LOCUS       sequence-210897-        6657 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  35SPromoter-LwaCas13a-HSPTerminator.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6657)
  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 6657)
  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: pJJB296
FEATURES             Location/Qualifiers
     source          1..6657
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      119..707
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     608..627
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     861..878
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        1405..1750
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     1693..1713
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     CDS             1775..5230
                     /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      5240..5488
                     /label=HSP terminator
                     /note="efficient transcription terminator from the 
                     Arabidopsis thaliana heat shock protein 18.2 gene (Nagaya 
                     et al., 2010)"
     promoter        5565..5669
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gtcctcttag gcggtccgat tagcgacata gtcatcaatg ccttgacggc aaaatccctt
       61 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
      121 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
      181 cggtggtttg tttgccggat caagagctac taactctttt tccgaaggta actggcttca
      241 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
      301 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
      361 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
      421 cgcagcggtc gggctgaacg gggggttcgt gctcacagcc cagcttggag cgaacgacct
      481 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
      541 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
      601 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
      661 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
      721 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgttc tttcctgcgt
      781 tatcccctga ttctgtggat aaccgtatta ccgcctttga gtgagctgat accgctcgcc
      841 gcagccgaac gaccgagcgc agcgagtcag tgagcgagga agcggaagag cgcccacctg
      901 ctcatgatcg gcgcgccaga tttgcctttt caatttcaga aagaatgcta acccacagat
      961 ggttagagag gcttacgcag caggtatcat caagacgatc tacccgagca ataatctcca
     1021 ggaaatcaaa taccttccca agaaggttaa agatgcagtc aaaagattca ggactaactg
     1081 catcaagaac acagagaaag atatatttct caagatcaga agtactattc cagtatggac
     1141 gattcaaggc ttgcttcaca aaccaaggca agtaatagag attggagtct ctaaaaaggt
     1201 agttcccact gaatcaaagg ccatggagtc aaagattcaa atagaggacc taacagaact
     1261 cgccgtaaag actggcgaac agttcataca gagtctctta cgactcaatg acaagaagaa
     1321 aatcttcgtc aacatggtgg agcacgacac acttgtctac tccaaaaata tcaaagatac
     1381 agtctcagaa gaccaaaggg caattgagac ttttcaacaa agggtaatat ccggaaacct
     1441 cctcggattc cattgcccag ctatctgtca ctttattgtg aagatagtgg aaaaggaagg
     1501 tggctcctac aaatgccatc attgcgataa aggaaaggcc atcgttgaag atgcctctgc
     1561 cgacagtggt cccaaagatg gacccccacc cacgaggagc atcgtggaaa aagaagacgt
