LOCUS       Exported                7582 bp ds-DNA    circular SYN 27-JUL-2024
DEFINITION  Inducible expression of PmcCAST proteins with crRNA targeting 
            PmcPSP1..
ACCESSION   .
VERSION     .
KEYWORDS    pPmcCascade
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7582)
  TITLE     Dual modes of CRISPR-associated transposon homing
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7582)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Saturday, Jul 27, 2024 from SnapGene 2.3.2
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7582
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     promoter        107..125
                     /note="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    126..150
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="lac operator"
                     /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             193..855
                     /codon_start=1
                     /note="Cas6"
                     /translation="MTAILIQSEEYVDLTFKLRGAPIPLDNGYLTYAALSRICPPLHEL
                     KSIGIHPIAGIPTRNNLLELTAQSRLKIRIYHQQIPLIYPYLAGQAFHIGQNFYQLDIP
                     DYKPLISSESVYSRLVIIKGFQDSTNFIEAVQRQMDNLGIQGKIELLTRQDGTPQRRQL
                     TINKEGKQFKVRGFGVKISELNPEDSLTLQEQGIGGKRKMMCGIFVPATRSKEEEET"
     CDS             855..2453
                     /codon_start=1
                     /note="Cas8"
                     /translation="MHHHHHHIVSTQPKISLSLHAADTTIMHRVGMTGLYMTLKRLEKQ
                     YPLSRQRGGHISWFLTADTIELFWEGSDFIALSWLINESFQLDDTGLIHLVGLDNDRID
                     LRQKIHIHEGICGVFLRLNKFYQAGEIINTELRFEEKQVEYQYKSLTWYAHQTFAEKLC
                     EADTQQLRHDYIQITSWLYLGGIVRHARTQNTTKLEEKPEYALALLFVPVVCHYCLLHI
                     PSEDLKERKPHRYLVVIPEIKDFEDASQRRWRLQQLETKQFHVSSLGEAGLLYYSLDDI
                     QPEVAYYQACQVWLYEKTNKASRQRTLMSIEEIKIDKNILITYQQVQKYFKTNYQIIKY
                     KQIFIKVNPIRSLIADNLVKGIHWWSNFWEKLVIEDSKEYLFNQLFSNREGFIIMAENS
                     EEDKQYLIFIKVFQQAMKGNFAKIYAKTEEGKDPPIKKKVERLRAELNYCYDELSFKEY
                     LSDFLVRGGLNKYFNEHQEEIALLIKKSPWQEIRIWSLLAIASYKPKDKLTNRDDSSLS
                     NNQKLEEVNDDSEEE"
     misc_feature    858..875
                     /note="His tag"
     misc_feature    2022..2030
                     /note="IRBS"
     misc_feature    2037..2390
                     /note="Cas11"
     misc_feature    2037..2039
                     /note="start codon"
     CDS             2431..3402
                     /codon_start=1
                     /note="Cas7"
                     /translation="MMTQKKNDSNIPNYYLYGTVLTRYGLASLNHDIRRGNKTILQKGY
                     WNNGKIHSFVGSSAIRWALRFYLQKQGYLVNRVWDEEEHINRLTSEDFDPEKFYDDDIF
                     GFALLESAETEEDTSTTKRKKKQTKTSTPNQRMGALGMNMAVSLTPYDGAVKLGAKSGR
                     EKDSTSLHFTEYHATRYQYYFGIDATHLKDFSRILPMIDGIMNLPKVGGSSNIFNYPFC
                     PDSLVFQWTNHFASYISYCFEYCDPKSKEAKLSQEFIDEVECGQIDPSKLWIGGTIVKD
                     LQQLDNFESSPLNKAHIYRNRNEMIEALKTVIKRDLGLEESK"
     CDS             3399..4037
                     /codon_start=1
                     /note="Cas5"
                     /translation="MIAPLILYLDVPFTTFRESHAREMGKTYPVPPPATVYGMLLSLVG
                     ETNVYRHCGVELAIAMLSSPKKSRILRQMRRFKNADFSHPENVIPCYQEILSNLKCLIW
                     VRSDEEKIQPSLRERIQLAFDHPELVRRFGCLFLGESDQLIKTIKLAREDYLEGVRQWA
                     IRDNRGRLTLPYWVDHVGSRNTRFLRYRIEEMDRLSPPDLAWTMVQSPI"
     terminator      4067..4114
                     /note="T7 terminator"
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(4235..5026)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /note="SmR"
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     promoter        complement(5027..5118)
                     /gene="bla"
                     /note="AmpR promoter"
     CDS             complement(6179..7261)
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /note="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"
     promoter        complement(7262..7339)
                     /gene="lacI"
                     /note="lacI promoter"
                     /note="
                     "
     promoter        7385..7403
                     /note="T7 promoter"
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    7404..7428
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="lac operator"
                     /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)."
