LOCUS       40924_8056        8829 bp DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pCAG-scFvGCN4sfGFPTET1CD DNA, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8829)
  AUTHORS   Morita S, Noguchi H, Horii T, Nakabayashi K, Kimura M, Okamura K, 
            Sakai A, Nakashima H, Hata K, Nakashima K, Hatada I.
  TITLE     Targeted DNA demethylation in vivo using dCas9-peptide repeat and 
            scFv-TET1 catalytic domain fusions
  JOURNAL   Nat. Biotechnol. 34 (10), 1060-1065 (2016)
  PUBMED    27571369
REFERENCE   2  (bases 1 to 8829)
  AUTHORS   Hatada I, Morita S.
  TITLE     Direct Submission
  JOURNAL   Submitted (19-JUL-2016) Contact:Izuho Hatada Gunma University, 
            Institute for Molecular and Cellular Regulation; 3-39-15 
            Showa-machi, Maebashi, Gunma 371-8512, Japan URL 
            :http://epigenome.dept.showa.gunma-u.ac.jp/~hatada/
REFERENCE   3  (bases 1 to 8829)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 8829)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat. 
            Biotechnol."; date: "2016"; volume: "34"; issue: "10"; pages: 
            "1060-1065"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (19-JUL-2016) Contact:Izuho Hatada Gunma University, Institute for 
            Molecular and Cellular Regulation"; volume: " 3-39-15 Showa-machi, 
            Maebashi, Gunma 371-8512, Japan URL :http"; pages: 
            "//epigenome.dept.showa.gunma-u.ac.jp/~hatada"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..8829
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        61..364
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        367..642
                     /label=chicken beta-actin promoter
     intron          643..1651
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and
                     rabbit beta-globin"
     CDS             2463..2489
                     /codon_start=1
                     /label=HA
                     /note="HA (human influenza hemagglutinin) epitope tag"
                     /translation="YPYDVPDYA"
     CDS             2580..3290
                     /codon_start=1
                     /label=superfolder GFP
                     /note="GFP variant that folds robustly even when fused to
                     poorly folded proteins (Nager et al., 2011)"
                     /translation="SKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGT
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKAN
                     FKIRHNVEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     polyA_signal    5617..5738
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      complement(5745..6200)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        6227..6331
                     /label=AmpR promoter
     promoter        6333..6690
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             6725..7516
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    7751..7798
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      8127..8715
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     promoter        8829
                     /label=CAG
                     /note="CMV early enhancer fused to modified chicken 
                     beta-actin promoter"
ORIGIN
        1 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
       61 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      121 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      181 atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      241 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      301 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      361 catgggtcga ggtgagcccc acgttctgct tcactctccc catctccccc ccctccccac
