LOCUS       sequence-338901-        7748 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of CRISPR-associated transposases.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7748)
  AUTHORS   Zhang Y, Yang J, Yang S, Zhang J, Chen J, Tao R, Jiang Y, Yang J, 
            Yang S
  TITLE     Programming Cells by Multicopy Chromosomal Integration Using 
            CRISPR-Associated Transposases.
  JOURNAL   CRISPR J. 2021 Jun;4(3):350-359. doi: 10.1089/crispr.2021.0018.
  PUBMED    34152213
REFERENCE   2  (bases 1 to 7748)
  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: 
            "10.1089/crispr.2021"; journalName: "CRISPR J"; date: "2021-06"; 
            volume: "4"; issue: "3"; pages: "350-359"
COMMENT     Sequence Label: pRha-tns
FEATURES             Location/Qualifiers
     source          1..7748
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(417..1265)
                     /codon_start=1
                     /gene="E. coli rhaR"
                     /product="transcriptional activator of rhaR and rhaS"
                     /label=rhaR
                     /note="needed for rhamnose-dependent activation of the rhaB
                     promoter"
                     /translation="MAHQLKLLKDDFFASDQQAVAVADRYPQDVFAEHTHDFCELVIVW
                     RGNGLHVLNDRPYRITRGDLFYIHADDKHSYASVNDLVLQNIIYCPERLKLNLDWQGAI
                     PGFNASAGQPHWRLGSMGMAQARQVIGQLEHESSQHVPFANEMAELLFGQLVMLLNRHR
                     YTSDSLPPTSSETLLDKLITRLAASLKSPFALDKFCDEASCSERVLRQQFRQQTGMTIN
                     QYLRQVRVCHAQYLLQHSRLLISDISTECGFEDSNYFSVVFTRETGMTPSQWRHLNSQK
                     D"
     CDS             complement(1339..2175)
                     /codon_start=1
                     /gene="E. coli rhaS"
                     /product="positive regulator of the rhaB promoter
                     "
                     /label=rhaS
                     /note="activated by L-rhamnose"
                     /translation="MTVLHSVDFFPSGNASVAIEPRLPQADFPEHHHDFHEIVIVEHGT
                     GIHVFNGQPYTITGGTVCFVRDHDRHLYEHTDNLCLTNVLYRSPDRFQFLAGLNQLLPQ
                     ELDGQYPSHWRVNHSVLQQVRQLVAQMEQQEGENDLPSTASREILFMQLLLLLRKSSLQ
                     ENLENSASRLNLLLAWLEDHFADEVNWDAVADQFSLSLRTLHRQLKQQTGLTPQRYLNR
                     LRLMKARHLLRHSEASVTDIAYRCGFSDSNHFSTLFRREFNWSPRDIRQGRDGFLQ"
     promoter        2321..2439
                     /label=rhaB promoter
                     /note="promoter of the E. coli rhaBAD operon, conferring 
                     tight induction with L-rhamnose and repression with 
                     D-glucose in the presence of RhaR and RhaS (Giacalone et 
                     al., 2006)"
     CDS             4561..4572
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(5999..6017)
                     /label=T7 Term
                     /note="T7 terminator, reverse primer"
     terminator      6013..6060
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(6290..7105)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(6290..7105)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     7013..7032
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     promoter        complement(7106..7197)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(join(7215..7748,1..102))
                     /direction=LEFT
                     /label=ColA ori
                     /note="Plasmids containing the ColA origin of replication 
                     can be propagated in E. coli cells that contain additional 
                     plasmids with compatible origins."
