LOCUS       Exported                8103 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pEFBOS-IRESGFPNeo, complete sequence.
ACCESSION   EF459695
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8103)
  AUTHORS   Lim SM, Stanley EG, Elefanty AG.
  TITLE     pEFBOS-IRESGFPNeo, a derivative of pEFBOS containing an IRESGFPNeo 
            cassette
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 8103)
  AUTHORS   Lim SM, Stanley EG, Elefanty AG.
  TITLE     Direct Submission
  JOURNAL   Submitted (27-FEB-2007) Monash Immunology and Stem Cell 
            Laboratories, Monash University, West Ring Road, Clayton, Vic 3800, 
            Australia
REFERENCE   3  (bases 1 to 8103)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..8103
                     /organism="Cloning vector pEFBOS-IRESGFPNeo"
                     /mol_type="other DNA"
                     /note="contains multiple cloning sites and IRES-GFPNeo 
                     cassette"
                     /db_xref="taxon:433942"
     protein_bind    107..128
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        143..173
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    181..197
                     /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)."
     primer_bind     205..221
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      239..1743
                     /regulatory_class="promoter"
                     /note="EF1-alpha promoter"
     promoter        complement(325..521)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     promoter        complement(333..543)
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      339..474
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     promoter        562..1743
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          792..1734
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     misc_feature    1826..2412
                     /label=IRES2
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     misc_structure  1837..2412
                     /label=internal ribosomal entry site (IRES)
                     /note="internal ribosomal entry site (IRES)"
     CDS             2413..3936
                     /codon_start=1
                     /product="GFPNeo fusion protein"
                     /label=GFPNeo fusion protein
                     /note="GFP-neomycin resistance fusion; confers GFP 
                     expression for detection under fluorescence microscopy and 
                     G418 (neo) resistance; GFPneo"
                     /protein_id="ABO65261.1"
                     /translation="MARQSKGEELFTGVVPILVELDGDVNGYKFSVSGEGEGDATYGKL
                     TLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFK
                     DDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNG
                     IKANFKTRHNIEDGGVQLADHYQQNTPIGDDPVLLPDNHYLSTQSALSKDPNEKRDHMV
                     LLEFVTAAGITHGMDELYKTRGIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFR
                     LSAQGRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEV
                     PGQDLLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQD
                     DLDEEHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVA
                     DRYQDIALATRDIAEELCGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             2425..3135
                     /codon_start=1
                     /gene="mgfp5"
                     /product="GFP with folding enhancement mutations"
                     /label=mgfp5
                     /note="suitable for expression in plants due to removal of 
                     a cryptic intron"
                     /translation="SKGEELFTGVVPILVELDGDVNGYKFSVSGEGEGDATYGKLTLKF
                     ICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKRHDFFKSAMPEGYVQERTIFFKDDGN
                     YKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKAN
                     FKTRHNIEDGGVQLADHYQQNTPIGDDPVLLPDNHYLSTQSALSKDPNEKRDHMVLLEF
                     VTAAGITHGMDELYK"
     CDS             3145..3936
                     /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="IEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRPV
                     LFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLSS
                     HLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQG
                     LAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIAL
                     ATRDIAEELCGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     3UTR           4523..5223
                     /label=bovine growth hormone 3 UTR
                     /note="bovine growth hormone 3 UTR"
     misc_feature    5226..8103
                     /label=plasmid backbone from pUC8
                     /note="plasmid backbone from pUC8"
     primer_bind     complement(5232..5248)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      5461..5916
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        6198..6302
                     /gene="bla"
                     /label=AmpR promoter
     CDS             6303..7163
                     /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"
     rep_origin      7334..7922
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"