      421 ccccaatttt gtatttattt attttttaat tattttgtgc agcgatgggg gcgggggggg
      481 ggggggcgcg cgccaggcgg ggcggggcgg ggcgaggggc ggggcggggc gaggcggaga
      541 ggtgcggcgg cagccaatca gagcggcgcg ctccgaaagt ttccttttat ggcgaggcgg
      601 cggcggcggc ggccctataa aaagcgaagc gcgcggcggg cgggagtcgc tgcgttgcct
      661 tcgccccgtg ccccgctccg cgccgcctcg cgccgcccgc cccggctctg actgaccgcg
      721 ttactcccac aggtgagcgg gcgggacggc ccttctcctc cgggctgtaa ttagcgcttg
      781 gtttaatgac ggctcgtttc ttttctgtgg ctgcgtgaaa gccttaaagg gctccgggag
      841 ggccctttgt gcggggggga gcggctcggg gggtgcgtgc gtgtgtgtgt gcgtggggag
      901 cgccgcgtgc ggcccgcgct gcccggcggc tgtgagcgct gcgggcgcgg cgcggggctt
      961 tgtgcgctcc gcgtgtgcgc gaggggagcg cggccggggg cggtgccccg cggtgcgggg
     1021 gggctgcgag gggaacaaag gctgcgtgcg gggtgtgtgc gtgggggggt gagcaggggg
     1081 tgtgggcgcg gcggtcgggc tgtaaccccc ccctgcaccc ccctccccga gttgctgagc
     1141 acggcccggc ttcgggtgcg gggctccgtg cggggcgtgg cgcggggctc gccgtgccgg
     1201 gcggggggtg gcggcaggtg ggggtgccgg gcggggcggg gccgcctcgg gccggggagg
     1261 gctcggggga ggggcgcggc ggccccggag cgccggcggc tgtcgaggcg cggcgagccg
     1321 cagccattgc cttttatggt aatcgtgcga gagggcgcag ggacttcctt tgtcccaaat
     1381 ctggcggagc cgaaatctgg gaggcgccgc cgcaccccct ctagcgggcg cgggcgaagc
     1441 ggtgcggcgc cggcaggaag gaaatgggcg gggagggcct tcgtgcgtcg ccgcgccgcc
     1501 gtccccttct ccatctccag cctcggggct gccgcagggg gacggctgcc ttcggggggg
     1561 acggggcagg gcggggttcg gcttctggcg tgtgaccggc ggctctagag cctctgctaa
     1621 ccatgttcat gccttcttct ttttcctaca gctcctgggc aacgtgctgg ttgttgtgct
     1681 gtctcatcat tttggcaaag aattctgcag tcgacggtac catgggcccc gacatcgtga
     1741 tgacccagag ccccagcagc ctgagcgcca gcgtgggcga ccgcgtgacc atcacctgcc
     1801 gcagcagcac cggcgccgtg accaccagca actacgccag ctgggtgcag gagaagcccg
     1861 gcaagctgtt caagggcctg atcggcggca ccaacaaccg cgcccccggc gtgcccagcc
     1921 gcttcagcgg cagcctgatc ggcgacaagg ccaccctgac catcagcagc ctgcagcccg
     1981 aggacttcgc cacctacttc tgcgccctgt ggtacagcaa ccactgggtg ttcggccagg
     2041 gcaccaaggt ggagctgaag cgcggcggcg gcggcagcgg cggcggcggc agcggcggcg
     2101 gcggcagcag cggcggcggc agcgaggtga agctgctgga gagcggcggc ggcctggtgc
     2161 agcccggcgg cagcctgaag ctgagctgcg ccgtgagcgg cttcagcctg accgactacg
     2221 gcgtgaactg ggtgcgccag gcccccggcc gcggcctgga gtggatcggc gtgatctggg
     2281 gcgacggcat caccgactac aacagcgccc tgaaggaccg cttcatcatc agcaaggaca
     2341 acggcaagaa caccgtgtac ctgcagatga gcaaggtgcg cagcgacgac accgccctgt
     2401 actactgcgt gaccggcctg ttcgactact ggggccaggg caccctggtg accgtgagca
     2461 gctacccata cgatgttcca gattacgctg gtggaggcgg aggttctggg ggaggaggta
     2521 gtggcggtgg tggttcagga ggcggcggaa gcttggatcc aggtggaggt ggaagcggta
     2581 gcaaaggaga agaacttttc actggagttg tcccaattct tgttgaatta gatggtgatg
     2641 ttaatgggca caaattttct gtccgtggag agggtgaagg tgatgctaca aacggaaaac
     2701 tcacccttaa atttatttgc actactggaa aactacctgt tccgtggcca acacttgtca
     2761 ctactctgac ctatggtgtt caatgctttt cccgttatcc ggatcacatg aaacggcatg
     2821 actttttcaa gagtgccatg cccgaaggtt atgtacagga acgcactata tctttcaaag
     2881 atgacgggac ctacaagacg cgtgctgaag tcaagtttga aggtgatacc cttgttaatc
     2941 gtatcgagtt aaagggtatt gattttaaag aagatggaaa cattcttgga cacaaactcg
     3001 agtacaactt taactcacac aatgtataca tcacggcaga caaacaaaag aatggaatca
     3061 aagctaactt caaaattcgc cacaacgttg aagatggttc cgttcaacta gcagaccatt
