LOCUS       Exported                5881 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Suicide vector pMRKO1, complete sequence.
ACCESSION   HQ122651
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5881)
  AUTHORS   Ramsey M, Rumbaugh K, Whiteley M.
  TITLE     Aggregatibacter actinomycetemcomitans suicide vector
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5881)
  AUTHORS   Ramsey M, Rumbaugh K, Whiteley M.
  TITLE     Direct Submission
  JOURNAL   Submitted (09-AUG-2010) Molecular Genetics and Microbiology, 
            University of Texas at Austin, 2506 Speedway Blvd, Austin, TX 78712,
            USA
REFERENCE   3  (bases 1 to 5881)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..5881
                     /organism="Suicide vector pMRKO1"
                     /mol_type="other DNA"
                     /db_xref="taxon:945966"
     rep_origin      171..759
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     regulatory      1034..1118
                     /regulatory_class="promoter"
                     /note="pLac"
     protein_bind    1047..1068
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        1083..1113
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1121..1137
                     /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)."
     primer_bind     1145..1161
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             1157..1939
                     /codon_start=1
                     /product="mCherry fluorescent protein"
                     /label=mCherry fluorescent protein
                     /protein_id="ADV78248.1"
                     /translation="MTMITPSLHACRSTLEDPRVPVATMVSKGEEDNMAIIKEFMRFKV
                     HMEGSVNGHEFEIEGEGEGRPYEGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKH
                     PADIPDYLKLSFPEGFKWERVMNFEDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGP
                     VMQKKTMGWEASSERMYPEDGALKGEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAY
                     NVNIKLDITSHNEDYTIVEQYERAEGRHSTGGMDELYK"
     regulatory      1223..1232
                     /regulatory_class="other"
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
     CDS             1229..1939
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMDELYK"
     CDS             2310..2984
                     /codon_start=1
                     /gene="SpcR"
                     /product="spectinomycin resistance protein"
                     /label=SpcR
                     /protein_id="ADV78249.1"
                     /translation="MFGSGVESGLKPNSDLDFLVVVSEPLTDQSKEILIQKIRPISKKI
                     GDKSNLRYIELTIIIQQEMVPWNHPPKQEFIYGEWLQELYEQGYIPQKELNSDLTIMLY
                     QAKRKNKRIYGNYDLEELLPDIPFSDVRRAIMDSSEELIDNYQDDETNSILTLCRMILT
                     MDTGKIIPKDIAGNAVAESSPLEHRERILLAVRSYLGENIEWTNENVNLTINYLNNRLK
                     KL"
     gene            2310..2984
                     /gene="SpcR"
                     /label=SpcR
     rep_origin      3169..3525
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K; 
                     requires the R6K initiator protein pi for replication"
     oriT            4169..4278
                     /note="incP origin of transfer"
     CDS             4311..4682
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     promoter        complement(5750..5854)
                     /gene="bla"
                     /label=AmpR promoter