     misc_feature    7475..7510
                     /note="DR"
     misc_feature    7503..7573
                     /note="mature crRNA"
     misc_feature    7511..7545
                     /note="PmcPSP1"
     misc_feature    7546..7581
                     /note="DR"
ORIGIN
        1 atccgaattc gagctcggcg cgcctgcagg tcgacaagct tgcggccgca taatgcttaa
       61 gtcgaacaga aagtaatcgt attgtacacg gccgcataat cgaaattaat acgactcact
      121 ataggggaat tgtgagcgga taacaattcc ccatcttagt atattagtta agtataagaa
      181 ggagatatac atatgacagc tatattaatt caaagtgagg aatatgtaga cttaacattt
      241 aagctaagag gcgcaccgat tcctcttgac aacggttatc ttacctacgc tgccttatcc
      301 cgtatttgtc ccccactaca tgaactaaag tctattggga ttcatccgat tgctggaata
      361 ccgaccagga acaatttgct tgagctaact gctcaatctc gtctaaaaat tcggatttat
      421 catcagcaaa ttcctttaat ttatccttat ttagcgggtc aagcttttca tataggtcaa
      481 aatttttatc aactagatat ccccgactac aaaccgttaa tttcttcaga aagtgtctat
      541 tccagattag tgataattaa agggttccaa gactctacta actttattga agctgtgcaa
      601 cggcaaatgg ataatttagg aatacaagga aaaattgaac ttcttacccg acaagatgga
      661 acacctcaac gaagacaatt aaccatcaat aaagaaggga aacagtttaa agttagagga
      721 tttggcgtaa aaatcagtga actcaatcct gaagattcct taaccttgca agaacaggga
      781 attggcggga aacgaaagat gatgtgtgga atatttgtcc cagcgactcg cagcaaggaa
      841 gaagaggaaa cctgatgcat catcatcatc atcatatagt ctctacccaa cccaaaattt
      901 ccctatctct ccatgctgcg gatacgacaa taatgcaccg cgtcggaatg accggacttt
      961 acatgacttt aaagcgattg gaaaagcaat atcccttatc tcgtcaacgg ggaggacata
     1021 tatcatggtt tttgactgct gatactattg aattattctg ggaaggaagt gatttcatcg
     1081 ccttatcttg gttaattaat gagtcttttc aattagatga tacaggttta attcatttag
     1141 taggattaga caatgatcga atagatttaa ggcagaaaat ccatattcat gagggaatat
     1201 gcggtgtttt tctacgtctt aataagtttt atcaagcagg agagataatt aacactgaac
     1261 taaggtttga agaaaaacaa gtcgagtatc aatataaatc cctaacttgg tacgcacatc
     1321 aaacctttgc agagaagtta tgcgaggcag atacccaaca actcagacat gattacattc
     1381 aaataaccag ttggttgtat ttagggggaa ttgttcgcca tgctcggacg cagaatacta
     1441 caaaactaga agagaaacct gaatacgctt tagcattatt atttgtacca gtagtttgtc
     1501 attattgctt acttcatata ccatcagaag acttaaaaga aaggaaaccc catcgctact
     1561 tggtggtgat tccagaaata aaggattttg aagacgcttc tcaaaggaga tggcgattac
     1621 agcaattaga aactaaacag tttcatgtta gtagccttgg ggaagcagga ttactttatt
     1681 acagcttaga cgatattcag cctgaagtgg cctactatca agcttgtcaa gtttggttat
