LOCUS       sequence-190014-        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 catalytically inactive 
            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-Ti
FEATURES             Location/Qualifiers
     source          1..6763
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..18
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             835..855
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1366..1386
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             1897..1917
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     CDS             2569..2592
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     terminator      2630..2661
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     CDS             complement(2678..3493)
                     /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     3401..3420
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     CDS             complement(3583..4248)
                     /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(4908..4979)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(5024..5650)
                     /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     5555..5574
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        5666..5721
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    5702..5720
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             5762..5782
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=7xHis
                     /translation="HHHHHHH"
     CDS             5783..6490
                     /codon_start=1
                     /product="tobacco etch virus protease (Kapust et al., 
                     2001)"
                     /label=TEV protease
                     /translation="GESLFKGPRDYNPISSTICHLTNESDGHTTSLYGIGFGPFIITNK
                     HLFRRNNGTLLVQSLHGVFKVKNTTTLQQHLIDGRDMIIIRMPKDFPPFPQKLKFREPQ
                     REERICLVTTNFQTKSMSSMVSDTSCTFPSSDGIFWKHWIQTKDGQAGSPLVSTRDGFI
                     VGIHSASNFTNTNNYFTSVPKNFMELLTNQEAQQWVSGWRLNADSVLWGGHKVFMVKPE
                     EPFQPVKEATQLMN"
     terminator      6577..6613
                     /label=soxR terminator
                     /note="bidirectional E. coli soxR transcription terminator"
     misc_feature    6664..6685
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     promoter        6686..6720
                     /label=BBa_J23102
                     /note="Anderson constitutive promoter 
                     (https://parts.igem.org/Promoters/Catalog/Anderson). 
                     Measured strength relative to BBa_J223100: 0.86"
ORIGIN
        1 catcaccatc atcaccatgt tgatctgcgt accctgggtt atagccagca gcagcaagaa
       61 aaaatcaaac cgaaagttcg tagcaccgtt gcacagcatc atgaagcact ggttggtcat
      121 ggttttaccc atgcacatat tgttgcactg agccagcatc cggcagcact gggcaccgtt
      181 gcagttaaat atcaggatat gattgcagca ctgccggaag caacccatga agcaattgtt
      241 ggtgttggta aacagtggtc aggtgcacgt gccctggaag cactgctgac cgttgccggt
      301 gaactgcgtg gtccgcctct gcagctggat accggtcagc tgctgaaaat tgcaaaacgt
      361 ggtggtgtta ccgcagttga agcagttcat gcatggcgta atgcactgac cggtgcaccg
      421 ctgaatctga caccggaaca ggttgttgca attgccagca atggtggtgg caaacaggca
      481 ctggaaaccg ttcagcgtct gctgccggtt ctgtgtcagg cacatggtct gacccctgaa
      541 caggtggtgg ccattgcaag caataatggc ggtaaacaag ccctggaaac agtgcagcgc
      601 ctgttaccgg tgctgtgcca ggcccatggc ttaactccgg aacaggtggt agcgatcgca
      661 tcaaatggcg gaggtaaaca ggccttagaa accgtacagc gcttactgcc ggtgttatgc
      721 caggcgcacg gcctgacgcc agaacaggta gtggcaatcg cctcaaataa tggtggaaaa
      781 caggcgttag agacagtcca gcgcctgctg cctgtattat gtcaagccca tggcgaaaac
