LOCUS       sequence-410873-        5473 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of ISCvi2 TnpB and its reRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5473)
  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 5473)
  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_ISCvi2_reRNA
FEATURES             Location/Qualifiers
     source          1..5473
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        5..39
                     /label=BBa_J23108
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.51"
     RBS             55..66
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     primer_bind     complement(1505..1522)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1675..1761
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1853..1880
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1899..1990
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2448..2903
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2535..2554)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2745..2766
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3459..4118)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     4033..4052
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(4119..4221)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(4613..4630)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4747..5292)
                     /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(4827..4846)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5368..5387
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 cgatctgaca gctagctcag tcctaggtat aatgctagcc ccgggaaaag atctaaagag
       61 gagaaaatga ataaagcaat caaatacaga gtgcatccca caactgaaca atctgttatg
      121 tttgctaaga cctttggctg ttgtcgtaag gtctataatc ttatgctttc cgataaaatt
      181 gacggctaca aagctacggg caaatttcct actgttacgc ctgccaaata taagaacgat
      241 tatccttatc ttagggaagt ggatagtctt gctcttgcca ataagcaaat ggacttacag
      301 gaagcgttcc gcaatacatt tagcaaatcc cgcaaaaaga aaaatggttt tcctaaattc
      361 aaatccgcaa agcatagccg gaagtcctat actacgaaca atcaacatgg gacagttgcc
      421 attatagaca acaaatatat caagcttcct aagattggca aagtaaaagc cgtcatccat
      481 cgcattcctg atagcgattg gattgtaaaa tcagctacgg tatcacagga atcagatggc
      541 aaatattata tttctgttct ctttgagttt gacaatgcca taaatcttta tgtagcagat
      601 aagaccaatg ctatcggatt agattatact tctaatggct tgtatgtgga cagtaacggt
      661 aacgtaggaa caaatcacaa atactatcgt gaaagccatg acaaacttgc caaggcacaa
      721 cgtaggctgt cacgtatgca aggctctaaa aagcatgaaa tcaaatccaa caattatata
      781 aaacaacttc gcaaagtgaa taagatacat aggtatatcg ctaatcagcg tctggataac
      841 ctgcataaaa tatctacgaa gatagccaat tcgtatgatg ttgtttgcgt ggaatcctta
      901 aacatgaggt ctatggcaaa tcatggcttc ggtaacggca aagctactct tgataacggt
      961 tatggtatgt tcctgtctat gcttgaatac aagctatctg ataggaataa gtatcttatc
     1021 aaggtagata agtggtttcc gtcatcacaa atatgtcatt gttgtggaaa gatacatccc
     1081 gagatgaaaa atctcacaat tcgcacaatg aaatgtgatt gtggtcttac gatgaatcgt
     1141 gaccagaatg ccgctatcaa tatcttacgt gagggattac gactactgac taaagtagcc
     1201 tgagagatct ttgacagcta gctcagtcct aggtataata gaatgccgct atcaatatct
     1261 tacgtgaggg attacgacta ctgactaaag tagcctaaaa ctaaatatta cagtagggat
     1321 ggaatagccc aaactcatac gcctgtggac actgtgtaag acgagccgtt acgtatcatc
     1381 gtagccagcg cagtagtggt tgaagcagga agctcggtta cttgtaaccg agtagttcac
     1441 gctcggagat catcattgcg ttttttaagc ttggctgttt tggcggatga gagaagattt
     1501 tcagcctgat acagattaaa tcagaacgca gaagcggtct gataaaacag aatttgcctg
     1561 gcggcagtag cgcggtggtc ccacctgacc ccatgccgaa ctcagaagtg aaacgccgta
     1621 gcgccgatgg tagtgtgggg tctccccatg cgagagtagg gaactgccag gcatcaaata
     1681 aaacgaaagg ctcagtcgaa agactgggcc tttcgtttta tctgttgttt gtcggtgaac
     1741 gctctcctga gtaggacaaa tccgccggga gcggatttga acgttgcgaa gcaacggccc
     1801 ggagggtggc gggcaggacg cccgccataa actgccaggc atcaaattaa gcagaaggcc
     1861 atcctgacgg atggcctttt tgcgtttcta caaactcttt tgtttatttt tctaaataca
     1921 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     1981 aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt ttgcggcatt
     2041 ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg ctgaagatca
