LOCUS       sequence-401151-        9521 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  type I-F Cascade over-expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9521)
  TITLE     Distinct horizontal transfer mechanisms for type I and type V 
            CRISPR-associated transposons
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 9521)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pTniQ-Cascade
FEATURES             Location/Qualifiers
     source          1..9521
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      172..760
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     661..680
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     914..931
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    946..1085
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(1011..1030)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1171..1193
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(1187..1378)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     primer_bind     2836..2855
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
     primer_bind     2939..2958
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        2939..2957
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             3046..3068
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             3088..3117
                     /codon_start=1
                     /product="10xHis affinity tag"
                     /label=10xHis
                     /translation="HHHHHHHHHH"
     CDS             3127..4227
                     /codon_start=1
                     /gene="malE (mutated)"
                     /product="maltose binding protein from E. coli"
                     /label=MBP
                     /note="This version of the gene does not encode a signal 
                     sequence, so MBP will remain in the cytosol."
                     /translation="MKIEEGKLVIWINGDKGYNGLAEVGKKFEKDTGIKVTVEHPDKLE
                     EKFPQVAATGDGPDIIFWAHDRFGGYAQSGLLAEITPDKAFQDKLYPFTWDAVRYNGKL
                     IAYPIAVEALSLIYNKDLLPNPPKTWEEIPALDKELKAKGKSALMFNLQEPYFTWPLIA
                     ADGGYAFKYENGKYDIKDVGVDNAGAKAGLTFLVDLIKNKHMNADTDYSIAEAAFNKGE
                     TAMTINGPWAWSNIDTSKVNYGVTVLPTFKGQPSKPFVGVLSAGINAASPNKELAKEFL
                     ENYLLTDEGLEAVNKDKPLGAVALKSYEEELAKDPRITATMENAQKGEIMPNIPQMSAF
                     WYAVRTAVINAASGRQTVDEALKDAQT"
     primer_bind     4201..4224
                     /label=MBP-F
                     /note="Maltose binding protein, forward primer"
     CDS             4282..4302
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     primer_bind     complement(7989..8007)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      8003..8050
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(join(8416..8421,8422..8438,8439..8444))
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(8471..8489)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        8557..8661
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(8662..8730,8731..9521,1)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     complement(8880..8899)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 actgtcagac caagtttact catatatact ttagattgat ttaaaacttc atttttaatt
       61 taaaaggatc taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga
      121 gttttcgttc cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc
      181 tttttttctg cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt
      241 ttgtttgccg gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc
      301 gcagatacca aatactgtcc ttctagtgta gccgtagtta ggccaccact tcaagaactc
      361 tgtagcaccg cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg
      421 cgataagtcg tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg
