LOCUS       sequence-177590-        6615 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the Cascade subunits Cse2, Cas7, Cas5, Cas6e and pre-crRNA
            containing a shorter version of the wt spacer (-3) truncated by 3 
            nucleotides at the leader-distal end..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6615)
  AUTHORS   Kuznedelov K, Mekler V, Lemak S, Tokmina-Lukaszewska M, Datsenko KA,
            Jain I, Savitskaya E, Mallon J, Shmakov S, Bothner B, Bailey S, 
            Yakunin AF, Severinov K, Semenova E
  TITLE     Altered stoichiometry Escherichia coli Cascade complexes with 
            shortened CRISPR RNA spacers are capable of interference and primed 
            adaptation.
  JOURNAL   Nucleic Acids Res. 2016 Dec 15;44(22):10849-10861. Epub 2016 Oct 13.
  PUBMED    27738137
REFERENCE   2  (bases 1 to 6615)
  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: "Nucleic 
            Acids Res."; date: "2016-12-15"; volume: "44(22)"; pages: 
            "10849-10861. Epub 2016 Oct 13"
COMMENT     Sequence Label: pCDF-casBCDE(-3)
FEATURES             Location/Qualifiers
     source          1..6615
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    7..23
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     CDS             80..103
                     /codon_start=1
                     /product="peptide that binds Strep-Tactin(R), an engineered
                     form of streptavidin"
                     /label=Strep-Tag II
                     /translation="WSHPQFEK"
     CDS             110..133
                     /codon_start=1
                     /product="recognition and cleavage site for human 
                     rhinovirus 3C and PreScission proteases"
                     /label=HRV 3C site
                     /translation="LEVLFQGP"
     primer_bind     complement(3217..3235)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3231..3278
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(3449..4240)
                     /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"
     promoter        complement(4241..4332)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(5393..6475)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=lacI
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     primer_bind     6437..6456
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6476..6553)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     join(6599..6615,1..3)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(6599..6615,1..2)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
ORIGIN
        1 gggaaattgt gagcggataa caattcccct gtagaaataa ttttgtttaa ctttaataag
       61 gagatatacc atggcaagct ggagccaccc gcagttcgaa aagggtgcac ttgaagtcct
      121 ctttcaggga cccgggtacc agatggctga tgaaattgat gcaatggctt tatatcgagc
      181 ctggcaacaa ctggataatg gatcatgtgc gcaaattaga cgtgtttcag aacctgatga
      241 attacgcgat atccctgcgt tttataggct ggtgcaacct tttggttggg aaaacccacg
      301 tcaccagcag gctcttttgc gcatggtgtt ttgcctgagc gcaggaaaga atgtcatccg
      361 acatcaggac aaaaaatcgg agcaaacaac aggtatctcg ttgggaagag ctttagccaa
      421 tagtggaaga attaacgagc gccgtatctt tcaattaatt cgggctgaca gaacagccga
      481 tatggtccag ttacgtcgat tacttactca cgccgaaccc gtacttgact ggccattaat
      541 ggccaggatg ttgacctggt ggggaaagcg cgaacgccag caacttctgg aagattttgt
      601 attgaccaca aacaaaaatg cgtaaggaaa cctttctatg tctaacttta tcaatattca
      661 tgttctgatc tctcacagcc cttcatgtct gaaccgcgac gatatgaaca tgcagaaaga
      721 cgctattttc ggcggcaaaa gacgagtaag aatttcaagt caaagcctta aacgtgcgat
      781 gcgtaaaagt ggttattacg cacaaaatat tggtgaatcc agtctcagaa ccattcatct
