LOCUS       sequence-14201-e        7530 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 7530)
  AUTHORS   Cermak T, Doyle EL, Christian M, Wang L, Zhang Y, Schmidt C, Baller 
            JA, Somia NV, Bogdanove AJ, Voytas DF
  TITLE     Efficient design and assembly of custom TALEN and other TAL 
            effector-based constructs for DNA targeting.
  JOURNAL   Nucleic Acids Res. 2011 Jul;39(12):e82. doi: 10.1093/nar/gkr218
  PUBMED    21493687
REFERENCE   2  (bases 1 to 7530)
  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: "Nucleic 
            Acids Res."; date: "2011-07"; volume: "39(12):e82. doi"; pages: " 
            10.1093/nar/gkr21"
COMMENT     Sequence Label: pTAL3
FEATURES             Location/Qualifiers
     source          1..7530
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      35..129
                     /label=lambda t0 terminator
                     /note="transcription terminator from phage lambda"
     promoter        136..154
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        231..629
                     /gene="S. cerevisiae TEF1"
                     /label=TEF1 promoter
                     /note="promoter for EF-1-alpha"
     CDS             701..721
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             722..748
                     /codon_start=1
                     /product="epitope tag from the baculovirus Autographa 
                     californica GP64 envelope fusion protein"
                     /label=AcV5 tag
                     /translation="SWKDASGWS"
     CDS             complement(1514..1921)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
                     QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGARLQGG
                     TRQCIARCMHCLHCGHAATFCNLHKLACVQL"
     primer_bind     1771..1793
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1785..1802
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1786..1802
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        1812..1830
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(1878..1896)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1917..1933)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1917..1933)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1930..1952)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1941..1957
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(1965..1971,1972..1989,1990..1995))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2010..2031
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             2727..3314
                     /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
                     MEFFMKVYGYRGKHLGGSRKPDGAIYTVGSPIDYGVIVDTKAYSGGYNLPIGQADEMQR
                     YVEENQTRNKHINPNEWWKVYPSSVTEFKFLFVSGHFKGNYKAQLTRLNHITNCNGAVL
                     SVEELLIGGEMIKAGTLTLEEVRRKFNNGEINF"
     terminator      3337..3589
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     promoter        complement(3631..3649)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     terminator      complement(3651..3690)
                     /label=fd terminator
                     /note="central terminator from bacteriophage fd (Otsuka and
                     Kunisawa, 1982)"
     CDS             complement(3990..4649)
                     /codon_start=1
                     /gene="S. cerevisiae HIS3"
                     /product="imidazoleglycerol-phosphate dehydratase, required
                     for histidine biosynthesis"
