LOCUS       Exported                6741 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector ATG-less TV, complete sequence.
ACCESSION   JQ394983
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6741)
  AUTHORS   Konishi Y, Karnan S, Takahashi M, Ota A, Damdindorj L, Hosokawa Y, 
            Konishi H.
  TITLE     A system for the measurement of gene targeting efficiency in human 
            cell lines using an antibiotic resistance-GFP fusion gene
  JOURNAL   BioTechniques 53 (3), 141-152 (2012)
  PUBMED    22963476
REFERENCE   2  (bases 1 to 6741)
  AUTHORS   Konishi H.
  TITLE     Direct Submission
  JOURNAL   Submitted (07-JAN-2012) Aichi Medical University School of Medicine,
            1-1 Yazako-Karimta, Nagakute, Aichi 480-1195, Japan
REFERENCE   3  (bases 1 to 6741)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6741
                     /organism="Cloning vector ATG-less TV"
                     /mol_type="other DNA"
                     /db_xref="taxon:1249020"
     misc_feature    1..141
                     /label=ITR
                     /note="ITR"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     gene            181..896
                     /gene="EGFP"
                     /label=EGFP
     CDS             183..899
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             183..896
                     /codon_start=1
                     /gene="EGFP"
                     /product="EGFP"
                     /label=EGFP
                     /protein_id="AFV73970.1"
                     /translation="MSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     900..916
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    918..951
                     /label=loxP
                     /note="loxP"
     protein_bind    918..951
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    953..1248
                     /label=synthetic intron
                     /note="synthetic intron"
     intron          989..1218
                     /label=chimeric intron
                     /note="chimera between introns from adenovirus and 
                     immunoglobulin heavy chain genes"
     misc_feature    1258..1809
                     /label=IRES
                     /note="IRES"
     misc_feature    1347..1809
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     CDS             1823..2617
                     /codon_start=1
                     /product="neomycin phosphotransferase"
                     /label=neomycin phosphotransferase
                     /note="neomycin resistance gene"
                     /protein_id="AFV73969.1"
                     /translation="MAEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    2639..2773
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     misc_feature    2639..2770
                     /label=SV40 early mRNA polyA
                     /note="SV40 early mRNA polyA"
     misc_feature    2870..2903
                     /label=loxP
                     /note="loxP"
     protein_bind    2870..2903
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     primer_bind     complement(2956..2972)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    2975..3993
                     /note="partial SV40 largeT; an internal portion of SV40 
                     largeT exon 2"
     misc_feature    4004..4144
                     /label=ITR
                     /note="ITR"
     repeat_region   4004..4144
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     rep_origin      4219..4674
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        4956..5060
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5061..5921
                     /codon_start=1
                     /gene="ampR"
                     /product="beta-lactamase"
                     /label=ampR
                     /note="ampicilin resistance protein"
                     /protein_id="AFV73968.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            5061..5921
                     /gene="ampR"
                     /label=ampR
     rep_origin      6092..6680
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"