      841 tgcacaatta cgtgatgttc ttcggcaaaa acttggtgaa cgttttgacc aaaaaatcat
      901 cgataagaca ttagcgctgc tctccggtaa atcagttgat gaagccgaaa agatttctgc
      961 cgatgcggtt actccctggg ttgtgggaga aatagcctgg ttctgtgagc aggttgcaaa
     1021 agcagaggct gataatctgg atgataaaaa gctgctcaaa gttcttaagg aagatattgc
     1081 cgccatacgt gtgaatttac agcagggtgt tgatattgcg cttagtggaa gaatggcaac
     1141 cagcggcatg atgactgagt tgggaaaagt tgatggtgca atgtccattg cgcatgcgat
     1201 cactactcat caggttgatt ctgatattga ctggttcacc gctgtagatg atttacagga
     1261 acaaggttct gcacatctgg gaactcagga attttcatcg ggtgtttttt atcgttatgc
     1321 caacattaac ctcgctcaac ttcaggaaaa tttaggtggt gcctccaggg agcaggctct
     1381 ggaaattgca acccatgttg ttcatatgct ggcaacagag gtccctggag caaaacagcg
     1441 tacttatgcc gcttttaacc ctgcggatat ggtaatggtt aatttctccg atatgccact
     1501 ttctatggca aatgcttttg aaaaagcggt taaagcgaaa gatggctttt tgcaaccgtc
     1561 tatacaggcg tttaatcaat attgggatcg cgttgccaat ggatatggtc tgaacggagc
     1621 tgctgcgcaa ttcagcttat ctgatgtaga cccaattact gctcaagtta aacaaatgcc
     1681 tactttagaa cagttaaaat cctgggttcg taataatggc gaggcgtgaa catgagatct
     1741 tatttgatct tgcggcttgc tgggccaatg caagcctggg ggcagccgac ctttgaagga
     1801 acgcgaccta ccggaagatt tccgacccga agcgggttat tagggctact cggggcttgt
     1861 cttgggatcc aacgtgatga tacttcttca ttacaggcgt tatcagagag tgtgcaattt
     1921 gcagtgcgct gcgatgaact cattcttgac gatcgtcgtg tgtctgtaac ggggttgcgt
     1981 gattaccata cagtccttgg agcgcgagaa gattaccgtg gtttgaaaag tcatgaaacg
     2041 attcaaacat ggcgcgaata tttatgtgat gcctccttta ccgtcgctct ctggttaaca
     2101 ccccatgcaa cgatggttat ctcagaactt gaaaaagcag tattaaagcc tcggtataca
     2161 ccttacctgg ggcggagaag ttgcccacta acacacccgc tttttttggg gacatgtcag
     2221 gcatcggatc ctcagaaggc gctattaaat tatgagcccg ttggcggcga tatatatagt
     2281 gaggaatcag ttacagggca tcatttaaaa tttacggcgc gcgacgaacc gatgatcacc
     2341 ttgcctcgac aatttgcttc ccgagaatgg tatgtgatta aaggaggtat ggatgtatct
     2401 cagtaaagtc atcattgcca gggcctggag cagggatctt taccaacttc accagggatt
     2461 atggcattta tttccaaaca gaccggatgc tgctcgtgat tttctttttc atgttgagaa
     2521 gcgaaacaca ccagaaggct gtcatgtttt attgcagtca gcgcaaatgc ctgtttcaac
     2581 tgccgttgcg acagtcatta aaactaaaca ggttgaattt caacttcagg ttggtgttcc
     2641 actctatttt cggcttcggg caaatccgat caaaactatt ctcgacaatc aaaagcgcct
     2701 ggacagtaaa gggaatatta aacgctgtcg ggttccgtta ataaaagaag cagaacaaat
     2761 cgcgtggttg caacgtaaat tgggcaatgc ggcgcgcgtt gaagatgtgc atcccatatc
     2821 ggaacggcca cagtattttt ctggtgatgg taaaagtgga aagatccaaa cggtttgctt
     2881 tgaaggtgtg ctcaccatca acgacgcgcc agcgttaata gatcttgtac agcaaggtat
     2941 tgggccagct aaatcgatgg gatgtggctt gctatctttg gctccactgt gagcggccgc
     3001 actcgagatt tggatggttt aaggttggtg ggttgttttt atgggaaaaa atgctttaag
     3061 aacaaatgta tacttttaga gagttccccg cgccagcggg gataaaccgc tgtctttcgc
     3121 tgctgagggt gacgatccga gttccccgcg ccagcgggga taaaccgagc tcccattttc
     3181 aaacccatca agacgccttc gcctaggctg ctgccaccgc tgagcaataa ctagcataac
     3241 cccttggggc ctctaaacgg gtcttgaggg gttttttgct gaaacctcag gcatttgaga
     3301 agcacacggt cacactgctt ccggtagtca ataaaccggt aaaccagcaa tagacataag
     3361 cggctattta acgaccctgc cctgaaccga cgaccgggtc atcgtggccg gatcttgcgg
     3421 cccctcggct tgaacgaatt gttagacatt atttgccgac taccttggtg atctcgcctt
     3481 tcacgtagtg gacaaattct tccaactgat ctgcgcgcga ggccaagcga tcttcttctt
     3541 gtccaagata agcctgtcta gcttcaagta tgacgggctg atactgggcc ggcaggcgct
     3601 ccattgccca gtcggcagcg acatccttcg gcgcgatttt gccggttact gcgctgtacc
     3661 aaatgcggga caacgtaagc actacatttc gctcatcgcc agcccagtcg ggcggcgagt
