LOCUS       Exported                8829 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pCAG-scFvGCN4sfGFPTET1CD DNA, complete sequence.
ACCESSION   LC169511
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)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..8829
                     /organism="Cloning vector pCAG-scFvGCN4sfGFPTET1CD"
                     /mol_type="other DNA"
                     /db_xref="taxon:1882229"
     regulatory      1..1699
                     /regulatory_class="promoter"
                     /note="CAG promoter"
     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             1722..5495
                     /codon_start=1
                     /product="scFvGCN4sfGFPTET1CD"
                     /label=scFvGCN4sfGFPTET1CD
                     /protein_id="BAV54122.1"
                     /translation="MGPDIVMTQSPSSLSASVGDRVTITCRSSTGAVTTSNYASWVQEK
                     PGKLFKGLIGGTNNRAPGVPSRFSGSLIGDKATLTISSLQPEDFATYFCALWYSNHWVF
                     GQGTKVELKRGGGGSGGGGSGGGGSSGGGSEVKLLESGGGLVQPGGSLKLSCAVSGFSL
                     TDYGVNWVRQAPGRGLEWIGVIWGDGITDYNSALKDRFIISKDNGKNTVYLQMSKVRSD
                     DTALYYCVTGLFDYWGQGTLVTVSSYPYDVPDYAGGGGGSGGGGSGGGGSGGGGSLDPG
                     GGGSGSKGEELFTGVVPILVELDGDVNGHKFSVRGEGEGDATNGKLTLKFICTTGKLPV
                     PWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTISFKDDGTYKTRAEVKF
                     EGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYITADKQKNGIKANFKIRHNVED
                     GSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSVLSKDPNEKRDHMVLLEFVTAAGITHG
                     MDELYKGGGRTGGGGGGASRSELPTCSCLDRVIQKDKGPYYTHLGAGPSVAAVREIMEN
                     RYGQKGNAIRIEIVVYTGKEGKSSHGCPIAKWVLRRSSDEEKVLCLVRQRTGHHCPTAV
                     MVVLIMVWDGIPLPMADRLYTELTENLKSYNGHPTDRRCTLNENRTCTCQGIDPETCGA
                     SFSFGCSWSMYFNGCKFGRSPSPRRFRIDPSSPLHEKNLEDNLQSLATRLAPIYKQYAP
                     VAYQNQVEYENVARECRLGSKEGRPFSGVTACLDFCAHPHRDIHNMNNGSTVVCTLTRE
                     DNRSLGVIPQDEQLHVLPLYKLSDTDEFGSKEGMEAKIKSGAIEVLAPRRKKRTCFTQP
                     VPRSGKKRAAMMTEVLAHKIRAVEKKPIPRIKRKNNSTTTNNSKPSSLPTLGSNTETVQ
                     PEVKSETEPHFILKSSDNTKTYSLMPSAPHPVKEASPGFSWSPKTASATPAPLKNDATA
                     SCGFSERSSTPHCTMPSGRLSGANAAAADGPGISQLGEVAPLPTLSAPVMEPLINSEPS
                     TGVTEPLTPHQPNHQPSFLTSPQDLASSPMEEDEQHSEADEPPSDEPLSDDPLSPAEEK
                     LPHIDEYWSDSEHIFLDANIGGVAIAPAHGSVLIECARRELHATTPVEHPNRNHPTRLS
                     LVFYQHKNLNKPQHGFELNKIKFEAKEAKNKKMKASEQKDQAANEGPEQSSEVNELNQI
                     PSHKALTLTHDNVVTVSPYALTHVAGPYNHWV"
     CDS             2463..2489
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2580..3290
                     /codon_start=1
                     /product="GFP variant that folds robustly even when fused 
                     to poorly folded proteins (Pedelacq et al., 2006)"
                     /label=superfolder GFP
                     /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
                     /gene="bla"
                     /label=AmpR promoter
     promoter        6333..6690
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6541..6676
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6725..7519
                     /codon_start=1
                     /product="neomycin/kanamycin resistance protein"
                     /label=neomycin/kanamycin resistance protein
                     /protein_id="BAV54123.1"
                     /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
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"