LOCUS       Exported                8196 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pAP-ACN, complete sequence.
ACCESSION   EU530628
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8196)
  AUTHORS   Wu S, Ying G, Wu Q, Capecchi MR.
  TITLE     A protocol for constructing gene targeting vectors: generating 
            knockout mice for the cadherin family and beyond
  JOURNAL   Nat Protoc 3 (6), 1056-1076 (2008)
  PUBMED    18546598
REFERENCE   2  (bases 1 to 8196)
  AUTHORS   Wu S, Capecchi MR.
  TITLE     Direct Submission
  JOURNAL   Submitted (29-FEB-2008) Howard Hughes Medical Institute, University 
            of Utah, 15N 2030E, Salt Lake City, UT 84112, USA
REFERENCE   3  (bases 1 to 8196)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..8196
                     /organism="Cloning vector pAP-ACN"
                     /mol_type="other DNA"
                     /db_xref="taxon:512274"
     promoter        34..52
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             69..1676
                     /codon_start=1
                     /gene="AP"
                     /product="alkaline phosphatase"
                     /label=AP
                     /note="human placental alkaline phosphatase; designed as 
                     fusion protein; translational start site to be provided by 
                     upstream fusion"
                     /protein_id="ACC91779.1"
                     /translation="ARLQDLLLLLLLLGLRLQLSLGIIPVEEENPDFWNREAAEALGAA
                     KKLQPAQTAAKNLIIFLGDGMGVSTVTAARILKGQKKDKLGPEIPLAMDRFPYVALSKT
                     YNVDKHVPDSGATATAYLCGVKGNFQTIGLSAAARFNQCNTTRGNEVISVMNRAKKAGK
                     SVGVVTTTRVQHASPAGTYAHTVNRNWYSDADVPASARQEGCQDIATQLISNMDIDVIL
                     GGGRKYMFRMGTPDPEYPDDYSQGGTRLDGKNLVQEWLAKRQGARYVWNRTELMQASLD
                     PSVTHLMGLFEPGDMKYEIHRDSTLDPSLMEMTEAALRLLSRNPRGFFLFVEGGRIDHG
                     HHESRAYRALTETIMFDDAIERAGQLTSEEDTLSLVTADHSHVFSFGGYPLRGSSIFGL
                     APGKARDRKAYTVLLYGNGPGYVLKDGARPDVTESESGSPEYRQQSAVPLDEETHAGED
                     VAVFARGPQAHLVHGVQEQTFIAHVMAFAACLEPYTACDLAPPAGTTDAAHPGRSVVPA
                     LLPLLAGTLLLLETATAP"
     gene            69..1676
                     /gene="AP"
                     /label=AP
     CDS             87..1586
                     /codon_start=1
                     /product="secreted alkaline phosphatase from human 
                     placenta"
                     /label=SEAP
                     /note="The natural enzyme has a C-terminal transmembrane 
                     sequence that is replaced by a GPI anchor."
                     /translation="LLLLLLLGLRLQLSLGIIPVEEENPDFWNREAAEALGAAKKLQPA
                     QTAAKNLIIFLGDGMGVSTVTAARILKGQKKDKLGPEIPLAMDRFPYVALSKTYNVDKH
                     VPDSGATATAYLCGVKGNFQTIGLSAAARFNQCNTTRGNEVISVMNRAKKAGKSVGVVT
                     TTRVQHASPAGTYAHTVNRNWYSDADVPASARQEGCQDIATQLISNMDIDVILGGGRKY
                     MFRMGTPDPEYPDDYSQGGTRLDGKNLVQEWLAKRQGARYVWNRTELMQASLDPSVTHL
                     MGLFEPGDMKYEIHRDSTLDPSLMEMTEAALRLLSRNPRGFFLFVEGGRIDHGHHESRA
                     YRALTETIMFDDAIERAGQLTSEEDTLSLVTADHSHVFSFGGYPLRGSSIFGLAPGKAR
                     DRKAYTVLLYGNGPGYVLKDGARPDVTESESGSPEYRQQSAVPLDEETHAGEDVAVFAR
                     GPQAHLVHGVQEQTFIAHVMAFAACLEPYTACDLAPPAGTTD"
     polyA_site      1705..2099
                     /label=-beta-globin poly(A)
                     /note="human beta-globin polyadenylation signal"
     misc_recomb     2111..2144
                     /label=loxP site
                     /note="loxP site"
     protein_bind    complement(2111..2144)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     polyA_signal    2171..2219
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     regulatory      complement(2171..2218)
                     /regulatory_class="polyA_signal_sequence"
                     /note="HSV thymidine kinase polyA signal"
     CDS             complement(2226..3029)
                     /codon_start=1
                     /gene="neo"
                     /product="neomycin resistance protein"
                     /label=neo
                     /protein_id="ACC91780.1"
                     /translation="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     gene            complement(2226..3029)
                     /gene="neo"
                     /label=neo
     regulatory      complement(3046..3752)
                     /regulatory_class="promoter"
                     /note="RNA polymerase II large subunit promoter"
     polyA_signal    3768..3816
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     regulatory      3768..3815
                     /regulatory_class="polyA_signal_sequence"
                     /note="HSV thymidine kinase polyA signal"
     CDS             complement(join(3962..4165,4232..5080))
                     /codon_start=1
                     /gene="cre"
                     /product="Cre recombinase"
                     /label=cre
                     /protein_id="ACC91781.1"
                     /translation="MPKKKRKVSNLLTVHQNLPALPVDATSDEVRKNLMDMFRDRQAFS
                     EHTWKMLLSVCRSWAAWCKLNNRKWFPAEPEDVRDYLLYLQARGLAVKTIQQHLGQLNM
                     LHRRSGLPRPSDSNAVSLVMRRIRKENVDAGERAKQALAFERTDFDQVRSLMENSDRCQ
                     DIRNLAFLGIAYNTLLRIAEIARIRVKDISRTDGGRMLIHIGRTKTLVSTAGVEKALSL
                     GVTKLVERWISVSGVADDPNNYLFCRVRKNGVAAPSATSQLSTRALEGIFEATHRLIYG
                     AKDDSGQRYLAWSGHSARVGAARDMARAGVSIPEIMQAGGWTNVNIVMNYIRNLDSETG
                     AMVRLLEDGD"
     gene            complement(3962..5080)
                     /gene="cre"
                     /label=cre
     intron          complement(4166..4231)
                     /note="SV40 T-antigen intron"
     CDS             complement(5057..5077)
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     regulatory      complement(5081..5779)
                     /regulatory_class="promoter"
                     /note="testis-specific angiotensin-converting enzyme 
                     promoter (tACE)"
     misc_recomb     5787..5820
                     /label=loxP site
                     /note="loxP site"
     protein_bind    complement(5787..5820)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."
     promoter        complement(5884..5902)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     promoter        6248..6352
                     /gene="bla"
                     /label=AmpR promoter
     CDS             6353..7213
                     /codon_start=1
                     /gene="bla"
                     /product="ampicillin resistance protein"
                     /label=bla
                     /protein_id="ACC91778.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            6353..7213
                     /gene="bla"
                     /label=bla
     rep_origin      7384..7972
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"