     1741 atgaaaaaac aaataaagct tctcgtcaac ggacattgat gagtatagaa gaaattaaaa
     1801 ttgataagaa tattttaatc acttatcaac aggtgcaaaa atactttaaa actaattatc
     1861 agataattaa atataagcaa atttttatta aggtcaatcc gattcgctcc ctgattgctg
     1921 ataatttagt gaaaggaatc cattggtggt ctaatttttg ggaaaaattg gtgatagagg
     1981 attcaaaaga atatttattt aaccagttat tttccaatcg agaaggattc atcataatgg
     2041 cagagaatag tgaagaagac aagcagtatc ttatttttat taaggtattt cagcaagcta
     2101 tgaagggcaa ctttgctaaa atttatgcca aaacagagga aggtaaagat cctccaatta
     2161 agaagaaagt tgaacgatta agggcagagc taaactattg ttatgatgaa ttgtcgttta
     2221 aggaatattt gtctgacttt ttagtaagag ggggattaaa taagtatttt aatgagcatc
     2281 aggaagagat tgcactacta attaaaaaat caccttggca agaaataaga atttggtcat
     2341 tgttagcgat cgccagttat aaacccaaag ataaactcac taatagggat gatagttcat
     2401 tgtcaaacaa tcaaaaacta gaagaagtga atgatgactc agaagaagaa tgacagtaat
     2461 atcccaaatt attatcttta cggaacagtc ctcactcgtt atggtttagc ttccttaaat
     2521 catgacatta gacgaggaaa taagactatt ctgcaaaaag gctattggaa taatggtaaa
     2581 attcattctt ttgtaggttc aagtgcaatt cgttgggcgt tacgttttta tcttcaaaag
     2641 caaggttatt tagttaatcg agtctgggat gaggaagaac atattaatcg gttaacaagt
     2701 gaagatttcg atccagaaaa attttatgat gatgatattt ttgggtttgc tttattagag
     2761 tctgctgaaa cagaggagga cacttcaaca acaaagagaa aaaagaaaca aacaaagacc
     2821 agcactccta atcagcgaat gggtgcttta gggatgaata tggctgtttc gctaacgcct
     2881 tatgatggtg cagttaagtt aggggcaaaa agtggcagag agaaggatag tacatctctc
     2941 cattttacag aatatcatgc aactcgatat caatattatt ttggaattga tgccactcat
     3001 ctcaaagatt tttcgcggat tttacctatg atagatggga ttatgaatct gcccaaggta
     3061 ggagggagta gtaatatttt taattatcct ttctgtcccg atagtttggt atttcaatgg
     3121 acaaatcatt ttgcttcata tatttcctat tgctttgaat attgcgatcc taagagtaag
     3181 gaagcaaaac tttcacaaga gtttatagat gaagtggaat gcggacagat tgaccctagt
     3241 aaattgtgga ttgggggaac aattgttaag gatttacagc aattggataa ttttgaaagt
     3301 tcccctttga acaaggcgca tatttatcgt aatcgaaatg aaatgattga agctttgaaa
     3361 acagtcatta aaagagattt gggcttggaa gaatcaaaat gattgcacca ttaattctct
     3421 atttggatgt tccatttacg acttttcggg agtcccatgc aagggaaatg ggaaaaacct
     3481 atcctgttcc tcctccagcg acagtgtatg ggatgttgtt gtctttggta ggagaaacga
     3541 atgtctatcg ccactgtggg gtagaattgg ccattgcaat gttatctagc cccaaaaagt
     3601 cgcgaatttt gcgtcaaatg agacggttta agaatgctga ttttagccat ccagagaatg
