LOCUS       Exported               13468 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Targeting vector pHPRT8.9-Neo2(-) DNA, complete sequence.
ACCESSION   LC269955
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 13468)
  AUTHORS   Saito S, Adachi N.
  TITLE     Gene-targeting assay at the HPRT locus
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 13468)
  AUTHORS   Adachi N, Saito S.
  TITLE     Direct Submission
  JOURNAL   Submitted (14-MAY-2017) Contact:Shinta Saito Yokohama City 
            University, Graduate school of Nanobioscience; 22-2 Seto, 
            Kanazawa-ku, Yokohama, Kanagawa 236-0027, Japan
REFERENCE   3  (bases 1 to 13468)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..13468
                     /organism="Targeting vector pHPRT8.9-Neo2(-)"
                     /mol_type="other DNA"
                     /db_xref="taxon:2003189"
     rep_origin      4..459
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     600..616
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        626..644
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    653..4474
                     /gene="HPRT"
                     /label=5 arm
                     /note="5 arm"
     gene            653..4474
                     /gene="HPRT"
                     /label=HPRT
     CDS             2966..2977
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     regulatory      complement(4489..4780)
                     /regulatory_class="polyA_signal_sequence"
                     /note="PGK poly(A) signal"
     CDS             complement(4789..5592)
                     /codon_start=1
                     /product="aminoglycoside 3-phosphotransferase"
                     /label=aminoglycoside 3-phosphotransferase
                     /note="neomycin-resistance gene"
                     /protein_id="BAX73945.1"
                     /translation="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGDDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     regulatory      complement(5606..6105)
                     /regulatory_class="promoter"
                     /note="PGK"
     promoter        complement(5606..6105)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     misc_feature    6115..11231
                     /gene="HPRT"
                     /label=3 arm
                     /note="3 arm"
     gene            6115..11231
                     /gene="HPRT"
                     /label=HPRT
     primer_bind     complement(11227..11243)
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        complement(11280..11298)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(11319..11335)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    11343..11359
                     /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)."
     promoter        complement(11367..11397)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    11412..11433
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(11721..12309)
                     /direction=LEFT
                     /label=pUC origin
                     /note="pUC origin"
     CDS             complement(12480..13340)
                     /codon_start=1
                     /product="beta-lactamase"
                     /label=beta-lactamase
                     /note="ampicillin-resistance gene"
                     /protein_id="BAX73946.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(13341..13445)
                     /gene="bla"
                     /label=AmpR promoter