LOCUS       sequence-102893-        6840 bp    DNA     circular SYN 15-APR-2025
DEFINITION  TALEN array receiving vector with half-site NN RVD.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6840)
  AUTHORS   Martinez RA, Stein JL, Krostag AR, Nelson AM, Marken JS, Menon V, 
            May RC, Yao Z, Kaykas A, Geschwind DH, Grimley JS
  TITLE     Genome engineering of isogenic human ES cells to model autism 
            disorders.
  JOURNAL   Nucleic Acids Res. 2015 Mar 11. pii: gkv164.
  PUBMED    25765640
REFERENCE   2  (bases 1 to 6840)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Apr 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nucleic 
            Acids Res. 2015 Mar 11. pii: gkv164."
COMMENT     Sequence Label: AI-CN491
FEATURES             Location/Qualifiers
     source          1..6840
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             907..972
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             979..999
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             complement(1134..1715)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWRTSRGGPVPNSPYSESYYARSLAVVL
                     QRRDWENPGVTQLNRLAAHPPFASWRNSEEARTDRPSQQLRSLNGEWEDVFKGAPVSAL
                     RQACTASTAVTPPLFAIFSSCPVSSWSGGPLSSPATVSSASSARAPDHCLPTPTIASCV
                     ASGRAAIISWYLTATVPSAAGCCESATICA"
     primer_bind     1565..1587
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     1579..1596
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     1580..1596
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        1606..1624
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1672..1692)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(1672..1690)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(1711..1727)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    1735..1751
                     /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(1759..1765,1766..1783,1784..1789))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1804..1825
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             2101..2688
                     /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"
     CDS             2704..2745
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2755..2772
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2795..2812)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2801..3025
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3071..3499
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3158..3177)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3368..3389
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3508..3528)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3513..3843
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3694..3829
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3756..3775
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     promoter        3891..3938
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter "
     CDS             3957..4355
                     /codon_start=1
                     /gene="Aspergillus terreus BSD"
                     /product="blasticidin S deaminase"
                     /label=BSD
                     /note="confers resistance to blasticidin"
                     /translation="MAKPLSQEESTLIERATATINSIPISEHYSVASAALSSDGRIFTG
                     VNVYHFTGGPCAELVVLGTAAAAAAGNLTCIVAIGNENRGILSPCGRCRQVLLDLHPGI
                     KAIVKDSDGQPTAVGIRELLPSGYVWEG"
     polyA_signal    4513..4634
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4550..4569)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4604..4623
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4683..4699)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4707..4723
                     /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(4731..4737,4738..4755,4756..4761))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4776..4797
