et al., 2015)"
                     /label=LbCpf1
                     /translation="SKLEKFTNCYSLSKTLRFKAIPVGKTQENIDNKRLLVEDEKRAED
                     YKGVKKLLDRYYLSFINDVLHSIKLKNLNNYISLFRKKTRTEKENKELENLEINLRKEI
                     AKAFKGNEGYKSLFKKDIIETILPEFLDDKDEIALVNSFNGFTTAFTGFFDNRENMFSE
                     EAKSTSIAFRCINENLTRYISNMDIFEKVDAIFDKHEVQEIKEKILNSDYDVEDFFEGE
                     FFNFVLTQEGIDVYNAIIGGFVTESGEKIKGLNEYINLYNQKTKQKLPKFKPLYKQVLS
                     DRESLSFYGEGYTSDEEVLEVFRNTLNKNSEIFSSIKKLEKLFKNFDEYSSAGIFVKNG
                     PAISTISKDIFGEWNVIRDKWNAEYDDIHLKKKAVVTEKYEDDRRKSFKKIGSFSLEQL
                     QEYADADLSVVEKLKEIIIQKVDEIYKVYGSSEKLFDADFVLEKSLKKNDAVVAIMKDL
                     LDSVKSFENYIKAFFGEGKETNRDESFYGDFVLAYDILLKVDHIYDAIRNYVTQKPYSK
                     DKFKLYFQNPQFMGGWDKDKETDYRATILRYGSKYYLAIMDKKYAKCLQKIDKDDVNGN
                     YEKINYKLLPGPNKMLPKVFFSKKWMAYYNPSEDIQKIYKNGTFKKGDMFNLNDCHKLI
                     DFFKDSISRYPKWSNAYDFNFSETEKYKDIAGFYREVEEQGYKVSFESASKKEVDKLVE
                     EGKLYMFQIYNKDFSDKSHGTPNLHTMYFKLLFDENNHGQIRLSGGAELFMRRASLKKE
                     ELVVHPANSPIANKNPDNPKKTTTLSYDVYKDKRFSEDQYELHIPIAINKCPKNIFKIN
                     TEVRVLLKHDDNPYVIGIDRGERNLLYIVVVDGKGNIVEQYSLNEIINNFNGIRIKTDY
                     HSLLDKKEKERFEARQNWTSIENIKELKAGYISQVVHKICELVEKYDAVIALEDLNSGF
                     KNSRVKVEKQVYQKFEKMLIDKLNYMVDKKSNPCATGGALKGYQITNKFESFKSMSTQN
                     GFIFYIPAWLTSKIDPSTGFVNLLKTKYTSIADSKKFISSFDRIMYVPEEDLFEFALDY
                     KNFSRTDADYIKKWKLYSYGNRIRIFRNPKKNNVFDWEEVCLTSAYKELFNKYGINYQQ
                     GDIRALLCEQSDKAFYSSFMALMSLMLQMRNSITGRTDVDFLISPVKNSDGIFYDSRNY
                     EAQENAILPKNADANGAYNIARKVLWAIGQFKKAEDEKLDKVKIAISNKEWLEYAQTSV
                     KH"
     CDS             5967..6014
                     /codon_start=1
                     /product="bipartite nuclear localization signal from 
                     nucleoplasmin"
                     /label=nucleoplasmin NLS
                     /translation="KRPAATKKAGQAKKKK"
ORIGIN
        1 gtggctcttc agtggacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat
       61 gataataatg gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc
      121 tatttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
      181 ataaatgctt caataatatt gaaaaaggaa gagtatgcgc tcacgcaact ggtccagaac
      241 cttgaccgaa cgcagcggtg gtaacggcgc agtggcggtt ttcatggctt gttatgactg
      301 tttttttttg gggtacagtc tatgcctcgg gcatccaagc agcaagcgcg ttacgccgtg
      361 ggtcgatgtt tgatgttatg gagcagcaac gatgttacgc agcagggcag tcgccctaaa
      421 acaaagttaa acatcatgag ggaagcggtg atcgccgaag tatcgactca actatcagag
      481 gtagttggcg tcatcgagcg ccatctcgaa ccgacgttgc tggccgtaca tttgtacggc
      541 tccgcagtgg atggcggcct gaagccacac agcgatattg atttgctggt tacggtgacc
      601 gtaaggcttg atgaaacaac gcggcgagct ttgatcaacg accttttgga aacttcggct
      661 tcccctggag agagcgagat tctccgcgct gtagaagtca ccattgttgt gcacgacgac
      721 atcattccgt ggcgttatcc agctaagcgc gaactgcaat ttggagaatg gcagcgcaat
      781 gacattcttg caggtatctt cgagccagcc acgatcgaca ttgatctggc tatcttgctg
      841 acaaaagcaa gagaacatag cgttgccttg gtaggtccag cggcggagga actctttgat
      901 ccggttcctg aacaggatct atttgaggcg ctaaatgaaa ccttaacgct atggaactcg
