LOCUS       sequence-203090-        7189 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  OsActin Promoter-LwaCas13a-HSP Terminator.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7189)
  AUTHORS   Abudayyeh OO, Gootenberg JS, Essletzbichler P, Han S, Joung J, 
            Belanto JJ, Verdine V, Cox DBT, Kellner MJ, Regev A, Lander ES, 
            Voytas DF, Ting AY, Zhang F
  TITLE     RNA targeting with CRISPR-Cas13.
  JOURNAL   Nature. 2017 Oct 4. doi: 10.1038/nature24049.
  PUBMED    28976959
REFERENCE   2  (bases 1 to 7189)
  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: "Nature. 
            2017 Oct 4. doi: 10.1038/nature24049."
COMMENT     Sequence Label: pJJB302
FEATURES             Location/Qualifiers
     source          1..7189
                     /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        880..1721
                     /label=Act1 promoter
                     /note="rice actin promoter"
     CDS             1724..1741
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             2309..5764
                     /codon_start=1
                     /product="Leptotrichia wadei CRISPR-Cas effector that acts 
                     as an RNA-guided RNAse (Abudayyeh et al., 2017)"
                     /label=LwCas13a
                     /note="previously called C2c2"
                     /translation="MKVTKVDGISHKKYIEEGKLVKSTSEENRTSERLSELLSIRLDIY
                     IKNPDNASEEENRIRRENLKKFFSNKVLHLKDSVLYLKNRKEKNAVQDKNYSEEDISEY
                     DLKNKNSFSVLKKILLNEDVNSEELEIFRKDVEAKLNKINSLKYSFEENKANYQKINEN
                     NVEKVGGKSKRNIIYDYYRESAKRNDYINNVQEAFDKLYKKEDIEKLFFLIENSKKHEK
                     YKIREYYHKIIGRKNDKENFAKIIYEEIQNVNNIKELIEKIPDMSELKKSQVFYKYYLD
                     KEELNDKNIKYAFCHFVEIEMSQLLKNYVYKRLSNISNDKIKRIFEYQNLKKLIENKLL
                     NKLDTYVRNCGKYNYYLQVGEIATSDFIARNRQNEAFLRNIIGVSSVAYFSLRNILETE
                     NENGITGRMRGKTVKNNKGEEKYVSGEVDKIYNENKQNEVKENLKMFYSYDFNMDNKNE
                     IEDFFANIDEAISSIRHGIVHFNLELEGKDIFAFKNIAPSEISKKMFQNEINEKKLKLK
                     IFKQLNSANVFNYYEKDVIIKYLKNTKFNFVNKNIPFVPSFTKLYNKIEDLRNTLKFFW
                     SVPKDKEEKDAQIYLLKNIYYGEFLNKFVKNSKVFFKITNEVIKINKQRNQKTGHYKYQ
                     KFENIEKTVPVEYLAIIQSREMINNQDKEEKNTYIDFIQQIFLKGFIDYLNKNNLKYIE
                     SNNNNDNNDIFSKIKIKKDNKEKYDKILKNYEKHNRNKEIPHEINEFVREIKLGKILKY
                     TENLNMFYLILKLLNHKELTNLKGSLEKYQSANKEETFSDELELINLLNLDNNRVTEDF
                     ELEANEIGKFLDFNENKIKDRKELKKFDTNKIYFDGENIIKHRAFYNIKKYGMLNLLEK
                     IADKAKYKISLKELKEYSNKKNEIEKNYTMQQNLHRKYARPKKDEKFNDEDYKEYEKAI
                     GNIQKYTHLKNKVEFNELNLLQGLLLKILHRLVGYTSIWERDLRFRLKGEFPENHYIEE
                     IFNFDNSKNVKYKSGQIVEKYINFYKELYKDNVEKRSIYSDKKVKKLKQEKKDLYIRNY
