LOCUS       sequence-330730-        7596 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of PmcCAST Cascade proteins. Two BsaI sites for
            spacer cloning..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7596)
  AUTHORS   Saito M, Ladha A, Strecker J, Faure G, Neumann E, Altae-Tran H, 
            Macrae RK, Zhang F
  TITLE     Dual modes of CRISPR-associated transposon homing.
  JOURNAL   Cell. 2021 Apr 29;184(9):2441-2453.e18. doi: 
            10.1016/j.cell.2021.03.006. Epub 2021 Mar 25.
  PUBMED    33770501
REFERENCE   2  (bases 1 to 7596)
  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"; doi: 
            "10.1016/j.cell.2021.03.006"; journalName: "Cell"; date: 
            "2021-04-29- 29"; volume: "184"; issue: "9"; pages: "2441-2453.e18"
COMMENT     Sequence Label: pMS24: pCascade(PmcCAST)_entry
FEATURES             Location/Qualifiers
     source          1..7596
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(334..1416)
                     /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     1378..1397
                     /label=LacI-R
                     /note="LacI, reverse primer"
     promoter        complement(1417..1494)
                     /gene="lacI"
                     /label=lacI promoter
     promoter        1540..1558
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1559..1583
                     /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)."
     promoter        1826..1844
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    1845..1869
                     /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)."
     primer_bind     complement(5754..5772)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      5768..5815
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(5986..6777)
                     /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(6778..6869)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cgaaggttaa gtcagttggg gaactgctta accttcgatc aaaccacctc cccaggtggt
       61 tttttcgttt acagggcaaa agattacgcg cagaaaaaaa ggatctcaag aagatccttt
      121 gatcttttct actgaaccgc tctagatttc agtgcaattt atctcttcaa atgtagcacc
      181 tgaagtcagc cccatacgat ataagttgta attctcatgt tagtcatgcc ccgcgcccac
      241 cggaaggagc tgactgggtt gaaggctctc aagggcatcg gtcgagatcc cggtgcctaa
      301 tgagtgagct aacttacatt aattgcgttg cgctcactgc ccgctttcca gtcgggaaac
      361 ctgtcgtgcc agctgcatta atgaatcggc caacgcgcgg ggagaggcgg tttgcgtatt
      421 gggcgccagg gtggtttttc ttttcaccag tgagacgggc aacagctgat tgcccttcac
      481 cgcctggccc tgagagagtt gcagcaagcg gtccacgctg gtttgcccca gcaggcgaaa
      541 atcctgtttg atggtggtta acggcgggat ataacatgag ctgtcttcgg tatcgtcgta
      601 tcccactacc gagatgtccg caccaacgcg cagcccggac tcggtaatgg cgcgcattgc
      661 gcccagcgcc atctgatcgt tggcaaccag catcgcagtg ggaacgatgc cctcattcag
      721 catttgcatg gtttgttgaa aaccggacat ggcactccag tcgccttccc gttccgctat
