LOCUS       sequence-177591-        6691 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses the Cascade subunits Cse2, Cas7, Cas5, Cas6e and pre-crRNA
            containing 33-nt spacer (referred to as 'wt') matching a protospacer
            in the M13 phage genome. .
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6691)
  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 6691)
  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(g8)
FEATURES             Location/Qualifiers
     source          1..6691
                     /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(3293..3311)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      3307..3354
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(3525..4316)
                     /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(4317..4408)
                     /gene="bla"
                     /label=AmpR promoter
     CDS             complement(5469..6551)
                     /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     6513..6532
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(6552..6629)
                     /gene="lacI"
                     /label=lacI promoter
     primer_bind     join(6675..6691,1..3)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        join(6675..6691,1..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 tcttttttac ctgtttgaaa acaaagaatt
     3061 agctgatctt taataataag gaaatgttac attaaggttg gtgggttgtt tttatgggaa
     3121 aaaatgcttt aagaacaaat gtatactttt agagagttcc ccgcgccagc ggggataaac
     3181 cgctgtcttt cgctgctgag ggtgacgatc ccgcgagttc cccgcgccag cggggataaa
     3241 ccgagctccc attttcaaac ccatcaagac gccttcgcct aggctgctgc caccgctgag
     3301 caataactag cataacccct tggggcctct aaacgggtct tgaggggttt tttgctgaaa
     3361 cctcaggcat ttgagaagca cacggtcaca ctgcttccgg tagtcaataa accggtaaac
     3421 cagcaataga cataagcggc tatttaacga ccctgccctg aaccgacgac cgggtcatcg
     3481 tggccggatc ttgcggcccc tcggcttgaa cgaattgtta gacattattt gccgactacc
     3541 ttggtgatct cgcctttcac gtagtggaca aattcttcca actgatctgc gcgcgaggcc
     3601 aagcgatctt cttcttgtcc aagataagcc tgtctagctt caagtatgac gggctgatac
     3661 tgggccggca ggcgctccat tgcccagtcg gcagcgacat ccttcggcgc gattttgccg
     3721 gttactgcgc tgtaccaaat gcgggacaac gtaagcacta catttcgctc atcgccagcc
     3781 cagtcgggcg gcgagttcca tagcgttaag gtttcattta gcgcctcaaa tagatcctgt
     3841 tcaggaaccg gatcaaagag ttcctccgcc gctggaccta ccaaggcaac gctatgttct
     3901 cttgcttttg tcagcaagat agccagatca atgtcgatcg tggctggctc gaagatacct
     3961 gcaagaatgt cattgcgctg ccattctcca aattgcagtt cgcgcttagc tggataacgc
     4021 cacggaatga tgtcgtcgtg cacaacaatg gtgacttcta cagcgcggag aatctcgctc
     4081 tctccagggg aagccgaagt ttccaaaagg tcgttgatca aagctcgccg cgttgtttca
     4141 tcaagcctta cggtcaccgt aaccagcaaa tcaatatcac tgtgtggctt caggccgcca
     4201 tccactgcgg agccgtacaa atgtacggcc agcaacgtcg gttcgagatg gcgctcgatg
     4261 acgccaacta cctctgatag ttgagtcgat acttcggcga tcaccgcttc cctcatactc
     4321 ttcctttttc aatattattg aagcatttat cagggttatt gtctcatgag cggatacata
     4381 tttgaatgta tttagaaaaa taaacaaata gctagctcac tcggtcgcta cgctccgggc
     4441 gtgagactgc ggcgggcgct gcggacacat acaaagttac ccacagattc cgtggataag
     4501 caggggacta acatgtgagg caaaacagca gggccgcgcc ggtggcgttt ttccataggc
     4561 tccgccctcc tgccagagtt cacataaaca gacgcttttc cggtgcatct gtgggagccg
     4621 tgaggctcaa ccatgaatct gacagtacgg gcgaaacccg acaggactta aagatcccca
     4681 ccgtttccgg cgggtcgctc cctcttgcgc tctcctgttc cgaccctgcc gtttaccgga
     4741 tacctgttcc gcctttctcc cttacgggaa gtgtggcgct ttctcatagc tcacacactg
     4801 gtatctcggc tcggtgtagg tcgttcgctc caagctgggc tgtaagcaag aactccccgt
     4861 tcagcccgac tgctgcgcct tatccggtaa ctgttcactt gagtccaacc cggaaaagca
     4921 cggtaaaacg ccactggcag cagccattgg taactgggag ttcgcagagg atttgtttag
     4981 ctaaacacgc ggttgctctt gaagtgtgcg ccaaagtccg gctacactgg aaggacagat
     5041 ttggttgctg tgctctgcga aagccagtta ccacggttaa gcagttcccc aactgactta
     5101 accttcgatc aaaccacctc cccaggtggt tttttcgaag gttaagtcag ttggggaact
     5161 gcttaacctt cgatcaaacc acctccccag gtggtttttt cgtttacagg gcaaaagatt
     5221 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctactga accgctctag
     5281 atttcagtgc aatttatctc ttcaaatgta gcacctgaag tcagccccat acgatataag
     5341 ttgtaattct catgttagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg
     5401 ctctcaaggg catcggtcga gatcccggtg cctaatgagt gagctaactt acattaattg
     5461 cgttgcgctc actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa
     5521 tcggccaacg cgcggggaga ggcggtttgc gtattgggcg ccagggtggt ttttcttttc
     5581 accagtgaga cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc
     5641 aagcggtcca cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttaacggc
     5701 gggatataac atgagctgtc ttcggtatcg tcgtatccca ctaccgagat gtccgcacca
     5761 acgcgcagcc cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca
     5821 accagcatcg cagtgggaac gatgccctca ttcagcattt gcatggtttg ttgaaaaccg
     5881 gacatggcac tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga
     5941 tatttatgcc agccagccag acgcagacgc gccgagacag aacttaatgg gcccgctaac
     6001 agcgcgattt gctggtgacc caatgcgacc agatgctcca cgcccagtcg cgtaccgtct
     6061 tcatgggaga aaataatact gttgatgggt gtctggtcag agacatcaag aaataacgcc
     6121 ggaacattag tgcaggcagc ttccacagca atggcatcct ggtcatccag cggatagtta
     6181 atgatcagcc cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg
     6241 acgccgcttc gttctaccat cgacaccacc acgctggcac ccagttgatc ggcgcgagat
     6301 ttaatcgccg cgacaatttg cgacggcgcg tgcagggcca gactggaggt ggcaacgcca
     6361 atcagcaacg actgtttgcc cgccagttgt tgtgccacgc ggttgggaat gtaattcagc
     6421 tccgccatcg ccgcttccac tttttcccgc gttttcgcag aaacgtggct ggcctggttc
     6481 accacgcggg aaacggtctg ataagagaca ccggcatact ctgcgacatc gtataacgtt
     6541 actggtttca cattcaccac cctgaattga ctctcttccg ggcgctatca tgccataccg
     6601 cgaaaggttt tgcgccattc gatggtgtcc gggatctcga cgctctccct tatgcgactc
     6661 ctgcattagg aaattaatac gactcactat a
//
