LOCUS       sequence-70557-e        7210 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TALEN expression in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7210)
  AUTHORS   Frank S, Skryabin BV, Greber B
  TITLE     A modified TALEN-based system for robust generation of knock-out 
            human pluripotent stem cell lines and disease models.
  JOURNAL   BMC Genomics. 2013 Nov 9;14(1):773.
  PUBMED    24206569
REFERENCE   2  (bases 1 to 7210)
  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: "BMC 
            Genomics."; date: "2013-11-9"; volume: "14(1)"; pages: "773"
COMMENT     Sequence Label: pTAL7b
FEATURES             Location/Qualifiers
     source          1..7210
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(2..796)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             complement(2..796)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(113..132)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     723..742
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        1139..1468
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1319..1454
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1381..1400
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        1516..1563
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             1582..1980
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEDYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     LTR             2067..2300
                     /label=3' LTR (Delta-U3)
                     /note="self-inactivating 3' long terminal repeat (LTR) from
                     HIV-1"
     polyA_signal    2372..2493
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      2587..3175
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3076..3095
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        3272..3651
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3653..3930
                     /label=chicken beta-actin promoter
     intron          3931..4948
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     4872..4894
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     CDS             complement(4957..5778)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQQKLEDLEGWPVTQFAARSLAVVLQRRDWENPGVTQL
                     NRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWDAPCSGARLQGGSCQSIPPSVHSFH
                     SCHSAAFSDLQQLSSVQSGLEWWTPQFPGDGEECFECSGTAPLLSDAHNGFVGCLRQSG
                     DHVLIFDGHSPECSGMLRQSHNMGVCETVPHQCFMVLSDGGPHLWFDLLLLLLAVAEGS
                     QIHLGCWTGGYPTSATTSATWFIVVILVVSGRHLPLFLRSHGGGPYSESYY"
     primer_bind     4961..4980
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4961..4979
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      4984..4993
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4996..5016
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             5026..5049
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     primer_bind     5667..5689
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     5681..5698
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     5682..5698
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5774..5790)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5774..5790)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5787..5809)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5798..5814
                     /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(5822..5828,5829..5846,5847..5852))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5867..5888
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             6065..6652
                     /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"
     primer_bind     complement(6673..6689)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6673..6689)
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     intron          6849..6914
                     /label=small t intron
                     /note="SV40 (simian virus 40) small t antigen intron"
     polyA_signal    6961..7095
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
ORIGIN
        1 ttcagaagaa ctcgtcaaga aggcgataga aggcgatgcg ctgcgaatcg ggagcggcga
       61 taccgtaaag cacgaggaag cggtcagccc attcgccgcc aagctcttca gcaatatcac
      121 gggtagccaa cgctatgtcc tgatagcggt ccgccacacc cagccggcca cagtcgatga
      181 atccagaaaa gcggccattt tccaccatga tattcggcaa gcaggcatcg ccatgggtca
