LOCUS       sequence-418002-        5540 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of ISCpe2 TnpB and its reRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5540)
  AUTHORS   Xiang G, Li Y, Sun J, Huo Y, Cao S, Cao Y, Guo Y, Yang L, Cai Y, 
            Zhang YE, Wang H
  TITLE     Evolutionary mining and functional characterization of TnpB 
            nucleases identify efficient miniature genome editors.
  JOURNAL   Nat Biotechnol. 2023 Jun 29. doi: 10.1038/s41587-023-01857-x.
  PUBMED    37386294
REFERENCE   2  (bases 1 to 5540)
  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: "Nat 
            Biotechnol. 2023 Jun 29. doi: 10.1038/s41587-023-01857-x."
COMMENT     Sequence Label: pBad33_ISCpe2_reRNA
FEATURES             Location/Qualifiers
     source          1..5540
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        17..51
                     /label=BBa_J23108
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.51"
     RBS             67..78
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     promoter        1253..1287
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     primer_bind     complement(1584..1601)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1754..1840
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1932..1959
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1978..2069
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2527..2982
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2614..2633)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2824..2845
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3538..4197)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     4112..4131
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(4198..4300)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(4692..4709)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4826..5371)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A 
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin.
                     "
     primer_bind     complement(4906..4925)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5447..5466
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 acagcttatc atcgatctga cagctagctc agtcctaggt ataatgctag ccccgggaaa
       61 agatctaaag aggagaaaga attcatgata agaaagaaag catataaatt tagaatatac
      121 ccaactaaga aacaagccga gctaattaac aagactattg gatgttgtag gtttgtattc
      181 aactattctt taggtgttca aaaaactaaa gataattatt ggaatatagt tgaggagatg
      241 gttcaacaag gctacttcca agagaataat tggaaaggcg agttctttaa taaagccaat
      301 tcaattaaag atatagctaa attaaagaaa agctatgatt ggcttaaaga agtagatagc
      361 atagcactac aggcttctgt tgaaaatctt gatgctggat atgataagta ttacaaaaaa
      421 acaggtagca gacctaaatt taagtctaag aaaaatgaaa tacaatctta tactactaaa
      481 ttagtaaaag ctaagggtaa tgtaaatatt gagatagtag gtaaaggaat taaactacct
      541 aaacttggat tagttaagat agaaaattca agaaatgtag atggaaatat taaaagagtt
      601 actgtgagca gaacacaatc tggtaaatat tttgcatcta ttctttgtga tgtaaatatt
      661 caagaattac ctaaaataga taaaaaagtt ggtgtagatg ttggattaaa aacttttgct
      721 gtttgttctg atggatatga agaagctaat cctaaacact ttagaaaggc agaaaaaaga
      781 cttattaaac ttcaaagaga tttagctcgt aaagaatata actctaaaaa ctatcacaaa
      841 aatagaatta tgattgctaa attacatgaa aggatagcaa atcaaagaat ggacttctta
      901 caaaaatttt caactaagct aattagagaa aaccaatcaa tagccattga agatttaaga
      961 gtttcaaata tgcttaaaaa ccacaaactt gctaaagtaa tttcagaagc aagttggtca
     1021 gaatttagaa gaatgctaga atataaggct gaatggtatg gtagagaaat agtaattgct
     1081 ccaccagact atgcaagctc tcaactatgt tctgaatgtg gctataagaa tatagaagtt
     1141 aaaaatctag gtctaagaga atgggtatgt cccgaatgtg gaacacacca tcaaagagac
     1201 cttaatgctt caataaactt ggaaaagctg atagctggat cctgagagat ctttgacagc
     1261 tagctcagtc ctaggtataa tactagtgaa tgtggctata agaatataga agttaaaaat
     1321 ctaggtctaa gagaatgggt atgtcccgaa tgtggaacac accatcaaag agaccttaat
     1381 gcttcaataa acttggaaaa gctgatagct taaactattg gtatccaacg aggtcggaac
     1441 gcccttgtta gcttggttaa tcttacaccg ttaggtgtgt tgcccaagaa tcttggcatt
     1501 tcaatgccaa gtagttcaag ctcggagatc atcattgcgt tttttaagct tggctgtttt
     1561 ggcggatgag agaagatttt cagcctgata cagattaaat cagaacgcag aagcggtctg
     1621 ataaaacaga atttgcctgg cggcagtagc gcggtggtcc cacctgaccc catgccgaac
     1681 tcagaagtga aacgccgtag cgccgatggt agtgtggggt ctccccatgc gagagtaggg
     1741 aactgccagg catcaaataa aacgaaaggc tcagtcgaaa gactgggcct ttcgttttat
     1801 ctgttgtttg tcggtgaacg ctctcctgag taggacaaat ccgccgggag cggatttgaa