                     IAHFNYIPHAEISLLEVLENLRKLLSYDRKLKNAIMKSIVDILKEYGFVATFKIGADKK
                     IEIQTLESEKIVHLKNLKKKKLMTDRNSEELCELVKVMFEYKALE"
     terminator      5775..6023
                     /label=HSP terminator
                     /note="efficient transcription terminator from the 
                     Arabidopsis thaliana heat shock protein 18.2 gene (Nagaya 
                     et al., 2010)"
     promoter        6097..6201
                     /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 ggcgcgccgt cgaggtcatt catatgcttg
      901 agaagagagt cgggatagtc caaaataaaa caaaggtaag attacctggt caaaagtgaa
      961 aacatcagtt aaaaggtggt ataaagtaaa atatcggtaa taaaaggtgg cccaaagtga
     1021 aatttactct tttctactat tataaaaatt gaggatgttt ttgtcggtac tttgatacgt
     1081 catttttgta tgaattggtt tttaagttta ttcgcttttg gaaatgcata tctgtatttg
     1141 agtcgggttt taagttcgtt tgcttttgta aatacagagg gatttgtata agaaatatct
     1201 ttaaaaaaac ccatatgcta atttgacata atttttgaga aaaatatata ttcaggcgaa
     1261 ttctcacaat gaacaataat aagattaaaa tagctttccc ccgttgcagc gcatgggtat
     1321 tttttctagt aaaaataaaa gataaactta gactcaaaac atttacaaaa acaaccccta
     1381 aagttcctaa agcccaaagt gctatccacg atccatagca agcccagccc aacccaaccc
     1441 aacccaaccc accccagtcc agccaactgg acaatagtct ccacaccccc ccactatcac
     1501 cgtgagttgt ccgcacgcac cgcacgtctc gcagccaaaa aaaaaaaaag aaagaaaaaa
     1561 aagaaaaaga aaaaacagca gggggtccgg gtcgtggggg ccggaaacgc gaggaggatc
     1621 gcgagccagc gacgaggccg gccctccctc cgcttccaaa gaaacgcccc ccatcgccac
     1681 tatatacata cccccccctc tcctcccatc cccccaaccc taccaccacc accaccacca
     1741 cctccacctc ctcccccctc gctgccggac gacgagctcc tcccccctcc ccctccgccg
     1801 ccgccgcgcc ggtaaccacc ccgcccctct cctctttctt tctccgtttt ttttttccgt
     1861 ctcggtctcg atctttggcc ttggtagttt gggtgggcga gaggcggctt cgtgcgcgcc
     1921 cagatcggtg cgcgggaggg gcgggatctc gcggctgggg ctctcgccgg cgtggatccg
     1981 gcccggatct cgcggggaat ggggctctcg gatgtagatc tgcgatccgc cgttgttggg
     2041 ggagatgatg gggggtttaa aatttccgcc atgctaaaca agatcaggaa gaggggaaaa
     2101 gggcactatg gtttatattt ttatatattt ctgctgcttc gtcaggctta gatgtgctag
     2161 atctttcttt cttctttttg tgggtagaat ttgaatccct cagcattgtt catcggtagt
     2221 ttttcttttc atgatttgtg acaaatgcag cctcgtgcgg agcttttttg taggtagacg
     2281 ataagcttcc tgcatcgaca tgggatccat gaaagtgacc aaggtcgacg gcatcagcca
     2341 caagaagtac atcgaagagg gcaagctcgt gaagtccacc agcgaggaaa accggaccag
     2401 cgagagactg agcgagctgc tgagcatccg gctggacatc tacatcaaga accccgacaa
     2461 cgcctccgag gaagagaacc ggatcagaag agagaacctg aagaagttct ttagcaacaa
     2521 ggtgctgcac ctgaaggaca gcgtgctgta tctgaagaac cggaaagaaa agaacgccgt
     2581 gcaggacaag aactatagcg aagaggacat cagcgagtac gacctgaaaa acaagaacag
     2641 cttctccgtg ctgaagaaga tcctgctgaa cgaggacgtg aactctgagg aactggaaat