     2101 gttgggtgca gcaaactatt aactggcgaa ctacttactc tagcttcccg gcaacaatta
     2161 atagactgga tggaggcgga taaagttgca ggaccacttc tgcgctcggc ccttccggct
     2221 ggctggttta ttgctgataa atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca
     2281 gcactggggc cagatggtaa gccctcccgt atcgtagtta tctacacgac ggggagtcag
     2341 gcaactatgg atgaacgaaa tagacagatc gctgagatag gtgcctcact gattaagcat
     2401 tggtaactgt cagaccaagt ttactcatat atactttaga ttgatttacg cgccctgtag
     2461 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     2521 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     2581 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     2641 cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata
     2701 gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca
     2761 aactggaaca acactcaacc ctatctcggg ctattctttt gatttataag ggattttgcc
     2821 gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa
     2881 caaaatatta acgtttacaa tttaaaagga tctaggtgaa gatccttttt gataatctca
     2941 tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga
     3001 tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa
     3061 aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga
     3121 aggtaactgg cttcagcaga gcgcagatac caaatactgt ccttctagtg tagccgtagt
     3181 taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt
     3241 taccagtcag gcatttgaga agcacacggt cacactgctt ccggtagtca ataaaccggt
     3301 aaaccagcaa tagacataag cggctattta acgaccctgc cctgaaccga cgaccgggtc
     3361 gaatttgctt tcgaatttct gccattcatc cgcttattat cacttattca ggcgtagcaa
     3421 ccaggcgttt aagggcacca ataactgcct taaaaaaatt acgccccgcc ctgccactca
     3481 tcgcagtact gttgtaattc attaagcatt ctgccgacat ggaagccatc acaaacggca
     3541 tgatgaacct gaatcgccag cggcatcagc accttgtcgc cttgcgtata atatttgccc
     3601 atggtgaaaa cgggggcgaa gaagttgtcc atattggcca cgtttaaatc aaaactggtg
     3661 aaactcaccc agggattggc tgagacgaaa aacatattct caataaaccc tttagggaaa
     3721 taggccaggt tttcaccgta acacgccaca tcttgcgaat atatgtgtag aaactgccgg
     3781 aaatcgtcgt ggtattcact ccagagcgat gaaaacgttt cagtttgctc atggaaaacg
     3841 gtgtaacaag ggtgaacact atcccatatc accagctcac cgtctttcat tgccatacgg
     3901 aactccggat gagcattcat caggcgggca agaatgtgaa taaaggccgg ataaaacttg
     3961 tgcttatttt tctttacggt ctttaaaaag gccgtaatat ccagctgaac ggtctggtta
     4021 taggtacatt gagcaactga ctgaaatgcc tcaaaatgtt ctttacgatg ccattgggat
     4081 atatcaacgg tggtatatcc agtgattttt ttctccattt tagcttcctt agctcctgaa
     4141 aatctcgata actcaaaaaa tacgcccggt agtgatctta tttcattatg gtgaaagttg
     4201 gaacctctta cgtgccgatc aacgtctcat tttcgccaaa agttggccca gggcttcccg
     4261 gtatcaacag ggacaccagg atttatttat tctgcgaagt gatcttccgt cacaggtatt
     4321 tattcggcgc aaagtgcgtc gggtgatgct gccaacttac tgatttagtg tatgatggtg
     4381 tttttgaggt gctccagtgg cttctgtttc tatcagctgt ccctcctgtt cagctactga
     4441 cggggtggtg cgtaacggca aaagcaccgc cggacatcag cgctagcgga gtgtatactg
     4501 gcttactatg ttggcactga tgagggtgtc agtgaagtgc ttcatgtggc aggagaaaaa
     4561 aggctgcacc ggtgcgtcag cagaatatgt gatacaggat atattccgct tcctcgctca
     4621 ctgactcgct acgctcggtc gttcgactgc ggcgagcgga aatggcttac gaacggggcg
     4681 gagatttcct ggaagatgcc aggaagatac ttaacaggga agtgagaggg ccgcggcaaa
     4741 gccgtttttc cataggctcc gcccccctga caagcatcac gaaatctgac gctcaaatca
     4801 gtggtggcga aacccgacag gactataaag ataccaggcg tttccccctg gcggctccct
     4861 cgtgcgctct cctgttcctg cctttcggtt taccggtgtc attccgctgt tatggccgcg
     4921 tttgtctcat tccacgcctg acactcagtt ccgggtaggc agttcgctcc aagctggact
     4981 gtatgcacga accccccgtt cagtccgacc gctgcgcctt atccggtaac tatcgtcttg
     5041 agtccaaccc ggaaagacat gcaaaagcac cactggcagc agccactggt aattgattta
     5101 gaggagttag tcttgaagtc atgcgccggt taaggctaaa ctgaaaggac aagttttggt
     5161 gactgcgctc ctccaagcca gttacctcgg ttcaaagagt tggtagctca gagaaccttc
     5221 gaaaaaccgc cctgcaaggc ggttttttcg ttttcagagc aagagattac gcgcagacca
     5281 aaacgatctc aagaagatca tcttattaat cagataaaat atttctaggc tcatgagccc
     5341 gaagtggcga gcccgatctt ccccatcggt gatgtcggcg atataggcgc cagcaaccgc
     5401 acctgtggcg ccggtgatgc cggccacgat gcgtccggcg tagaggatct gctcatgttt
     5461 gacagcttat cat
//