      781 cggctgaatt tgattgcgag tgagatattt atgccagcca gccagacgca gacgcgccga
      841 gacagaactt aatgggcccg ctaacagcgc gatttgctgg tgacccaatg cgaccagatg
      901 ctccacgccc agtcgcgtac cgtcttcatg ggagaaaata atactgttga tgggtgtctg
      961 gtcagagaca tcaagaaata acgccggaac attagtgcag gcagcttcca cagcaatggc
     1021 atcctggtca tccagcggat agttaatgat cagcccactg acgcgttgcg cgagaagatt
     1081 gtgcaccgcc gctttacagg cttcgacgcc gcttcgttct accatcgaca ccaccacgct
     1141 ggcacccagt tgatcggcgc gagatttaat cgccgcgaca atttgcgacg gcgcgtgcag
     1201 ggccagactg gaggtggcaa cgccaatcag caacgactgt ttgcccgcca gttgttgtgc
     1261 cacgcggttg ggaatgtaat tcagctccgc catcgccgct tccacttttt cccgcgtttt
     1321 cgcagaaacg tggctggcct ggttcaccac gcgggaaacg gtctgataag agacaccggc
     1381 atactctgcg acatcgtata acgttactgg tttcacattc accaccctga attgactctc
     1441 ttccgggcgc tatcatgcca taccgcgaaa ggttttgcgc cattcgatgg tgtccgggat
     1501 ctcgacgctc tcccttatgc gactcctgca ttaggaaatt aatacgactc actatagggg
     1561 aattgtgagc ggataacaat tcccctgtag aaataatttt gtttaacttt aataaggaga
     1621 tataccatgg tgagtcgtgg ctttatgccg ttaggcgttg ctcaaagaga cctctggtct
     1681 cagtgagtcg tggctttatg ccgttaggcg ttgctcaaga tccgaattcg agctcggcgc
     1741 gcctgcaggt cgacaagctt gcggccgcat aatgcttaag tcgaacagaa agtaatcgta
     1801 ttgtacacgg ccgcataatc gaaattaata cgactcacta taggggaatt gtgagcggat
     1861 aacaattccc catcttagta tattagttaa gtataagaag gagatataca tatgacagct
     1921 atattaattc aaagtgagga atatgtagac ttaacattta agctaagagg cgcaccgatt
     1981 cctcttgaca acggttatct tacctacgct gccttatccc gtatttgtcc cccactacat
     2041 gaactaaagt ctattgggat tcatccgatt gctggaatac cgaccaggaa caatttgctt
     2101 gagctaactg ctcaatctcg tctaaaaatt cggatttatc atcagcaaat tcctttaatt
     2161 tatccttatt tagcgggtca agcttttcat ataggtcaaa atttttatca actagatatc
     2221 cccgactaca aaccgttaat ttcttcagaa agtgtctatt ccagattagt gataattaaa
     2281 gggttccaag actctactaa ctttattgaa gctgtgcaac ggcaaatgga taatttagga
     2341 atacaaggaa aaattgaact tcttacccga caagatggaa cacctcaacg aagacaatta
     2401 accatcaata aagaagggaa acagtttaaa gttagaggat ttggcgtaaa aatcagtgaa
     2461 ctcaatcctg aagattcctt aaccttgcaa gaacagggaa ttggcgggaa acgaaagatg
     2521 atgtgtggaa tatttgtccc agcgactcgc agcaaggaag aagaggaaac ctgatgatag
     2581 tctctaccca acccaaaatt tccctatctc tccatgctgc ggatacgaca ataatgcacc
     2641 gcgtcggaat gaccggactt tacatgactt taaagcgatt ggaaaagcaa tatcccttat
     2701 ctcgtcaacg gggaggacat atatcatggt ttttgactgc tgatactatt gaattattct
     2761 gggaaggaag tgatttcatc gccttatctt ggttaattaa tgagtctttt caattagatg
     2821 atacaggttt aattcattta gtaggattag acaatgatcg aatagattta aggcagaaaa
     2881 tccatattca tgagggaata tgcggtgttt ttctacgtct taataagttt tatcaagcag
     2941 gagagataat taacactgaa ctaaggtttg aagaaaaaca agtcgagtat caatataaat
     3001 ccctaacttg gtacgcacat caaacctttg cagagaagtt atgcgaggca gatacccaac
