LOCUS       sequence-410882-        5515 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of ISEfa4 TnpB and its reRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5515)
  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 5515)
  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_ISEfa4_reRNA
FEATURES             Location/Qualifiers
     source          1..5515
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        19..53
                     /label=BBa_J23108
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.51"
     RBS             69..80
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     promoter        1249..1283
                     /label=J23119(SpeI) promoter
                     /note="bacterial promoter (Registry of Standard Biological 
                     Parts BBa_J23119) modified to end with an SpeI site"
     primer_bind     complement(1561..1578)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1731..1817
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1909..1936
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1955..2046
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2504..2959
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2591..2610)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2801..2822
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3515..4174)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     4089..4108
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(4175..4277)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(4669..4686)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4803..5348)
                     /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(4883..4902)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5424..5443
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 tgacagctta tcatcgatct gacagctagc tcagtcctag gtataatgct agccccggga
       61 aaagatctaa agaggagaaa gaattcatgg aacaactaaa agcatacaaa ttcagaatct
      121 atccaacaga ggaacaagaa atcttctttg ccaaatcttt tggttgtgtc cgtaaggtct
      181 acaatctaat gcttgatgac cgaaaaaagg cgtatgaaga agttaaaaat gattcttcta
      241 aaaaaatgac ttttccgaca ccagctaaat acaagaaaga atttccgttt ttgaaagaaa
      301 ttgatagcct tgctttagcc aatgcccaac ttaatttaga taaagcatac aagaattttt
      361 ttcgggataa atccgttgga tttccacgtt tcaaaagtaa gaaaaatcct gttcagagtt
      421 atacaacgaa taatcaaaat ggtacagttg ctttgattga tagtaaattc atcaaagttc
      481 ctaaattgaa gtcattagta agaattaagc ttcatagaca accaaaaggg attatcaaat
      541 ctgctacaat atcacgtcat tctagcggta aatactatat ttctttgttg tgtaaagaag
      601 aaattatcga attacctaaa actaattcca caattggtat tgacttaggt attatggact
      661 ttgcgattct ttctgatgga caaaaaattg ataataataa attcacgtcc aaaatggaaa
      721 agaaactaaa gcgtgaacaa cgcaaattgt ctaggcgtgc tttattggct aaaaataaag
      781 gtaccaatct atttgaagct aaaaattatc aaaaacaaaa acgcaaagta gctagattgc
      841 atgaaaaagt aatgaatcaa cgtactgatt ttctgaataa gttgagtaca gaaatgatca
      901 aaaatcacga tattatctgt attgaagact taaacacaaa aggtatgttg cgaaatcata
      961 aattagctaa aagtatttct gatgtttctt ggtcaaagtt tgtgactaaa ttacaataca
     1021 aggctgactg gtatggacga aaaatcatca aagtagataa atggtttccg tctagtcaga
     1081 tttgttcaga gtgcggacat aaagatggca agaaatctct tgaaattcga gaatggactt
     1141 gtcctgtttg tcatactcat catgatcgtg atatcaatgc tagtatcaat attctgaccg
     1201 aaggtctaag actacattct atgggattgg ctggatcctg agagatcttt gacagctagc
     1261 tcagtcctag gtataatact agtgtatcaa tattctgacc gaaggtctaa gactacattc
     1321 tatgggattg gcttagatcc aatcatcaag aaccgtagga actacgggga tagcttggta
     1381 aataagagac accgctgtta gtaaagaaat acgctatcaa gtatgctcta ttcccaagaa
     1441 gctcccactt caagcgttaa gaaccgtcag gtttagctaa gtggtgagta gttcacgctc
     1501 ggagatcatc attgcgtttt ttaagcttgg ctgttttggc ggatgagaga agattttcag
     1561 cctgatacag attaaatcag aacgcagaag cggtctgata aaacagaatt tgcctggcgg
     1621 cagtagcgcg gtggtcccac ctgaccccat gccgaactca gaagtgaaac gccgtagcgc
     1681 cgatggtagt gtggggtctc cccatgcgag agtagggaac tgccaggcat caaataaaac
     1741 gaaaggctca gtcgaaagac tgggcctttc gttttatctg ttgtttgtcg gtgaacgctc
     1801 tcctgagtag gacaaatccg ccgggagcgg atttgaacgt tgcgaagcaa cggcccggag
     1861 ggtggcgggc aggacgcccg ccataaactg ccaggcatca aattaagcag aaggccatcc
     1921 tgacggatgg cctttttgcg tttctacaaa ctcttttgtt tatttttcta aatacattca
     1981 aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata ttgaaaaagg
