LOCUS       Exported                8438 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  ShRNA expression vector pSiRPG, complete sequence.
ACCESSION   DQ465352
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8438)
  AUTHORS   Storvold GL, Gjernes E, Askautrud HA, Borresen-Dale AL, Perou CM, 
            Frengen E.
  TITLE     A retroviral vector for siRNA expression in mammalian cells
  JOURNAL   Mol. Biotechnol. 35 (3), 275-282 (2007)
  PUBMED    17652791
REFERENCE   2  (bases 1 to 8438)
  AUTHORS   Stoervold GL.
  TITLE     Direct Submission
  JOURNAL   Submitted (28-MAR-2006) Institute for Medical Genetics, University 
            of Oslo, Norway, PO Box 1036 Blindern, Oslo N-0315, Norway
REFERENCE   3  (bases 1 to 8438)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..8438
                     /organism="shRNA expression vector pSiRPG"
                     /mol_type="other DNA"
                     /note="sequence was created by fusing the sequence of 
                     pSUPERIOR.retro.puro, pSUPERIOR.retro.neo+gfp (OligoEngine,
                     Inc.) and pBabe Puro."
                     /db_xref="taxon:380637"
     misc_feature    1..6711
                     /label=from pSUPERIOR.retro.puro
                     /note="from pSUPERIOR.retro.puro"
     regulatory      5..226
                     /regulatory_class="promoter"
                     /note="H1 promoter"
     protein_bind    211..229
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_feature    complement(233..1209)
                     /label=stuffer sequence
                     /note="stuffer sequence"
     enhancer        450..829
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        830..1033
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     promoter        1078..1096
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(1218..1234)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(1253..1669)
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag 
                     gene lacking the start codon"
     misc_feature    1736..2077
                     /label=MESV Psi
                     /note="packaging signal of murine embryonic stem cell 
                     virus"
     LTR             2141..2657
                     /label=5 LTR
                     /note="5 long terminal repeat from murine embryonic stem 
                     cell virus"
     repeat_region   2143..2658
                     /label=5 LTR
                     /note="5 LTR"
     promoter        3485..3589
                     /gene="bla"
                     /label=AmpR promoter
     gene            3587..4450
                     /gene="Amp"
                     /label=Amp
     CDS             3590..4450
                     /codon_start=1
                     /gene="Amp"
                     /product="beta-lactamase"
                     /label=Amp
                     /note="ampicillin resistance"
                     /protein_id="ABE96682.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      complement(4595..5268)
                     /direction=LEFT
                     /label=pUC Ori
                     /note="pUC Ori"
     rep_origin      4621..5209
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    5497..5518
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        5533..5563
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5571..5587
                     /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)."
     repeat_region   5750..6103
                     /label=3 LTR
                     /note="3 LTR"
     CDS             complement(6173..6772)
                     /codon_start=1
                     /gene="Puro"
                     /product="puromycin N-acetyl transferase"
                     /label=Puro
                     /note="puromycin resistance"
                     /protein_id="ABE96683.1"
                     /translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
                     VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
                     AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
                     APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
     gene            complement(6173..6772)
                     /gene="Puro"
                     /label=Puro
     misc_feature    6712..7124
                     /label=from pBabe Puro
                     /note="from pBabe Puro"
     regulatory      complement(6777..7119)
                     /regulatory_class="promoter"
                     /note="SV40 early promoter"
     promoter        complement(6782..7111)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6796..6931
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     misc_feature    7125..8009
                     /label=from pSUPERIOR.retro.neo+GFP
                     /note="from pSUPERIOR.retro.neo+GFP"
     CDS             complement(7167..7904)
                     /codon_start=1
                     /gene="EGPF"
                     /product="enhanced green flurorescent protein"
                     /label=EGPF
                     /protein_id="ABE96684.1"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYKSGLRSA"
     gene            complement(7167..7904)
                     /gene="EGPF"
                     /label=EGPF
     CDS             complement(7188..7904)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     regulatory      7901..7910
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     regulatory      complement(7928..8326)
                     /regulatory_class="promoter"
                     /note="PGK promoter"
     promoter        complement(7939..8438)
                     /label=PGK promoter
                     /note="mouse phosphoglycerate kinase 1 promoter"
     misc_feature    8010..8438
                     /label=from pSUPERIOR.retro.puro
                     /note="from pSUPERIOR.retro.puro"