     3061 aactcagaca tgattacatt caaataacca gttggttgta tttaggggga attgttcgcc
     3121 atgctcggac gcagaatact acaaaactag aagagaaacc tgaatacgct ttagcattat
     3181 tatttgtacc agtagtttgt cattattgct tacttcatat accatcagaa gacttaaaag
     3241 aaaggaaacc ccatcgctac ttggtggtga ttccagaaat aaaggatttt gaagacgctt
     3301 ctcaaaggag atggcgatta cagcaattag aaactaaaca gtttcatgtt agtagccttg
     3361 gggaagcagg attactttat tacagcttag acgatattca gcctgaagtg gcctactatc
     3421 aagcttgtca agtttggtta tatgaaaaaa caaataaagc ttctcgtcaa cggacattga
     3481 tgagtataga agaaattaaa attgataaga atattttaat cacttatcaa caggtgcaaa
     3541 aatactttaa aactaattat cagataatta aatataagca aatttttatt aaggtcaatc
     3601 cgattcgctc cctgattgct gataatttag tgaaaggaat ccattggtgg tctaattttt
     3661 gggaaaaatt ggtgatagag gattcaaaag aatatttatt taaccagtta ttttccaatc
     3721 gagaaggatt catcataatg gcagagaata gtgaagaaga caagcagtat cttattttta
     3781 ttaaggtatt tcagcaagct atgaagggca actttgctaa aatttatgcc aaaacagagg
     3841 aaggtaaaga tcctccaatt aagaagaaag ttgaacgatt aagggcagag ctaaactatt
     3901 gttatgatga attgtcgttt aaggaatatt tgtctgactt tttagtaaga gggggattaa
     3961 ataagtattt taatgagcat caggaagaga ttgcactact aattaaaaaa tcaccttggc
     4021 aagaaataag aatttggtca ttgttagcga tcgccagtta taaacccaaa gataaactca
     4081 ctaataggga tgatagttca ttgtcaaaca atcaaaaact agaagaagtg aatgatgact
     4141 cagaagaaga atgacagtaa tatcccaaat tattatcttt acggaacagt cctcactcgt
     4201 tatggtttag cttccttaaa tcatgacatt agacgaggaa ataagactat tctgcaaaaa
     4261 ggctattgga ataatggtaa aattcattct tttgtaggtt caagtgcaat tcgttgggcg
     4321 ttacgttttt atcttcaaaa gcaaggttat ttagttaatc gagtctggga tgaggaagaa
     4381 catattaatc ggttaacaag tgaagatttc gatccagaaa aattttatga tgatgatatt
     4441 tttgggtttg ctttattaga gtctgctgaa acagaggagg acacttcaac aacaaagaga
     4501 aaaaagaaac aaacaaagac cagcactcct aatcagcgaa tgggtgcttt agggatgaat
     4561 atggctgttt cgctaacgcc ttatgatggt gcagttaagt taggggcaaa aagtggcaga
     4621 gagaaggata gtacatctct ccattttaca gaatatcatg caactcgata tcaatattat
     4681 tttggaattg atgccactca tctcaaagat ttttcgcgga ttttacctat gatagatggg
     4741 attatgaatc tgcccaaggt aggagggagt agtaatattt ttaattatcc tttctgtccc
     4801 gatagtttgg tatttcaatg gacaaatcat tttgcttcat atatttccta ttgctttgaa
     4861 tattgcgatc ctaagagtaa ggaagcaaaa ctttcacaag agtttataga tgaagtggaa
     4921 tgcggacaga ttgaccctag taaattgtgg attgggggaa caattgttaa ggatttacag
     4981 caattggata attttgaaag ttcccctttg aacaaggcgc atatttatcg taatcgaaat
     5041 gaaatgattg aagctttgaa aacagtcatt aaaagagatt tgggcttgga agaatcaaaa
     5101 tgattgcacc attaattctc tatttggatg ttccatttac gacttttcgg gagtcccatg
     5161 caagggaaat gggaaaaacc tatcctgttc ctcctccagc gacagtgtat gggatgttgt
     5221 tgtctttggt aggagaaacg aatgtctatc gccactgtgg ggtagaattg gccattgcaa
     5281 tgttatctag ccccaaaaag tcgcgaattt tgcgtcaaat gagacggttt aagaatgctg
