LOCUS       sequence-201852-        6763 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses TALE targeted to lac operator with 3 TEV cut sites in its 
            repeat domain, anhydrotetracycline inducible TEV protease.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6763)
  AUTHORS   Copeland MF, Politz MC, Johnson CB, Markley AL, Pfleger BF
  TITLE     A transcription activator-like effector (TALE) induction system 
            mediated by proteolysis.
  JOURNAL   Nat Chem Biol. 2016 Apr;12(4):254-60. doi: 10.1038/nchembio.2021. 
            Epub 2016 Feb 8.
  PUBMED    26854666
REFERENCE   2  (bases 1 to 6763)
  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"; doi: 
            "10.1038/nchembio.2021"; journalName: "Nat Chem Biol"; date: 
            "2016-04"; volume: "12"; issue: "4"; pages: "254-60"
COMMENT     Sequence Label: p5A1-T
FEATURES             Location/Qualifiers
     source          1..6763
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    1..22
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     promoter        23..57
                     /label=BBa_J23102
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.86"
     CDS             101..118
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             935..955
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1466..1486
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1997..2017
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2669..2692
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     terminator      2730..2761
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     CDS             complement(2778..3593)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNGDRVFRLAQAQSRMNNGLVGA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     3501..3520
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     CDS             complement(3683..4348)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRRT"
     terminator      complement(5008..5079)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(5124..5750)
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor TetR"
                     /label=TetR
                     /note="TetR binds to the tetracycline operator tetO to 
                     inhibit transcription. This inhibition can be relieved by 
                     adding tetracycline or doxycycline."
                     /translation="MMSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWH
                     VKNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGT
                     RPTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPT
                     TDSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     primer_bind     5655..5674
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        5766..5821
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    5802..5820
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             5862..5882
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=7xHis
                     /translation="HHHHHHH"
     CDS             5883..6590