     primer_bind     complement(7360..7379)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgatcaaacc gcctccccag gcggtttttt cgtttacaga gcaggagatt acgacgatcg
       61 taaaaggatc tcaagaagat cctttacgga ttcccgacac catcactcta gatttcagtg
      121 caatttatct cttcaaatgt agcacctgaa gtcagcccca tacgatataa gttgtaattc
      181 tcatgttagt catgccccgc gcccaccgga aggagctgac tgggttgaag gctctcaagg
      241 gcatcggtcg agatcccggt gcctaatgag tgagctaact tacattaatt gcgttgcgcg
      301 ttcttatggc tcttgtatct atcagtgaag catcaagact aacaaacaaa agtagaacaa
      361 ctgttcaccg ttacatatca aagggaaaac tgtccatacc catggggcat ggcgaattaa
      421 tctttctgcg aattgagatg acgccactgg ctgggcgtca tcccggtttc ccgggtaaac
      481 accaccgaaa aatagttact atcttcaaag ccacattcgg tcgaaatatc actgattaac
      541 aggcggctat gctggagaag atattgcgca tgacacactc tgacctgtcg cagatattga
      601 ttgatggtca ttccagtctg ctggcgaaat tgctgacgca aaacgcgctc actgcacgat
      661 gcctcatcac aaaatttatc cagcgcaaag ggacttttca ggctagccgc cagccgggta
      721 atcagcttat ccagcaacgt ttcgctggat gttggcggca acgaatcact ggtgtaacga
      781 tggcgattca gcaacatcac caactgcccg aacagcaact cagccatttc gttagcaaac
      841 ggcacatgct gactactttc atgctcaagc tgaccgataa cctgccgcgc ctgcgccatc
      901 cccatgctac ctaagcgcca gtgtggttgc cctgcgctgg cgttaaatcc cggaatcgcc
      961 ccctgccagt caagattcag cttcagacgc tccgggcaat aaataatatt ctgcaaaacc
     1021 agatcgttaa cggaagcgta ggagtgttta tcgtcagcat gaatgtaaaa gagatcgcca
     1081 cgggtaatgc gataagggcg atcgttgagt acatgcaggc cattaccgcg ccagacaatc
     1141 accagctcac aaaaatcatg tgtatgttca gcaaagacat cttgcggata acggtcagcc
     1201 acagcgactg cctgctggtc gctggcaaaa aaatcatctt tgagaagttt taactgatgc
     1261 gccaccgtgg ctacctcggc cagagaacga agttgattat tcgcaatatg gcgtacaaat
     1321 acgttgagaa gattcgcgtt attgcagaaa gccatcccgt ccctggcgaa tatcacgcgg
     1381 tgaccagtta aactctcggc gaaaaagcgt cgaaaagtgg ttactgtcgc tgaatccaca
     1441 gcgataggcg atgtcagtaa cgctggcctc gctgtggcgt agcagatgtc gggctttcat
     1501 cagtcgcagg cggttcaggt atcgctgagg cgtcagtccc gtttgctgct taagctgccg
     1561 atgtagcgta cgcagtgaaa gagaaaattg atccgccacg gcatcccaat tcacctcatc
     1621 ggcaaaatgg tcctccagcc aggccagaag caagttgaga cgtgatgcgc tgttttccag
     1681 gttctcctgc aaactgcttt tacgcagcaa gagcagtaat tgcataaaca agatctcgcg
     1741 actggcggtc gagggtaaat cattttcccc ttcctgctgt tccatctgtg caaccagctg
     1801 tcgcacctgc tgcaatacgc tgtggttaac gcgccagtga gacggatact gcccatccag
     1861 ctcttgtggc agcaactgat tcagcccggc gagaaactga aatcgatccg gcgagcgata
     1921 cagcacattg gtcagacaca gattatcggt atgttcatac agatgccgat catgatcgcg
     1981 tacgaaacag accgtgccac cggtgatggt atagggctgc ccattaaaca catgaatacc
     2041 cgtgccatgt tcgacaatca caatttcatg aaaatcatga tgatgttcag gaaaatccgc
     2101 ctgcgggagc cggggttcta tcgccacgga cgcgttacca gacggaaaaa aatccacact
     2161 atgtaatacg gtcatactgg cctcctgatg tcgtcaacac ggcgaaatag taatcacgag
     2221 gtcaggttct taccttaaat tttcgacgga aaaccacgta aaaaacgtcg atttttcaag
     2281 atacagcgtg aattttcagg aaatgcggtg agcatcacat caccacaatt cagcaaattg
     2341 tgaacatcat cacgttcatc tttccctggt tgccaatggc ccattttcct gtcagtaacg
     2401 agaaggtcgc gaattcaggc gctttttaga ctggtcgtac ctgtagaaat aattttgttt
     2461 aactttaata aggagatata ccatggcgac aagtttacct acgccctcag caattacgac