      481 gtcgggctga acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga
      541 actgagatac ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc
      601 ggacaggtat ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg
      661 gggaaacgcc tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg
      721 atttttgtga tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt
      781 tttacggttc ctggcctttt gctggccttt tgctcacatg ttctttcctg cgttatcccc
      841 tgattctgtg gataaccgta ttaccgcctt tgagtgagct gataccgctc gccgcagccg
      901 aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa gagcgcctga tgcggtattt
      961 tctccttacg catctgtgcg gtatttcaca ccgcaatggt gcactctcag tacaatctgc
     1021 tctgatgccg catagttaag ccagtataca ctccgctatc gctacgtgac tgggtcatgg
     1081 ctgcgccccg acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg
     1141 catccgctta cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac
     1201 cgtcatcacc gaaacgcgcg aggcagctgc ggtaaagctc atcagcgtgg tcgtgaagcg
     1261 attcacagat gtctgcctgt tcatccgcgt ccagctcgtt gagtttctcc agaagcgtta
     1321 atgtctggct tctgataaag cgggccatgt taagggcggt tttttcctgt ttggtcactg
     1381 atgcctccgt gtaaggggga tttctgttca tgggggtaat gataccgatg aaacgagaga
     1441 ggatgctcac gatacgggtt actgatgatg aacatgcccg gttactggaa cgttgtgagg
     1501 gtaaacaact ggcggtatgg atgcggcggg accagagaaa aatcactcag ggtcaatgcc
     1561 agcgcttcgt taatacagat gtaggtgttc cacagggtag ccagcagcat cctgcgatgc
     1621 agatccggaa cataatggtg cagggcgctg acttccgcgt ttccagactt tacgaaacac
     1681 ggaaaccgaa gaccattcat gttgttgctc aggtcgcaga cgttttgcag cagcagtcgc
     1741 ttcacgttcg ctcgcgtatc ggtgattcat tctgctaacc agtaaggcaa ccccgccagc
     1801 ctagccgggt cctcaacgac aggagcacga tcatgcgcac ccgtggccag gacccaacgc
     1861 tgcccgagat gcgccgcgtg cggctgctgg agatggcgga cgcgatggat atgttctgcc
     1921 aagggttggt ttgcgcattc acagttctcc gcaagaattg attggctcca attcttggag
     1981 tggtgaatcc gttagcgagg tgccgccggc ttccattcag gtcgaggtgg cccggctcca
     2041 tgcaccgcga cgcaacgcgg ggaggcagac aaggtatagg gcggcgccta caatccatgc
     2101 caacccgttc catgtgctcg ccgaggcggc ataaatcgcc gtgacgatca gcggtccaat
     2161 gatcgaagtt aggctggtaa gagccgcgag cgatccttga agctgtccct gatggtcgtc
     2221 atctacctgc ctggacagca tggcctgcaa cgcgggcatc ccgatgccgc cggaagcgag
     2281 aagaatcata atggggaagg ccatccagcc tcgcgtcgcg aacgccagca agacgtagcc
     2341 cagcgcgtcg gccgccatgc cggcgataat ggcctgcttc tcgccgaaac gtttggtggc
     2401 gggaccagtg acgaaggctt gagcgagggc gtgcaagatt ccgaataccg caagcgacag
     2461 gccgatcatc gtcgcgctcc agcgaaagcg gtcctcgccg aaaatgaccc agagcgctgc
     2521 cggcacctgt cctacgagtt gcatgataaa gaagacagtc ataagtgcgg cgacgatagt
     2581 catgccccgc gcccaccgga aggagctgac tgggttgaag gctctcaagg gcatcggtcg
     2641 acgctctccc ttatgcgact cctgcattag gaagcagccc agtagtaggt tgaggccgtt
     2701 gagcaccgcc gccgcaagga atggtgcatg caaggagatg gcgcccaaca gtcccccggc
     2761 cacggggcct gccaccatac ccacgccgaa acaagcgctc atgagcccga agtggcgagc
     2821 ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca gcaaccgcac ctgtggcgcc
     2881 ggtgatgccg gccacgatgc gtccggcgta gaggatcgag atctcgatcc cgcgaaatta
     2941 atacgactca ctatagggag accacaacgg tttccctcta gtgccggctc cggagagctc
     3001 tttaattaag cggccgccct gcaggactcg agttctagaa ataattttgt ttaactttaa
     3061 gaaggagata tacatatgaa atcttctcac catcaccatc accatcacca tcaccatggt
     3121 tcttctatga aaatcgaaga aggtaaactg gtaatctgga ttaacggcga taaaggctat
     3181 aacggtctcg ctgaagtcgg taagaaattc gagaaagata ccggaattaa agtcaccgtt
     3241 gagcatccgg ataaactgga agagaaattc ccacaggttg cggcaactgg cgatggccct
     3301 gacattatct tctgggcaca cgaccgcttt ggtggctacg ctcaatctgg cctgttggct
     3361 gaaatcaccc cggacaaagc gttccaggac aagctgtatc cgtttacctg ggatgccgta
     3421 cgttacaacg gcaagctgat tgcttacccg atcgctgttg aagcgttatc gctgatttat
