LOCUS       sequence-410888-        5474 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of ISMac7 TnpB and its reRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5474)
  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 5474)
  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_ISMac7_reRNA
FEATURES             Location/Qualifiers
     source          1..5474
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        12..46
                     /label=BBa_J23108
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.51"
     RBS             62..73
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     promoter        1206..1240
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     CDS             1393..1404
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(1513..1530)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1683..1769
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1861..1888
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1907..1998
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2456..2911
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2543..2562)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2753..2774
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3467..4126)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     4041..4060
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(4127..4229)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(4621..4638)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4755..5300)
                     /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(4835..4854)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5376..5395
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 ttatcatcga tctgacagct agctcagtcc taggtataat gctagccccg ggaaaagatc
       61 taaagaggag aaagaattca tgatgaaagc gttcaaattt aggctctatc ctacaactac
      121 acaagctgat caactcaatc aacatatagg tagctgtagg tttgtctata attgggcact
      181 tgatcagaaa attaaaacat acgaacaaac agggaaatca atttccagat ttgacttaaa
      241 caaattaatt ccagctttga agacctctaa cgaatggtta ggagaagtca attctcaatc
      301 attacagggg atgactaagc aggttgagtc tgctttcact cgatttttcc gagagaagaa
      361 tggatttccg aagtttaagt caaagaaaaa tcctatccag tcttttcccg taccccaaca
      421 ctattctgtg aattttgaaa accataccgt taaacttcct aaaattggtg agattaaagc
      481 agttcttcac cgaaaatttg atggagagct taaaacagct agggtatcaa ggacttgtaa
      541 agggcattac tacatcagcg tccttgttga agacggtaaa gaacttcctg aaaaacaggg
      601 attctcagaa tcaacaacag taggaataga tgtgggtatc aaagattttg ccgtcctttc
      661 cacaggagaa aagattgaga atcctaaata cctgaaaaac tctttgcaaa ggttaaaggt
      721 ccttcagaaa agagtatcaa ggaaacaaaa aggctctaaa aatagggcaa aagccagaca
      781 gaggcttgct gtactccacg acaaaataac aaatcagagg aatgacttcc agaacaaact
      841 ctcttttaga cttgttagcg aaaaccaagc tgtagctctg gaaactctga atgttaaagg
      901 tatggttaaa aatcatcatt tggcacaggc tataagtgat tctgcatgga gcagctttgt
      961 aacaaagtta gaatataaag cagaatggtt tggaaaaggc gtattaagga taggacaatt
     1021 tgaaccttct tctaagctct gtaacgtgtg tggataccat aataaagagc ttcagttaaa
     1081 agacagagaa tggacttgtc ctgattgtaa aacaaaacat gatagagaca taaatgccgc
     1141 tatcaatatc aagaaattcg ctctcataga tcaaaatcta attggattgg gatcctgaga
     1201 gatctttgac agctagctca gtcctaggta taatactagt gacagagaat ggacttgtcc
     1261 tgattgtaaa acaaaacatg atagagacat aaatgccgct atcaatatca agaaattcgc
     1321 tctcatagat caaaatctaa ttggattgta acacctacgg aacgtgggga agagcttggg
     1381 gacttgccct caatagaggg aagaatgaac caagaagcaa ctcagtcttt agctgagtgg
     1441 tagttcacgc tcggagatca tcattgcgtt ttttaagctt ggctgttttg gcggatgaga
     1501 gaagattttc agcctgatac agattaaatc agaacgcaga agcggtctga taaaacagaa
     1561 tttgcctggc ggcagtagcg cggtggtccc acctgacccc atgccgaact cagaagtgaa
     1621 acgccgtagc gccgatggta gtgtggggtc tccccatgcg agagtaggga actgccaggc
     1681 atcaaataaa acgaaaggct cagtcgaaag actgggcctt tcgttttatc tgttgtttgt
     1741 cggtgaacgc tctcctgagt aggacaaatc cgccgggagc ggatttgaac gttgcgaagc
     1801 aacggcccgg agggtggcgg gcaggacgcc cgccataaac tgccaggcat caaattaagc
     1861 agaaggccat cctgacggat ggcctttttg cgtttctaca aactcttttg tttatttttc
     1921 taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa
     1981 tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt
