LOCUS       sequence-411111-        5451 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of ISAam1 TnpB and its reRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5451)
  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 5451)
  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_ISAam1_reRNA
FEATURES             Location/Qualifiers
     source          1..5451
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        7..41
                     /label=BBa_J23108
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.51"
     RBS             57..68
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     primer_bind     complement(1485..1502)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1655..1741
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1833..1860
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1879..1970
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2428..2883
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2515..2534)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2725..2746
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3439..4098)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     4013..4032
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(4099..4201)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(4593..4610)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4727..5272)
                     /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(4807..4826)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5348..5367
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 atcgatctga cagctagctc agtcctaggt ataatgctag ccccgggaaa agatctaaag
       61 aggagaaaga attcatggta aacaaatcat ataaattccg tctgtaccca acaaaagaac
      121 aagaacagct gcttgccaaa accttcggtt gtgtccgttt cgtttataac aaaatgcttg
      181 aagaacgcat acaaatttat gaaaagttca aagacgacaa agaagctttg aaaaaacaaa
      241 catttccgac ccctgccaag tacaaaaagg agtttccttg gcttaaagaa gtggatagcc
      301 ttgcgttagc aaacgcccaa ttgaatttgc agaaagcgtt tcaaaatttc ttttcgggtc
      361 gtgctggatt tccaaagttc aaaaaccgca aggcgaaaca gtcgtacacc acaaatgtgg
      421 tcaatggcaa tattcaactt tcagatggct atatcaagtt acccaaactg aaatgggtca
      481 agttcaagca acatcgggag attcctgccc accatatcat caaggcttgt acgatcaaga
      541 aaacaaaaac aggaaaatac tatgtttcta ttctcaccga atacgaacat caacctgtcc
      601 ccaaagaaat acaaacggtt gttgggcttg atttttccat gaatggttta tttgtcgata
      661 gcgagggtaa gagagccaat tatcctcgtt tctatcggca agcattggaa aaattagcga
      721 aagaacaacg gatattatca cgccgaaaga agggttctaa tcgttggcac aaacagcgtt
      781 tgaaagtagc gaagctacat gagaaaatag cgaaccaacg aaaagacttt ctacacaaaa
      841 aatcatacga attagcgaaa cagtatgatt gcgtggtcat cgaaaacctc aacatgaaag
      901 ggatgtcgca agtcctgaat ttcggcaaga gcgttcatga caacggctgg gggatgttca
      961 cgacatttct ccaatacaag ttagaggagc aagggaaaaa gcttatcaaa atagataagt
     1021 ggtttccgtc atctaaaact tgttcatgtt gcgatcaagt aaaggaatct ttatcgcttt
     1081 ctgagcgtac attccgctgt gaatgtggat tcgagagcga cagggacgtc aatgcggcaa
     1141 tcaatatcaa acatgagggc atgaaacgat tagcaatagc ctgagagatc tttgacagct
     1201 agctcagtcc taggtaaata gaatgtggat tcgagagcga cagggacgtc aatgcggcaa
     1261 tcaatatcaa acatgagggc atgaaacgat tagcaatagc ctaacttgtc ctcgaaccgt
     1321 gggacacacg gggatcgctc agtcaacttc ccgtcatgag atgggattac ctgagaagcc
     1381 cccacctcta agcgaagcgt aggtggtggg agcatgtcac gctcggagat catcattgcg
     1441 ttttttaagc ttggctgttt tggcgcatga gagaagattt tcagcctgat acagattaaa
     1501 tcagaacgca gaagcggtct gataaaacag aatttgcctg gcggcagtag cgcggtggtc
     1561 ccacctgacc ccatgccgaa ctcagaagtg aaacgccgta gcgccgatgg tagtgtgggg
     1621 tctccccatg cgagagtagg gaactgccag gcatcaaata aaacgaaagg ctcagtcgaa
     1681 agactgggcc tttcgtttta tctgttgttt gtcggtgaac gctctcctga gtaggacaaa
     1741 tccgccggga gcggatttga acgttgcgaa gcaacggccc ggagggtggc gggcaggacg
     1801 cccgccataa actgccaggc atcaaattaa gcagaaggcc atcctgacgg atggcctttt
     1861 tgcgtttcta caaactcttt tgtttatttt tctaaataca ttcaaatatg tatccgctca
     1921 tgagacaata accctgataa atgcttcaat aatattgaaa aaggaagagt atgagtattc
     1981 aacatttccg tgtcgccctt attccctttt ttgcggcatt ttgccttcct gtttttgctc