     5341 attttagcca tccagagaat gttattccct gttatcaaga aattttgtct aatctgaagt
     5401 gtttgatttg ggttcgttct gacgaggaaa agatacaacc aagtttaagg gagaggatac
     5461 agttagcgtt tgaccatcct gagctagtaa gacggtttgg ttgtttattt ttgggagaaa
     5521 gtgaccagtt gattaaaacg atcaaacttg caagggaaga ttatctagag ggagtgagac
     5581 agtgggcgat tagagataat cgaggcaggt taactttacc ctattgggtt gaccatgtgg
     5641 ggtcaagaaa tacccgattt ttgaggtatc ggattgagga aatggataga ttatcacctc
     5701 ctgatttagc ctggacaatg gttcaatctc cgatttgacc taggctgctg ccaccgctga
     5761 gcaataacta gcataacccc ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa
     5821 acctcaggca tttgagaagc acacggtcac actgcttccg gtagtcaata aaccggtaaa
     5881 ccagcaatag acataagcgg ctatttaacg accctgccct gaaccgacga ccgggtcatc
     5941 gtggccggat cttgcggccc ctcggcttga acgaattgtt agacattatt tgccgactac
     6001 cttggtgatc tcgcctttca cgtagtggac aaattcttcc aactgatctg cgcgcgaggc
     6061 caagcgatct tcttcttgtc caagataagc ctgtctagct tcaagtatga cgggctgata
     6121 ctgggccggc aggcgctcca ttgcccagtc ggcagcgaca tccttcggcg cgattttgcc
     6181 ggttactgcg ctgtaccaaa tgcgggacaa cgtaagcact acatttcgct catcgccagc
     6241 ccagtcgggc ggcgagttcc atagcgttaa ggtttcattt agcgcctcaa atagatcctg
     6301 ttcaggaacc ggatcaaaga gttcctccgc cgctggacct accaaggcaa cgctatgttc
     6361 tcttgctttt gtcagcaaga tagccagatc aatgtcgatc gtggctggct cgaagatacc
     6421 tgcaagaatg tcattgcgct gccattctcc aaattgcagt tcgcgcttag ctggataacg
     6481 ccacggaatg atgtcgtcgt gcacaacaat ggtgacttct acagcgcgga gaatctcgct
     6541 ctctccaggg gaagccgaag tttccaaaag gtcgttgatc aaagctcgcc gcgttgtttc
     6601 atcaagcctt acggtcaccg taaccagcaa atcaatatca ctgtgtggct tcaggccgcc
     6661 atccactgcg gagccgtaca aatgtacggc cagcaacgtc ggttcgagat ggcgctcgat
     6721 gacgccaact acctctgata gttgagtcga tacttcggcg atcaccgctt ccctcatact
     6781 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     6841 atttgaatgt atttagaaaa ataaacaaat agctagctca ctcggtcgct acgctccggg
     6901 cgtgagactg cggcgggcgc tgcggacaca tacaaagtta cccacagatt ccgtggataa
     6961 gcaggggact aacatgtgag gcaaaacagc agggccgcgc cggtggcgtt tttccatagg
     7021 ctccgccctc ctgccagagt tcacataaac agacgctttt ccggtgcatc tgtgggagcc
     7081 gtgaggctca accatgaatc tgacagtacg ggcgaaaccc gacaggactt aaagatcccc
     7141 accgtttccg gcgggtcgct ccctcttgcg ctctcctgtt ccgaccctgc cgtttaccgg
     7201 atacctgttc cgcctttctc ccttacggga agtgtggcgc tttctcatag ctcacacact
     7261 ggtatctcgg ctcggtgtag gtcgttcgct ccaagctggg ctgtaagcaa gaactccccg
     7321 ttcagcccga ctgctgcgcc ttatccggta actgttcact tgagtccaac ccggaaaagc
     7381 acggtaaaac gccactggca gcagccattg gtaactggga gttcgcagag gatttgttta
     7441 gctaaacacg cggttgctct tgaagtgtgc gccaaagtcc ggctacactg gaaggacaga
     7501 tttggttgct gtgctctgcg aaagccagtt accacggtta agcagttccc caactgactt
     7561 aaccttcgat caaaccacct ccccaggtgg tttttt
//