      961 ccgcccgact gggctggcga tgagcgaaat gtagtgctta cgttgtcccg catttggtac
     1021 agcgcagtaa ccggcaaaat cgcgccgaag gatgtcgctg ccgactgggc aatggagcgc
     1081 ctgccggccc agtatcagcc cgtcatactt gaagctagac aggcttatct tggacaagaa
     1141 gaagatcgct tggcctcgcg cgcagatcag ttggaagaat ttgtccatta cgtaaaaggc
     1201 gagatcacca aggtagtcgg caaataactg tcagaccaag tttactcata tatactttag
     1261 attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct ttttgataat
     1321 ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga ccccgtagaa
     1381 aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg cttgcaaaca
     1441 aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc aactcttttt
     1501 ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct agtgtagccg
     1561 tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc tctgctaatc
     1621 ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt ggactcaaga
     1681 cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg cacacagccc
     1741 agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct atgagaaagc
     1801 gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag ggtcggaaca
     1861 ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag tcctgtcggg
     1921 tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg gcggagccta
     1981 tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg gccttttgct
     2041 cacatgttct ttcctgcgtt atcccctgat tctgtggata accgtattac cgcctttgag
     2101 tgagctgata ccgctcgccg cagccgaacg accgagcgca gcgagtcagt gagcgaggaa
     2161 gcggaagagc gcccaatacg caaaccgcct ctccccgcgc gttggccgat tcattaatca
     2221 ctctgtggtc tcaaatggct cctaagaaga agcggaaggt tggtattcac ggggtgcctg
     2281 cggcttcaaa gctcgagaaa ttcaccaact gttattcgtt gagcaaaaca ctgcggttta
     2341 aagcgattcc agtcggcaag actcaagaga atatagacaa taagcggctg ttggtggaag
     2401 atgaaaagcg cgcggaagat tacaaagggg tgaagaagtt gttggacaga tactacctct
     2461 cttttatcaa tgatgtcttg cactcaatca aattgaagaa tctgaacaac tacatctccc
     2521 tcttcagaaa gaaaacaagg acagaaaagg agaataagga acttgaaaat ttggagatca
     2581 atctgaggaa agagatcgcg aaagccttta aaggcaacga aggatacaaa agtctgttca
     2641 agaaggatat aattgagaca attttgccag agttcctcga tgacaaggac gagattgcgc
     2701 tggtcaattc gttcaacgga ttcacaacag cattcacagg cttctttgat aatcgggaaa
     2761 atatgttctc tgaggaggca aagtccactt ctattgcgtt caggtgtatc aatgagaatc
     2821 tcactaggta catttccaac atggatatct ttgagaaggt tgacgcaatt tttgacaagc
     2881 acgaagttca ggagattaag gagaagatcc tcaattccga ttatgacgtt gaggacttct
     2941 tcgaaggtga gttttttaat ttcgtgctca ctcaagaggg tatcgacgtg tataatgcga
     3001 tcatcggtgg gttcgtgact gagtccggtg aaaagattaa gggattgaac gagtatatca
     3061 acctttacaa ccaaaagacg aaacagaagc tgccaaagtt caagcctctt tacaaacagg
     3121 ttctttcaga ccgcgagtca ctctcgttct atggggaggg ctacacttcg gatgaggaag
     3181 tcctggaggt gttcaggaat actctcaata agaattcgga gattttctct tctataaaaa
     3241 aactggaaaa gttgtttaag aattttgacg aatactctag cgccggcata tttgtgaaaa
     3301 acggcccggc catatcaacg ataagtaaag atatcttcgg cgaatggaac gtgatcagag
     3361 acaaatggaa cgcggagtat gacgatattc acctgaagaa gaaggctgtc gtaacggaga
     3421 agtacgagga tgatcgcagg aaaagcttca aaaagatcgg aagtttcagc ctggaacagt