      241 cgacgagatc ctcgccgtcg ggcatgctcg ccttgagcct ggcgaacagt tcggctggcg
      301 cgagcccctg atgctcttcg tccagatcat cctgatcgac aagaccggct tccatccgag
      361 tacgtgctcg ctcgatgcga tgtttcgctt ggtggtcgaa tgggcaggta gccggatcaa
      421 gcgtatgcag ccgccgcatt gcatcagcca tgatggatac tttctcggca ggagcaaggt
      481 gagatgacag gagatcctgc cccggcactt cgcccaatag cagccagtcc cttcccgctt
      541 cagtgacaac gtcgagcaca gctgcgcaag gaacgcccgt cgtggccagc cacgatagcc
      601 gcgctgcctc gtcttgcagt tcattcaggg caccggacag gtcggtcttg acaaaaagaa
      661 ccgggcgccc ctgcgctgac agccggaaca cggcggcatc agagcagccg attgtctgtt
      721 gtgcccagtc atagccgaat agcctctcca cccaagcggc cggagaacct gcgtgcaatc
      781 catcttgttc aatcatgcga aacgatcctc atcctgtctc ttgatcagat cttgatcccc
      841 tgcgccatca gatccttggc ggcgagaaag ccatccagtt tactttgcag ggcttcccaa
      901 ccttaccaga gggcgcccca gctggcaatt ccggttcgct tgctgtccat aaaaccgccc
      961 agtctagcta tcgccatgta agcccactgc aagctacctg ctttctcttt gcgcttgcgt
     1021 tttcccttgt ccagatagcc cagtagctga cattcatccg gggtcagcac cgtttctgcg
     1081 gactggcttt ctacgtgaaa aggatctagg tgaagatcct ttttgataat ctcatgatgt
     1141 gtgtcagtta gggtgtggaa agtccccagg ctccccagca ggcagaagta tgcaaagcat
     1201 gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca ggctccccag caggcagaag
     1261 tatgcaaagc atgcatctca attagtcagc aaccatagtc ccgcccctaa ctccgcccat
     1321 cccgccccta actccgccca gttccgccca ttctccgccc catggctgac taattttttt
     1381 tatttatgca gaggccgagg ccgcctctgc ctctgagcta ttccagaagt agtgaggagg
     1441 cttttttgga ggcctaggct tttgcaaaaa gctcccggga gcttgtatat ccattttcgg
     1501 atctgatcag cacgtgttga caattaatca tcggcatagt atatcggcat agtataatac
     1561 gacaaggtga ggaactaaac catggccaag cctttgtctc aagaagaatc caccctcatt
     1621 gaaagagcaa cggctacaat caacagcatc cccatctctg aagactacag cgtcgccagc
     1681 gcagctctct ctagcgacgg ccgcatcttc actggtgtca atgtatatca ttttactggg
     1741 ggaccttgtg cagaactcgt ggtgctgggc actgctgctg ctgcggcagc tggcaacctg
     1801 acttgtatcg tcgcgatcgg aaatgagaac aggggcatct tgagcccctg cggacggtgc
     1861 cgacaggtgc ttctcgatct gcatcctggg atcaaagcca tagtgaagga cagtgatgga
     1921 cagccgacgg cagttgggat tcgtgaattg ctgccctctg gttatgtgtg ggagggctaa
     1981 gcacaattcg agctcggtac ctttaagacc aatgacttac aaggcagctg tagatcttag
     2041 ccacttttta aaagaaaagg ggggactgga agggctaatt cactcccaac gaagacaaga
     2101 tctgcttttt gcttgtactg ggtctctctg gttagaccag atctgagcct gggagctctc
     2161 tggctaacta gggaacccac tgcttaagcc tcaataaagc ttgccttgag tgcttcaagt
     2221 agtgtgtgcc cgtctgttgt gtgactctgg taactagaga tccctcagac ccttttagtc
     2281 agtgtggaaa atctctagca gtagtagttc atgtcatctt attattcagt atttataact
     2341 tgcaaagaaa tgaatatcag agagtgagag gaacttgttt attgcagctt ataatggtta
     2401 caaataaagc aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag
     2461 ttgtggtttg tccaaactca tcaatgtatc ttatcatgtc tggctctagc tcatgaccaa
     2521 aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa agatcaaagg
     2581 atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc
     2641 gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc cgaaggtaac
     2701 tggcttcagc agagcgcaga taccaaatac tgtccttcta gtgtagccgt agttaggcca
     2761 ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt
     2821 ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc
     2881 ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg
     2941 aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc
     3001 cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac
     3061 gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct
     3121 ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc
     3181 cagcaacgcg gcctttttac ggttcctggg cttttgctgg ccttttgctc acatgttctt
     3241 tcctgcgtta tcccctgggc gcgccggatt cgacattgat tattgactag ttattaatag
     3301 taatcaatta cggggtcatt agttcatagc ccatatatgg agttccgcgt tacataactt
     3361 acggtaaatg gcccgcctgg ctgaccgccc aacgaccccc gcccattgac gtcaataatg
     3421 acgtatgttc ccatagtaac gccaataggg actttccatt gacgtcaatg ggtggagtat
     3481 ttacggtaaa ctgcccactt ggcagtacat caagtgtatc atatgccaag tacgccccct
     3541 attgacgtca atgacggtaa atggcccgcc tggcattatg cccagtacat gaccttatgg
     3601 gactttccta cttggcagta catctacgta ttagtcatcg ctattaccat ggtcgaggtg
     3661 agccccacgt tctgcttcac tctccccatc tcccccccct ccccaccccc aattttgtat