     3661 ttattccctg ttatcaagaa attttgtcta atctgaagtg tttgatttgg gttcgttctg
     3721 acgaggaaaa gatacaacca agtttaaggg agaggataca gttagcgttt gaccatcctg
     3781 agctagtaag acggtttggt tgtttatttt tgggagaaag tgaccagttg attaaaacga
     3841 tcaaacttgc aagggaagat tatctagagg gagtgagaca gtgggcgatt agagataatc
     3901 gaggcaggtt aactttaccc tattgggttg accatgtggg gtcaagaaat acccgatttt
     3961 tgaggtatcg gattgaggaa atggatagat tatcacctcc tgatttagcc tggacaatgg
     4021 ttcaatctcc gatttgacct aggctgctgc caccgctgag caataactag cataacccct
     4081 tggggcctct aaacgggtct tgaggggttt tttgctgaaa cctcaggcat ttgagaagca
     4141 cacggtcaca ctgcttccgg tagtgccctg aaccgacgac cgggtcatcg tggccggatc
     4201 ttgcggcccc tcggcttgaa cgaattgtta gacattattt gccgactacc ttggtgatct
     4261 cgcctttcac gtagtggaca aattcttcca actgatctgc gcgcgaggcc aagcgatctt
     4321 cttcttgtcc aagataagcc tgtctagctt caagtatgac gggctgatac tgggccggca
     4381 ggcgctccat tgcccagtcg gcagcgacat ccttcggcgc gattttgccg gttactgcgc
     4441 tgtaccaaat gcgggacaac gtaagcacta catttcgctc atcgccagcc cagtcgggcg
     4501 gcgagttcca tagcgttaag gtttcattta gcgcctcaaa tagatcctgt tcaggaaccg
     4561 gatcaaagag ttcctccgcc gctggaccta ccaaggcaac gctatgttct cttgcttttg
     4621 tcagcaagat agccagatca atgtcgatcg tggctggctc gaagatacct gcaagaatgt
     4681 cattgcgctg ccattctcca aattgcagtt cgcgcttagc tggataacgc cacggaatga
     4741 tgtcgtcgtg cacaacaatg gtgacttcta cagcgcggag aatctcgctc tctccagggg
     4801 aagccgaagt ttccaaaagg tcgttgatca aagctcgccg cgttgtttca tcaagcctta
     4861 cggtcaccgt aaccagcaaa tcaatatcac tgtgtggctt caggccgcca tccactgcgg
     4921 agccgtacaa atgtacggcc agcaacgtcg gttcgagatg gcgctcgatg acgccaacta
     4981 cctctgatag ttgagtcgat acttcggcga tcaccgcttc cctcatactc ttcctttttc
     5041 aatattattg aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta
     5101 tttagaaaaa taaacaaata gctagctcac tcggtcgcta cgctccgggc gtgagactgc
     5161 ggcgggcgct gcggacacat acaaagttac ccacagattc cgtggataag caggggacta
     5221 acatgtgagg caaaacagca gggccgcgcc ggtggcgttt ttccataggc tccgccctcc
     5281 tgccagagtt cacataaaca gacgcttttc cggtgcatct gtgggagccg tgaggctcaa
     5341 ccatgaatct gacagtacgg gcgaaacccg acaggactta aagatcccca ccgtttccgg
     5401 cgggtcgctc cctcttgcgc tctcctgttc cgaccctgcc gtttaccgga tacctgttcc
     5461 gcctttctcc cttacgggaa gtgtggcgct ttctcatagc tcacacactg gtatctcggc
     5521 tcggtgtagg tcgttcgctc caagctgggc tgtaagcaag aactccccgt tcagcccgac
     5581 tgctgcgcct tatccggtaa ctgttcactt gagtccaacc cggaaaagca cggtaaaacg