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4914..4931)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5085..5673)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5165..5184)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5844..6635,6636..6704))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     6467..6486
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6705..6809)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 accatggact acaaagacca tgacggtgat tataaagatc atgacatcga ttacaaggat
      961 gacgatgaca agatggcccc caagaagaag aggaaggtgg gcattcaccg cggggtacct
     1021 atggtggact tgaggacact cggttattcg caacagcaac aggagaaaat caagcctaag
     1081 gtcaggagca ccgtcgcgca acaccacgag gcgcttgtgg ggcatggctt cactcatgcg
     1141 catattgtcg cgctttcaca gcaccctgcg gcgcttggga cggtggctgt caaataccaa
     1201 gatatgattg cggccctgcc cgaagccacg cacgaggcaa ttgtaggggt cggtaaacag
     1261 tggtcgggag cgcgagcact tgaggcgctg ctgactgtgg cgggtgagct tagggggcct
     1321 ccgctccagc tcgacaccgg gcagctgctg aagatcgcga agagaggggg agtaacagcg
     1381 gtagaggcag tgcacgcctg gcgcaatgcg ctcaccgggg cccccttgaa cacgtcttcc
     1441 cattcgccat tcaggctgcg caactgttgg gaagggcgat cggtgcgggc ctcttcgcta
     1501 ttacgccagc tggcgaaagg gggatgtgct gcaaggcgat taagttgggt aacgccaggg
     1561 ttttcccagt cacgacgttg taaaacgacg gccagtgagc gcgcgtaata cgactcacta
     1621 tagggcgaat tgggtaccgg gccccccctc gaggtcctcc agcttttgtt ccctttagtg
     1681 agggttaatt gcgcgcttgg cgtaatcatg gtcatagctg tttcctgtgt gaaattgtta
     1741 tccgctcaca attccacaca acatacgagc cggaagcata aagtgtaaag cctggggtgc
     1801 ctaatgagtg agctaactca cattaattgc gttgcgcgaa gacacctgac acccgaacag
     1861 gtggtcgcca ttgctaataa taacggagga cggccagcct tggagtccat cgtagcccaa
     1921 ttgtccaggc ccgatcccgc gttggctgcg ttaacgaatg accatctggt ggcgttggca
     1981 tgtcttggtg gacgacccgc gctcgatgca gtcaaaaagg gtctgcctca tgctcccgca
     2041 ttgatcaaaa gaaccaaccg gcggattccc gagagaactt cccatcgagt cgcgggatcc
     2101 caactagtca aaagtgaact ggaggagaag aaatctgaac ttcgtcataa attgaaatat
     2161 gtgcctcatg aatatattga attaattgaa attgccagaa attccactca ggatagaatt
     2221 cttgaaatga aggtaatgga attttttatg aaagtttatg gatatagagg taaacatttg
     2281 ggtggatcaa ggaaaccgga cggagcaatt tatactgtcg gatctcctat tgattacggt
     2341 gtgatcgtgg atactaaagc ttatagcgga ggttataatc tgccaattgg ccaagcagat
     2401 gaaatgcaac gatatgtcga agaaaatcaa acacgaaaca aacatatcaa ccctaatgaa
     2461 tggtggaaag tctatccatc ttctgtaacg gaatttaagt ttttatttgt gagtggtcac
     2521 tttaaaggaa actacaaagc tcagcttaca cgattaaatc atatcactaa ttgtaatgga
     2581 gctgttctta gtgtagaaga gcttttaatt ggtggagaaa tgattaaagc cggcacatta
     2641 accttagagg aagtcagacg gaaatttaat aacggcgaga taaactttta agggcccttc
     2701 gaaggtaagc ctatccctaa ccctctcctc ggtctcgatt ctacgcgtac cggtcatcat
     2761 caccatcacc attgagttta aacccgctga tcagcctcga ctgtgccttc tagttgccag
     2821 ccatctgttg tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact
     2881 gtcctttcct aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt
     2941 ctggggggtg gggtggggca ggacagcaag ggggaggatt gggaacacaa tagcaggcat
     3001 gctggggatg cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg
     3061 gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     3121 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     3181 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     3241 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     3301 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     3361 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     3421 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     3481 caaaaattta acgcgaatta attctgtgga atgtgtgtca gttagggtgt ggaaagtccc
     3541 caggctcccc aggcaggcag aagtatgcaa agcatgcatc tcaattagtc agcaaccagg
     3601 tgtggaaagt ccccaggctc cccagcaggc agaagtatgc aaagcatgca tctcaattag
     3661 tcagcaacca tagtcccgcc cctaactccg cccatcccgc ccctaactcc gcccagttcc
     3721 gcccattctc cgccccatgg ctgactaatt ttttttattt atgcagaggc cgaggccgcc