                     /label=HIS3
                     /note="yeast auxotrophic marker"
                     /translation="MTEQKALVKRITNETKIQIAISLKGGPLAIEHSIFPEKEAEAVAE
                     QATQSQVINVHTGIGFLDHMIHALAKHSGWSLIVECIGDLHIDDHHTTEDCGIALGQAF
                     KEALLARGVKRFGSGFAPLDEALSRAVVDLSNRPYAVVELGLQREKVGDLSCEMIPHFL
                     ESFAEASRITLHVDCLRGKNDHHRSESAFKALAVAIREATSPNGTNDVPSTKGVLM"
     promoter        complement(4650..4837)
                     /gene="S. cerevisiae HIS3"
                     /label=HIS3 promoter
     misc_feature    4988..5491
                     /label=CEN/ARS
                     /note="S. cerevisiae CEN6 centromere fused to an 
                     autonomously replicating sequence"
     promoter        5533..5637
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      6705..7293
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7194..7213
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     7447..7464
                     /label=L4440
                     /note="L4440 vector, forward primer"
ORIGIN
        1 tcaacgacat cacctggttg cgagccttct gaaggactcc tgttgataga tccagtaatg
       61 acctcagaac tccatctgga tttgttcaga acgctcggtt gccgccgggc gttttttatt
      121 ggtgagaatc aggcgtaata cgactcacta tagggcgaat ggaactggac gctcgcctgt
      181 ggtcgcaggg tggctcgtac gaacttaccg cggagacata tcgataatca catagcttca
      241 aaatgtttct actccttttt tactcttcca gattttctcg gactccgcgc atcgccgtac
      301 cacttcaaaa cacccaagca cagcatacta aatttcccct ctttcttcct ctagggtgtc
      361 gttaattacc cgtactaaag gtttggaaaa gaaaaaagag accgcctcgt ttctttttct
      421 tcgtcgaaaa aggcaataaa aatttttatc acgtttcttt ttcttgaaaa tttttttttt
      481 gatttttttc tctttcgatg acctcccatt gatatttaag ttaataaacg gtcttcaatt
      541 tctcaagttt cagtttcatt tttcttgttc tattacaact ttttttactt cttgctcatt
      601 agaaagaaag catagcaatc taatctaagt ttagatctaa ctcgagaaag atattgtata
      661 tatcgtaaca ataggaggtt caacaatggc ttcctcccct ccaaagaaaa agagaaaggt
      721 tagttggaag gacgcaagtg gttggtctag aatgcatgcg gatcccattc gtccgcgcag
      781 gccaagtcct gcccgcgagc ttctgcccgg accccaaccg gatagggttc agccgactgc
      841 agatcgtggg gtgtctgcgc ctgctggcag ccctctggat ggcttgcccg ctcggcggac
      901 ggtgtcccgg acccggctgc catctccccc tgcgccctca cctgcgttct cggcgggcag
      961 cttcagcgat ctgctccgtc cgttcgatcc gtcgcttctt gatacatcgc ttcttgattc
     1021 gatgcctgcc gtcggcacgc cgcatacagc ggctgcccca gcagagtggg atgaggcgca
     1081 atcggctctg cgtgcagccg atgacccgcc acccaccgtg cgtgtcgctg tcactgccgc
     1141 gcggccgccg cgcgccaagc cggccccgcg acggcgtgct gcgcaaccct ccgacgcttc
     1201 gccggccgcg caggtggatc tacgcacgct cggctacagt cagcagcagc aagagaagat
     1261 caaaccgaag gtgcgttcga cagtggcgca gcaccacgag gcactggtgg gccatgggtt
     1321 tacacacgcg cacatcgttg cgctcagcca acacccggca gcgttaggga ccgtcgctgt
     1381 cacgtatcag cacataatca cggcgttgcc agaggcgaca cacgaagaca tcgttggcgt
     1441 cggcaaacag tggtccggcg cacgcgccct ggaggccttg ctcacggatg cgggggagtt
     1501 gagaggtccg ccgttacagt tggacacagg ccaacttgtg aagattgcaa aacgtggcgg
     1561 cgtgaccgca atggaggcag tgcatgcatc gcgcaatgca ctgacgggtg cccccctgga
     1621 gacgggcgcc gctacagggc gcgtcccatt cgccattcag gctgcgcaac tgttgggaag
     1681 ggcgatcggt gcgggcctct tcgctattac gccagctggc gaaaggggga tgtgctgcaa
     1741 ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca
     1801 gtgagcgcgc gtaatacgac tcactatagg gcgaattggg taccgggccc cccctcgagg
     1861 tcctccagct tttgttccct ttagtgaggg ttaattgcgc gcttggcgta atcatggtca