     3721 ttatttattt tttaattatt ttgtgcagcg atgggggcgg gggggggggg ggggcgcgcg
     3781 ccaggcgggg cggggcgggg cgaggggcgg ggcggggcga ggcggagagg tgcggcggca
     3841 gccaatcaga gcggcgcgct ccgaaagttt ccttttatgg cgaggcggcg gcggcggcgg
     3901 ccctataaaa agcgaagcgc gcggcgggcg ggagtcgctg cgcgctgcct tcgccccgtg
     3961 ccccgctccg ccgccgcctc gcgccgcccg ccccggctct gactgaccgc gttactccca
     4021 caggtgagcg ggcgggacgg cccttctcct ccgggctgta attagcgctt ggtttaatga
     4081 cggcttgttt cttttctgtg gctgcgtgaa agccttgagg ggctccggga gggccctttg
     4141 tgcggggggg agcggctcgg ggggtgcgtg cgtgtgtgtg tgcgtgggga gcgccgcgtg
     4201 cggctccgcg ctgcccggcg gctgtgagcg ctgcgggcgc ggcgcggggc tttgtgcgct
     4261 ccgcagtgtg cgcgagggga gcgcggccgg gggcggtgcc ccgcggtgcg gggggggctg
     4321 cgaggggaac aaaggctgcg tgcggggtgt gtgcgtgggg gggtgagcag ggggtgtggg
     4381 cgcgtcggtc gggctgcaac cccccctgca cccccctccc cgagttgctg agcacggccc
     4441 ggcttcgggt gcggggctcc gtacggggcg tggcgcgggg ctcgccgtgc cgggcggggg
     4501 gtggcggcag gtgggggtgc cgggcggggc ggggccgcct cgggccgggg agggctcggg
     4561 ggaggggcgc ggcggccccc ggagcgccgg cggctgtcga ggcgcggcga gccgcagcca
     4621 ttgcctttta tggtaatcgt gcgagagggc gcagggactt cctttgtccc aaatctgtgc
     4681 ggagccgaaa tctgggaggc gccgccgcac cccctctagc gggcgcgggg cgaagcggtg
     4741 cggcgccggc aggaaggaaa tgggcgggga gggccttcgt gcgtcgccgc gccgccgtcc
     4801 ccttctccct ctccagcctc ggggctgtcc gcggggggac ggctgccttc gggggggacg
     4861 gggcagggcg gggttcggct tctggcgtgt gaccggcggc tctagagcct ctgctaacca
     4921 tgttcatgcc ttcttctttt tcctacagat ccttaattaa taatacgact cactataggg
     4981 gccgccacca tgggacctaa gaaaaagagg aaggtggcgg ccgctgacta caaggatgac
     5041 gacgataaac caggtggcgg aggtagtggc ggaggtgggg tacccgccag tccagcagcc
     5101 caggtggatc tgagaaccct cggctacagc cagcagcagc aggagaagat caaaccaaag
     5161 gtgcggtcca ccgtcgctca gcaccatgaa gcactggtgg ggcacggttt cacacacgcc
     5221 catattgtgg ctctgtctca gcatcccgct gcactcggga ctgtggccgt caaatatcag
     5281 gacatgatcg ccgctctgcc tgaggcaacc cacgaagcca ttgtgggcgt cggaaagcag
     5341 tggagcggtg ccagagcact cgaagcactc ctcaccgtcg ccggggaact gcggggtcca
     5401 ccactccagt ccggactgga cactggacag ctgctgaaga tcgctaaacg cggcggagtg
     5461 acagctgtgg aagctgtgca cgcttggagg aatgctctga caggagcccc cctggagacg
     5521 ggcgccgcta cagggcgcgt cccattcgcc attcaggctg cgcaactgtt gggaagggcg
     5581 atcggtgcgg gcctcttcgc tattacgcca gctggcgaaa gggggatgtg ctgcaaggcg
     5641 attaagttgg gtaacgccag ggttttccca gtcacgacgt tgtaaaacga cggccagtga
     5701 gcgcgcggcg aattgggtaa cgggccaccc ctcgaggtcc tccagctttt gttgcgcgct
     5761 tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac
     5821 acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac
     5881 tcacattaat tgcgttgcgc tcactgcccg ctttccaccg gtcgtctcca acgatcatct
     5941 ggtcgctctc gcttgcctcg gtggacggcc cgctctggac gcagtcaaaa agggtctccc
     6001 ccatgctccc gcactgatca agagaaccaa caggagaatt cctgagggat ccgatcgttt
     6061 aaaccagctc gtgaaaagcg aactcgaaga aaagaaaagt gaactgcggc acaaactgaa
     6121 atacgtccca catgaataca ttgagctgat cgagattgct aggaactcca cccaggacag
     6181 aatcctcgag atgaaagtga tggaattctt tatgaaagtc tacgggtatc ggggcaagca
     6241 cctgggcgga tctcgcaaac cagatggggc aatctacact gtgggtagtc ccatcgacta
     6301 tggcgtgatt gtcgatacca aggcctacag tgggggttat aatctgccca ttggacaggc
     6361 tgacgagatg cagcgatacg tggaggaaaa ccagacaaga aataagcata tcaaccccaa
     6421 tgagtggtgg aaagtgtatc ctagctccgt cactgaattc aagtttctct tcgtgtcagg
     6481 ccactttaag ggaaactaca aagcacagct gaccaggctc aatcatatta caaactgcaa
     6541 tggcgccgtg ctgagcgtcg aggaactgct catcggcgga gagatgatca aggccggcac
     6601 actcaccctg gaggaggtcc gccgaaaatt caataacggg gaaatcaact tctgaacgcg
     6661 taaatgattg cagatccact agttctagaa ttccagctga gcgccggtcg ctaccattac
     6721 cagttggtct ggtgtcaaaa ataataataa ccgggcaggg gggatctgca tggatctttg
     6781 tgaaggaacc ttacttctgt ggtgtgacat aattggacaa actacctaca gagatttaaa
     6841 gctctaaggt aaatataaaa tttttaagtg tataatgtgt taaactactg attctaattg
     6901 tttgtgtatt ttagattcca acctatggaa ctgatgaatg ggagcagtgg tggaatgcca
     6961 gatccagaca tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga
     7021 aaaaaatgct ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc
     7081 tgcaataaac aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag
     7141 gtgtgggagg ttttttaaag caagtaaaac ctctacaaat gtggtatggc tgattatgat
     7201 ctgcggccgc
//