     5641 ccactggcag cagccattgg taactgggag ttcgcagagg atttgtttag ctaaacacgc
     5701 ggttgctctt gaagtgtgcg ccaaagtccg gctacactgg aaggacagat ttggttgctg
     5761 tgctctgcga aagccagtta ccacggttaa gcagttcccc aactgactta accttcgatc
     5821 aaaccacctc cccaggtggt tttttcgaag gttaagtcag ttggggaact gcttaacctt
     5881 cgatcaaacc acctccccag gtggtttttt cgtttacagg gcaaaagatt acgcgcagaa
     5941 aaaaaggatc tcaagaagat cctttgatct tttctactga accgctctag atttcagtgc
     6001 aatttatctc ttcaaatgta gcacctgaag tcagccccat acgatataag ttgtaattct
     6061 catgttagtc atgccccgcg cccaccggaa ggagctgact gggttgaagg ctctcaaggg
     6121 catcggtcga gatcccggtg cctaatgagt gagctaactt acattaattg cgttgcgctc
     6181 actgcccgct ttccagtcgg gaaacctgtc gtgccagctg cattaatgaa tcggccaacg
     6241 cgcggggaga ggcggtttgc gtattgggcg ccagggtggt ttttcttttc accagtgaga
     6301 cgggcaacag ctgattgccc ttcaccgcct ggccctgaga gagttgcagc aagcggtcca
     6361 cgctggtttg ccccagcagg cgaaaatcct gtttgatggt ggttaacggc gggatataac
     6421 atgagctgtc ttcggtatcg tcgtatccca ctaccgagat gtccgcacca acgcgcagcc
     6481 cggactcggt aatggcgcgc attgcgccca gcgccatctg atcgttggca accagcatcg
     6541 cagtgggaac gatgccctca ttcagcattt gcatggtttg ttgaaaaccg gacatggcac
     6601 tccagtcgcc ttcccgttcc gctatcggct gaatttgatt gcgagtgaga tatttatgcc
     6661 agccagccag acgcagacgc gccgagacag aacttaatgg gcccgctaac agcgcgattt
     6721 gctggtgacc caatgcgacc agatgctcca cgcccagtcg cgtaccgtct tcatgggaga
     6781 aaataatact gttgatgggt gtctggtcag agacatcaag aaataacgcc ggaacattag
     6841 tgcaggcagc ttccacagca atggcatcct ggtcatccag cggatagtta atgatcagcc
     6901 cactgacgcg ttgcgcgaga agattgtgca ccgccgcttt acaggcttcg acgccgcttc
     6961 gttctaccat cgacaccacc acgctggcac ccagttgatc ggcgcgagat ttaatcgccg
     7021 cgacaatttg cgacggcgcg tgcagggcca gactggaggt ggcaacgcca atcagcaacg
     7081 actgtttgcc cgccagttgt tgtgccacgc ggttgggaat gtaattcagc tccgccatcg
     7141 ccgcttccac tttttcccgc gttttcgcag aaacgtggct ggcctggttc accacgcggg
     7201 aaacggtctg ataagagaca ccggcatact ctgcgacatc gtataacgtt actggtttca
     7261 cattcaccac cctgaattga ctctcttccg ggcgctatca tgccataccg cgaaaggttt
     7321 tgcgccattc gatggtgtcc gggatctcga cgctctccct tatgcgactc ctgcattagg
     7381 aaattaatac gactcactat aggggaattg tgagcggata acaattcccc tgtagaaata
     7441 attttgttta actttaataa ggagatatac catggtgagt cgtggcttta tgccgttagg
     7501 cgttgctcaa gagaagtcat ttaataaggc cactgttaaa cgtaggtgag tcgtggcttt
     7561 atgccgttag gcgttgctca ag
//