     2041 gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct
     2101 gaagatcagt tgggtgcagc aaactattaa ctggcgaact acttactcta gcttcccggc
     2161 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     2221 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     2281 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     2341 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     2401 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttacgcg
     2461 ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca
     2521 cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc
     2581 gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct
     2641 ttacggcacc tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag tgggccatcg
     2701 ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc
     2761 ttgttccaaa ctggaacaac actcaaccct atctcgggct attcttttga tttataaggg
     2821 attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg
     2881 aattttaaca aaatattaac gtttacaatt taaaaggatc taggtgaaga tcctttttga
     2941 taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt cagaccccgt
     3001 agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct gctgcttgca
     3061 aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc taccaactct
     3121 ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgtcc ttctagtgta
     3181 gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc tcgctctgct
     3241 aatcctgtta ccagtcaggc atttgagaag cacacggtca cactgcttcc ggtagtcaat
     3301 aaaccggtaa accagcaata gacataagcg gctatttaac gaccctgccc tgaaccgacg
     3361 accgggtcga atttgctttc gaatttctgc cattcatccg cttattatca cttattcagg
     3421 cgtagcaacc aggcgtttaa gggcaccaat aactgcctta aaaaaattac gccccgccct
     3481 gccactcatc gcagtactgt tgtaattcat taagcattct gccgacatgg aagccatcac
     3541 aaacggcatg atgaacctga atcgccagcg gcatcagcac cttgtcgcct tgcgtataat
     3601 atttgcccat ggtgaaaacg ggggcgaaga agttgtccat attggccacg tttaaatcaa
     3661 aactggtgaa actcacccag ggattggctg agacgaaaaa catattctca ataaaccctt
     3721 tagggaaata ggccaggttt tcaccgtaac acgccacatc ttgcgaatat atgtgtagaa
     3781 actgccggaa atcgtcgtgg tattcactcc agagcgatga aaacgtttca gtttgctcat
     3841 ggaaaacggt gtaacaaggg tgaacactat cccatatcac cagctcaccg tctttcattg
     3901 ccatacggaa ctccggatga gcattcatca ggcgggcaag aatgtgaata aaggccggat
     3961 aaaacttgtg cttatttttc tttacggtct ttaaaaaggc cgtaatatcc agctgaacgg
     4021 tctggttata ggtacattga gcaactgact gaaatgcctc aaaatgttct ttacgatgcc
     4081 attgggatat atcaacggtg gtatatccag tgattttttt ctccatttta gcttccttag
     4141 ctcctgaaaa tctcgataac tcaaaaaata cgcccggtag tgatcttatt tcattatggt
     4201 gaaagttgga acctcttacg tgccgatcaa cgtctcattt tcgccaaaag ttggcccagg
     4261 gcttcccggt atcaacaggg acaccaggat ttatttattc tgcgaagtga tcttccgtca
     4321 caggtattta ttcggcgcaa agtgcgtcgg gtgatgctgc caacttactg atttagtgta
     4381 tgatggtgtt tttgaggtgc tccagtggct tctgtttcta tcagctgtcc ctcctgttca
     4441 gctactgacg gggtggtgcg taacggcaaa agcaccgccg gacatcagcg ctagcggagt
     4501 gtatactggc ttactatgtt ggcactgatg agggtgtcag tgaagtgctt catgtggcag
     4561 gagaaaaaag gctgcaccgg tgcgtcagca gaatatgtga tacaggatat attccgcttc
     4621 ctcgctcact gactcgctac gctcggtcgt tcgactgcgg cgagcggaaa tggcttacga
     4681 acggggcgga gatttcctgg aagatgccag gaagatactt aacagggaag tgagagggcc
     4741 gcggcaaagc cgtttttcca taggctccgc ccccctgaca agcatcacga aatctgacgc
     4801 tcaaatcagt ggtggcgaaa cccgacagga ctataaagat accaggcgtt tccccctggc
     4861 ggctccctcg tgcgctctcc tgttcctgcc tttcggttta ccggtgtcat tccgctgtta
     4921 tggccgcgtt tgtctcattc cacgcctgac actcagttcc gggtaggcag ttcgctccaa
     4981 gctggactgt atgcacgaac cccccgttca gtccgaccgc tgcgccttat ccggtaacta
     5041 tcgtcttgag tccaacccgg aaagacatgc aaaagcacca ctggcagcag ccactggtaa
     5101 ttgatttaga ggagttagtc ttgaagtcat gcgccggtta aggctaaact gaaaggacaa
     5161 gttttggtga ctgcgctcct ccaagccagt tacctcggtt caaagagttg gtagctcaga
     5221 gaaccttcga aaaaccgccc tgcaaggcgg ttttttcgtt ttcagagcaa gagattacgc
     5281 gcagaccaaa acgatctcaa gaagatcatc ttattaatca gataaaatat ttctaggctc
     5341 atgagcccga agtggcgagc ccgatcttcc ccatcggtga tgtcggcgat ataggcgcca
     5401 gcaaccgcac ctgtggcgcc ggtgatgccg gccacgatgc gtccggcgta gaggatctgc
     5461 tcatgtttga cagc
//