                     /codon_start=1
                     /product="tobacco etch virus protease (Kapust et al., 
                     2001)"
                     /label=TEV protease
                     /translation="GESLFKGPRDYNPISSTICHLTNESDGHTTSLYGIGFGPFIITNK
                     HLFRRNNGTLLVQSLHGVFKVKNTTTLQQHLIDGRDMIIIRMPKDFPPFPQKLKFREPQ
                     REERICLVTTNFQTKSMSSMVSDTSCTFPSSDGIFWKHWIQTKDGQCGSPLVSTRDGFI
                     VGIHSASNFTNTNNYFTSVPKNFMELLTNQEAQQWVSGWRLNADSVLWGGHKVFMVKPE
                     EPFQPVKEATQLMN"
     terminator      6677..6713
                     /label=soxR terminator
                     /note="bidirectional E. coli soxR transcription terminator"
ORIGIN
        1 gaattcgcgg ccgcttctag agttgacagc tagctcagtc ctaggtactg tgctagctac
       61 tagatatcgc tcaatactga ccaaggaggt atctacaatg catcaccatc atcaccatgt
      121 tgatctgcgt accctgggtt atagccagca gcagcaagaa aaaatcaaac cgaaagttcg
      181 tagcaccgtt gcacagcatc atgaagcact ggttggtcat ggttttaccc atgcacatat
      241 tgttgcactg agccagcatc cggcagcact gggcaccgtt gcagttaaat atcaggatat
      301 gattgcagca ctgccggaag caacccatga agcaattgtt ggtgttggta aacagtggtc
      361 aggtgcacgt gccctggaag cactgctgac cgttgccggt gaactgcgtg gtccgcctct
      421 gcagctggat accggtcagc tgctgaaaat tgcaaaacgt ggtggtgtta ccgcagttga
      481 agcagttcat gcatggcgta atgcactgac cggtgcaccg ctgaatctga caccggaaca
      541 ggttgttgca attgccagca atggtggtgg caaacaggca ctggaaaccg ttcagcgtct
      601 gctgccggtt ctgtgtcagg cacatggtct gacccctgaa caggtggtgg ccattgcaag
      661 caataatggc ggtaaacaag ccctggaaac agtgcagcgc ctgttaccgg tgctgtgcca
      721 ggcccatggc ttaactccgg aacaggtggt agcgatcgca tcaaatggcg gaggtaaaca
      781 ggccttagaa accgtacagc gcttactgcc ggtgttatgc caggcgcacg gcctgacgcc
      841 agaacaggta gtggcaatcg cctcaaataa tggtggaaaa caggcgttag agacagtcca
      901 gcgcctgctg cctgtattat gtcaagccca tggcgaaaac ctgtattttc aaggcctgac
      961 cccagagcaa gttgttgcga ttgcaagtaa cattgggggt aaacaggcac ttgagacagt
     1021 tcaacgttta ctgcctgtac tgtgccaagc tcacggtctg actccggaac aagtcgtcgc
     1081 gattgcgagt aataacggtg gcaaacaagc attagaaacg gtgcaacgcc tgctgccagt
     1141 tctttgccag gctcacggtt taacccctga gcaggttgta gctattgcga gtcatgatgg
     1201 tggtaagcag gcgttggaaa ctgtgcaaag actgctgccc gtgttgtgcc aagcacatgg
     1261 tttaacccca gaacaagtcg tagcaatcgc aagcaatcat ggtggcaagc aagcgcttga
     1321 aacagtacag cgtttattac cggtactttg tcaggcccac ggtcttacac cagaacaagt
     1381 tgtggccata gccagtaaca atggcggaaa gcaggctctg gaaacggtac aacgtctgtt
     1441 acctgttctg tgtcaagcgc acggagaaaa tttatacttt cagggtttaa cacctgaaca
     1501 agtagttgcc attgcgtcaa atatcggagg caagcaggcc ttggagacag tgcagagatt
     1561 actgccagtg ttgtgtcagg ctcatggcct tacacccgag caggtcgtgg caattgcatc
     1621 taatggtggc ggtaagcaag ctttagagac tgttcagaga ctgcttcctg tcctgtgcca
     1681 ggcacacgga cttacgcctg agcaagtggt tgcaatcgcc tctaacatcg gtggtaagca
     1741 agcactggaa actgtccaac gcttacttcc ggtgctttgt caagcacacg gcttaacgcc
     1801 agagcaggtc gtcgccatag ccagcaatat cggtggtaaa caggcccttg aaacggtcca