     2521 ttcggcgtta gagtatgcat tccatactcc cgctcgcaat ctaacgaaat ctcgcggaaa
     2581 aaacattcat cgttatgtca gtgtaaagat gagtaagagg attacggtag aatctactct
     2641 agagtgtgat gcctgctatc actttgattt tgagccaagt attgttcgct tttgcgctca
     2701 accgattcga tttttatatt atctcaatgg tcagtctcac tcctatgttc ctgactttct
     2761 agttcaattt gataccaacg agtttgttct atatgaagta aagtcagctt atgctaagaa
     2821 caaacctgat tttgatgttg aatgggaggc gaaagtaaaa gcagcaactg aactagggct
     2881 agaattggag cttgttgaag agagtgatat tagggatacg gttgtattaa ataatcttaa
     2941 gcgcatgcat cgttatgctt cgaaagatga gctgaataac gtacataact ctctcttaaa
     3001 aataataaag tacaatggcg cccaatctgc aagatgcttg ggagaacagt tgggtttaaa
     3061 aggccgaact gttttaccaa ttttgtgcga tttgctgtca aggtgtttac tcgatacacg
     3121 tttggataag cctctatctc ttgaatctcg atttgagttg gccagttatg gctaagaaag
     3181 ggttctcaag tttccataga aaagcagtct cttcccaaga tacgcttgaa tcgatagagc
     3241 ttgtctctag cgctaattgt ctagaaagtg ttacgtatca agatatatca gcatttcccg
     3301 aaacaattgc ggtagagatt aatttccgat taagcattct tcgcttttta gcgcgaaagt
     3361 gcgaaaccat tgtggccaaa tcaattgaac cacatcgtgt agagctacag caaaactata
     3421 gtagaaaaat acccagtgca ataacgatat atcgatggtg gcttgctttt cgaaaatcag
     3481 actacaaccc cattagctta gcacctaata tcaaggatag aggtaataga gagacaaaag
     3541 tgtcaacagt tgttgattct attatggaac aggcagttga aagagttata tctggacgaa
     3601 aagtcaatgt tagctctgca tataaacgtg ttcgacgaaa agttcgtcaa tacaatctaa
     3661 ctcatggaac gaaatacacg tatcctaagt acgaatctgt aagaaagcga gtaaaaaaga
     3721 aaaccccatt tgagttatta gccgcaggga aaggggagag agtagctaag agagagtttc
     3781 gccgaatggg aaaaaagatc ctcacgtcta gcgtgctaga gagggttgaa atagatcaca
     3841 ctgtcgttga cctttttgca gtacacgaag agtatcgaat cccattgggc cgaccttggc
     3901 ttactcaatt ggttgattgt tacagtaaag ctgtaatcgg tttttattta ggtttcgagc
     3961 ctcctagcta tgtgtcggtt tcccttgcac ttaagaatgc aatacaacgc aaagatgact
     4021 taatctcctc gtatgaatcg atcgagaatg aatggctatg ttatggcatc ccagacctac
     4081 tcgtaactga taatggtaaa gagtttttgt cgaaagcatt tgatcaagca tgtgaatcac
     4141 tattgatcaa tgtgcatcaa aataaagttg agacgcccga caacaaacct catgttgaac
     4201 gtaactacgg gactattaat acttctctgt tagacgattt acctgggaaa tccttcagcc
     4261 agtaccttca aagagaaggg tacgactctg tgggagaagc tacccttaca ctcaatgaga
     4321 ttagagaaat ttacttaatt tggttggtgg atatttatca taaaaaaccc aatcagagag
     4381 gcactaattg tcctaatgtt gcgtggaaaa agggttgtca agaatgggaa ccagaggagt
     4441 tctctggttc taaagacgaa ttagacttta aatttgctat tgttgattac aaacaactta
     4501 ctaaagtagg gataactgtc tacaaagaac tgagttatag caatgaccgt ttagctgaat
     4561 atagagggaa gaaaggaaac cataaagttc agttcaagta taaccctgag tgtatggcag
     4621 ttatttgggt gttggatgag gatatgaatg agtactttac agttaatgcg attgactacg
     4681 aatatgcaag tagagtatca ctttggcaac ataaatataa catgaaatat caagcagaac
     4741 taaattcagc agaatatgat gaggacaagg aaattgatgc agaaataaaa attgaagaaa
     4801 tcgcagatcg ttcaattgtt aagactaaca aaatcagagc tcggaggcgt ggcgctaggc
     4861 atcaagagaa tagcgcaagg gctaagtcaa tcagtaatgc gaacccggcc tcgatacaaa
     4921 aacatgaaga tgaaatcgtt agtgcagata atgacgattg ggatattgat tatgtctgag
     4981 aaatcgtcaa atgagtgaaa cgcgtgaggc tcgtatatca agagctaaaa gggcatttgt
     5041 atccacaccg agcgttagga aaatcttgag ttacatggat agatgtagag atctatcaga
     5101 cctagagtct gagcctacat gcatgatggt ctatggtgct tcgggtgtag gtaaaacgac