     1861 cgttgcgaag caacggcccg gagggtggcg ggcaggacgc ccgccataaa ctgccaggca
     1921 tcaaattaag cagaaggcca tcctgacgga tggccttttt gcgtttctac aaactctttt
     1981 gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa ccctgataaa
     2041 tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt gtcgccctta
     2101 ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg ctggtgaaag
     2161 taaaagatgc tgaagatcag ttgggtgcag caaactatta actggcgaac tacttactct
     2221 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     2281 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     2341 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     2401 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     2461 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     2521 tgatttacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     2581 tgaccgctac acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc
     2641 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     2701 gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat ggttcacgta
     2761 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     2821 atagtggact cttgttccaa actggaacaa cactcaaccc tatctcgggc tattcttttg
     2881 atttataagg gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa
     2941 aatttaacgc gaattttaac aaaatattaa cgtttacaat ttaaaaggat ctaggtgaag
     3001 atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg
     3061 tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct gcgcgtaatc
     3121 tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc ggatcaagag
     3181 ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc gaatactgtc
     3241 cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc gcctacatac
     3301 ctcgctctgc taatcctgtt accagtcagg catttgagaa gcacacggtc acactgcttc
     3361 cggtagtcaa taaaccggta aaccagcaat agacataagc ggctatttaa cgaccctgcc
     3421 ctgaaccgac gaccgggtcg aatttgcttt cgaatttctg ccattcatcc gcttattatc
     3481 acttattcag gcgtagcaac caggcgttta agggcaccaa taactgcctt aaaaaaatta
     3541 cgccccgccc tgccactcat cgcagtactg ttgtaattca ttaagcattc tgccgacatg
     3601 gaagccatca caaacggcat gatgaacctg aatcgccagc ggcatcagca ccttgtcgcc
     3661 ttgcgtataa tatttgccca tggtgaaaac gggggcgaag aagttgtcca tattggccac
     3721 gtttaaatca aaactggtga aactcaccca gggattggct gagacgaaaa acatattctc
     3781 aataaaccct ttagggaaat aggccaggtt ttcaccgtaa cacgccacat cttgcgaata
     3841 tatgtgtaga aactgccgga aatcgtcgtg gtattcactc cagagcgatg aaaacgtttc
     3901 agtttgctca tggaaaacgg tgtaacaagg gtgaacacta tcccatatca ccagctcacc
     3961 gtctttcatt gccatacgga actccggatg agcattcatc aggcgggcaa gaatgtgaat
     4021 aaaggccgga taaaacttgt gcttattttt ctttacggtc tttaaaaagg ccgtaatatc
     4081 cagctgaacg gtctggttat aggtacattg agcaactgac tgaaatgcct caaaatgttc
     4141 tttacgatgc cattgggata tatcaacggt ggtatatcca gtgatttttt tctccatttt
     4201 agcttcctta gctcctgaaa atctcgataa ctcaaaaaat acgcccggta gtgatcttat
     4261 ttcattatgg tgaaagttgg aacctcttac gtgccgatca acgtctcatt ttcgccaaaa
     4321 gttggcccag ggcttcccgg tatcaacagg gacaccagga tttatttatt ctgcgaagtg
     4381 atcttccgtc acaggtattt attcggcgca aagtgcgtcg ggtgatgctg ccaacttact
     4441 gatttagtgt atgatggtgt ttttgaggtg ctccagtggc ttctgtttct atcagctgtc
     4501 cctcctgttc agctactgac ggggtggtgc gtaacggcaa aagcaccgcc ggacatcagc
     4561 gctagcggag tgtatactgg cttactatgt tggcactgat gagggtgtca gtgaagtgct
     4621 tcatgtggca ggagaaaaaa ggctgcaccg gtgcgtcagc agaatatgtg atacaggata
     4681 tattccgctt cctcgctcac tgactcgcta cgctcggtcg ttcgactgcg gcgagcggaa
     4741 atggcttacg aacggggcgg agatttcctg gaagatgcca ggaagatact taacagggaa
     4801 gtgagagggc cgcggcaaag ccgtttttcc ataggctccg cccccctgac aagcatcacg
     4861 aaatctgacg ctcaaatcag tggtggcgaa acccgacagg actataaaga taccaggcgt
     4921 ttccccctgg cggctccctc gtgcgctctc ctgttcctgc ctttcggttt accggtgtca
     4981 ttccgctgtt atggccgcgt ttgtctcatt ccacgcctga cactcagttc cgggtaggca
     5041 gttcgctcca agctggactg tatgcacgaa ccccccgttc agtccgaccg ctgcgcctta
     5101 tccggtaact atcgtcttga gtccaacccg gaaagacatg caaaagcacc actggcagca
     5161 gccactggta attgatttag aggagttagt cttgaagtca tgcgccggtt aaggctaaac
     5221 tgaaaggaca agttttggtg actgcgctcc tccaagccag ttacctcggt tcaaagagtt
     5281 ggtagctcag agaaccttcg aaaaaccgcc ctgcaaggcg gttttttcgt tttcagagca
     5341 agagattacg cgcagaccaa aacgatctca agaagatcat cttattaatc agataaaata
     5401 tttctaggct catgagcccg aagtggcgag cccgatcttc cccatcggtg atgtcggcga
     5461 tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc ggccacgatg cgtccggcgt
     5521 agaggatctg ctcatgtttg
//
