LOCUS       sequence-410877-        5486 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bacterial expression of ISVgsp2 TnpB and its reRNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5486)
  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 5486)
  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_ISVgsp2_reRNA
FEATURES             Location/Qualifiers
     source          1..5486
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        16..50
                     /label=BBa_J23108
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.51"
     RBS             66..77
                     /note="strong bacterial ribosome binding site (Elowitz and 
                     Leibler, 2000)"
     primer_bind     complement(1529..1546)
                     /label=pBAD Reverse
                     /note="For vectors with E. coli araBAD promoter, reverse 
                     primer"
     terminator      1699..1785
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     terminator      1877..1904
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     promoter        1923..2014
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      2472..2927
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2559..2578)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2769..2790
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             complement(3483..4142)
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     4057..4076
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     promoter        complement(4143..4245)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     primer_bind     complement(4637..4654)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4771..5316)
                     /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(4851..4870)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5392..5411
                     /label=pBRrevBam
                     /note="pBR322 vectors, tet region, downstream of BamHI, 
                     reverse primer"
ORIGIN
        1 cagcttatca tcgatctgac agctagctca gtcctaggta taatgctagc cccgggaaaa
       61 gatctaaaga ggagaaaatg ttacgacata aagcgtataa atttagagtg tacccgaacg
      121 tcgagcagga aatcttaatc aataaaacaa tcgggtcatc tagatttgtc tataattata
      181 tgttaaatgc ttggaagaca tcctttaagg aaacgggtaa aggtttatct tatggcaagt
      241 gttctgccat gttgccttct atgaaaaaat cagaagagac atcatggtta aaagaagttg
      301 atagtatcgc gttacaatca tctattaaga atttagctga atcatttgat cggtttttca
      361 acaaacaaac tcaatcccct aagtttaaaa gcaagaaaaa tcctgttcaa agttatacca
      421 caaaaaacac aaacaataat gtggctattg taggcaataa ggtgaaatta cctaagctag
      481 gattggttaa attttctaag agtcgtgacg ttaatggtcg tattttaagt gccactgtcc
      541 gtagaaatgc cagtggtaaa tttttcgtat cgttacaatg tgaagaggaa ataactgagt
      601 tcccaacagc aaacaaaaca gtggggattg atctaggtat tacagatttt gcgattcttt
      661 ctaatggtca aaaaatcact aacaataaat tcgcagctaa aatggaaaag aaactaaagc
      721 gtgaacaacg gaagttatca aaacgggcat cattagctaa acagaacggg attaagttgt
      781 ttgaagctaa aaattaccaa aaacaaaagc gaaaagtagc tagactgtct gaaaaagtaa
      841 tgaatcaacg cagtgacttt ctaaataagt tgagtacaga aatcatcaaa aatcacgata
      901 ttatttgtat tgaagactta agtacaaaag gaatgttgcg taatcataag ctagccaaaa
      961 gtatatccga tgtctcttgg tccagttttg tgactaactt acagtacaag gctgactggt
     1021 atgggcgtga aataatcaaa gtagacaaat ggtttgcatc tagtcaaatt tgttcagaat
     1081 gtgcccataa agacggcaag aaatctcttg aaatgagaga atggacttgt cctatttgtc
     1141 atactcatca cgatcgagat attaatgcca gtgttaatat cttgaccgaa ggtctaagat
     1201 tacaagcttt agcctgagag atctttgaca gctagctcag tcctaggtat aatagtcata
     1261 ctcatcacga tcgagatatt aatgccagtg ttaatatctt gaccgaaggt ctaagattac
     1321 aagctttagc ctagataaac tcaataaccg taggaactgc ggggatagct tggtaaataa
     1381 aagacaccgc tgatagtaaa gaaatacgct atcaagtatg cttttttccc aagaatcttg
     1441 tgactttagt catgagaggt tcaagctcgg agatcatcat tgcgtttttt aagcttggct
     1501 gttttggcgg atgagagaag attttcagcc tgatacagat taaatcagaa cgcagaagcg
     1561 gtctgataaa acagaatttg cctggcggca gtagcgcggt ggtcccacct gaccccatgc
     1621 cgaactcaga agtgaaacgc cgtagcgccg atggtagtgt ggggtctccc catgcgagag
     1681 tagggaactg ccaggcatca aataaaacga aaggctcagt cgaaagactg ggcctttcgt
     1741 tttatctgtt gtttgtcggt gaacgctctc ctgagtagga caaatccgcc gggagcggat
     1801 ttgaacgttg cgaagcaacg gcccggaggg tggcgggcag gacgcccgcc ataaactgcc
     1861 aggcatcaaa ttaagcagaa ggccatcctg acggatggcc tttttgcgtt tctacaaact