     5161 cgtcatcaag aaatacttaa atcaaaacag aagagagtcc gaagccgggg gcgatataat
     5221 accggttttg catattgagt tgccagacaa tgcgaagcca gtagatgcag caagggaatt
     5281 gctggttgaa atgggtgacc cgctagcact ttatgaaact gacttagcta gattgacgaa
     5341 aagactgact gaattaatcc ctgcggtcgg cgtgaagctg attattatcg atgagttcca
     5401 acatttggtg gaagaaaggt caaatcgggt tcttacccaa gtaggtaatt ggctaaaaat
     5461 gatacttaac aaaacgaaat gtccaattgt tatatttggt atgccatact caaaagttgt
     5521 actgcaagca aactcgcaac ttcacgggcg attttccatt caggttgaac tgcgcccctt
     5581 tagctaccag ggaggtagag gtgtatttaa aacttttttg gaataccttg ataaagccct
     5641 accttttgaa aaacaggctg gcttagccaa cgaaagtttg cagaaaaaat tgtatgcatt
     5701 ctctcaggga aacatgcgtt cgttgagaaa ccttatttat caagcatcta tcgaagcaat
     5761 tgataatcag catgagacga taaccgaaga agatttcgtt tttgcatcga agttgacatc
     5821 gggcgataaa cccaactcat ggaaaaatcc ttttgaggag ggtgttgagg taacagaaga
     5881 tatgttacga ccgccaccaa aagatattgg ttgggaggac tatttgagac attcaacccc
     5941 gagagtgagt aaaccaggta gaaataaaaa ctttttcgaa taacctaggc tgctgccacc
     6001 gctgagcaat aactagcata accccttggg gcctctaaac gggtcttgag gggttttttg
     6061 ctgaaacctc aggcatttga gaagcacacg gtcacactgc ttccggtagt caataaaccg
     6121 gtaaaccagc aatagacata agcggctatt taacgaccct gccctgaacc gacgacaagc
     6181 tgacgaccgg gtctccgcaa gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     6241 tttatttttc taaatacatt caaatatgta tccgctcatg aattaattct tagaaaaact
     6301 catcgagcat caaatgaaac tgcaatttat tcatatcagg attatcaata ccatattttt
     6361 gaaaaagccg tttctgtaat gaaggagaaa actcaccgag gcagttccat aggatggcaa
     6421 gatcctggta tcggtctgcg attccgactc gtccaacatc aatacaacct attaatttcc
     6481 cctcgtcaaa aataaggtta tcaagtgaga aatcaccatg agtgacgact gaatccggtg
     6541 agaatggcaa aagtttatgc atttctttcc agacttgttc aacaggccag ccattacgct
     6601 cgtcatcaaa atcactcgca tcaaccaaac cgttattcat tcgtgattgc gcctgagcga
     6661 gacgaaatac gcggtcgctg ttaaaaggac aattacaaac aggaatcgaa tgcaaccggc
     6721 gcaggaacac tgccagcgca tcaacaatat tttcacctga atcaggatat tcttctaata
     6781 cctggaatgc tgttttcccg gggatcgcag tggtgagtaa ccatgcatca tcaggagtac
     6841 ggataaaatg cttgatggtc ggaagaggca taaattccgt cagccagttt agtctgacca
     6901 tctcatctgt aacatcattg gcaacgctac ctttgccatg tttcagaaac aactctggcg
     6961 catcgggctt cccatacaat cgatagattg tcgcacctga ttgcccgaca ttatcgcgag
     7021 cccatttata cccatataaa tcagcatcca tgttggaatt taatcgcggc ctagagcaag
     7081 acgtttcccg ttgaatatgg ctcatactct tcctttttca atattattga agcatttatc
     7141 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     7201 gcatgctagc gcagaaacgt cctagaagat gccaggagga tacttagcag agagacaata
     7261 aggccggagc gaagccgttt ttccataggc tccgcccccc tgacgaacat cacgaaatct
     7321 gacgctcaaa tcagtggtgg cgaaacccga caggactata aagataccag gcgtttcccc
     7381 ctgatggctc cctcttgcgc tctcctgttc ccgtcctgcg gcgtccgtgt tgtggtggag
     7441 gctttaccca aatcaccacg tcccgttccg tgtagacagt tcgctccaag ctgggctgtg
     7501 tgcaagaacc ccccgttcag cccgactgct gcgccttatc cggtaactat catcttgagt
     7561 ccaacccgga aagacacgac aaaacgccac tggcagcagc cattggtaac tgagaattag
     7621 tggatttaga tatcgagagt cttgaagtgg tggcctaaca gaggctacac tgaaaggaca
     7681 gtatttggta tctgcgctcc actaaagcca gttaccaggt taagcagttc cccaactgac
     7741 ttaacctt
//