     3481 aacaaagatc tgctgccgaa cccgccaaaa acctgggaag agatcccggc gctggataaa
     3541 gaactgaaag cgaaaggtaa gagcgcgctg atgttcaacc tgcaagaacc gtacttcacc
     3601 tggccgctga ttgctgctga cgggggttat gcgttcaagt atgaaaacgg caagtacgac
     3661 attaaagacg tgggcgtgga taacgctggc gcgaaagcgg gtctgacctt cctggttgac
     3721 ctgattaaaa acaaacacat gaatgcagac accgattact ccatcgcaga agctgccttt
     3781 aataaaggcg aaacagcgat gaccatcaac ggcccgtggg catggtccaa catcgacacc
     3841 agcaaagtga attatggtgt aacggtactg ccgaccttca agggtcaacc atccaaaccg
     3901 ttcgttggcg tgctgagcgc aggtattaac gccgccagtc cgaacaaaga gctggcaaaa
     3961 gagttcctcg aaaactatct gctgactgat gaaggtctgg aagcggttaa taaagacaaa
     4021 ccgctgggtg ccgtagcgct gaagtcttac gaggaagagt tggcgaaaga tccacgtatt
     4081 accgccacta tggaaaacgc ccagaaaggt gaaatcatgc cgaacatccc gcagatgtcc
     4141 gctttctggt atgccgtgcg tactgcggtg atcaacgccg ccagcggtcg tcagactgtc
     4201 gatgaagccc tgaaagacgc gcagactaat tcgagctcga acaacaacaa caataacaat
     4261 aacaacaacc tcgggatcga ggaaaacctg tacttccaat ccaatgcaat gcaaactctg
     4321 aaagaactaa tcgcatccaa tcctgacgac ttaacaactg aacttaagag agcatttcgt
     4381 ccactcacac cgcatattgc aattgatggt aatgaacttg acgcactgac gatattagtc
     4441 aatttaaccg ataagactga tgatcagaaa gacctgctcg atcgagccaa atgcaagcaa
     4501 aaacttcgag atgaaaaatg gtgggctagc tgcataaatt gcgttaatta cagacaaagc
     4561 cataacccaa aattcccgga tatacgttct gaaggcgtga ttcgaaccca agccctgggt
     4621 gaattaccga gtttcctact ctcgtcctca aaaatcccac cataccattg gtcatatagc
     4681 catgattcaa aatacgtcaa caaaagcgca ttcctcacca atgagttttg ttgggatggt
     4741 gagatctcat gtttgggtga gcttcttaaa gatgcagatc acccactttg gaatactttg
     4801 aaaaagttag gttgttctca aaaaacttgc aaagcaatgg caaaacaact agctgatatt
     4861 actctcacga ctatcaatgt cacactcgca ccaaattacc tgactcaaat atctctcccc
     4921 gatagtgata catcctacat ttcactctct cctgttgcat cgctatcgat gcaaagccac
     4981 tttcatcaga ggctccaaga tgaaaatcgg catagcgcga taacgcggtt tagccgaact
     5041 accaacatgg gggtcactgc gatgacatgt ggcggcgcat ttagaatgtt aaagtctggc
     5101 gctaagtttt ccagcccccc tcatcaccga ttaaacagta aacgaagttg gttgacgtca
     5161 gagcatgttc agtcattaaa acagtaccag cgcctcaaca aaagcctcat acctgaaaac
     5221 tctcggattg cactccgtag aaaatacaaa atcgagcttc aaaatatggt cagatcttgg
     5281 tttgcaatgc aagaccatac ccttgattcg aacatactta tccaacattt gaatcatgac
     5341 ctatcttact taggagccac aaaacgtttt gcatacgatc cagcgatgac caagctcttt
     5401 actgagcttt tgaaacgaga gttatcaaat tcaatcaata atggtgagca acacactaat
     5461 ggatcgtttt tagtcctacc gaatatcagg gtttgtggcg caacagcttt aagctccccg
     5521 gtaacggtgg ggattccatc acttacagct ttctttggct tcgttcacgc atttgaacgg
     5581 aatataaatc gcaccacctc atcgtttcgt gttgaatcct ttgcgatatg cgtccatcaa
     5641 ctacatgtcg agaagcgagg tttgacagca gagtttgtgg aaaaaggcga cgggactata
     5701 tccgctcccg cgacccggga tgactggcag tgtgatgtcg tatttagcct tattttgaac
     5761 accaactttg ctcaacatat tgaccaagat acgttagtta catcactacc aaagcgattg
     5821 gctcggggtt cagcaaaaat tgcgattgat gactttaaac atatcaactc attctcgaca
     5881 ttagaaacag cgatcgaatc tctgccaata gaagctggta ggtggttatc actttacgca
     5941 cagtcaaaca ataatctaag tgatctatta gcagccatga cagaggacca tcagctcatg
     6001 gcaagctgcg tcggttacca cttgttagaa gagcccaaag ataaaccaaa ctccctcaga
     6061 ggttacaaac acgctatcgc cgagtgcatc attggactca ttaactcaat cacctttagc
     6121 tcagagactg atcccaacac aatcttttgg tcgctaaaga actatcaaaa ctacctagtg
     6181 gtacagccaa ggagtatcaa cgatgaaact accgacaaat ctagcctatg agcgctctat
     6241 cgacccatca gatgtctgtt tttttgtcgt ctggcccgat gatagaaaaa cacctttaac
     6301 ctacaattct cgtactctgc tcgggcaaat ggaagcggca tcattagcct atgatgtctc
     6361 aggtcaacca ataaaaagtg ccaccgctga ggcgttagct caagggaacc ctcatcaagt
     6421 tgatttctgc cacgttccat acggtgcgag tcatattgaa tgcagtttct ccgtctcgtt
     6481 ttcttctgaa ctacgtcaac catataagtg taactcaagc aaagttaaac aaacgctagt