     1621 tccaaccacg tcttcaaagc aagtggattg atgtgatatc tccactgacg taagggatga
     1681 cgcacaatcc cactatcctt cgcaagaccc ttcctctata taaggaagtt catttcattt
     1741 ggagagaaca cgggggactc ccgacatggg atccatgaaa gtgaccaagg tcgacggcat
     1801 cagccacaag aagtacatcg aagagggcaa gctcgtgaag tccaccagcg aggaaaaccg
     1861 gaccagcgag agactgagcg agctgctgag catccggctg gacatctaca tcaagaaccc
     1921 cgacaacgcc tccgaggaag agaaccggat cagaagagag aacctgaaga agttctttag
     1981 caacaaggtg ctgcacctga aggacagcgt gctgtatctg aagaaccgga aagaaaagaa
     2041 cgccgtgcag gacaagaact atagcgaaga ggacatcagc gagtacgacc tgaaaaacaa
     2101 gaacagcttc tccgtgctga agaagatcct gctgaacgag gacgtgaact ctgaggaact
     2161 ggaaatcttt cggaaggacg tggaagccaa gctgaacaag atcaacagcc tgaagtacag
     2221 cttcgaagag aacaaggcca actaccagaa gatcaacgag aacaacgtgg aaaaagtggg
     2281 cggcaagagc aagcggaaca tcatctacga ctactacaga gagagcgcca agcgcaacga
     2341 ctacatcaac aacgtgcagg aagccttcga caagctgtat aagaaagagg atatcgagaa
     2401 actgtttttc ctgatcgaga acagcaagaa gcacgagaag tacaagatcc gcgagtacta
     2461 tcacaagatc atcggccgga agaacgacaa agagaacttc gccaagatta tctacgaaga
     2521 gatccagaac gtgaacaaca tcaaagagct gattgagaag atccccgaca tgtctgagct
     2581 gaagaaaagc caggtgttct acaagtacta cctggacaaa gaggaactga acgacaagaa
     2641 tattaagtac gccttctgcc acttcgtgga aatcgagatg tcccagctgc tgaaaaacta
     2701 cgtgtacaag cggctgagca acatcagcaa cgataagatc aagcggatct tcgagtacca
     2761 gaatctgaaa aagctgatcg aaaacaaact gctgaacaag ctggacacct acgtgcggaa
     2821 ctgcggcaag tacaactact atctgcaagt gggcgagatc gccacctccg actttatcgc
     2881 ccggaaccgg cagaacgagg ccttcctgag aaacatcatc ggcgtgtcca gcgtggccta
     2941 cttcagcctg aggaacatcc tggaaaccga gaacgagaac ggtatcaccg gccggatgcg
     3001 gggcaagacc gtgaagaaca acaagggcga agagaaatac gtgtccggcg aggtggacaa
     3061 gatctacaat gagaacaagc agaacgaagt gaaagaaaat ctgaagatgt tctacagcta
     3121 cgacttcaac atggacaaca agaacgagat cgaggacttc ttcgccaaca tcgacgaggc
     3181 catcagcagc atcagacacg gcatcgtgca cttcaacctg gaactggaag gcaaggacat
     3241 cttcgccttc aagaatatcg cccccagcga gatctccaag aagatgtttc agaacgaaat
     3301 caacgaaaag aagctgaagc tgaaaatctt caagcagctg aacagcgcca acgtgttcaa
     3361 ctactacgag aaggatgtga tcatcaagta cctgaagaat accaagttca acttcgtgaa
     3421 caaaaacatc cccttcgtgc ccagcttcac caagctgtac aacaagattg aggacctgcg
     3481 gaataccctg aagttttttt ggagcgtgcc caaggacaaa gaagagaagg acgcccagat
     3541 ctacctgctg aagaatatct actacggcga gttcctgaac aagttcgtga aaaactccaa
     3601 ggtgttcttt aagatcacca atgaagtgat caagattaac aagcagcgga accagaaaac
     3661 cggccactac aagtatcaga agttcgagaa catcgagaaa accgtgcccg tggaatacct
     3721 ggccatcatc cagagcagag agatgatcaa caaccaggac aaagaggaaa agaataccta
     3781 catcgacttt attcagcaga ttttcctgaa gggcttcatc gactacctga acaagaacaa
     3841 tctgaagtat atcgagagca acaacaacaa tgacaacaac gacatcttct ccaagatcaa
     3901 gatcaaaaag gataacaaag agaagtacga caagatcctg aagaactatg agaagcacaa
     3961 tcggaacaaa gaaatccctc acgagatcaa tgagttcgtg cgcgagatca agctggggaa
     4021 gattctgaag tacaccgaga atctgaacat gttttacctg atcctgaagc tgctgaacca
     4081 caaagagctg accaacctga agggcagcct ggaaaagtac cagtccgcca acaaagaaga
