LOCUS       sequence-190017-        6433 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses TALE targeted to sucA with one TEV cut site in its 
            N-terminal domain, anhydrotetracycline inducible TEV protease.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6433)
  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 6433)
  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: p5C3-T
FEATURES             Location/Qualifiers
     source          1..6433
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             1..18
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             436..456
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             2239..2262
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     terminator      2300..2331
                     /label=tonB terminator
                     /note="bidirectional E. coli tonB-P14 transcription 
                     terminator"
     CDS             complement(2348..3163)
                     /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     3071..3090
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     CDS             complement(3253..3918)
                     /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(4578..4649)
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     CDS             complement(4694..5320)
                     /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     5225..5244
                     /label=Tet-R
                     /note="Tetracycline resistance gene, reverse primer"
     promoter        5336..5391
                     /gene="tetR"
                     /label=tetR/tetA promoters
                     /note="overlapping promoters for bacterial tetR and tetA"
     protein_bind    5372..5390
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             5432..5452
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=7xHis
                     /translation="HHHHHHH"
     CDS             5453..6160
                     /codon_start=1
                     /product="tobacco etch virus protease (Kapust et al., 
                     2001)"
                     /label=TEV protease
                     /translation="GESLFKGPRDYNPISSTICHLTNESDGHTTSLYGIGFGPFIITNK
                     HLFRRNNGTLLVQSLHGVFKVKNTTTLQQHLIDGRDMIIIRMPKDFPPFPQKLKFREPQ
                     REERICLVTTNFQTKSMSSMVSDTSCTFPSSDGIFWKHWIQTKDGQCGSPLVSTRDGFI
                     VGIHSASNFTNTNNYFTSVPKNFMELLTNQEAQQWVSGWRLNADSVLWGGHKVFMVKPE
                     EPFQPVKEATQLMN"
     terminator      6247..6283
                     /label=soxR terminator
                     /note="bidirectional E. coli soxR transcription terminator"
     misc_feature    6334..6355
                     /label=BioBrick prefix
                     /note="BioBrick prefix for parts that do not start with 
                     ""ATG"""
     promoter        6356..6390
                     /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 ctgaatggtt ctggcgaaaa cctgtacttc cagtctggct cgggtctgac accggaacag
      481 gtggtggcta ttgccagcaa taatggtggc aaacaggcac tggaaaccgt tcagcgtctg
      541 ctgccggtgc tgtgccaggc acatggttta accccggcgc aggtggttgc gattgcaagc
      601 aatattggcg gtaaacaagc cctggaaaca gtgcagcgcc tgttacctgt cttatgccaa
      661 gatcatggct tgacccctga ccaagtggtc gccatcgcat cacatgatgg aggtaaacag
      721 gccttagaaa ccgtacagcg cttactgccg gtactgtgcc aggaccacgg cctgacgcca
      781 gagcaggtgg tagcgatcgc ctcaaatatt ggtggaaaac aggcgttaga gacagtccag