     2041 aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc ggcattttgc
     2101 cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga agatcagttg
     2161 ggtgcagcaa actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag
     2221 actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct
     2281 ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac
     2341 tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa
     2401 ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt
     2461 aactgtcaga ccaagtttac tcatatatac tttagattga tttacgcgcc ctgtagcggc
     2521 gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga ccgctacact tgccagcgcc
     2581 ctagcgcccg ctcctttcgc tttcttccct tcctttctcg ccacgttcgc cggctttccc
     2641 cgtcaagctc taaatcgggg gctcccttta gggttccgat ttagtgcttt acggcacctc
     2701 gaccccaaaa aacttgattt gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg
     2761 gtttttcgcc ctttgacgtt ggagtccacg ttctttaata gtggactctt gttccaaact
     2821 ggaacaacac tcaaccctat ctcgggctat tcttttgatt tataagggat tttgccgatt
     2881 tcggcctatt ggttaaaaaa tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa
     2941 atattaacgt ttacaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     3001 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     3061 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     3121 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     3181 aactggcttc agcagagcgc agataccaaa tactgtcctt ctagtgtagc cgtagttagg
     3241 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     3301 agtcaggcat ttgagaagca cacggtcaca ctgcttccgg tagtcaataa accggtaaac
     3361 cagcaataga cataagcggc tatttaacga ccctgccctg aaccgacgac cgggtcgaat
     3421 ttgctttcga atttctgcca ttcatccgct tattatcact tattcaggcg tagcaaccag
     3481 gcgtttaagg gcaccaataa ctgccttaaa aaaattacgc cccgccctgc cactcatcgc
     3541 agtactgttg taattcatta agcattctgc cgacatggaa gccatcacaa acggcatgat
     3601 gaacctgaat cgccagcggc atcagcacct tgtcgccttg cgtataatat ttgcccatgg
     3661 tgaaaacggg ggcgaagaag ttgtccatat tggccacgtt taaatcaaaa ctggtgaaac
     3721 tcacccaggg attggctgag acgaaaaaca tattctcaat aaacccttta gggaaatagg
     3781 ccaggttttc accgtaacac gccacatctt gcgaatatat gtgtagaaac tgccggaaat
     3841 cgtcgtggta ttcactccag agcgatgaaa acgtttcagt ttgctcatgg aaaacggtgt
     3901 aacaagggtg aacactatcc catatcacca gctcaccgtc tttcattgcc atacggaact
     3961 ccggatgagc attcatcagg cgggcaagaa tgtgaataaa ggccggataa aacttgtgct
     4021 tatttttctt tacggtcttt aaaaaggccg taatatccag ctgaacggtc tggttatagg
     4081 tacattgagc aactgactga aatgcctcaa aatgttcttt acgatgccat tgggatatat
     4141 caacggtggt atatccagtg atttttttct ccattttagc ttccttagct cctgaaaatc
     4201 tcgataactc aaaaaatacg cccggtagtg atcttatttc attatggtga aagttggaac
     4261 ctcttacgtg ccgatcaacg tctcattttc gccaaaagtt ggcccagggc ttcccggtat
     4321 caacagggac accaggattt atttattctg cgaagtgatc ttccgtcaca ggtatttatt
     4381 cggcgcaaag tgcgtcgggt gatgctgcca acttactgat ttagtgtatg atggtgtttt
     4441 tgaggtgctc cagtggcttc tgtttctatc agctgtccct cctgttcagc tactgacggg
     4501 gtggtgcgta acggcaaaag caccgccgga catcagcgct agcggagtgt atactggctt
     4561 actatgttgg cactgatgag ggtgtcagtg aagtgcttca tgtggcagga gaaaaaaggc
     4621 tgcaccggtg cgtcagcaga atatgtgata caggatatat tccgcttcct cgctcactga
     4681 ctcgctacgc tcggtcgttc gactgcggcg agcggaaatg gcttacgaac ggggcggaga
     4741 tttcctggaa gatgccagga agatacttaa cagggaagtg agagggccgc ggcaaagccg
     4801 tttttccata ggctccgccc ccctgacaag catcacgaaa tctgacgctc aaatcagtgg
     4861 tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggcgg ctccctcgtg
     4921 cgctctcctg ttcctgcctt tcggtttacc ggtgtcattc cgctgttatg gccgcgtttg
     4981 tctcattcca cgcctgacac tcagttccgg gtaggcagtt cgctccaagc tggactgtat
     5041 gcacgaaccc cccgttcagt ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     5101 caacccggaa agacatgcaa aagcaccact ggcagcagcc actggtaatt gatttagagg
     5161 agttagtctt gaagtcatgc gccggttaag gctaaactga aaggacaagt tttggtgact
     5221 gcgctcctcc aagccagtta cctcggttca aagagttggt agctcagaga accttcgaaa
     5281 aaccgccctg caaggcggtt ttttcgtttt cagagcaaga gattacgcgc agaccaaaac
     5341 gatctcaaga agatcatctt attaatcaga taaaatattt ctaggctcat gagcccgaag
     5401 tggcgagccc gatcttcccc atcggtgatg tcggcgatat aggcgccagc aaccgcacct
     5461 gtggcgccgg tgatgccggc cacgatgcgt ccggcgtaga ggatctgctc atgtt
//