     2701 ctttcggaag gacgtggaag ccaagctgaa caagatcaac agcctgaagt acagcttcga
     2761 agagaacaag gccaactacc agaagatcaa cgagaacaac gtggaaaaag tgggcggcaa
     2821 gagcaagcgg aacatcatct acgactacta cagagagagc gccaagcgca acgactacat
     2881 caacaacgtg caggaagcct tcgacaagct gtataagaaa gaggatatcg agaaactgtt
     2941 tttcctgatc gagaacagca agaagcacga gaagtacaag atccgcgagt actatcacaa
     3001 gatcatcggc cggaagaacg acaaagagaa cttcgccaag attatctacg aagagatcca
     3061 gaacgtgaac aacatcaaag agctgattga gaagatcccc gacatgtctg agctgaagaa
     3121 aagccaggtg ttctacaagt actacctgga caaagaggaa ctgaacgaca agaatattaa
     3181 gtacgccttc tgccacttcg tggaaatcga gatgtcccag ctgctgaaaa actacgtgta
     3241 caagcggctg agcaacatca gcaacgataa gatcaagcgg atcttcgagt accagaatct
     3301 gaaaaagctg atcgaaaaca aactgctgaa caagctggac acctacgtgc ggaactgcgg
     3361 caagtacaac tactatctgc aagtgggcga gatcgccacc tccgacttta tcgcccggaa
     3421 ccggcagaac gaggccttcc tgagaaacat catcggcgtg tccagcgtgg cctacttcag
     3481 cctgaggaac atcctggaaa ccgagaacga gaacggtatc accggccgga tgcggggcaa
     3541 gaccgtgaag aacaacaagg gcgaagagaa atacgtgtcc ggcgaggtgg acaagatcta
     3601 caatgagaac aagcagaacg aagtgaaaga aaatctgaag atgttctaca gctacgactt
     3661 caacatggac aacaagaacg agatcgagga cttcttcgcc aacatcgacg aggccatcag
     3721 cagcatcaga cacggcatcg tgcacttcaa cctggaactg gaaggcaagg acatcttcgc
     3781 cttcaagaat atcgccccca gcgagatctc caagaagatg tttcagaacg aaatcaacga
     3841 aaagaagctg aagctgaaaa tcttcaagca gctgaacagc gccaacgtgt tcaactacta
     3901 cgagaaggat gtgatcatca agtacctgaa gaataccaag ttcaacttcg tgaacaaaaa
     3961 catccccttc gtgcccagct tcaccaagct gtacaacaag attgaggacc tgcggaatac
     4021 cctgaagttt ttttggagcg tgcccaagga caaagaagag aaggacgccc agatctacct
     4081 gctgaagaat atctactacg gcgagttcct gaacaagttc gtgaaaaact ccaaggtgtt
     4141 ctttaagatc accaatgaag tgatcaagat taacaagcag cggaaccaga aaaccggcca
     4201 ctacaagtat cagaagttcg agaacatcga gaaaaccgtg cccgtggaat acctggccat
     4261 catccagagc agagagatga tcaacaacca ggacaaagag gaaaagaata cctacatcga
     4321 ctttattcag cagattttcc tgaagggctt catcgactac ctgaacaaga acaatctgaa
     4381 gtatatcgag agcaacaaca acaatgacaa caacgacatc ttctccaaga tcaagatcaa
     4441 aaaggataac aaagagaagt acgacaagat cctgaagaac tatgagaagc acaatcggaa
     4501 caaagaaatc cctcacgaga tcaatgagtt cgtgcgcgag atcaagctgg ggaagattct
     4561 gaagtacacc gagaatctga acatgtttta cctgatcctg aagctgctga accacaaaga
     4621 gctgaccaac ctgaagggca gcctggaaaa gtaccagtcc gccaacaaag aagaaacctt
     4681 cagcgacgag ttggaactga tcaacctgct gaacctggac aacaacagag tgaccgagga
     4741 cttcgagctg gaagccaacg agatcggcaa gttcctggac ttcaacgaaa acaaaatcaa
     4801 ggaccggaaa gagctgaaaa agttcgacac caacaagatc tatttcgacg gcgagaacat
     4861 catcaagcac cgggccttct acaatatcaa gaaatacggc atgctgaatc tgctggaaaa