     1921 tagctgtttc ctgtgtgaaa ttgttatccg ctcacaattc cacacaacat acgagccgga
     1981 agcataaagt gtaaagcctg gggtgcctaa tgagtgagct aactcacatt aattgcgttg
     2041 cgctcactgc ccgctttcca ccggtcgtct ccaacgacca cctcgtcgcc ttggcctgcc
     2101 tcggcggacg tcctgccatg gatgcagtga aaaagggatt gccgcacgcg ccggaattga
     2161 tcagaagagt caatcgccgt attggcgaac gcacgtccca tcgcgttgcc gactacgcgc
     2221 aagtggttcg cgtgctggag tttttccagt gccactccca cccagcgtac gcatttgatg
     2281 aggccatgac gcagttcggg atgagcagga acgggttggt acagctcttt cgcagagtgg
     2341 gcgtcaccga actcgaagcc cgcggtggaa cgctcccccc agcctcgcag cgttgggacc
     2401 gtatcctcca ggcatcaggg atgaaaaggg ccaaaccgtc ccctacttca gctcaaacac
     2461 cggatcaggc gtctttgcat gcattcgccg attcgctgga gcgtgacctt gatgcgccca
     2521 gcccaatgca cgagggagat cagacgcggg caagcagccg taaacggtcc cgatcggatc
     2581 gtgctgtcac cggcccctcc gcacagcagg ctgtcgaggt gcgcgttccc gaacagcgcg
     2641 atgcgctgca tttgcccctc agctggaggg taaaacgccc gcgtaccagg atctggggcg
     2701 gcctcccgga tccgatatct agatcccagc tagtgaaatc tgaattggaa gagaagaaat
     2761 ctgaacttag acataaattg aaatatgtgc cacatgaata tattgaattg attgaaatcg
     2821 caagaaattc aactcaggat agaatccttg aaatgaaggt gatggagttc tttatgaagg
     2881 tttatggtta tcgtggtaaa catttgggtg gatcaaggaa accagacgga gcaatttata
     2941 ctgtcggatc tcctattgat tacggtgtga tcgttgatac taaggcatat tcaggaggtt
     3001 ataatcttcc aattggtcaa gcagatgaaa tgcaaagata tgtcgaagag aatcaaacaa
     3061 gaaacaagca tatcaaccct aatgaatggt ggaaagtcta tccatcttca gtaacagaat
     3121 ttaagttctt gtttgtgagt ggtcatttca aaggaaacta caaagctcag cttacaagat
     3181 tgaatcatat cactaattgt aatggagctg ttcttagtgt agaagagctt ttgattggtg
     3241 gagaaatgat taaagctggt acattgacac ttgaggaagt gagaaggaaa tttaataacg
     3301 gtgagataaa cttttaatag gagctcgaat ttccccgatc gttcaaacat ttggcaataa
     3361 agtttcttaa gattgaatcc tgttgccggt cttgcgatga ttatcatata atttctgttg
     3421 aattacgtta agcatgtaat aattaacatg taatgcatga cgttatttat gagatgggtt
     3481 tttatgatta gagtcccgca attatacatt taatacgcga tagaaaacaa aatatagcgc
     3541 gcaaactagg ataaattatc gcgcgcggtg tcatctatgt tactagatcg tgggcccaca
     3601 ttcctgtaga actcggtgag acttttgttc cctttagtga gggttaattg tccaaaaaaa
     3661 aaggctccaa aaggagcctt taattgtttc ggtttatgct taggacacca ttgattaacg
     3721 cttacaattt cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca
     3781 tagatccgtc gagttcaaga gaaaaaaaaa gaaaaagcaa aaagaaaaaa ggaaagcgcg
     3841 cctcgttcag aatgacacgt atagaatgat gcattacctt gtcatcttca gtatcatact
     3901 gttcgtataa atacttactg acattcatag ctatacatat atacacatgt atatatatcg
     3961 tatgctggag ctttaaataa tcggtgtcac tacataagaa cacctttggt ggagggaaca
     4021 tcgttggttc cattgggcga ggtggcttct cttatggcaa ccgcaagagc cttgaacgca
     4081 ctctcactac ggtgatggtc attcttgcct cgcagacaat caacgtggag ggtaattctg
     4141 cttgcctctg caaaactttc aagaaaatgc gggatcatct cgcaagagag gtctcctact
     4201 ttctcccttt gcaaaccaag ttcgacaact gcgtatggcc tgttagaaag gtctaccacc
     4261 gctctggaaa gtgcctcatc caaaggcgca aatcctgatc caaacctttt tactccacgc
     4321 gccagtagag cctctttaaa agcctgaccg agagcaatcc cgcagtcttc agtggtgtga
     4381 tggtcgtcta tgtgtaagtc accaatgcac tcaacgatta gcgaccagcc ggaatgcttg
     4441 gccagagcat gtatcatgtg gtccagaaac cctatacctg tgtggacgtt aatcacttgc
     4501 gattgtgtgg cctgttctgc tactgcttct gcctcttttt ctgggaagat cgagtgctct
     4561 atcgctaggg gaccaccctt taaagagata gcaatctgaa tcttggtttc atttgtaata
     4621 cgctttacta gggctttctg ctctgtcatc tttgccttcg tttatcttgc ctgctcattt