      841 ctgtattttc aaggcctgac cccagagcaa gttgttgcga ttgcaagtaa cattgggggt
      901 aaacaggcac ttgagacagt tcaacgttta ctgcctgtac tgtgccaagc tcacggtctg
      961 actccggaac aagtcgtcgc gattgcgagt aataacggtg gcaaacaagc attagaaacg
     1021 gtgcaacgcc tgctgccagt tctttgccag gctcacggtt taacccctga gcaggttgta
     1081 gctattgcga gtcatgatgg tggtaagcag gcgttggaaa ctgtgcaaag actgctgccc
     1141 gtgttgtgcc aagcacatgg tttaacccca gaacaagtcg tagcaatcgc aagcaatcat
     1201 ggtggcaagc aagcgcttga aacagtacag cgtttattac cggtactttg tcaggcccac
     1261 ggtcttacac cagaacaagt tgtggccata gccagtaaca atggcggaaa gcaggctctg
     1321 gaaacggtac aacgtctgtt acctgttctg tgtcaagcgc acggagaaaa tttatacttt
     1381 cagggtttaa cacctgaaca agtagttgcc attgcgtcaa atatcggagg caagcaggcc
     1441 ttggagacag tgcagagatt actgccagtg ttgtgtcagg ctcatggcct tacacccgag
     1501 caggtcgtgg caattgcatc taatggtggc ggtaagcaag ctttagagac tgttcagaga
     1561 ctgcttcctg tcctgtgcca ggcacacgga cttacgcctg agcaagtggt tgcaatcgcc
     1621 tctaacatcg gtggtaagca agcactggaa actgtccaac gcttacttcc ggtgctttgt
     1681 caagcacacg gcttaacgcc agagcaggtc gtcgccatag ccagcaatat cggtggtaaa
     1741 caggcccttg aaacggtcca aagacttctg ccggtccttt gccaagcgca tgggctgaca
     1801 cctgagcagg tagtcgcgat tgcctcacat gacggtggga agcaggcatt agaaacagtt
     1861 caaagattat taccagtcct gtgtcaggcg catggggaga acttgtattt ccagggctta
     1921 accccagagc aggtagttgc aatagcatcc aacattggcg gaaaacaagc gttggaaacg
     1981 gttcagcggt tattgcctgt tttgtgccag gcgcatggtt tgacacccga gcaagtggta
     2041 gccatagcct caaatatagg gggtaaacaa gctttggaga cagtacaacg gctgcttcca
     2101 gttttatgtc aggcccatgg attgacgcct gaacaagttg tcgctatcgc aagtaatggg
     2161 ggtggtaaac aagcgcttga aaccgttcaa cgccttctgc ctgtgctttg tcaggcacat
     2221 ggattaacac ccgaacaggt tgtcgcgata gcttcaaacg gtggtggtcg tccggcactg
     2281 gaaagcattg ttgcacagct gagccgtccg gatccggcac tggcagcact gaccaatgat
     2341 catctggttg cactggcatg tctgggtggt cgccctgccc tggatgcagt taaaaaaggt
     2401 ctgccgcatg caccggcact gattaaacgt accaatcgtc gtattccgga acgtaccagc
     2461 catcgtgttg cagatcatgc acaggttgtt cgtgttctgg gtttttttca gtgtcatagc
     2521 catccggcac aggcatttga tgatgcaatg acccagtttg gtatgagcga ttataaagat
     2581 gatgatgata aataaggtac cctaatcata cctgacctcc atagcagaaa gtcaaaaacc
     2641 tccgaccgga ggcttttgac ttgaggggga tccgccctca gaaaaactca tcgagcatca
     2701 agtgaaactg caatttattc atatcaggat tatcaatacc atatttttga aaaagccgtt
     2761 tctgtaatga aggagaaaac tcaccgaggc agttccatag gatggcaaga tcctggtatc
     2821 ggtctgcgat tccgactcgt ccaacatcaa tacaacctat taatttcccc tcgtcaaaaa
     2881 taaggttatc aagtgagaaa tcaccatgag tgacgactga atccggtgag aatggcaaaa
     2941 gcttatgcat ttctttccag acttgttcaa caggccagcc attacgctcg tcatcaaaat
     3001 cactcgcacc aaccaaaccg ttattcattc gtgattgcgc ctgagcgaga cgaaatacgc
     3061 gatcgccgtt aaaaggacaa ttacaaacag gaatcgaatg caaccggcgc aggaacactg
     3121 ccagcgcatc aacaatattt tcacctgaat caggatattc ttctaatacc tggaatgctg
     3181 ttttccctgg gatcgcagtg gtgagtaacc atgcatcatc aggagtacgg ataaaatgct
     3241 tgatggtcgg aagaggcata aattccgtca gccagtttag cctgaccatc tcatctgtaa
     3301 catcattggc aacgctacct ttgccatgtt tcagaaacaa ctctggcgca tcgggcttcc
     3361 catacaatcg atagattgtc gcacctgatt gcccgacatt atcgcgagcc catttatacc
     3421 catataaatc agcatccatg ttggaattta atcgcggcct cgagcaagac gtttcccgtt
     3481 gaatatggct cattttagct tccttagctc ctgaaaatct cgataactca aaaaatacgc
     3541 ccggtagtga tcttatttca ttatggtgaa agttggaacc tcttacgtac ggcgcggcag
     3601 cgtgacccgt gtcggcggct ccaacggctc gccatcgtcc agaaaacacg gctcatcggg
     3661 catcggcagg cgctgctgcc cgcgtcgttc ccattcctcc gtttcggtca aggctggcag
     3721 gtctggttcc atgcccggaa tgccgggctg gctgggcggc tcctcgccgg ggccggtcgg
     3781 tagttgctgc tcgcccggat acagggtcgg gatgcggcgc aggtcgccat gccccaacag