     4921 gatcgccgat aaggccaagt ataagatcag cctgaaagaa ctgaaagagt acagcaacaa
     4981 gaagaatgag attgaaaaga actacaccat gcagcagaac ctgcaccgga agtacgccag
     5041 acccaagaag gacgaaaagt tcaacgacga ggactacaaa gagtatgaga aggccatcgg
     5101 caacatccag aagtacaccc acctgaagaa caaggtggaa ttcaatgagc tgaacctgct
     5161 gcagggcctg ctgctgaaga tcctgcaccg gctcgtgggc tacaccagca tctgggagcg
     5221 ggacctgaga ttccggctga agggcgagtt tcccgagaac cactacatcg aggaaatttt
     5281 caatttcgac aactccaaga atgtgaagta caaaagcggc cagatcgtgg aaaagtatat
     5341 caacttctac aaagaactgt acaaggacaa tgtggaaaag cggagcatct actccgacaa
     5401 gaaagtgaag aaactgaagc aggaaaaaaa ggacctgtac atccggaact acattgccca
     5461 cttcaactac atcccccacg ccgagattag cctgctggaa gtgctggaaa acctgcggaa
     5521 gctgctgtcc tacgaccgga agctgaagaa cgccatcatg aagtccatcg tggacattct
     5581 gaaagaatac ggcttcgtgg ccaccttcaa gatcggcgct gacaagaaga tcgaaatcca
     5641 gaccctggaa tcagagaaga tcgtgcacct gaagaatctg aagaaaaaga aactgatgac
     5701 cgaccggaac agcgaggaac tgtgcgaact cgtgaaagtc atgttcgagt acaaggccct
     5761 ggaatgactc gagatatgaa gatgaagatg aaatatttgg tgtgtcaaat aaaaagcttg
     5821 tgtgcttaag tttgtgtttt tttcttggct tgttgtgtta tgaatttgtg gctttttcta
     5881 atattaaatg aatgtaagat cacattataa tgaataaaca aatgtttcta taatccattg
     5941 tgaatgtttt gttggatctc ttctgcagca tataactact gtatgtgcta tggtatggac
     6001 tatggaatat gattaaagat aaggagctcc ggtgacggac gtgggcaggt gtgatgaaaa
     6061 gccttagggt caggtggcac ttttcgggga aatgtgcgat gaacccctat ttgtttattt
     6121 ttctaaatac attcaaatat gtatccgcta atgagacaat aaccctgata aatgcttcaa
     6181 ttatattgaa aaaggaagag tatgagtatc caacatttcc gtgtcgccct tattcccttt
     6241 tttgcggcct tctgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat
     6301 gctgaagatc agttgggtgc ccgagtgggt tacattgaac tggatctcaa cagcggcaaa
     6361 atcttagaga gttttcgccc cgaagaacgc ttcccaatga tgagcacttt caaagttctg
     6421 ctatgtggcg cggtgttatc ccgtattgat gctgggcaag agcaactcgg tcgccgcata
     6481 cactattctc agaatgactt ggttgagtat tcaccagtca cagaaaagca tcttacggat
     6541 ggcatgacag taagagaatt atgtagtgct gccataacca tgagtgataa cactgcggcc
     6601 aacttacttc tgacaacgat tggaggaccg aaggagctaa ccgctttttt gcacaacatg
     6661 ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac
     6721 gacgagcgtg acaccacgat gcctgtagca atggcaacaa ctctccgcaa actattaact
     6781 ggcgaactac ttactcttgc ttcacgccag caactcattg actggatgga ggcggacaaa
     6841 gttgcaggac cacttctgcg ctcggcactt ccggctggct ggtttattgc tgataaatct
     6901 ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc
     6961 tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga
     7021 cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga cgaagggaaa
     7081 taaatagtag cccgcctgat gtgcgggctt ttttttggtg gtggttacca gtgtcgctta
     7141 gctaccgtag gtcctcttag gcggtccgat tagcgacata gtcatcaat
//