     3121 atcaacaaaa tactccaatt ggcgatggcc ctgtcctttt accagacaac cattacctgt
     3181 cgacacaatc tgtcctttcg aaagatccca acgaaaagcg tgaccacatg gtccttcttg
     3241 agtttgtaac tgctgctggg attacacatg gcatggatga gctctacaaa ggtggaggtc
     3301 ggaccggtgg cggtggcgga ggggctagca gatccgaact gcccacctgc agctgtcttg
     3361 atcgagttat acaaaaagac aaaggcccat attatacaca ccttggggca ggaccaagtg
     3421 ttgctgctgt cagggaaatc atggagaata ggtatggtca aaaaggaaac gcaataagga
     3481 tagaaatagt agtgtacacc ggtaaagaag ggaaaagctc tcatgggtgt ccaattgcta
     3541 agtgggtttt aagaagaagc agtgatgaag aaaaagttct ttgtttggtc cggcagcgta
     3601 caggccacca ctgtccaact gctgtgatgg tggtgctcat catggtgtgg gatggcatcc
     3661 ctcttccaat ggccgaccgg ctatacacag agctcacaga gaatctaaag tcatacaatg
     3721 ggcaccctac cgacagaaga tgcaccctca atgaaaatcg tacctgtaca tgtcaaggaa
     3781 ttgatccaga gacttgtgga gcttcattct cttttggctg ttcatggagt atgtacttta
     3841 atggctgtaa gtttggtaga agcccaagcc ccagaagatt tagaattgat ccaagctctc
     3901 ccttacatga aaaaaacctt gaagataact tacagagttt ggctacacga ttagctccaa
     3961 tttataagca gtatgctcca gtagcttacc aaaatcaggt ggaatatgaa aatgttgccc
     4021 gagaatgtcg gcttggcagc aaggaaggtc gtcccttctc tggggtcact gcttgcctgg
     4081 acttctgtgc tcatccccac agggacattc acaacatgaa taatggaagc actgtggttt
     4141 gtaccttaac tcgagaagat aaccgctctt tgggtgttat tcctcaagat gagcagctcc
     4201 atgtgctacc tctttataag ctttcagaca cagatgagtt tggctccaag gaaggaatgg
     4261 aagccaagat caaatctggg gccatcgagg tcctggcacc ccgccgcaaa aaaagaacgt
     4321 gtttcactca gcctgttccc cgttctggaa agaagagggc tgcgatgatg acagaggttc
     4381 ttgcacataa gataagggca gtggaaaaga aacctattcc ccgaatcaag cggaagaata
     4441 actcaacaac aacaaacaac agtaagcctt cgtcactgcc aaccttaggg agtaacactg
     4501 agaccgtgca acctgaagta aaaagtgaaa ccgaacccca ttttatctta aaaagttcag
     4561 acaacactaa aacttattcg ctgatgccat ccgctcctca cccagtgaaa gaggcatctc
     4621 caggcttctc ctggtccccg aagactgctt cagccacacc agctccactg aagaatgacg
     4681 caacagcctc atgcgggttt tcagaaagaa gcagcactcc ccactgtacg atgccttcgg
     4741 gaagactcag tggtgccaat gcagctgctg ctgatggccc tggcatttca cagcttggcg
     4801 aagtggctcc tctccccacc ctgtctgctc ctgtgatgga gcccctcatt aattctgagc
     4861 cttccactgg tgtgactgag ccgctaacgc ctcatcagcc aaaccaccag ccctccttcc
     4921 tcacctctcc tcaagacctt gcctcttctc caatggaaga agatgagcag cattctgaag
     4981 cagatgagcc tccatcagac gaacccctat ctgatgaccc cctgtcacct gctgaggaga
     5041 aattgcccca cattgatgag tattggtcag acagtgagca catctttttg gatgcaaata
     5101 ttggtggggt ggccatcgca cctgctcacg gctcggtttt gattgagtgt gcccggcgag
     5161 agctgcacgc taccactcct gttgagcacc ccaaccgtaa tcatccaacc cgcctctccc
     5221 ttgtctttta ccagcacaaa aacctaaata agccccaaca tggttttgaa ctaaacaaga
     5281 ttaagtttga ggctaaagaa gctaagaata agaaaatgaa ggcctcagag caaaaagacc
     5341 aggcagctaa tgaaggtcca gaacagtcct ctgaagtaaa tgaattgaac caaattcctt
     5401 ctcataaagc attaacatta acccatgaca atgttgtcac cgtgtcccct tatgctctca
     5461 cacacgttgc ggggccctat aaccattggg tctgagcggc cgcgactcta gatcataatc
     5521 agccatacca catttgtaga ggttttactt gctttaaaaa acctcccaca cctccccctg
     5581 aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat
     5641 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     5701 tctagttgtg gtttgtccaa actcatcaat gtatcttaag gcgtaaattg taagcgttaa
     5761 tattttgtta aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaataggc
     5821 cgaaatcggc aaaatccctt ataaatcaaa agaatagacc gagatagggt tgagtgttgt
     5881 tccagtttgg aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa
