LOCUS       Exported                6737 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pNK076, complete sequence.
ACCESSION   KY412814
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6737)
  AUTHORS   Kandul NP, Guo M, Hay BA.
  TITLE     A positive readout single transcript reporter for site-specific mRNA
            cleavage
  JOURNAL   PeerJ (2017) In press
REFERENCE   2  (bases 1 to 6737)
  AUTHORS   Kandul NP, Guo M, Hay BA.
  TITLE     Direct Submission
  JOURNAL   Submitted (02-JAN-2017) Biology and Biological Engineering, 
            California Institute of Technology, 1200 E. California Blvd., 
            Pasadena, CA 91125, USA
REFERENCE   3  (bases 1 to 6737)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..6737
                     /organism="Cloning vector pNK076"
                     /mol_type="other DNA"
                     /db_xref="taxon:2020921"
     rep_origin      4..460
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     601..617
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        627..645
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      688..2531
                     /regulatory_class="promoter"
     promoter        699..3208
                     /label=Ac5 promoter
                     /note="Drosophila melanogaster actin 5C promoter"
     CDS             3236..4171
                     /codon_start=1
                     /product="RLuc"
                     /label=RLuc
                     /note="renila luciferase"
                     /protein_id="ASL70199.1"
                     /translation="MTSKVYDPEQRKRMITGPQWWARCKQMNVLDSFINYYDSEKHAEN
                     AVIFLHGNAASSYLWRHVVPHIEPVARCIIPDLIGMGKSGKSGNGSYRLLDHYKYLTAW
                     FELLNLPKKIIFVGHDWGACLAFHYSYEHQDKIKAIVHAESVVDVIESWDEWPDIEEDI
                     ALIKSEEGEKMVLENNFFVETMLPSKIMRKLEPEEFAAYLEPFKEKGEVRRPTLSWPRE
                     IPLVKGGKPDVVQIVRNYNAYLRASDDLPKMFIESDPGFFSNAIVEGAKKFPNTEFVKV
                     KGLHFSQEDAPDEMGKYIKSFVERVLKNEQ"
     promoter        complement(4549..4567)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(4588..4604)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    4612..4628
                     /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(4636..4666)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4681..4702
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(4990..5578)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5749..6609)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     promoter        complement(6610..6714)
                     /gene="bla"
                     /label=AmpR promoter