     3841 cgattcgtcc tggtcgtcgt gatcaaccac cacggcggca ctgaacaccg acaggcgcaa
     3901 ctggtcgcgg ggctggcccc acgccacgcg gtcattgacc acgtaggccg acacggtgcc
     3961 ggggccgttg agcttcacga cggagatcca gcgctcggcc accaagtcct tgactgcgta
     4021 ttggaccgtc cgcaaagaac gtccgatgag cttggaaagt gtcttctggc tgaccaccac
     4081 ggcgttctgg tggcccatct gcgccacgag gtgatgcagc agcattgccg ccgtgggttt
     4141 cctcgcaata agcccggccc acgcctcatg cgctttgcgt tccgtttgca cccagtgacc
     4201 gggcttgttc ttggcttgaa tgccgatttc tctggactgc gtggccatgc ttatctccat
     4261 gcggtagggg tgccgcacgg ttgcggcacc atgcgcaatc agctgcaact tttcggcagc
     4321 gcgacaacaa ttatgcgttg cgtaaaagtg gcagtcaatt acagattttc tttaacctac
     4381 gcaatgagct attgcggggg gtgccgcaat gagctgttgc gtacccccct tttttaagtt
     4441 gttgattttt aagtctttcg catttcgccc tatatctagt tctttggtgc ccaaagaagg
     4501 gcacccctgc ggggttcccc cacgccttcg gcgcggctcc ccctccggca aaaagtggcc
     4561 cctccggggc ttgttgatcg actgcgcggc cttcggcctt gcccaaggtg gcgctgcccc
     4621 cttggaaccc ccgcactcgc cgccgtgagg ctcggggggc aggcgggcgg gcttcgccct
     4681 tcgactgccc ccactcgcat aggcttgggt cgttccaggc gcgtcaaggc caagccgctg
     4741 cgcggtcgct gcgcgagcct tgacccgcct tccacttggt gtccaaccgg caagcgaagc
     4801 gcgcaggccg caggccggag gcttttcccc agagaaaatt aaaaaaattg atggggcaag
     4861 gccgcaggcc gcgcagttgg agccggtggg tatgtggtcg aaggctgggt agcgttcacc
     4921 gacaaacaac agataaaacg aaaggcccag tctttcgact gagcctttcg ttttatttga
     4981 tgcctggctc tagtagtgat ctacactagc actatcagtg ttattaagac ccactttcac
     5041 atttaagttg tttttctaat ccgcatatga tcaattcaag gccgaataag aaggctggct
     5101 ctgcaccttg gtgatcaaat aattcgatag cttgtcgtaa taatggcggc atactatcag
     5161 tagtaggtgt ttccctttct tctttagcga cttgatgctc ttgatcttcc aatacgcaac
     5221 ctaaagtaaa atgccccaca gcgctgagtg catataatgc attctctagt gaaaaacctt
     5281 gttggcataa aaaggctaat tgattttcga gagtttcata ctgtttttct gtaggccgtg
     5341 tacctaaatg tacttttgct ccatcgcgat gacttagtaa agcacatcta aaacttttag
     5401 cgttattacg taaaaaatct tgccagcttt ccccttctaa agggcaaaag tgagtatggt
     5461 gcctatctaa catctcaatg gctaaggcgt cgagcaaagc ccgcttattt tttacatgcc
     5521 aatacaatgt aggctgctct acacctagct tctgggcgag tttacgggtt gttaaacctt
     5581 cgattccgac ctcattaagc agctctaatg cgctgttaat cactttactt ttatctaatc
     5641 tagacatcat taattcctaa tttttgttga cactctatcg ttgatagagt tattttacca
     5701 ctccctatca gtgatagaga aaagaattca aaagatcttt taagaaggag atatacatat
     5761 gcatcatcat catcatcatc atggagaaag cttgtttaag gggccgcgtg attacaaccc
     5821 gatatcgagc accatttgtc atttgacgaa tgaatctgat gggcacacaa catcgttgta
     5881 tggtattgga tttggtccct tcatcattac aaacaagcac ttgtttagaa gaaataatgg
     5941 aacactgttg gtccaatcac tacatggtgt attcaaggtc aagaacacca cgactttgca
     6001 acaacacctc attgatggga gggacatgat aattattcgc atgcctaagg atttcccacc
     6061 atttcctcaa aagctgaaat ttagagagcc acaaagggaa gagcgcatat gtcttgtgac
     6121 aaccaacttc caaactaaga gcatgtctag catggtgtca gacactagtt gcacattccc
     6181 ttcatctgat ggcatattct ggaagcattg gattcaaacc aaggatgggc aggcgggcag
     6241 tccattagta tcaactagag atgggttcat tgttggtata cactcagcat cgaatttcac
     6301 caacacaaac aattatttca caagcgtgcc gaaaaacttc atggaattgt tgacaaatca
     6361 ggaggcgcag cagtgggtta gtggttggcg attaaatgct gactcagtat tgtggggggg
     6421 ccataaagtt ttcatggtga aacctgaaga gccttttcag ccagttaagg aagcgactca
     6481 actcatgaat cgtcgtcgcc gtcgctaagt ccatcagctt gtccagcagg gttgtccacg
     6541 ggccgagcga agcgagccag ccggtggccg cggaacaaaa ctaaagcgcc acaagggcgc
     6601 tttagtttgt tttcagtcca gttacgctgg agtctgaggc tcgtcctgaa tgatatcaag
     6661 cttgaattcg cggccgcttc tagagttgac agctagctca gtcctaggta ctgtgctagc
     6721 tactagatat cgctcaatac tgaccaagga ggtatctaca atg
//