     4141 aaccttcagc gacgagttgg aactgatcaa cctgctgaac ctggacaaca acagagtgac
     4201 cgaggacttc gagctggaag ccaacgagat cggcaagttc ctggacttca acgaaaacaa
     4261 aatcaaggac cggaaagagc tgaaaaagtt cgacaccaac aagatctatt tcgacggcga
     4321 gaacatcatc aagcaccggg ccttctacaa tatcaagaaa tacggcatgc tgaatctgct
     4381 ggaaaagatc gccgataagg ccaagtataa gatcagcctg aaagaactga aagagtacag
     4441 caacaagaag aatgagattg aaaagaacta caccatgcag cagaacctgc accggaagta
     4501 cgccagaccc aagaaggacg aaaagttcaa cgacgaggac tacaaagagt atgagaaggc
     4561 catcggcaac atccagaagt acacccacct gaagaacaag gtggaattca atgagctgaa
     4621 cctgctgcag ggcctgctgc tgaagatcct gcaccggctc gtgggctaca ccagcatctg
     4681 ggagcgggac ctgagattcc ggctgaaggg cgagtttccc gagaaccact acatcgagga
     4741 aattttcaat ttcgacaact ccaagaatgt gaagtacaaa agcggccaga tcgtggaaaa
     4801 gtatatcaac ttctacaaag aactgtacaa ggacaatgtg gaaaagcgga gcatctactc
     4861 cgacaagaaa gtgaagaaac tgaagcagga aaaaaaggac ctgtacatcc ggaactacat
     4921 tgcccacttc aactacatcc cccacgccga gattagcctg ctggaagtgc tggaaaacct
     4981 gcggaagctg ctgtcctacg accggaagct gaagaacgcc atcatgaagt ccatcgtgga
     5041 cattctgaaa gaatacggct tcgtggccac cttcaagatc ggcgctgaca agaagatcga
     5101 aatccagacc ctggaatcag agaagatcgt gcacctgaag aatctgaaga aaaagaaact
     5161 gatgaccgac cggaacagcg aggaactgtg cgaactcgtg aaagtcatgt tcgagtacaa
     5221 ggccctggaa tgacgacgat atgaagatga agatgaaata tttggtgtgt caaataaaaa
     5281 gcttgtgtgc ttaagtttgt gtttttttct tggcttgttg tgttatgaat ttgtggcttt
     5341 ttctaatatt aaatgaatgt aagatcacat tataatgaat aaacaaatgt ttctataatc
     5401 cattgtgaat gttttgttgg atctcttctg cagcatataa ctactgtatg tgctatggta
     5461 tggactatgg aatatgatta aagataagcc agagctctgg tgacggacgt gggcaggtgt
     5521 gatgaaaagc cttagggtca ggtggcactt ttcggggaaa tgtgcgatga acccctattt
     5581 gtttattttt ctaaatacat tcaaatatgt atccgctaat gagacaataa ccctgataaa
     5641 tgcttcaatt atattgaaaa aggaagagta tgagtatcca acatttccgt gtcgccctta
     5701 ttcccttttt tgcggccttc tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     5761 taaaagatgc tgaagatcag ttgggtgccc gagtgggtta cattgaactg gatctcaaca
     5821 gcggcaaaat cttagagagt tttcgccccg aagaacgctt cccaatgatg agcactttca
     5881 aagttctgct atgtggcgcg gtgttatccc gtattgatgc tgggcaagag caactcggtc
     5941 gccgcataca ctattctcag aatgacttgg ttgagtattc accagtcaca gaaaagcatc
     6001 ttacggatgg catgacagta agagaattat gtagtgctgc cataaccatg agtgataaca
     6061 ctgcggccaa cttacttctg acaacgattg gaggaccgaa ggagctaacc gcttttttgc
     6121 acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg aatgaagcca
     6181 taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaact ctccgcaaac
     6241 tattaactgg cgaactactt actcttgctt cacgccagca actcattgac tggatggagg
     6301 cggacaaagt tgcaggacca cttctgcgct cggcacttcc ggctggctgg tttattgctg
     6361 ataaatctgg agccggtgag cgtgggtctc gcggtatcat tgcagcactg gggccagatg
     6421 gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact atggatgaac
     6481 gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa ctgtcagacg
     6541 aagggaaata aatagtagcc cgcctgatgt gcgggctttt ttttggtggt ggttaccagt
     6601 gtcgcttagc taccgtaggt cctcttaggc ggtccgatta gcgacatagt catcaat
//