     6541 gcaattagtc gagctctacg aaacgaaaat cggctggact gagctagcaa cccgatattt
     6601 gatgaacatt tgcaacggta aatggctgtg gaaaaatacc cgtaaagcct attgctggaa
     6661 cattgtactt acaccttggc cttggaacgg ggaaaaggtt ggatttgaag atattcgtac
     6721 taactacacc tcacggcaag actttaaaaa taataaaaat tggtctgcta tagttgaaat
     6781 gatcaaaacc gcattttcta gtactgatgg gctggcgata tttgaagtca gggccacctt
     6841 gcacttgcca acgaatgcta tggtgcggcc aagccaagtt ttcacagaaa aagaaagtgg
     6901 cagtaaaagt aaatctaaaa ctcaaaacag tcgagttttt cagagtacaa ctattgatgg
     6961 tgaacgatcg ccaatactag gggcctttaa aacgggagca gctattgcaa ccattgacga
     7021 ctggtatcct gaagccactg agccactaag ggtcggacgg tttggggttc atcgcgaaga
     7081 tgtcacttgc taccgtcatc cgtctaccgg aaaagatttt ttctcgatat tacaacaagc
     7141 agagcactat attgaagtgt tgagcgccaa caaaactccc gctcaagaaa ctatcaacga
     7201 catgcacttt ttaatggcta acctgattaa gggtgggatg ttccagcata aaggagactg
     7261 actgtgaaat ggtattataa gacaatcacc tttctgccag agttgtgcaa caacgagtca
     7321 ctggctgcaa agtgtctccg cgttctgcat ggatttaact atcagtatga gacacgaaat
     7381 atcggcgttt catttccgct ttggtgtgat gcaacggttg gaaaaaagat ttcatttgtc
     7441 agcaagaaca agatagaact cgacttacta cttaaacaac actatttcgt ccaaatggaa
     7501 caacttcaat actttcatat atccaacact gttctcgtcc cagaagattg tacatacgtt
     7561 tcctttagac gctgtcaatc tatagataag ctcacagcag cagggctggc aaggaaaatc
     7621 agacgcctgg agaaacgtgc tctatctaga ggcgagcaat ttgacccatc atcttttgct
     7681 caaaaagagc atactgcaat agcgcactac cactcacttg gggagtccag caaacagacg
     7741 aaccgcaact ttcgactcaa tatcaggatg ctctcggagc aaccccgtga ggggaactcg
     7801 atttttagta gctatggctt atcaaattca gaaaactcgt ttcagcctgt acccttaatc
     7861 taatagcatt ggaagtggat aacggatccg cgatcgcggc gcgccacctg gtggccggcc
     7921 ggtaccacgc gtgcgcgctg atccggctgc taacaaagcc cgaaaggaag ctgagttggc
     7981 tgctgccacc gctgagcaat aactagcata accccttggg gcctctaaac gggtcttgag
     8041 gggttttttg ctgaaaggag gaactatatc cggatatcca caggacgggt gtggtcgcca
     8101 tgatcgcgta gtcgatagtg gctccaagta gcgaagcgag caggactggg cggcggccaa
     8161 agcggtcgga cagtgctccg agaacgggtg cgcatagaaa ttgcatcaac gcatatagcg
     8221 ctagcagcac gccatagtga ctggcgatgc tgtcggaatg gacgatatcc cgcaagaggc
     8281 ccggcagtac cggcataacc aagcctatgc ctacagcatc cagggtgacg gtgccgagga
     8341 tgacgatgag cgcattgtta gatttcatac acggtgcctg actgcgttag caatttaact
     8401 gtgataaact accgcattaa agcttatcga tgataagctg tcaaacatga gaattcttga
     8461 agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat aataatggtt
     8521 tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat ttgtttattt
     8581 ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata aatgcttcaa
     8641 taacattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct tattcccttt
     8701 tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa agtaaaagat
     8761 gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa cagcggtaag
     8821 atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt taaagttctg
     8881 ctatgtggcg cggtattatc ccgtgttgac gccgggcaag agcaactcgg tcgccgcata
     8941 cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca tcttacggat
     9001 ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa cactgcggcc
     9061 aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt gcacaacatg
     9121 ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc cataccaaac
     9181 gacgagcgtg acaccacgat gcctgcagca atggcaacaa cgttgcgcaa actattaact
     9241 ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga ggcggataaa
     9301 gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc tgataaatct
     9361 ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga tggtaagccc
     9421 tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga acgaaataga
     9481 cagatcgctg agataggtgc ctcactgatt aagcattggt a
//