     2041 acccagaaac gctggtgaaa gtaaaagatg ctgaagatca gttgggtgca gcaaactatt
     2101 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     2161 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     2221 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     2281 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     2341 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     2401 ttactcatat atactttaga ttgatttacg cgccctgtag cggcgcatta agcgcggcgg
     2461 gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt
     2521 tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc
     2581 gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg
     2641 atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga
     2701 cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc
     2761 ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa
     2821 aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta acgtttacaa
     2881 tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt
     2941 gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat
     3001 cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg
     3061 gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga
     3121 gcgcagatac caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac
     3181 tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtcag gcatttgaga
     3241 agcacacggt cacactgctt ccggtagtca ataaaccggt aaaccagcaa tagacataag
     3301 cggctattta acgaccctgc cctgaaccga cgaccgggtc gaatttgctt tcgaatttct
     3361 gccattcatc cgcttattat cacttattca ggcgtagcaa ccaggcgttt aagggcacca
     3421 ataactgcct taaaaaaatt acgccccgcc ctgccactca tcgcagtact gttgtaattc
     3481 attaagcatt ctgccgacat ggaagccatc acaaacggca tgatgaacct gaatcgccag
     3541 cggcatcagc accttgtcgc cttgcgtata atatttgccc atggtgaaaa cgggggcgaa
     3601 gaagttgtcc atattggcca cgtttaaatc aaaactggtg aaactcaccc agggattggc
     3661 tgagacgaaa aacatattct caataaaccc tttagggaaa taggccaggt tttcaccgta
     3721 acacgccaca tcttgcgaat atatgtgtag aaactgccgg aaatcgtcgt ggtattcact
     3781 ccagagcgat gaaaacgttt cagtttgctc atggaaaacg gtgtaacaag ggtgaacact
     3841 atcccatatc accagctcac cgtctttcat tgccatacgg aactccggat gagcattcat
     3901 caggcgggca agaatgtgaa taaaggccgg ataaaacttg tgcttatttt tctttacggt
     3961 ctttaaaaag gccgtaatat ccagctgaac ggtctggtta taggtacatt gagcaactga
     4021 ctgaaatgcc tcaaaatgtt ctttacgatg ccattgggat atatcaacgg tggtatatcc
     4081 agtgattttt ttctccattt tagcttcctt agctcctgaa aatctcgata actcaaaaaa
     4141 tacgcccggt agtgatctta tttcattatg gtgaaagttg gaacctctta cgtgccgatc
     4201 aacgtctcat tttcgccaaa agttggccca gggcttcccg gtatcaacag ggacaccagg
     4261 atttatttat tctgcgaagt gatcttccgt cacaggtatt tattcggctc aaagtgcgtc
     4321 gggtgatgct gccaacttac tgatttagtg tatgatggtg tttttgaggt gctccagtgg
     4381 cttctgtttc tatcagctgt ccctcctgtt cagctactga cggggtggtg cgtaacggca
     4441 aaagcaccgc cggacatcag cgctagcgga gtgtatactg gcttactatg ttggcactga
     4501 tgagggtgtc agtgaagtgc ttcatgtggc aggagaaaaa aggctgcacc ggtgcgtcag
     4561 cagaatatgt gatacaggat atattccgct tcctcgctca ctgactcgct acgctcggtc
     4621 gttcgactgc ggcgagcgga aatggcttac gaacggggcg gagatttcct ggaagatgcc
     4681 aggaagatac ttaacaggga agtgagaggg ccgcggcaaa gccgtttttc cataggctcc
     4741 gcccccctga caagcatcac gaaatctgac gctcaaatca gtggtggcga aacccgacag
     4801 gactataaag ataccaggcg tttccccctg gcggctccct cgtgcgctct cctgttcctg
     4861 cctttcggtt taccggtgtc attccgctgt tatggccgcg tttgtctcat tccacgcctg
     4921 acactcagtt ccgggtaggc agttcgctcc aagctggact gtatgcacga accccccgtt
     4981 cagtccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggaaagacat
     5041 gcaaaagcac cactggcagc agccactggt aattgattta gaggagttag tcttgaagtc
     5101 atgcgccggt taaggctaaa ctgaaaggac aagttttggt gactgcgctc ctccaagcca
     5161 gttacctcgg ttcaaagagt tggtagctca gagaaccttc gaaaaaccgc cctgcaaggc
     5221 ggttttttcg ttttcagagc aagagattac gcgcagacca aaacgatctc aagaagatca
     5281 tcttattaat cagataaaat atttctaggc tcatgagccc gaagtggcga gcccgatctt
     5341 ccccatcggt gatgtcggcg atataggcgc cagcaaccgc acctgtggcg ccggtgatgc
     5401 cggccacgat gcgtccggcg tagaggatct gctcatgttt gacagcttat c
//