     3481 tgcaggagta tgctgacgcc gatcttagcg tcgtcgagaa gttgaaggag ataatcatcc
     3541 aaaaggtcga cgagatatat aaagtctatg gatcaagtga aaaactgttc gacgccgact
     3601 tcgttttgga gaagtccctg aagaagaacg acgctgttgt tgccattatg aaggatctgc
     3661 tcgacagcgt gaagagtttc gagaactata ttaaggcttt tttcggggag gggaaggaga
     3721 ctaacagaga tgagtccttc tacggagact tcgtcctcgc gtacgatata ctccttaagg
     3781 tagaccacat ctacgacgca atcagaaatt acgtgacaca aaagccgtac agcaaggaca
     3841 agttcaaact ctacttccag aacccccagt tcatgggcgg ctgggacaag gacaaggaaa
     3901 cggattacag ggctacgatc ctgaggtatg gttcaaaata ctacttggcg attatggaca
     3961 agaagtacgc caagtgtctc cagaagattg acaaagacga tgtcaatggc aattatgaga
     4021 agatcaacta caagctgctt ccgggtccga acaagatgct cccaaaggtt ttcttcagca
     4081 agaaatggat ggcctactat aacccaagcg aggacatcca gaagatttat aagaacggta
     4141 cgttcaagaa gggcgacatg ttcaatctta acgactgtca caagctgatc gacttcttca
     4201 aagactcaat tagccggtac ccaaagtggt ctaacgccta tgacttcaac ttttcggaaa
     4261 ccgagaagta caaggatata gccggatttt atagagaggt ggaagagcag ggctacaagg
     4321 tgtcattcga gtccgccagc aagaaggaag tggacaagct cgtggaagag ggtaagctct
     4381 acatgttcca gatttataat aaagacttta gcgataagag ccacgggaca cctaatctcc
     4441 acacaatgta tttcaagctg ctcttcgacg agaataacca cggccaaatc aggttgtcag
     4501 gaggggctga actcttcatg cggcgcgcta gccttaagaa ggaggagctt gtagtccacc
     4561 ctgcgaatag tccaattgcg aataagaacc cggacaatcc taaaaagact acaacattga
     4621 gctacgacgt gtacaaggat aagaggtttt ccgaggatca gtacgagctc cacatcccga
     4681 ttgcgatcaa caagtgccca aagaatattt tcaagataaa cacagaggtg cgtgtactcc
     4741 tgaagcatga cgacaatcct tacgtcattg ggattgatcg gggcgagagg aacctcctct
     4801 atattgtggt ggtggacggg aaggggaaca tagtcgaaca gtactccctt aacgaaataa
     4861 ttaacaattt caacggcatc cgtatcaaga ccgactacca ttcgttgctg gacaagaagg
     4921 agaaggagag atttgaggcg cggcaaaatt ggacaagtat cgagaacatc aaggaactca
     4981 aagcaggtta tatctctcaa gttgtgcata agatatgcga gctggttgag aagtatgacg
     5041 cagtgatcgc tcttgaggac ctcaactcgg gctttaagaa ttctagagtt aaagtggaga
     5101 agcaggtcta tcaaaagttc gagaagatgc ttatagataa gctcaactac atggtcgata
     5161 agaaatcgaa cccatgtgcc accggcggcg cactcaaagg ttaccaaata acaaacaaat
     5221 tcgagtcctt caaatcgatg agtactcaga atgggttcat attttatata ccggcgtggc
     5281 ttacgtctaa gatcgacccg tcaactggtt ttgtcaacct gttaaagacg aaatacacgt
     5341 ccattgccga ttcgaaaaag ttcatatcta gttttgatcg tattatgtac gtcccagagg
     5401 aagatctttt cgagtttgct ctcgactaca aaaacttttc gcggaccgat gcggattaca
     5461 ttaaaaaatg gaaactctat tcgtacggca acagaatcag gatttttcgc aaccctaaga
     5521 agaataacgt ctttgattgg gaggaagttt gcttgactag cgcgtacaag gagctcttta
     5581 ataagtatgg cattaactac caacagggtg atatcagagc actgctttgc gaacaatctg
     5641 acaaggcttt ctactcatcc ttcatggctt tgatgagcct gatgctccag atgagaaatt
     5701 caattacagg cagaaccgac gtggatttct tgatctcccc ggttaaaaat tctgatggca
     5761 tcttttacga tagcaggaac tatgaagcgc aagagaatgc gattctgcca aaaaatgcag
     5821 acgccaacgg tgcctataac atcgccagga aagtcctgtg ggcgatcggc cagttcaaaa
     5881 aggccgagga cgaaaaattg gacaaggtca aaatcgctat cagcaacaaa gagtggctgg
     5941 agtatgctca gacatccgta aagcataagc gtcctgctgc caccaaaaag gccggacagg
     6001 ctaagaaaaa gaagtgagct ttgagaccac gaa
//
