LOCUS       Exported                5568 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pPGKneo-II, complete sequence.
ACCESSION   AF335420
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5568)
  AUTHORS   Graubert TA, Lu ZH, Ley TJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (11-JAN-2001) Division of Oncology, Section of Stem Cell 
            Biology, Washington University School of Medicine, 660 South Euclid 
            Avenue, Campus Box 8007, St. Louis, MO 63110-1093, USA
REFERENCE   2  (bases 1 to 5568)
  AUTHORS   Graubert TA, Lu ZH, Ley TJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (12-MAR-2002) Division of Oncology, Section of Stem Cell 
            Biology, Washington University School of Medicine, 660 South Euclid 
            Avenue, Campus Box 8007, St. Louis, MO 63110-1093, USA
REFERENCE   3  (bases 1 to 5568)
  AUTHORS   Graubert TA, Lu ZH, Ley TJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (25-JUL-2002) Division of Oncology, Section of Stem Cell 
            Biology, Washington University School of Medicine, 660 South Euclid 
            Avenue, Campus Box 8007, St. Louis, MO 63110-1093, USA
REFERENCE   4  (bases 1 to 5568)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     On Jul 25, 2002 this sequence version replaced AF335420.2.
FEATURES             Location/Qualifiers
     source          1..5568
                     /organism="Cloning vector pPGKneo-II"
                     /mol_type="other DNA"
                     /db_xref="taxon:155139"
     misc_feature    1..5568
                     /note="loxP-PGKneo-loxP cassette was generated by 
                     amplification of original Soriano PGKneo template with PCR 
                     primers containing 34 bp loxP sequence and cloned into 
                     pCRII with PGK promoter on the side of T7 priming site"
     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"
     promoter        239..257
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     misc_feature    complement(343..376)
                     /label=lox-P site
                     /note="lox-P site"
     protein_bind    343..376
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     misc_feature    complement(378..574)
                     /label=bovine growth hormone polyA signal
                     /note="bovine growth hormone polyA signal"
     polyA_signal    464..575
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     CDS             complement(613..1416)
                     /codon_start=1
                     /product="neomycin phosphotransferase"
                     /label=neomycin phosphotransferase
                     /protein_id="AAK28136.1"
                     /translation="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     regulatory      complement(1425..1920)
                     /regulatory_class="promoter"
                     /note="PGK"
     misc_feature    complement(1921..1954)
                     /label=lox-P site
                     /note="lox-P site"
     protein_bind    1921..1954
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     promoter        complement(2025..2043)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(2050..2066)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      2208..2588
                     /direction=RIGHT
                     /label=M13 ori
                     /note="M13 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     CDS             2956..3750
                     /codon_start=1
                     /product="aminoglycoside 3-phosphotransferase"
                     /function="kanamycin resistance"
                     /label=aminoglycoside 3-phosphotransferase
                     /protein_id="AAK28134.1"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SRLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVIHGDACLPNIMVENGRFSGFNDCGRLGVADRYQDIA
                     LDTRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             3768..4628
                     /codon_start=1
                     /product="beta-lactamase"
                     /function="ampicillin resistance"
                     /label=beta-lactamase
                     /protein_id="AAK28135.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCHTLLSRIDAGQEQLGRRARYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDESDTTMPVAMPTTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      4799..5387
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"