      841 cgcctgctgc cagtcctttg ccaagcccac ggcctgactc cggcacaggt ggtggcaatt
      901 gcaagtcatg atgggggtaa acaggcactt gagacagttc aacgtttact gccggtgctg
      961 tgccaggctc atggtctgac gcccgatcaa gtggttgcaa ttgcgagtaa tggcggtggc
     1021 aaacaagcat tagaaacggt gcaacgcctg ctgcccgtcc tttgccagga ccatggttta
     1081 accccggatc aggttgtagc aattgcgagt aatattggtg gtaagcaggc gttggaaact
     1141 gtgcaaagac tgctgccggt tctgtgccaa gcacacggtt taacgccaga acaggttgtg
     1201 gccatcgcaa gcaacatcgg tggcaagcaa gcgcttgaaa cagtacagcg tttattacct
     1261 gtgctctgtc aggaccacgg tcttaccccg gcccaagttg ttgccatagc cagtaatcat
     1321 ggcggaaagc aggctctgga aacggtacaa cgtctgttac cggtactgtg tcaggatcat
     1381 ggattgaccc cggcacaggt tgtcgcgatt gcgtcaaata tcggaggcaa gcaggccttg
     1441 gagacagtgc agagattact gccagtttta tgtcaagcac atggcctgac accggaacag
     1501 gttgtggcga ttgcatctca tgatggcggt aagcaagctt tagagactgt tcagagactg
     1561 cttccggtgc tgtgtcagga ccacggactt acgcctgacc aagttgttgc tatcgcctct
     1621 aacatcggtg gtaagcaagc actggaaact gtccaacgct tacttccagt cctttgtcaa
     1681 gaccacggct taacgccgga gcaggtcgta gcgatagcca gcaataatgg tggtaaacag
     1741 gcccttgaaa cggtccaaag acttctgcct gtcctgtgtc aagaccatgg gctgacacct
     1801 gcccaggtag tggccattgc ctcaaatggc ggtgggaagc aggcattaga aacagttcaa
     1861 agattattac cggtcttgtg tcaggcgcat gggttgacac ccgaacaagt cgtggcgata
     1921 gcttcaaacg gtggtggtcg tccggcactg gaaagcattg ttgcacagct gagccgtccg
     1981 gatccggcac tggcagcact gaccaatgat catctggttg cactggcatg tctgggtggt
     2041 cgccctgccc tggatgcagt taaaaaaggt ctgccgcatg caccggcact gattaaacgt
     2101 accaatcgtc gtattccgga acgtaccagc catcgtgttg cagatcatgc acaggttgtt
     2161 cgtgttctgg gtttttttca gtgtcatagc catccggcac aggcatttga tgatgcaatg
     2221 acccagtttg gtatgagcga ttataaagat gatgatgata aataaggtac cctaatcata
     2281 cctgacctcc atagcagaaa gtcaaaaacc tccgaccgga ggcttttgac ttgaggggga
     2341 tccgccctca gaaaaactca tcgagcatca agtgaaactg caatttattc atatcaggat
     2401 tatcaatacc atatttttga aaaagccgtt tctgtaatga aggagaaaac tcaccgaggc
     2461 agttccatag gatggcaaga tcctggtatc ggtctgcgat tccgactcgt ccaacatcaa
     2521 tacaacctat taatttcccc tcgtcaaaaa taaggttatc aagtgagaaa tcaccatgag
     2581 tgacgactga atccggtgag aatggcaaaa gcttatgcat ttctttccag acttgttcaa
     2641 caggccagcc attacgctcg tcatcaaaat cactcgcacc aaccaaaccg ttattcattc
     2701 gtgattgcgc ctgagcgaga cgaaatacgc gatcgccgtt aaaaggacaa ttacaaacag
     2761 gaatcgaatg caaccggcgc aggaacactg ccagcgcatc aacaatattt tcacctgaat
     2821 caggatattc ttctaatacc tggaatgctg ttttccctgg gatcgcagtg gtgagtaacc
     2881 atgcatcatc aggagtacgg ataaaatgct tgatggtcgg aagaggcata aattccgtca
     2941 gccagtttag cctgaccatc tcatctgtaa catcattggc aacgctacct ttgccatgtt
     3001 tcagaaacaa ctctggcgca tcgggcttcc catacaatcg atagattgtc gcacctgatt
     3061 gcccgacatt atcgcgagcc catttatacc catataaatc agcatccatg ttggaattta
     3121 atcgcggcct cgagcaagac gtttcccgtt gaatatggct cattttagct tccttagctc
     3181 ctgaaaatct cgataactca aaaaatacgc ccggtagtga tcttatttca ttatggtgaa
     3241 agttggaacc tcttacgtac ggcgcggcag cgtgacccgt gtcggcggct ccaacggctc
     3301 gccatcgtcc agaaaacacg gctcatcggg catcggcagg cgctgctgcc cgcgtcgttc
     3361 ccattcctcc gtttcggtca aggctggcag gtctggttcc atgcccggaa tgccgggctg
     3421 gctgggcggc tcctcgccgg ggccggtcgg tagttgctgc tcgcccggat acagggtcgg
     3481 gatgcggcgc aggtcgccat gccccaacag cgattcgtcc tggtcgtcgt gatcaaccac
     3541 cacggcggca ctgaacaccg acaggcgcaa ctggtcgcgg ggctggcccc acgccacgcg
     3601 gtcattgacc acgtaggccg acacggtgcc ggggccgttg agcttcacga cggagatcca
