LOCUS       Exported                6486 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Reporter vector pFila, complete sequence.
ACCESSION   HQ425563
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6486)
  AUTHORS   Bi Y, Zheng X, Shao C, Pan W, Jiang L, Ouyang H.
  TITLE     Construction and application of a built-in dual luciferase reporter 
            for microRNA functional analysis
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6486)
  AUTHORS   Bi Y, Zheng X, Shao C, Pan W, Jiang L, Ouyang H.
  TITLE     Direct Submission
  JOURNAL   Submitted (19-OCT-2010) College of Life Science, Wuhan University, 
            Bayi Road, Wuhan, Hubei 430072, P.R. China
REFERENCE   3  (bases 1 to 6486)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6486
                     /organism="Reporter vector pFila"
                     /mol_type="other DNA"
                     /label=for microRNA target identification and screening
                     /note="for microRNA target identification and screening"
                     /db_xref="taxon:931607"
     regulatory      7..759
                     /regulatory_class="promoter"
                     /note="HSV-TK promoter"
     promoter        8..759
                     /label=HSV TK promoter
                     /note="herpes simplex virus thymidine kinase promoter"
     intron          826..962
                     /note="chimeric intron"
     intron          829..961
                     /label=chimeric intron
                     /note="chimera between introns from human beta-globin and 
                     immunoglobulin heavy chain genes"
     regulatory      1006..1024
                     /regulatory_class="promoter"
                     /note="T7 RNA polymerase promoter"
     promoter        1006..1024
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      1023
                     /regulatory_class="ribosome_binding_site"
                     /note="T7 RNA polymerase transcription initiation site"
     CDS             1034..1969
                     /codon_start=1
                     /product="Renilla luciferase protein"
                     /label=Renilla luciferase protein
                     /note="Rluc"
                     /protein_id="ADR65117.1"
                     /translation="MTSKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     misc_feature    1971..1976
                     /label=XbaI restriction site
                     /note="XbaI restriction site"
     misc_recomb     1995..2000
                     /label=ApaI restriction site
                     /note="ApaI restriction site"
     polyA_signal    2028..2149
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     regulatory      2039..2240
                     /regulatory_class="polyA_signal_sequence"
                     /note="SV40 late polyadenylation signal (upstream)"
     regulatory      2279..2481
                     /regulatory_class="promoter"
                     /note="SV40 promoter"
     promoter        2279..2475
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      2326..2461
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     regulatory      2505..2514
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             2511..4163
                     /codon_start=1
                     /product="firefly luciferase protein"
                     /label=firefly luciferase protein
                     /note="Fluc"
                     /protein_id="ADR65118.1"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     regulatory      4195..4416
                     /regulatory_class="polyA_signal_sequence"
                     /note="SV40 late polyadenylation signal (downstream)"
     polyA_signal    4204..4325
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     promoter        4695..4799
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4800..5660
                     /codon_start=1
                     /product="beta-lactamase"
                     /label=beta-lactamase
                     /note="resistance to ampicillin"
                     /protein_id="ADR65119.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      5802..6445
                     /label=pBR322 plasmid replication origin
                     /note="pBR322 plasmid replication origin"
     rep_origin      5831..6419
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"