     3781 tctgcctctg agctattcca gaagtagtga ggaggctttt ttggaggcct aggcttttgc
     3841 aaaaagctcc cgggagcttg tatatccatt ttcggatctg atcagcacgt gttgacaatt
     3901 aatcatcggc atagtatatc ggcatagtat aatacgacaa ggtgaggaac taaaccatgg
     3961 ccaagccttt gtctcaagaa gaatccaccc tcattgaaag agcaacggct acaatcaaca
     4021 gcatccccat ctctgaacac tacagcgtcg ccagcgcagc tctctctagc gacggccgca
     4081 tcttcactgg tgtcaatgta tatcatttta ctgggggacc ttgtgcagaa ctcgtggtgc
     4141 tgggcactgc tgctgctgcg gcagctggca acctgacttg tatcgtcgcg atcggaaatg
     4201 agaacagggg catcttgagc ccctgcggac ggtgccgaca ggtgcttctc gatctgcatc
     4261 ctgggatcaa agccatagtg aaggacagtg atggacagcc gacggcagtt gggattcgtg
     4321 aattgctgcc ctctggttat gtgtgggagg gctaagcact tcgtggccga ggagcaggac
     4381 tgacacgtgc tacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga
     4441 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc
     4501 ttcgcccacc ccaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     4561 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     4621 atcaatgtat cttatcatgt ctgtataccg tcgacctcta gctagagctt ggcgtaatca
     4681 tggtcatagc tgtttcctgt gtgaaattgt tatccgctca caattccaca caacatacga
     4741 gccggaagca taaagtgtaa agcctggggt gcctaatgag tgagctaact cacattaatt
     4801 gcgttgcgct cactgcccgc tttccagtcg ggaaacctgt cgtgccagct gcattaatga
     4861 atcggccaac gcgcggggag aggcggtttg cgtattgggc gctcttccgc ttcctcgctc
     4921 actgactcgc tgcgctcggt cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg
     4981 gtaatacggt tatccacaga atcaggggat aacgcaggaa agaacatgtg agcaaaaggc
     5041 cagcaaaagg ccaggaaccg taaaaaggcc gcgttgctgg cgtttttcca taggctccgc
     5101 ccccctgacg agcatcacaa aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga
     5161 ctataaagat accaggcgtt tccccctgga agctccctcg tgcgctctcc tgttccgacc
     5221 ctgccgctta ccggatacct gtccgccttt ctcccttcgg gaagcgtggc gctttctcaa
     5281 tgctcacgct gtaggtatct cagttcggtg taggtcgttc gctccaagct gggctgtgtg
     5341 cacgaacccc ccgttcagcc cgaccgctgc gccttatccg gtaactatcg tcttgagtcc
     5401 aacccggtaa gacacgactt atcgccactg gcagcagcca ctggtaacag gattagcaga
     5461 gcgaggtatg taggcggtgc tacagagttc ttgaagtggt ggcctaacta cggctacact
     5521 agaaggacag tatttggtat ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt
     5581 ggtagctctt gatccggcaa acaaaccacc gctggtagcg gtggtttttt tgtttgcaag
     5641 cagcagatta cgcgcagaaa aaaaggatct caagaagatc ctttgatctt ttctacgggg
     5701 tctgacgctc agtggaacga aaactcacgt taagggattt tggtcatgag attatcaaaa
     5761 aggatcttca cctagatcct tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata
     5821 tatgagtaaa cttggtctga cagttaccaa tgcttaatca gtgaggcacc tatctcagcg
     5881 atctgtctat ttcgttcatc catagttgcc tgactccccg tcgtgtagat aactacgata
     5941 cgggagggct taccatctgg ccccagtgct gcaatgatac cgcgagaccc acgctcaccg
     6001 gctccagatt tatcagcaat aaaccagcca gccggaaggg ccgagcgcag aagtggtcct
     6061 gcaactttat ccgcctccat ccagtctatt aattgttgcc gggaagctag agtaagtagt
     6121 tcgccagtta atagtttgcg caacgttgtt gccattgcta caggcatcgt ggtgtcacgc
     6181 tcgtcgtttg gtatggcttc attcagctcc ggttcccaac gatcaaggcg agttacatga
     6241 tcccccatgt tgtgcaaaaa agcggttagc tccttcggtc ctccgatcgt tgtcagaagt
     6301 aagttggccg cagtgttatc actcatggtt atggcagcac tgcataattc tcttactgtc
     6361 atgccatccg taagatgctt ttctgtgact ggtgagtact caaccaagtc attctgagaa
     6421 tagtgtatgc ggcgaccgag ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca
     6481 catagcagaa ctttaaaagt gctcatcatt ggaaaacgtt cttcggggcg aaaactctca
     6541 aggatcttac cgctgttgag atccagttcg atgtaaccca ctcgtgcacc caactgatct
     6601 tcagcatctt ttactttcac cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc
     6661 gcaaaaaagg gaataagggc gacacggaaa tgttgaatac tcatactctt cctttttcaa
     6721 tattattgaa gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt
     6781 tagaaaaata aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc
//