     1921 cttttgttta tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg
     1981 ataaatgctt caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc
     2041 ccttattccc ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt
     2101 gaaagtaaaa gatgctgaag atcagttggg tgcagcaaac tattaactgg cgaactactt
     2161 actctagctt cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca
     2221 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     2281 cgtgggtctc gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta
     2341 gttatctaca cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag
     2401 ataggtgcct cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt
     2461 tagattgatt tacgcgccct gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg
     2521 cagcgtgacc gctacacttg ccagcgccct agcgcccgct cctttcgctt tcttcccttc
     2581 ctttctcgcc acgttcgccg gctttccccg tcaagctcta aatcgggggc tccctttagg
     2641 gttccgattt agtgctttac ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc
     2701 acgtagtggg ccatcgccct gatagacggt ttttcgccct ttgacgttgg agtccacgtt
     2761 ctttaatagt ggactcttgt tccaaactgg aacaacactc aaccctatct cgggctattc
     2821 ttttgattta taagggattt tgccgatttc ggcctattgg ttaaaaaatg agctgattta
     2881 acaaaaattt aacgcgaatt ttaacaaaat attaacgttt acaatttaaa aggatctagg
     2941 tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact
     3001 gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg
     3061 taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc
     3121 aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata
     3181 ctgtccttct agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta
     3241 catacctcgc tctgctaatc ctgttaccag tcaggcattt gagaagcaca cggtcacact
     3301 gcttccggta gtcaataaac cggtaaacca gcaatagaca taagcggcta tttaacgacc
     3361 ctgccctgaa ccgacgaccg ggtcgaattt gctttcgaat ttctgccatt catccgctta
     3421 ttatcactta ttcaggcgta gcaaccaggc gtttaagggc accaataact gccttaaaaa
     3481 aattacgccc cgccctgcca ctcatcgcag tactgttgta attcattaag cattctgccg
     3541 acatggaagc catcacaaac ggcatgatga acctgaatcg ccagcggcat cagcaccttg
     3601 tcgccttgcg tataatattt gcccatggtg aaaacggggg cgaagaagtt gtccatattg
     3661 gccacgttta aatcaaaact ggtgaaactc acccagggat tggctgagac gaaaaacata
     3721 ttctcaataa accctttagg gaaataggcc aggttttcac cgtaacacgc cacatcttgc
     3781 gaatatatgt gtagaaactg ccggaaatcg tcgtggtatt cactccagag cgatgaaaac
     3841 gtttcagttt gctcatggaa aacggtgtaa caagggtgaa cactatccca tatcaccagc
     3901 tcaccgtctt tcattgccat acggaactcc ggatgagcat tcatcaggcg ggcaagaatg
     3961 tgaataaagg ccggataaaa cttgtgctta tttttcttta cggtctttaa aaaggccgta
     4021 atatccagct gaacggtctg gttataggta cattgagcaa ctgactgaaa tgcctcaaaa
     4081 tgttctttac gatgccattg ggatatatca acggtggtat atccagtgat ttttttctcc
     4141 attttagctt ccttagctcc tgaaaatctc gataactcaa aaaatacgcc cggtagtgat
     4201 cttatttcat tatggtgaaa gttggaacct cttacgtgcc gatcaacgtc tcattttcgc
     4261 caaaagttgg cccagggctt cccggtatca acagggacac caggatttat ttattctgcg
     4321 aagtgatctt ccgtcacagg tatttattcg gcgcaaagtg cgtcgggtga tgctgccaac
     4381 ttactgattt agtgtatgat ggtgtttttg aggtgctcca gtggcttctg tttctatcag
     4441 ctgtccctcc tgttcagcta ctgacggggt ggtgcgtaac ggcaaaagca ccgccggaca
     4501 tcagcgctag cggagtgtat actggcttac tatgttggca ctgatgaggg tgtcagtgaa
     4561 gtgcttcatg tggcaggaga aaaaaggctg caccggtgcg tcagcagaat atgtgataca
     4621 ggatatattc cgcttcctcg ctcactgact cgctacgctc ggtcgttcga ctgcggcgag
     4681 cggaaatggc ttacgaacgg ggcggagatt tcctggaaga tgccaggaag atacttaaca
     4741 gggaagtgag agggccgcgg caaagccgtt tttccatagg ctccgccccc ctgacaagca
     4801 tcacgaaatc tgacgctcaa atcagtggtg gcgaaacccg acaggactat aaagatacca
     4861 ggcgtttccc cctggcggct ccctcgtgcg ctctcctgtt cctgcctttc ggtttaccgg
     4921 tgtcattccg ctgttatggc cgcgtttgtc tcattccacg cctgacactc agttccgggt
     4981 aggcagttcg ctccaagctg gactgtatgc acgaaccccc cgttcagtcc gaccgctgcg
     5041 ccttatccgg taactatcgt cttgagtcca acccggaaag acatgcaaaa gcaccactgg
     5101 cagcagccac tggtaattga tttagaggag ttagtcttga agtcatgcgc cggttaaggc
     5161 taaactgaaa ggacaagttt tggtgactgc gctcctccaa gccagttacc tcggttcaaa
     5221 gagttggtag ctcagagaac cttcgaaaaa ccgccctgca aggcggtttt ttcgttttca
     5281 gagcaagaga ttacgcgcag accaaaacga tctcaagaag atcatcttat taatcagata
     5341 aaatatttct aggctcatga gcccgaagtg gcgagcccga tcttccccat cggtgatgtc
     5401 ggcgatatag gcgccagcaa ccgcacctgt ggcgccggtg atgccggcca cgatgcgtcc
     5461 ggcgtagagg atctgctcat gtttga
//