     1861 aagacttctg ccggtccttt gccaagcgca tgggctgaca cctgagcagg tagtcgcgat
     1921 tgcctcacat gacggtggga agcaggcatt agaaacagtt caaagattat taccagtcct
     1981 gtgtcaggcg catggggaga acttgtattt ccagggctta accccagagc aggtagttgc
     2041 aatagcatcc aacattggcg gaaaacaagc gttggaaacg gttcagcggt tattgcctgt
     2101 tttgtgccag gcgcatggtt tgacacccga gcaagtggta gccatagcct caaatatagg
     2161 gggtaaacaa gctttggaga cagtacaacg gctgcttcca gttttatgtc aggcccatgg
     2221 attgacgcct gaacaagttg tcgctatcgc aagtaatggg ggtggtaaac aagcgcttga
     2281 aaccgttcaa cgccttctgc ctgtgctttg tcaggcacat ggattaacac ccgaacaggt
     2341 tgtcgcgata gcttcaaacg gtggtggtcg tccggcactg gaaagcattg ttgcacagct
     2401 gagccgtccg gatccggcac tggcagcact gaccaatgat catctggttg cactggcatg
     2461 tctgggtggt cgccctgccc tggatgcagt taaaaaaggt ctgccgcatg caccggcact
     2521 gattaaacgt accaatcgtc gtattccgga acgtaccagc catcgtgttg cagatcatgc
     2581 acaggttgtt cgtgttctgg gtttttttca gtgtcatagc catccggcac aggcatttga
     2641 tgatgcaatg acccagtttg gtatgagcga ttataaagat gatgatgata aataaggtac
     2701 cctaatcata cctgacctcc atagcagaaa gtcaaaaacc tccgaccgga ggcttttgac
     2761 ttgaggggga tccgccctca gaaaaactca tcgagcatca agtgaaactg caatttattc
     2821 atatcaggat tatcaatacc atatttttga aaaagccgtt tctgtaatga aggagaaaac
     2881 tcaccgaggc agttccatag gatggcaaga tcctggtatc ggtctgcgat tccgactcgt
     2941 ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa
     3001 tcaccatgag tgacgactga atccggtgag aatggcaaaa gcttatgcat ttctttccag
     3061 acttgttcaa caggccagcc attacgctcg tcatcaaaat cactcgcacc aaccaaaccg
     3121 ttattcattc gtgattgcgc ctgagcgaga cgaaatacgc gatcgccgtt aaaaggacaa
     3181 ttacaaacag gaatcgaatg caaccggcgc aggaacactg ccagcgcatc aacaatattt
     3241 tcacctgaat caggatattc ttctaatacc tggaatgctg ttttccctgg gatcgcagtg
     3301 gtgagtaacc atgcatcatc aggagtacgg ataaaatgct tgatggtcgg aagaggcata
     3361 aattccgtca gccagtttag cctgaccatc tcatctgtaa catcattggc aacgctacct
     3421 ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc catacaatcg atagattgtc
     3481 gcacctgatt gcccgacatt atcgcgagcc catttatacc catataaatc agcatccatg
     3541 ttggaattta atcgcggcct cgagcaagac gtttcccgtt gaatatggct cattttagct
     3601 tccttagctc ctgaaaatct cgataactca aaaaatacgc ccggtagtga tcttatttca
     3661 ttatggtgaa agttggaacc tcttacgtac ggcgcggcag cgtgacccgt gtcggcggct
     3721 ccaacggctc gccatcgtcc agaaaacacg gctcatcggg catcggcagg cgctgctgcc
     3781 cgcgtcgttc ccattcctcc gtttcggtca aggctggcag gtctggttcc atgcccggaa
     3841 tgccgggctg gctgggcggc tcctcgccgg ggccggtcgg tagttgctgc tcgcccggat
     3901 acagggtcgg gatgcggcgc aggtcgccat gccccaacag cgattcgtcc tggtcgtcgt
     3961 gatcaaccac cacggcggca ctgaacaccg acaggcgcaa ctggtcgcgg ggctggcccc
     4021 acgccacgcg gtcattgacc acgtaggccg acacggtgcc ggggccgttg agcttcacga
     4081 cggagatcca gcgctcggcc accaagtcct tgactgcgta ttggaccgtc cgcaaagaac
     4141 gtccgatgag cttggaaagt gtcttctggc tgaccaccac ggcgttctgg tggcccatct
     4201 gcgccacgag gtgatgcagc agcattgccg ccgtgggttt cctcgcaata agcccggccc
     4261 acgcctcatg cgctttgcgt tccgtttgca cccagtgacc gggcttgttc ttggcttgaa
