LOCUS       sequence-411096-        5449 bp    DNA     circular SYN 15-APR-2025
DEFINITION  Bacterial expression of ISAepa1 TnpB and its reRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5449)
  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 5449)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 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_ISAepa1_reRNA
FEATURES             Location/Qualifiers
     source          1..5449
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        8..42
                     /label=BBa_J23108
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.51"
     RBS             58..69
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     primer_bind     complement(1484..1501)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1654..1740
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1832..1859
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1878..1969
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2427..2882
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2514..2533)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2724..2745
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3438..4097)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     4012..4031
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(4098..4200)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(4592..4609)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4726..5271)
                     /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(4806..4825)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5347..5366
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 catcgatctg acagctagct cagtcctagg tataatgcta gccccgggaa aagatctaaa
       61 gaggagaaaa tggccaaaca aaacaaagcc ttcaaatttc gtttgtatcc gaataaagaa
      121 cagaaaatcc aattagccaa aacatttggt tgtgttcgct ttgtctacaa taagatgctg
      181 gaagaacgta aggaaacgta tgagaaattc aaaaacgata aagaaacgct taaacaacaa
      241 gcattcccga caccagcgaa atacaaagat gaatttccat tcctaaaaga agtggattct
      301 ttagctttag caaacgcaca gatcaatttg caaaatgctt ataagaactt ttttgaaggt
      361 cgtgcggagt ttcctaaatt caaaagcaag aaaaataaac aatcttacac aaccaatatg
      421 gtcaacggca atattaagct agaaaatggt catatcaaac taccaaaaat caaaaagcca
      481 atgaaaatga accgacatag agaaatacct gctgattata aaatcaaatc ttgtactatt
      541 tctaaaacaa agacaggtaa atattatatt tcgattttga cagaatacga gaaagatatt
      601 cagtcagtaa agattcaaaa agtagttggc ttagattttg ctatggacgg tctatatgtc
      661 gagagtgaac agggtgagaa agccaattac ccacggtact atcgacaagc attagacaaa
      721 ctggcaaaag cacaacgaat cctttctcgc agaaaaaaag gttctgcacg ttgggaaaaa
      781 caacgtttga aagtagcaaa gttacaagag aaaattgcca atcaacgcaa gaattttctt
      841 caccataaat caaaagaatt agcatcaacc tatgacgcag ttatcattga ggacttagac
      901 atgaaaggaa tgtctcaagc cctcaatttt ggaaaaagtg ttcatgataa tggctggggt
      961 atgttcacta ctttcttaga atacaagcta aaagaacaag ggaaacaact tgtaaaaatc
     1021 gataagtggt tcccttctac taagaaatgt tcttgttgtg gcaaggaaaa agaaatgcca
     1081 ttatctaaac gtgtatatga atgttcttgc ggttttgtat tagatcgtga tcacaattca
     1141 gcaatcaata tcaaaaatga aggattacgc atattagcat gagagatctt tgacagctag
     1201 ctcagtccta ggtataatag ttcttgcggt tttgtattag atcgtgatca caattcagca
     1261 atcaatatca aaaatgaagg attacgcata ttagcataaa gaataataac tttcggaacg
     1321 agagggtaag ctcggtcaat ttcttatcgt tagataggat tacccgagaa gcccccactt
     1381 caagcgaccc gcaagggtgg taagtggagg gtagttcacg ctcggagatc atcattgcgt
     1441 tttttaagct tggctgtttt ggcggatgag agaagatttt cagcctgata cagattaaat
     1501 cagaacgcag aagcggtctg ataaaacaga atttgcctgg cggcagtagc gcggtggtcc
     1561 cacctgaccc catgccgaac tcagaagtga aacgccgtag cgccgatggt agtgtggggt
     1621 ctccccatgc gagagtaggg aactgccagg catcaaataa aacgaaaggc tcagtcgaaa
     1681 gactgggcct ttcgttttat ctgttgtttg tcggtgaacg ctctcctgag taggacaaat
     1741 ccgccgggag cggatttgaa cgttgcgaag caacggcccg gagggtggcg ggcaggacgc
     1801 ccgccataaa ctgccaggca tcaaattaag cagaaggcca tcctgacgga tggccttttt
     1861 gcgtttctac aaactctttt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     1921 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     1981 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     2041 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcag caaactatta
