LOCUS       Exported               10437 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Gene trap vector pCMT-SAhygpA-NP21 DNA, complete sequence.
ACCESSION   AB609713
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10437)
  AUTHORS   Horie K, Kokubu C, Yoshida J, Akagi K, Isotani A, Oshitani A, Yusa 
            K, Ikeda R, Huang Y, Bradley A, Takeda J.
  TITLE     A homozygous mutant embryonic stem cell bank applicable for 
            phenotype-driven genetic screening
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 10437)
  AUTHORS   Horie K, Takeda J.
  TITLE     Direct Submission
  JOURNAL   Submitted (19-JAN-2011) Contact:Kyoji Horie Osaka University, 
            Department of Social and Environmental Medicine, Graduate School of 
            Medicine; 2-2 Yamadaoka, Suita, Osaka 565-0871, Japan
REFERENCE   3  (bases 1 to 10437)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..10437
                     /organism="Gene trap vector pCMT-SAhygpA-NP21"
                     /mol_type="other DNA"
                     /db_xref="taxon:947494"
     rep_origin      complement(3..458)
                     /direction=LEFT
                     /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"
     regulatory      669..1341
                     /regulatory_class="promoter"
                     /note="human cytomegalovirus promoter"
     enhancer        745..1124
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1125..1328
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     misc_feature    1342..1409
                     /label=R region of Moloney Murine Leukemia Virus
                     /note="R region of Moloney Murine Leukemia Virus"
     misc_feature    1410..1484
                     /label=U5 region of Moloney Murine Leukemia Virus
                     /note="U5 region of Moloney Murine Leukemia Virus"
     misc_feature    1485..2765
                     /label=packaging signal of Moloney Murine Leukemia Virus
                     /note="packaging signal of Moloney Murine Leukemia Virus"
     misc_feature    1549..1906
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus 
                     (MMLV)"
     CDS             1965..2381
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     misc_feature    2391..2765
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     misc_feature    complement(2801..3025)
                     /label=contains bovine growth hormone polyA signal
                     /note="contains bovine growth hormone polyA signal"
     polyA_signal    2801..3025
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     misc_feature    complement(3036..3069)
                     /label=lox2272 site
                     /note="lox2272 site"
     protein_bind    3036..3069
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites."
     CDS             3080..4681
                     /codon_start=1
                     /gene="puro-delta-tk"
                     /product="fusion protein between puromycin 
                     N-acetyltransferase and a truncated version of herpes 
                     simplex virus type 1 thymidine kinase"
                     /label=puro-delta-tk
                     /protein_id="BAL04097.1"
                     /translation="MGTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIE
                     RVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRL
                     AAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLET
                     SAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGAGSMPTLLRVYIDGPHGMGKTTT
                     TQLLVALGSRDDIVYVPEPMTYWRVLGASETIANIYTTQHRLDQGEISAGDAAVVMTSA
                     QITMGMPYAVTDAVLAPHIGGEAGSSHAPPPALTLIFDRHPIAALLCYPAARYLMGSMT
                     PQAVLAFVALIPPTLPGTNIVLGALPEDRHIDRLAKRQRPGERLDLAMLAAIRRVYGLL
                     ANTVRYLQCGGSWREDWGQLSGTAVPPQGAEPQSNAGPRPHIGDTLFTLFRAPELLAPN
                     GDLYNVFAWALDVLAKRLRSMHVFILDYDQSPAGCRDALLQLTSGMVQTHVTTPGSIPT
                     ICDLARTFAREMGEAN"
     gene            3080..4681
                     /gene="puro-delta-tk"
                     /label=puro-delta-tk
     CDS             3080..3679
                     /codon_start=1
                     /gene="pac from Streptomyces alboniger"
                     /product="puromycin N-acetyltransferase"
                     /label=PuroR
                     /note="confers resistance to puromycin"
                     /translation="MGTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIE
                     RVTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRL
                     AAQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLET
                     SAPRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     CDS             3686..4681
                     /codon_start=1
                     /product="truncated herpes simplex virus thymidine kinase 
                     (Salomon et al., 1995)"
                     /label=-Delta-TK
                     /note="confers sensitivity to nucleoside analogs such as 
                     acyclovir or ganciclovir"
                     /translation="MPTLLRVYIDGPHGMGKTTTTQLLVALGSRDDIVYVPEPMTYWRV
                     LGASETIANIYTTQHRLDQGEISAGDAAVVMTSAQITMGMPYAVTDAVLAPHIGGEAGS
                     SHAPPPALTLIFDRHPIAALLCYPAARYLMGSMTPQAVLAFVALIPPTLPGTNIVLGAL
                     PEDRHIDRLAKRQRPGERLDLAMLAAIRRVYGLLANTVRYLQCGGSWREDWGQLSGTAV
                     PPQGAEPQSNAGPRPHIGDTLFTLFRAPELLAPNGDLYNVFAWALDVLAKRLRSMHVFI
                     LDYDQSPAGCRDALLQLTSGMVQTHVTTPGSIPTICDLARTFAREMGEAN"
     CDS             complement(4682..5485)
                     /codon_start=1
                     /gene="neo"
                     /product="neomycin phosphotransferase"
                     /label=neo
                     /protein_id="BAL04098.1"
                     /translation="MGSAIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQ
                     GRPVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQD
                     LLSSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDE
                     EHQGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQ
                     DIALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     gene            complement(4682..5485)
                     /gene="neo"
                     /label=neo
     regulatory      5482..5491
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     misc_feature    5502..5535
                     /label=lox2272 site
                     /note="lox2272 site"
     protein_bind    complement(5502..5535)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GGATACTT). lox2272 sites are compatible with each
                     other, but incompatible with loxP or loxN sites."
     regulatory      complement(5536..6047)
                     /regulatory_class="promoter"
                     /note="phosphoglycerate kinase 1 promoter"
     promoter        complement(5540..6038)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     misc_feature    complement(6065..6275)
                     /label=contains phosphoglycerate kinase 1 polyA signal
                     /note="contains phosphoglycerate kinase 1 polyA signal"
     CDS             complement(6279..7304)
                     /codon_start=1
                     /gene="hph"
                     /product="hygromycin phosphotransferase"
                     /label=hph
                     /protein_id="BAL04099.1"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     gene            complement(6279..7304)
                     /gene="hph"
                     /label=hph
     misc_feature    complement(7337..7440)
                     /label=Adenovirus splice acceptor
                     /note="Adenovirus splice acceptor"
     misc_feature    7620..8048
                     /note="U3 region with an enhancer deletion of Moloney 
                     Murine Leukemia Virus"
     protein_bind    complement(7817..7850)
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     misc_feature    8050..8117
                     /label=R region of Moloney Murine Leukemia Virus
                     /note="R region of Moloney Murine Leukemia Virus"
     misc_feature    8118..8190
                     /label=U5 region of Moloney Murine Leukemia Virus
                     /note="U5 region of Moloney Murine Leukemia Virus"
     promoter        complement(8249..8267)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(8288..8304)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    8312..8328
                     /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(8336..8366)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    8381..8402
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(8690..9278)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(9449..10309)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=bla
                     /protein_id="BAL04100.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            complement(9449..10309)
                     /gene="bla"
                     /label=bla
     promoter        complement(10310..10414)
                     /gene="bla"
                     /label=AmpR promoter