     4321 tgccgatttc tctggactgc gtggccatgc ttatctccat gcggtagggg tgccgcacgg
     4381 ttgcggcacc atgcgcaatc agctgcaact tttcggcagc gcgacaacaa ttatgcgttg
     4441 cgtaaaagtg gcagtcaatt acagattttc tttaacctac gcaatgagct attgcggggg
     4501 gtgccgcaat gagctgttgc gtacccccct tttttaagtt gttgattttt aagtctttcg
     4561 catttcgccc tatatctagt tctttggtgc ccaaagaagg gcacccctgc ggggttcccc
     4621 cacgccttcg gcgcggctcc ccctccggca aaaagtggcc cctccggggc ttgttgatcg
     4681 actgcgcggc cttcggcctt gcccaaggtg gcgctgcccc cttggaaccc ccgcactcgc
     4741 cgccgtgagg ctcggggggc aggcgggcgg gcttcgccct tcgactgccc ccactcgcat
     4801 aggcttgggt cgttccaggc gcgtcaaggc caagccgctg cgcggtcgct gcgcgagcct
     4861 tgacccgcct tccacttggt gtccaaccgg caagcgaagc gcgcaggccg caggccggag
     4921 gcttttcccc agagaaaatt aaaaaaattg atggggcaag gccgcaggcc gcgcagttgg
     4981 agccggtggg tatgtggtcg aaggctgggt agcgttcacc gacaaacaac agataaaacg
     5041 aaaggcccag tctttcgact gagcctttcg ttttatttga tgcctggctc tagtagtgat
     5101 ctacactagc actatcagtg ttattaagac ccactttcac atttaagttg tttttctaat
     5161 ccgcatatga tcaattcaag gccgaataag aaggctggct ctgcaccttg gtgatcaaat
     5221 aattcgatag cttgtcgtaa taatggcggc atactatcag tagtaggtgt ttccctttct
     5281 tctttagcga cttgatgctc ttgatcttcc aatacgcaac ctaaagtaaa atgccccaca
     5341 gcgctgagtg catataatgc attctctagt gaaaaacctt gttggcataa aaaggctaat
     5401 tgattttcga gagtttcata ctgtttttct gtaggccgtg tacctaaatg tacttttgct
     5461 ccatcgcgat gacttagtaa agcacatcta aaacttttag cgttattacg taaaaaatct
     5521 tgccagcttt ccccttctaa agggcaaaag tgagtatggt gcctatctaa catctcaatg
     5581 gctaaggcgt cgagcaaagc ccgcttattt tttacatgcc aatacaatgt aggctgctct
     5641 acacctagct tctgggcgag tttacgggtt gttaaacctt cgattccgac ctcattaagc
     5701 agctctaatg cgctgttaat cactttactt ttatctaatc tagacatcat taattcctaa
     5761 tttttgttga cactctatcg ttgatagagt tattttacca ctccctatca gtgatagaga
     5821 aaagaattca aaagatcttt taagaaggag atatacatat gcatcatcat catcatcatc
     5881 atggagaaag cttgtttaag gggccgcgtg attacaaccc gatatcgagc accatttgtc
     5941 atttgacgaa tgaatctgat gggcacacaa catcgttgta tggtattgga tttggtccct
     6001 tcatcattac aaacaagcac ttgtttagaa gaaataatgg aacactgttg gtccaatcac
     6061 tacatggtgt attcaaggtc aagaacacca cgactttgca acaacacctc attgatggga
     6121 gggacatgat aattattcgc atgcctaagg atttcccacc atttcctcaa aagctgaaat
     6181 ttagagagcc acaaagggaa gagcgcatat gtcttgtgac aaccaacttc caaactaaga
     6241 gcatgtctag catggtgtca gacactagtt gcacattccc ttcatctgat ggcatattct
     6301 ggaagcattg gattcaaacc aaggatgggc agtgtggcag tccattagta tcaactagag
     6361 atgggttcat tgttggtata cactcagcat cgaatttcac caacacaaac aattatttca
     6421 caagcgtgcc gaaaaacttc atggaattgt tgacaaatca ggaggcgcag cagtgggtta
     6481 gtggttggcg attaaatgct gactcagtat tgtggggggg ccataaagtt ttcatggtga
     6541 aacctgaaga gccttttcag ccagttaagg aagcgactca actcatgaat cgtcgtcgcc
     6601 gtcgctaagt ccatcagctt gtccagcagg gttgtccacg ggccgagcga agcgagccag
     6661 ccggtggccg cggaacaaaa ctaaagcgcc acaagggcgc tttagtttgt tttcagtcca
     6721 gttacgctgg agtctgaggc tcgtcctgaa tgatatcaag ctt
//