     3721 tccatagcgt taaggtttca tttagcgcct caaatagatc ctgttcagga accggatcaa
     3781 agagttcctc cgccgctgga cctaccaagg caacgctatg ttctcttgct tttgtcagca
     3841 agatagccag atcaatgtcg atcgtggctg gctcgaagat acctgcaaga atgtcattgc
     3901 gctgccattc tccaaattgc agttcgcgct tagctggata acgccacgga atgatgtcgt
     3961 cgtgcacaac aatggtgact tctacagcgc ggagaatctc gctctctcca ggggaagccg
     4021 aagtttccaa aaggtcgttg atcaaagctc gccgcgttgt ttcatcaagc cttacggtca
     4081 ccgtaaccag caaatcaata tcactgtgtg gcttcaggcc gccatccact gcggagccgt
     4141 acaaatgtac ggccagcaac gtcggttcga gatggcgctc gatgacgcca actacctctg
     4201 atagttgagt cgatacttcg gcgatcaccg cttccctcat actcttcctt tttcaatatt
     4261 attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa tgtatttaga
     4321 aaaataaaca aatagctagc tcactcggtc gctacgctcc gggcgtgaga ctgcggcggg
     4381 cgctgcggac acatacaaag ttacccacag attccgtgga taagcagggg actaacatgt
     4441 gaggcaaaac agcagggccg cgccggtggc gtttttccat aggctccgcc ctcctgccag
     4501 agttcacata aacagacgct tttccggtgc atctgtggga gccgtgaggc tcaaccatga
     4561 atctgacagt acgggcgaaa cccgacagga cttaaagatc cccaccgttt ccggcgggtc
     4621 gctccctctt gcgctctcct gttccgaccc tgccgtttac cggatacctg ttccgccttt
     4681 ctcccttacg ggaagtgtgg cgctttctca tagctcacac actggtatct cggctcggtg
     4741 taggtcgttc gctccaagct gggctgtaag caagaactcc ccgttcagcc cgactgctgc
     4801 gccttatccg gtaactgttc acttgagtcc aacccggaaa agcacggtaa aacgccactg
     4861 gcagcagcca ttggtaactg ggagttcgca gaggatttgt ttagctaaac acgcggttgc
     4921 tcttgaagtg tgcgccaaag tccggctaca ctggaaggac agatttggtt gctgtgctct
     4981 gcgaaagcca gttaccacgg ttaagcagtt ccccaactga cttaaccttc gatcaaacca
     5041 cctccccagg tggttttttc gaaggttaag tcagttgggg aactgcttaa ccttcgatca
     5101 aaccacctcc ccaggtggtt ttttcgttta cagggcaaaa gattacgcgc agaaaaaaag
     5161 gatctcaaga agatcctttg atcttttcta ctgaaccgct ctagatttca gtgcaattta
     5221 tctcttcaaa tgtagcacct gaagtcagcc ccatacgata taagttgtaa ttctcatgtt
     5281 agtcatgccc cgcgcccacc ggaaggagct gactgggttg aaggctctca agggcatcgg
     5341 tcgagatccc ggtgcctaat gagtgagcta acttacatta attgcgttgc gctcactgcc
     5401 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     5461 gagaggcggt ttgcgtattg ggcgccaggg tggtttttct tttcaccagt gagacgggca
     5521 acagctgatt gcccttcacc gcctggccct gagagagttg cagcaagcgg tccacgctgg
     5581 tttgccccag caggcgaaaa tcctgtttga tggtggttaa cggcgggata taacatgagc
     5641 tgtcttcggt atcgtcgtat cccactaccg agatgtccgc accaacgcgc agcccggact
     5701 cggtaatggc gcgcattgcg cccagcgcca tctgatcgtt ggcaaccagc atcgcagtgg
     5761 gaacgatgcc ctcattcagc atttgcatgg tttgttgaaa accggacatg gcactccagt
     5821 cgccttcccg ttccgctatc ggctgaattt gattgcgagt gagatattta tgccagccag
     5881 ccagacgcag acgcgccgag acagaactta atgggcccgc taacagcgcg atttgctggt
     5941 gacccaatgc gaccagatgc tccacgccca gtcgcgtacc gtcttcatgg gagaaaataa
     6001 tactgttgat gggtgtctgg tcagagacat caagaaataa cgccggaaca ttagtgcagg
     6061 cagcttccac agcaatggca tcctggtcat ccagcggata gttaatgatc agcccactga
     6121 cgcgttgcgc gagaagattg tgcaccgccg ctttacaggc ttcgacgccg cttcgttcta
     6181 ccatcgacac caccacgctg gcacccagtt gatcggcgcg agatttaatc gccgcgacaa
     6241 tttgcgacgg cgcgtgcagg gccagactgg aggtggcaac gccaatcagc aacgactgtt
     6301 tgcccgccag ttgttgtgcc acgcggttgg gaatgtaatt cagctccgcc atcgccgctt
     6361 ccactttttc ccgcgttttc gcagaaacgt ggctggcctg gttcaccacg cgggaaacgg
     6421 tctgataaga gacaccggca tactctgcga catcgtataa cgttactggt ttcacattca
     6481 ccaccctgaa ttgactctct tccgggcgct atcatgccat accgcgaaag gttttgcgcc
     6541 attcgatggt gtccgggatc tcgacgctct cccttatgcg actcctgcat taggaaatta
     6601 atacgactca ctata
//
