LOCUS       sequence-44895-e        4344 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4344)
  AUTHORS   Zhang Y, Zhang F, Li X, Baller JA, Qi Y, Starker CG, Bogdanove AJ, 
            Voytas DF
  TITLE     TALENs enable efficient plant genome engineering.
  JOURNAL   Plant Physiol. 2012 Nov 2.
  PUBMED    23124327
REFERENCE   2  (bases 1 to 4344)
  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: "Plant 
            Physiol. 2012 Nov 2."
COMMENT     Sequence Label: pZHY013
FEATURES             Location/Qualifiers
     source          1..4344
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    complement(14..113)
                     /gene="mutant version of attL"
                     /label=attL2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     primer_bind     complement(130..149)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(131..149)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(154..170)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(154..170)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     CDS             601..1392
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      1485..2073
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1974..1993
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     2227..2244
                     /label=L4440
                     /note="L4440 vector, forward primer"
     terminator      complement(2403..2430)
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      complement(2522..2608)
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     2657..2679
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2671..2688
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2672..2688
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    2704..2803
                     /gene="mutant version of attL"
                     /label=attL1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             2825..2890
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             2897..2917
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3558..3611
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             3618..3683
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             3690..3710
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3751..4338
                     /codon_start=1
                     /product="nonspecific DNA cleavage domain of the FokI 
                     endonuclease (Li et al., 1992)"
                     /label=FokI cleavage domain
                     /note="must dimerize to be catalytically active"
                     /translation="QLVKSELEEKKSELRHKLKYVPHEYIELIEIARNSTQDRILEMKV
                     MEFFMKVYGYRGKHLGGSRKPDGAIYTVGSPIDYGVIVDTKAYSGGYNLPIGQADEMER
                     YVEENQTRNKHLNPNEWWKVYPSSVTEFKFLFVSGHFKGNYKAQLTRLNHITNCNGAVL
                     SVEELLIGGEMIKAGTLTLEEVRRKFNNGEINF"
     CDS             join(4341..4344,1..8)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
ORIGIN
        1 aagggcgaat tcgacccagc tttcttgtac aaagttggca ttataaaaaa taattgctca
       61 tcaatttgtt gcaacgaaca ggtcactatc agtcaaaata aaatcattat ttgccatcca
      121 gctgatatcc cctatagtga gtcgtattac atggtcatag ctgtttcctg gcagctctgg
      181 cccgtgtctc aaaatctctg atgttacatt gcacaagata aaaatatatc atcatgcctc
      241 ctctggacca gccaggacag aaatgcctcg acttcgctgc tacccaaggt tgccgggtga
      301 cgcacaccgt ggaaacggat gaaggcacga acccagtgga cataagcctg ttcggttcgt
      361 aagctgtaat gcaagtagcg tatgcgctca cgcaactggt ccagaacctt gaccgaacgc
      421 agcggtggta acggcgcagt ggcggttttc atggcttgtt atgactgttt ttttggggta
      481 cagtctatgc ctcgggcatc caagcagcaa gcgcgttacg ccgtgggtcg atgtttgatg
      541 ttatggagca gcaacgatgt tacgcagcag ggcagtcgcc ctaaaacaaa gttaaacatc
      601 atgagggaag cggtgatcgc cgaagtatcg actcaactat cagaggtagt tggcgtcatc
      661 gagcgccatc tcgaaccgac gttgctggcc gtacatttgt acggctccgc agtggatggc
      721 ggcctgaagc cacacagtga tattgatttg ctggttacgg tgaccgtaag gcttgatgaa
      781 acaacgcggc gagctttgat caacgacctt ttggaaactt cggcttcccc tggagagagc
      841 gagattctcc gcgctgtaga agtcaccatt gttgtgcacg acgacatcat tccgtggcgt
      901 tatccagcta agcgcgaact gcaatttgga gaatggcagc gcaatgacat tcttgcaggt