     3661 gcgctcggcc accaagtcct tgactgcgta ttggaccgtc cgcaaagaac gtccgatgag
     3721 cttggaaagt gtcttctggc tgaccaccac ggcgttctgg tggcccatct gcgccacgag
     3781 gtgatgcagc agcattgccg ccgtgggttt cctcgcaata agcccggccc acgcctcatg
     3841 cgctttgcgt tccgtttgca cccagtgacc gggcttgttc ttggcttgaa tgccgatttc
     3901 tctggactgc gtggccatgc ttatctccat gcggtagggg tgccgcacgg ttgcggcacc
     3961 atgcgcaatc agctgcaact tttcggcagc gcgacaacaa ttatgcgttg cgtaaaagtg
     4021 gcagtcaatt acagattttc tttaacctac gcaatgagct attgcggggg gtgccgcaat
     4081 gagctgttgc gtacccccct tttttaagtt gttgattttt aagtctttcg catttcgccc
     4141 tatatctagt tctttggtgc ccaaagaagg gcacccctgc ggggttcccc cacgccttcg
     4201 gcgcggctcc ccctccggca aaaagtggcc cctccggggc ttgttgatcg actgcgcggc
     4261 cttcggcctt gcccaaggtg gcgctgcccc cttggaaccc ccgcactcgc cgccgtgagg
     4321 ctcggggggc aggcgggcgg gcttcgccct tcgactgccc ccactcgcat rggcttgggt
     4381 cgttccaggc gcgtcaaggc caagccgctg cgcggtcgct gcgcgagcct tgacccgcct
     4441 tccacttggt gtccaaccgg caagcgaagc gcgcaggccg caggccggag gcttttcccc
     4501 agagaaaatt aaaaaaattg atggggcaag gccgcaggcc gcgcagttgg agccggtggg
     4561 tatgtggtcg aaggctgggt agcgttcacc gacaaacaac agataaaacg aaaggcccag
     4621 tctttcgact gagcctttcg ttttatttga tgcctggctc tagtagtgat ctacactagc
     4681 actatcagtg ttattaagac ccactttcac atttaagttg tttttctaat ccgcatatga
     4741 tcaattcaag gccgaataag aaggctggct ctgcaccttg gtgatcaaat aattcgatag
     4801 cttgtcgtaa taatggcggc atactatcag tagtaggtgt ttccctttct tctttagcga
     4861 cttgatgctc ttgatcttcc aatacgcaac ctaaagtaaa atgccccaca gcgctgagtg
     4921 catataatgc attctctagt gaaaaacctt gttggcataa aaaggctaat tgattttcga
     4981 gagtttcata ctgtttttct gtaggccgtg tacctaaatg tacttttgct ccatcgcgat
     5041 gacttagtaa agcacatcta aaacttttag cgttattacg taaaaaatct tgccagcttt
     5101 ccccttctaa agggcaaaag tgagtatggt gcctatctaa catctcaatg gctaaggcgt
     5161 cgagcaaagc ccgcttattt tttacatgcc aatacaatgt aggctgctct acacctagct
     5221 tctgggcgag tttacgggtt gttaaacctt cgattccgac ctcattaagc agctctaatg
     5281 cgctgttaat cactttactt ttatctaatc tagacatcat taattcctaa tttttgttga
     5341 cactctatcg ttgatagagt tattttacca ctccctatca gtgatagaga aaagaattca
     5401 aaagatcttt taagaaggag atatacatat gcatcatcat catcatcatc atggagaaag
     5461 cttgtttaag gggccgcgtg attacaaccc gatatcgagc accatttgtc atttgacgaa
     5521 tgaatctgat gggcacacaa catcgttgta tggtattgga tttggtccct tcatcattac
     5581 aaacaagcac ttgtttagaa gaaataatgg aacactgttg gtccaatcac tacatggtgt
     5641 attcaaggtc aagaacacca cgactttgca acaacacctc attgatggga gggacatgat
     5701 aattattcgc atgcctaagg atttcccacc atttcctcaa aagctgaaat ttagagagcc
     5761 acaaagggaa gagcgcatat gtcttgtgac aaccaacttc caaactaaga gcatgtctag
     5821 catggtgtca gacactagtt gcacattccc ttcatctgat ggcatattct ggaagcattg
     5881 gattcaaacc aaggatgggc agtgtggcag tccattagta tcaactagag atgggttcat
     5941 tgttggtata cactcagcat cgaatttcac caacacaaac aattatttca caagcgtgcc
     6001 gaaaaacttc atggaattgt tgacaaatca ggaggcgcag cagtgggtta gtggttggcg
     6061 attaaatgct gactcagtat tgtggggggg ccataaagtt ttcatggtga aacctgaaga
     6121 gccttttcag ccagttaagg aagcgactca actcatgaat cgtcgtcgcc gtcgctaagt
     6181 ccatcagctt gtccagcagg gttgtccacg ggccgagcga agcgagccag ccggtggccg
     6241 cggaacaaaa ctaaagcgcc acaagggcgc tttagtttgt tttcagtcca gttacgctgg
     6301 agtctgaggc tcgtcctgaa tgatatcaag cttgaattcg cggccgcttc tagagttgac
     6361 agctagctca gtcctaggta ctgtgctagc tactagatat cgctcaatac tgaccaagga
     6421 ggtatctaca atg
//