     2101 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     2161 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     2221 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     2281 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     2341 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     2401 tactcatata tactttagat tgatttacgc gccctgtagc ggcgcattaa gcgcggcggg
     2461 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccctagcgc ccgctccttt
     2521 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     2581 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     2641 tttgggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     2701 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaaccc
     2761 tatctcgggc tattcttttg atttataagg gattttgccg atttcggcct attggttaaa
     2821 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat
     2881 ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg
     2941 agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc
     3001 ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg
     3061 tttgtttgcc ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag
     3121 cgcagatacc aaatactgtc cttctagtgt agccgtagtt aggccaccac ttcaagaact
     3181 ctgtagcacc gcctacatac ctcgctctgc taatcctgtt accagtcagg catttgagaa
     3241 gcacacggtc acactgcttc cggtagtcaa taaaccggta aaccagcaat agacataagc
     3301 ggctatttaa cgaccctgcc ctgaaccgac gaccgggtcg aatttgcttt cgaatttctg
     3361 ccattcatcc gcttattatc acttattcag gcgtagcaac caggcgttta agggcaccaa
     3421 taactgcctt aaaaaaatta cgccccgccc tgccactcat cgcagtactg ttgtaattca
     3481 ttaagcattc tgccgacatg gaagccatca caaacggcat gatgaacctg aatcgccagc
     3541 ggcatcagca ccttgtcgcc ttgcgtataa tatttgccca tggtgaaaac gggggcgaag
     3601 aagttgtcca tattggccac gtttaaatca aaactggtga aactcaccca gggattggct
     3661 gagacgaaaa acatattctc aataaaccct ttagggaaat aggccaggtt ttcaccgtaa
     3721 cacgccacat cttgcgaata tatgtgtaga aactgccgga aatcgtcgtg gtattcactc
     3781 cagagcgatg aaaacgtttc agtttgctca tggaaaacgg tgtaacaagg gtgaacacta
     3841 tcccatatca ccagctcacc gtctttcatt gccatacgga actccggatg agcattcatc
     3901 aggcgggcaa gaatgtgaat aaaggccgga taaaacttgt gcttattttt ctttacggtc
     3961 tttaaaaagg ccgtaatatc cagctgaacg gtctggttat aggtacattg agcaactgac
     4021 tgaaatgcct caaaatgttc tttacgatgc cattgggata tatcaacggt ggtatatcca
     4081 gtgatttttt tctccatttt agcttcctta gctcctgaaa atctcgataa ctcaaaaaat
     4141 acgcccggta gtgatcttat ttcattatgg tgaaagttgg aacctcttac gtgccgatca
     4201 acgtctcatt ttcgccaaaa gttggcccag ggcttcccgg tatcaacagg gacaccagga
     4261 tttatttatt ctgcgaagtg atcttccgtc acaggtattt attcggcgca aagtgcgtcg
     4321 ggtgatgctg ccaacttact gatttagtgt atgatggtgt ttttgaggtg ctccagtggc
     4381 ttctgtttct atcagctgtc cctcctgttc agctactgac ggggtggtgc gtaacggcaa
     4441 aagcaccgcc ggacatcagc gctagcggag tgtatactgg cttactatgt tggcactgat
     4501 gagggtgtca gtgaagtgct tcatgtggca ggagaaaaaa ggctgcaccg gtgcgtcagc
     4561 agaatatgtg atacaggata tattccgctt cctcgctcac tgactcgcta cgctcggtcg
     4621 ttcgactgcg gcgagcggaa atggcttacg aacggggcgg agatttcctg gaagatgcca
     4681 ggaagatact taacagggaa gtgagagggc cgcggcaaag ccgtttttcc ataggctccg
     4741 cccccctgac aagcatcacg aaatctgacg ctcaaatcag tggtggcgaa acccgacagg
     4801 actataaaga taccaggcgt ttccccctgg cggctccctc gtgcgctctc ctgttcctgc
     4861 ctttcggttt accggtgtca ttccgctgtt atggccgcgt ttgtctcatt ccacgcctga
     4921 cactcagttc cgggtaggca gttcgctcca agctggactg tatgcacgaa ccccccgttc
     4981 agtccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg gaaagacatg
     5041 caaaagcacc actggcagca gccactggta attgatttag aggagttagt cttgaagtca
     5101 tgcgccggtt aaggctaaac tgaaaggaca agttttggtg actgcgctcc tccaagccag
     5161 ttacctcggt tcaaagagtt ggtagctcag agaaccttcg aaaaaccgcc ctgcaaggcg
     5221 gttttttcgt tttcagagca agagattacg cgcagaccaa aacgatctca agaagatcat
     5281 cttattaatc agataaaata tttctaggct catgagcccg aagtggcgag cccgatcttc
     5341 cccatcggtg atgtcggcga tataggcgcc agcaaccgca cctgtggcgc cggtgatgcc
     5401 ggccacgatg cgtccggcgt agaggatctg ctcatgtttg acagcttat
//