      961 atcttcgagc cagccacgat cgacattgat ctggctatct tgctgacaaa agcaagagaa
     1021 catagcgttg ccttggtagg tccagcggcg gaggaactct ttgatccggt tcctgaacag
     1081 gatctatttg aggcgctaaa tgaaacctta acgctatgga actcgccgcc cgactgggct
     1141 ggcgatgagc gaaatgtagt gcttacgttg tcccgcattt ggtacagcgc agtaaccggc
     1201 aaaatcgcgc cgaaggatgt cgctgccgac tgggcaatgg agcgcctgcc ggcccagtat
     1261 cagcccgtca tacttgaagc tagacaggct tatcttggac aagaagaaga tcgcttggcc
     1321 tcgcgcgcag atcagttgga agaatttgtc cactacgtga aaggcgagat caccaaggta
     1381 gtcggcaaat aaccctcgag ccacccatga ccaaaatccc ttaacgtgag ttacgcgtcg
     1441 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     1501 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     1561 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     1621 ccaaatactg tccttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     1681 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     1741 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     1801 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     1861 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     1921 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     1981 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     2041 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     2101 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     2161 gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc
     2221 gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc
     2281 cccgcgcgtt ggccgattca ttaatgcagc tggcacgaca ggtttcccga ctggaaagcg
     2341 ggcagtgagc gcaacgcaat taatacgcgt accgcgagcc aggaagagtt tgtagaaacg
     2401 caaaaaggcc atccgtcagg atggccttct gcttagtttg atgcctggca gtttatggcg
     2461 ggcgtcctgc ccgccaccct ccgggccgtt gcttcacaac gttcaaatcc gctcccggcg
     2521 gatttgtcct actcaggaga gcgttcaccg acaaacaaca gataaaacga aaggcccagt
     2581 cttccgactg agcctttcgt tttatttgat gcctggcagt tccctactct cgcgttaacg
     2641 cttgcatgga tgttttccca gtcacgacgt tgtaaaacga cggccagtct taagctcggg
     2701 ccccaaataa tgattttatt ttgactgata gtgacctgtt cgttgcaaca aattgatgag
     2761 caatgctttt ttataatgcc aactttgtac aaaaaagcag gctccgaatt cgcccttcac
     2821 catggattat aaggatcacg atggcgacta caaggaccac gatattgact acaaagacga
     2881 tgacgataaa atggctccta agaaaaagcg caaagtcggt atccatggcg ttccctctag
     2941 ataacgcagg atcccagctg gtgaagtccg agctggaaga aaaaaagagc gagctgcgcc
     3001 acaagctcaa gtacgtgccc cacgagtaca tcgagctgat cgagatcgcc cgcaacagca
     3061 cccaagaccg catcctggag atgaaagtga tggagttctt catgaaggtg tacggctacc
     3121 gcggcaagca cctgggcggc tcccgcaagc ccgatggcgc catctacacc gtgggctccc
     3181 ccatcgacta tggcgtcatt gtcgacacca aggcctactc cggcggctac aacttaccca
     3241 tcggtcaggc cgacgagatg caacgctacg tgaaggagaa ccagacccgc aataagcaca
     3301 ttaatcccaa cgagtggtgg aaggtgtacc cctcctccgt gaccgagttc aaattcctgt
     3361 tcgtgtccgg ccacttcaag ggcaattata aggcccaact gacccgcctg aaccacaaga
     3421 ccaactgcaa cggcgccgtg ctgtccgtgg aggaactgct gatcggcggc gagatgatca
     3481 aggctggtac cctgaccctg gaagaggtgc gccgcaagtt caacaatggt gaaatcaatt
     3541 tcaggtccgg cggcggagag ggcagaggaa gtcttctaac atgcggtgac gtggaggaga
     3601 atcccggccc taggatggac tacaaagacc atgacggtga ttataaagat catgacatcg
     3661 attacaagga tgacgatgac aagatggccc ccaagaagaa gaggaaggtg ggcattcacg
     3721 gggtgccggc tagctagtaa gcttagatct caactagtca aaagtgaact ggaggagaag
     3781 aaatctgaac ttcgtcataa attgaaatat gtgcctcatg aatatattga attaattgaa
     3841 attgccagaa attccactca ggatagaatt cttgaaatga aggtaatgga attttttatg
     3901 aaagtttatg gatatagagg taaacatttg ggtggatcaa ggaaaccgga cggagcaatt
     3961 tatactgtcg gatctcctat tgattacggt gtgatcgtgg atactaaagc ttatagcgga
     4021 ggttataatc tgccaattgg ccaagcagat gaaatggagc gatatgtcga agaaaatcaa
     4081 acacgaaaca aacatctcaa ccctaatgaa tggtggaaag tctatccatc ttctgtaacg
     4141 gaatttaagt ttttatttgt gagtggtcac tttaaaggaa actacaaagc tcagcttaca
     4201 cgattaaatc atatcactaa ttgtaatgga gctgttctta gtgtagaaga gcttttaatt
     4261 ggtggagaaa tgattaaagc cggcacatta accttagagg aagtgagacg gaaatttaat
     4321 aacggcgaga taaactttta atag
//