     4681 tttagtatat tcttcgaaga aatcacatta ctttatataa tgtataattc attatgtgat
     4741 aatgccaatc gctaagaaaa aaaaagagtc atccgctagg tggaaaaaaa aaaatgaaaa
     4801 tcattaccga ggcataaaaa aatatagagt gtactagagg aggccaagag taatagaaaa
     4861 agaaaattgc gggaaaggac tgtgttatga cttccctgac taatgccgtg ttcaaacgat
     4921 acctggcagt gactcctagc gctcaccaag ctcttaaaac gggaatttat ggatcttaat
     4981 taagacggat cgcttgcctg taacttacac gcgcctcgta tcttttaatg atggaataat
     5041 ttgggaattt actctgtgtt tatttatttt tatgttttgt atttggattt tagaaagtaa
     5101 ataaagaagg tagaagagtt acggaatgaa gaaaaaaaaa taaacaaagg tttaaaaaat
     5161 ttcaacaaaa agcgtacttt acatatatat ttattagaca agaaaagcag attaaataga
     5221 tatacattcg attaacgata agtaaaatgt aaaatcacag gattttcgtg tgtggtcttc
     5281 tacacagaca agatgaaaca attcggcatt aatacctgag agcaggaaga gcaagataaa
     5341 aggtagtatt tgttggcgat ccccctagag tcttttacat cttcggaaaa caaaaactat
     5401 tttttcttta atttcttttt ttactttcta tttttaattt atatatttat attaaaaaat
     5461 ttaaattata attattttta tagcacgtga tgaaaagcct tagggtcagg tggcactttt
     5521 cggggaaatg tgcgatgaac ccctatttgt ttatttttct aaatacattc aaatatgtat
     5581 ccgctaatga gacaataacc ctgataaatg cttcaattat attgaaaaag gaagagtatg
     5641 agtatccaac atttccgtgt cgcccttatt cccttttttg cggccttctg ccttcctgtt
     5701 tttgctcacc cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcccga
     5761 gtgggttaca ttgaactgga tctcaacagc ggcaaaatct tagagagttt tcgccccgaa
     5821 gaacgcttcc caatgatgag cactttcaaa gttctgctat gtggcgcggt gttatcccgt
     5881 attgatgctg ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt
     5941 gagtattcac cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgt
     6001 agtgctgcca taaccatgag tgataacact gcggccaact tacttctgac aacgattgga
     6061 ggaccgaagg agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat
     6121 cgttgggaac cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct
     6181 gtagcaatgg caacaactct ccgcaaacta ttaactggcg aactacttac tcttgcttca
     6241 cgccagcaac tcattgactg gatggaggcg gacaaagttg caggaccact tctgcgctcg
     6301 gcacttccgg ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc
     6361 ggtatcattg cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg
     6421 acggggagtc aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca
     6481 ctgattaagc attggtaact gtcagacgaa gggaaataaa tagtagcccg cctgatgtgc
     6541 gggctttttt ttggtggtgg ttaccagtgt cgcttagcta ccgtaggtcc tcttaggcgg
     6601 tccgattagc gacatagtca tcaatgcctt gacggcaaaa tcccttaacg tgagttttcg
     6661 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     6721 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     6781 ccggatcaag agctactaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     6841 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     6901 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     6961 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     7021 tgaacggggg gttcgtgctc acagcccagc ttggagcgaa cgacctacac cgaactgaga
     7081 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     7141 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     7201 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     7261 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     7321 ttcctggcct tttgctggcc ttttgctcac atgttctttc ctgcgttatc ccctgattct
     7381 gtggataacc gtattaccgc ctttgagtga gctgataccg ctcgccgcag ccgaacgacc
     7441 gagcgcagcg agtcagtgag cgaggaagcg gaagagcgcc caatacgcaa accgcctctc
     7501 cccgcgcgtt ggccgattca ttgatgcagg
//