     5941 aaccgtctat cagggcgatg gcccactacg tgaaccatca ccctaatcaa gttttttggg
     6001 gtcgaggtgc cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg
     6061 acggggaaag ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc
     6121 tagggcgctg gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa
     6181 tgcgccgcta cagggcgcgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     6241 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     6301 aatgcttcaa taatattgaa aaaggaagag tcctgaggcg gaaagaacca gctgtggaat
     6361 gtgtgtcagt tagggtgtgg aaagtcccca ggctccccag caggcagaag tatgcaaagc
     6421 atgcatctca attagtcagc aaccaggtgt ggaaagtccc caggctcccc agcaggcaga
     6481 agtatgcaaa gcatgcatct caattagtca gcaaccatag tcccgcccct aactccgccc
     6541 atcccgcccc taactccgcc cagttccgcc cattctccgc cccatggctg actaattttt
     6601 tttatttatg cagaggccga ggccgcctcg gcctctgagc tattccagaa gtagtgagga
     6661 ggcttttttg gaggcctagg cttttgcaaa gatcgatcaa gagacaggat gaggatcgtt
     6721 tcgcatgatt gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct
     6781 attcggctat gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct
     6841 gtcagcgcag gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga
     6901 actgcaagac gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc
     6961 tgtgctcgac gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg
     7021 gcaggatctc ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc
     7081 aatgcggcgg ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca
     7141 tcgcatcgag cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga
     7201 cgaagagcat caggggctcg cgccagccga actgttcgcc aggctcaagg cgagcatgcc
     7261 cgacggcgag gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga
     7321 aaatggccgc ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca
     7381 ggacatagcg ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg
     7441 cttcctcgtg ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct
     7501 tcttgacgag ttcttctgag cgggactctg gggttcgaaa tgaccgacca agcgacgccc
     7561 aacctgccat cacgagattt cgattccacc gccgccttct atgaaaggtt gggcttcgga
     7621 atcgttttcc gggacgccgg ctggatgatc ctccagcgcg gggatctcat gctggagttc
     7681 ttcgcccacc ctagggggag gctaactgaa acacggaagg agacaatacc ggaaggaacc
     7741 cgcgctatga cggcaataaa aagacagaat aaaacgcacg gtgttgggtc gtttgttcat
     7801 aaacgcgggg ttcggtccca gggctggcac tctgtcgata ccccaccgag accccattgg
     7861 ggccaatacg cccgcgtttc ttccttttcc ccaccccacc ccccaagttc gggtgaaggc
     7921 ccagggctcg cagccaacgt cggggcggca ggccctgcca tagcctcagg ttactcatat
     7981 atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt
     8041 tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac
     8101 cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc
     8161 ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca
     8221 actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgtccttcta
     8281 gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct
     8341 ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg
     8401 gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc
     8461 acacagccca gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta
     8521 tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg
     8581 gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt
     8641 cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg
     8701 cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg
     8761 ccttttgctc acatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc
     8821 gccatgcat
//
